DETAILED CORRESPONDENCE
Status of the Application
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Claims 1-30, 32, and 34-40 are pending.
Election/Restrictions
Applicant’s election without traverse of
Invention IA, corresponding to linking claim 1 and claims 2-30 and 36-38, drawn to a method for identifying, quantifying, and/or characterizing a protein of interest, comprising: (a) contacting a sample including a protein of interest to a solid surface to form an immobilized protein of interest; (b) eluting said immobilized protein of interest or a fragment thereof to form an enriched protein of interest or fragment thereof; (c) subjecting said enriched protein of interest or fragment thereof to size exclusion chromatography under native conditions to form a size exclusion chromatography eluate; (d) contacting said size exclusion chromatography eluate to a denaturing solution to form a denatured eluate; and (e) subjecting said denatured eluate to mass spectrometry analysis to identify, quantify, and/or characterize said protein of interest, and a method for producing a pharmacokinetic profile of a protein of interest, comprising: (a) quantifying a concentration of said protein of interest at a first time point after administration of said protein of interest to a subject by: (i) obtaining a sample from said subject including said protein of interest at a first time point after administration of said protein of interest to said subject; and (ii) quantifying said protein of interest according to the method of claim 1; and (b) repeating step (a) for at least one additional time point to produce a pharmacokinetic profile of said protein of interest.
in the reply filed 06/09/2026 is acknowledged.
Claims 32, 34-35 and 39-40 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on 06/09/2026.
Claims 1-30 and 36-38 are being examined on the merits.
Priority
The instant application is a CIP of 17/863,332 filed 07/12/2022 which claims domestic priority to U.S. Provisional Application No. 63/221,439 filed 07/13/2021.
Information Disclosure Statement
The Information Disclosure Statement (IDS) submitted on 08/11/2023 is in compliance with the provisions of 37 CFR 1.97, as the documents cited on the IDS are submitted with the parent application 17/863,332. Accordingly, the IDS has been considered by the examiner.
Objections to Specification
The disclosure is objected to because of the following informalities.
The use of the terms BIO-REX, MACRO-PREP, DOWEX, PARTISPHERE, HYPERD, AMBERLITE, SEPHAROSE, BAKERBOND, CAPTO, TOYOPEARL, BIO-GEL, BIOPRO, BIORESOLVE, ACQUITY, and Q EXACTIVE, which are trade names or marks used in commerce, have been noted in this application in paragraphs 0148-0149, 0153, 0180, 0182, 0192. The terms should be accompanied by the generic terminology; furthermore the terms should be capitalized wherever they appears or, where appropriate, include a proper symbol indicating use in commerce such as ™, SM , or ® following the terms.
Although the use of trade names and marks used in commerce (i.e., trademarks, service marks, certification marks, and collective marks) are permissible in patent applications, the proprietary nature of the marks should be respected and every effort made to prevent their use in any manner which might adversely affect their validity as commercial marks.
Appropriate correction is required.
Claim Objections
Claim 15 is objected to for the phrase “wherein the fragment is selected from the group consisting of a Fab fragment, a Fab’ fragment, a Fab2 fragment, a F(ab’)2 fragment, a Fv fragment, a Fd’ fragment, and a Fd fragment”. In the interest of improving claim form, Applicant should consider an amendment to recite “wherein the fragment is an antibody fragment selected from the group consisting of a Fab fragment, a Fab’ fragment, a Fab2 fragment, a F(ab’)2 fragment, a Fv fragment, a Fd’ fragment, and a Fd fragment”.
Claim Rejections - 35 USC § 112(b)
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
Claims 18 and 22 are rejected under 35 U.S.C. 112(b) as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor regards as the invention.
Claim 18 is indefinite for the phrase “about 150 mM ammonium acetate”, as “about” is a relative term according to MPEP 2173.05(b). The term "about" is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. As such, the limitation of the amount of ammonium acetate is rendered indefinite by the use of the term.
Claim 22 is indefinite for the phrase “about 60% acetonitrile and about 4% formic acid”, as “about” is a relative term according to MPEP 2173.05(b). The term "about" is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. As such, the limitations of the amounts of acetonitrile and formic acid are rendered indefinite by the use of the term.
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claims 1-12, 16-21 and 24-26 are rejected under 35 U.S.C. 103 as being unpatentable over Li et al. (Analytic Chem, 2012, 83:1267; cited on the IDS submitted 08/11/2023; herein Li) in view of Haberger et al. (mAbs, 2016, 8:331; cited on the attached Form PTO-892; herein Haberger).
Claim 1 (claims 2-12, 16-21 and 24-26 dependent therefrom) is drawn to a method for identifying, quantifying, and/or characterizing a protein of interest, comprising:
(i) contacting a sample including a protein of interest to a solid surface to form an immobilized protein of interest;
(ii) eluting the immobilized protein of interest of a fragment thereof to form an enriched protein of interest of fragment thereof;
(iii) subjecting the enriched protein of interest or a fragment thereof to size exclusion chromatography (SEC) under native conditions to form an SEC eluate;
(iv) contacting the SEC eluate to a denaturing solution to form a denatured eluate; and
(v) subjecting the denatured eluate to mass spectrometry (MS) analysis to identify, quantify, and/or characterize the protein of interest.
The term “native conditions” recited in the claim is not specifically defined in instant specification. The term “native” is discussed at [para 0159] in reference to examples of native MS and native folded states of biological analytes, but not in the context of native conditions for SEC as the term is used in step (iii) as claim 1. Therefore the term “native conditions” is being interpreted as conditions without a denaturant.
Li relates to general LC-MS/MS to quantify therapeutic monoclonal antibodies (mAbs) [title].
Regarding claim 1, Li teaches a method of quantifying mAbs using tandem liquid chromatographic mass spectrometry (LC-MS/MS) [abstract] and that LC-MS/MS methods can be applied to mAbs for the bioanalysis of preclinical samples, wherein mAbs are enzymatically digested to generate a signature peptide for MS quantification [p 1267, col 1, para 2]. Li further teaches multidimensional chromatography can be used to separate mAb signature peptides from the peptides of the endogenous matrix at the expense of time, and that the use of immunocapture to clean up and concentrate a sample overcome such drawbacks and enhance sensitivity [p 1267, col 1, para 2].
Specifically, Li teaches the addition of the biotinylated human Fc mAb clone 35 (b-Ab35) to streptavidin magnetic beads to produce b-Ab35-coated magnetic beads, followed by the addition of a sample containing alpha-DA-G2, wherein the sample and beads were incubated for 1 h at ambient temperature, washed with 0.01% Tween 20 in PBS, and eluted with a methanol/formic acid mixture, and the resulting supernatant was denatured in the presence of a buffer comprising 8M urea and 20 mM DTT before being subjected to LC-MS/MS analysis [p 1268, col 1, final paragraph; col 2, final paragraph to p 1269, col 1, para 1]. These teachings of Li correspond to steps (i), (ii) and (v), and the subjecting of eluate to a denaturing solution of step (iv) of claim 1.
Li does not teach the SEC limitations of steps (iii) and (iv).
Haberger relates to characterization of biotherapeutic proteins by SEC coupled to native MS [title], and discusses that SEC is the method of choice for routine product testing to detect and quantify high-molecular aggregates and to determine the content of low molecular forms such as fragments, but does not enable determination of molecular mass of an analyte [p 331, col 1, beginning at final paragraph].
Regarding claim 1, Haberger teaches a method of analyzing CrossMAb samples that are in-house bispecific antibodies [p 331, col 2, final paragraph] comprising SEC of CrossMAb samples with a mobile phase of KH2PO4 and KCl and pH 7 [p 338, col 2, para 2], followed by analysis of the eluate via ESI-UV/MS [Figure 5], wherein the SEC protocol was able to resolve the CrossMAb size variants [p 336, col 1, final paragraph, and Figure 5]. The teachings of Haberger are considered to correspond to subjecting a sample to SEC under native conditions to form an SEC eluate as recited in part (iii) of claim 1.
It would have been prima facie obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to combine Li and Haberger to modify the method of Li by using SEC on the sample before MS analysis, as taught by Haberger, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the method of Li because Haberger teaches SEC can detect and quantify high-molecular aggregates and to determine the content of low molecular forms such as fragments, and Haberger teaches a method resolves size variants of mAbs before MS analysis. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Haberger relate to methods of analyzing mAbs via MS.
Regarding claim 2, Li teaches the method to analyze alpha-DA-G2 as described above, which is an isotype of the human antidinitrophenol IgG2 mAb [p 1268, col 1, para 2], which is considered to correspond to a drug product.
Regarding claim 3, Li teaches the method was carried out on alpha-DA-G2 prepared in cynomolgus monkey serum [p 1271, col 2, final paragraph].
Regarding claim 4-5, Li teaches the method carried out on the mAb alpha-DA-G2 as discussed above [p 1268, col 1, final paragraph; col 2, final paragraph to p 1269, col 1, para 1], which is an antibody. As the instant specification does not specifically define a therapeutic antibody, but states in [para 0140] “an anti-human Fc antibody will preferentially bind to the Fc domain of any human antibody, such as, for example, a therapeutic antibody”, and the method of Li uses a biotinylated human Fc mAb clone 35 (b-Ab35) to pull down and analyze the mAb alpha-DA-G2, the alpha-DA-G2 of Li is considered to correspond to a therapeutic antibody.
Regarding claims 6-9, the solid surface of the method of Li consists of streptavidin magnetic beads coated with biotinylated human Fc mAb clone 35 (b-Ab35) as discussed in the rejection of claim 1 above.
Regarding claim 10, the method of Li comprises a washing step as discussed in the rejection of claim 1 above.
Regarding claims 11-12, Li teaches LC-MS/MS methods can be applied to mAbs for the bioanalysis of preclinical samples, wherein mAbs are enzymatically digested, usually with trypsin, to generate a signature peptide for MS quantification [p 1267, col 1, para 2].
Regarding claim 16, Haberger teaches the use of an SEC system directly coupled to a mass spectrometer [p 338, col 1, para 3].
Regarding claims 17-18, Haberger teaches the use of potassium phosphate as a the SEC mobile phase as discussed above in the rejection of claim 1, and also teaches the use of 25-100 mM ammonium acetate buffer as the SEC mobile phase [p 333, col 2, para 1], and MPEP 2144.05.I states in the case where the claimed ranges overlap or lie inside ranges disclosed by the prior art, a prima facie case of obviousness exists.
Regarding claims 19-20, Li teaches the analysis of captured fragments via LC-MS/MS with a mobile phase of 0.1% formic acid in acetonitrile/water, wherein the ratios of acetonitrile/water in mobile phases A and B are 5/95 and 95/5 respectively, and a gradient profile was carried out from 0% to 95% mobile phase B over time [p 1268, col 2, para 3], which is considered to correspond to a denaturing solution from 40% to 80% acetonitrile.
Regarding claim 21, Li teaches the use of 3% formic acid on samples to elute from the magnetic beads followed by the transfer of eluate in supernatant to a well plate to dry [p 1269, col 1, para 1]. As the supernatant is understood to contain the eluting solution of 3% formic acid and the eluate, the teachings of Li are considered to correspond to contacting eluate with a denaturing solution comprising 3% formic acid.
Regarding claim 24, Haberger teaches an SEC flow rate of 0.2 ml/min [p 338, col 1, para 3].
Regarding claim 25, Haberger teaches an SEC flow rate of 0.2 ml/min [p 338, col 1, para 3] that is coupled directly to MS analysis while Li teaches MS analysis with a mobile phase of 0.1% formic acid in acetonitrile/water that is considered a denaturing solution with a flow rate of 0.6 ml/min [p 1268, col 2, para 3]. It would be obvious to one of ordinary skill in the art in practicing the combined method of Haberger and Li to match the flow rates of the coupled protocols of SEC and MS analysis and maintain a mobile phase of 0.1% formic acid in acetonitrile/water with a flowrate of 0.2 ml/min, as MPEP 2144.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation.
Regarding claim 26, Haberger teaches the use of an electrospray ionization mass spectrometer (ESI/MS) [p 338, col 1, para 3].
Therefore, the invention of claims 1-12, 16-21 and 24-26 would have been obvious to one of ordinary skill in the art before the effective filing date.
Claims 13 and 15 are rejected under 35 U.S.C. 103 as being unpatentable over Li in view of Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Pawel-Rammingen et al. (Curr Opin Microbiol, 2003, 6:50; cited on the attached Form PTO-892; herein PR).
Claim 13 is drawn to the method of claim 12, wherein the at least one digestive enzyme comprises IdeS or a variant thereof.
Claim 15 is drawn to the method of claim 1, wherein the fragment is selected from the group consisting of a Fab fragment, a Fab’ fragment, a Fab2 fragment, a F(ab’)2 fragment, a Fv fragment, and a Fd fragment.
The teachings of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. These references do not teach the enzyme IdeS or a variant thereof.
PR relates to IdeS as an immunoglobulin-degrading cysteine proteinase of Streptococcus pyogenes [title].
Regarding claim 13, PR teaches S. pyogenes expresses proteolytic activity towards IgG via the activity of the cysteine proteinase enzyme IdeS, which cleaves the heavy chain of IgG to form two stable Fab fragments and one Fc fragment [p 52, col 2, paras 2-3].
In view of PR, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date to modify the combined method of Li and Haberger by using IdeS, since the simple substitution of one known element for another results in a predictable result. One of ordinary skill in the art would have recognized that both trypsin and IdeS are digestive enzymes, and as such both are capable of being incorporated into such methods compositions as described by Li. Thus it would have been obvious to one of ordinary skill in the art to replace trypsin with IdeS, as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because Li describes a method of capturing and analyzing mAbs using the digestive enzyme trypsin, and PR describes a digestive enzyme specific for IgGs.
Regarding claim 15, as the combined method of Li, Haberger and PR uses the IdeS enzyme to digest an immobilized mAb as described above, and PR teaches the activity of IdeS produces Fab and Fc fragments [p 52, col 2, para 3], the combined teachings of Li, Haberger and PR are considered to satisfy the limitations of claim 15.
Therefore, the invention of claims 13 and 15 would have been obvious to one of ordinary skill in the art before the effective filing date.
Claim 14 is rejected under 35 U.S.C. 103 as being unpatentable over Li in view of Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Rosli et al. (Methods Mol Biol, 2008, 418:89; cited on the attached Form PTO-892; herein Rosli).
Claim 14 is drawn to the method of claim 1, wherein eluting the immobilized protein of interest or fragment thereof comprises subjecting the solid surface to centrifugation.
The teachings of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. These references do not teach the use of centrifugation during elution.
Rosli relates to the recovery of biotinylated proteins from streptavidin-based affinity chromatography resins [title], and discusses the release of biotinylated proteins from streptavidin based reagents is a major problem due to the stability of the complex [abstract].
Regarding claim 14, Rosli teaches a protocol for the release of biotinylated proteins from streptavidin Sepharose comprising the use of a centrifuge [p 91, para 2 and p 94, Section 3.3].
In view of Rosli, it would have been prima facie obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of Li and Haberger by using centrifugation in the elution step, as taught by Rosli, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of Li and Haberger because Rosli teaches the release of biotinylated proteins from streptavidin is a major problem in streptavidin affinity methods and therefore teaches a quantitative release strategy using centrifugation. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Rosli relate to methods comprising streptavidin affinity capture and release.
Therefore, the invention of claim 14 would have been obvious to one of ordinary skill in the art before the effective filing date.
Claim 22 is rejected under 35 U.S.C. 103 as being unpatentable over Li in view of Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Donnelly et al. (Nat Method, 2019, 16:587; cited on the attached Form PTO-892; herein Donnelly).
Claim 22 is drawn to the method of claim 1, wherein the denaturing solution comprises about 60% acetonitrile and about 4% formic acid.
The teachings of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. These references do not teach a denaturing solution comprising about 60% acetonitrile and about 4% formic acid.
Donnelly reviews best practices and benchmarks for intact protein analysis for top-down MS [title], and discusses robust protocols for mass analysis of intact proteins such as antibodies from mixtures of varying complexity [abstract].
Regarding claim 22, Donnelly teaches denaturing direct-infusion MS comprising the treatment of samples to 49.95% acetonitrile, 49.95% water and 0.1% formic acid, wherein a 60:35:5 ratio of HPLC grade methanol:water:acetic acid may be used as an alternative resulting in improved signal to noise ratios, and discusses how the use of organic solvents and acids results in efficient ionization [p 591, col 1, para 3]. Therefore Donnelly teaches the result effective variable of organic solvent and acid ratio according to MPEP 2144.05.II.B which effects signal to noise ratio in MS, and thereby quality of the MS analysis. While Donnelly does not teach the 60% acetonitrile and 4% formic acid recited in the claim, MPEP 2144.05.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. Therefore one of ordinary skill in the art would be motivated to modify the combined method of Li, Haberger and Donnelly to get to the claimed 60% acetonitrile and 4% formic acid because Donnelly teaches the result effective variable of organic solvent and acid ratios, and gives an example of using an increased amount of organic solvent and acid can result in improved signal to noise ratio and therefore improved MS analysis.
In view of Donnelly, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date to modify the combined method of Li and Haberger by using a denaturing solution of 60% acetonitrile and 4% formic acid because MPEP 2144.05.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. One of ordinary skill in the art would have been motivated to modify the combined method of Li and Haberger because Donnelly teaches organic solvent and acid ratios are a result effective variable according to MPEP 2144.05.II.B which effects signal to noise ratio in MS, and thereby quality of the MS analysis. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Donnelly relate to methods of MS analysis comprising treatment of protein samples with acetonitrile and formic acid.
Therefore, the invention of claim 22 would have been obvious to one of ordinary skill in the art before the effective filing date.
Claim 23 is rejected under 35 U.S.C. 103 as being unpatentable over Li in view of Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Wong et al. (Sensors Actuators B, 2004, 100:359; cited on the attached Form PTO-892; herein Wong).
Claim 23 is drawn to the method of claim 1, wherein the denaturing solution is contacted to the SEC eluate using a T-mixer.
The teachings of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. These references do not teach the use of a T-mixer.
Wong relates to a T-mixer as a rapid mixing micromixer [title], and discusses that micromixers are essential components to microfluidic systems because some biochemical analysis require two reagents to be completely mixed before a reaction has proceeded considerably, and can require sub-millisecond mixing times, and therefore micromixers must be capable of mixing two reagents within the shortest time possible [p 359, col 1, paras 1-2 and col 2, paras 1-2].
Regarding claim 23, Wong teaches a T-mixer that can sufficiently cause complete mixing of two solutions within less than a millisecond after the two solutions make contact [abstract].
In view of Wong, it would have been prima facie obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of Li and Haberger by a T-mixer to contact the SEC eluate with denaturing solution, as taught by Wong, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of Li and Haberger because Wong teaches micromixers such as T-mixers are essential components of microfluidic systems in order to insure reagents are completely mixed in sub-millisecond times. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Wong both relate to methods requiring the efficient mixing of biochemical reagents for downstream analysis of the product.
Therefore, the invention of claim 23 would have been obvious to one of ordinary skill in the art before the effective filing date.
Claims 27-30 are rejected under 35 U.S.C. 103 as being unpatentable over Li in view of Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Ehkirch et al. (Anal Chem, 2018, 90:1578; cited on the attached Form PTO-892; herein Ehkirch).
Claim 27 (claims 28-30 dependent therefrom) is drawn to the method of claim 16, wherein a flow splitter is used to couple the SEC system with the MS and a detector.
The teachings of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. These references do not teach the use of a flow splitter.
Ehkirch relates to an online four-dimensional HIC x SEC-IM x MS methodology for characterization of antibody drug conjugates [title] and describes an analytical system that combines multiple chromatography separation methods with MS analysis for a comprehensive and streamlined characterization of native and forced degraded sample conformations [abstract].
Regarding claim 27, Ehkirch teaches the multidimensional analysis method above comprising a first chromatographic dimension, a second chromatographic dimension which is SEC, and a flow splitter connecting SEC to MS which divides the flow rate by a factor of 7 prior to entering MS [p 1581, col 1, paras 2-3; Figure 2].
In view of Ehkirch, it would have been prima facie obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of Li and Haberger using a flow splitter between the SEC and MS, as taught by Ehkirch, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of Li and Haberger because Ehkirch teaches a multidimensional chromatography-MS analytical method for antibody drug conjugate characterization comprising a flow splitter that provides streamlined characterization of native and forced degraded sample conformations. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Ehkirch relate to methods of analyzing samples comprising antibodies via MS.
Regarding claim 28, Haberger teaches the use of a detector to observe absorbance at 280 nm [Figure 1], which is considered to correspond to a UV detector.
Regarding claim 29, Ehkirch teaches the use of a flow splitter to divide the flow rate by a factor of 7 before entering MS but downstream of a detector [p 1581, col 1, paras 2-3; Figure 2], therefore indicating the flow to the MS to be a low flow relative to the high flow in the detector.
Regarding claim 30, Ehkirch teaches reducing the flow rate by a factor of 7 from an inlet flow rate of 100 µL/min [p 1581, col 1, para 3], which is considered to correspond to a low flow of less than 20 µL/min.
Therefore, the invention of claims 27-30 would have been obvious to one of ordinary skill in the art before the effective filing date.
Claims 36-38 are rejected under 35 U.S.C. 103 as being unpatentable over Li in view of Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Schaller et al. (J Proteome Res, 2019, 18:3032; cited on the attached Form PTO-892; herein Schaller).
Claim 36 (claims 37-38 dependent therefrom) is drawn to a method for producing a pharmacokinetic profile of a protein of interest, comprising:
(i) quantifying a first concentration of the protein of interest at a first time point after administration of the protein of interest to a subject by:
(a) obtaining a first sample from the subject including the protein of interest at a first time point after administration of the protein of interest to the subject; and
(b) quantifying the protein of interest in the first sample by subjecting the first sample according to the method of claim 1; and
(ii) quantifying a second concentration of the protein of interest at a second time point after administration of the protein of interest to the subject by:
(a) obtaining a second sample from the subject including the protein of interest at the second time point after administration of the protein of interest to the subject; and
(b) quantifying the protein of interest in the second sample by subjecting the second sample to the method of claim 1; and
(iii) producing the pharmacokinetic profile based on the first concentration and the second concentration.
The teachings of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. These references do not teach the production of a pharmacokinetic profile.
Schaller relates to pharmacokinetic analysis of a bispecific antibody in plasma and whole blood using MS [title], and discusses that drug development of biologics such as bispecific antibodies requires formidable characterization alongside preclinical efficacy [p 3, para 1].
Regarding claim 36, Schaller teaches the administration of a drug comprising a protein of interest to a subject (EGFRvIII:CD3 bi-scFv), and the collection of blood at multiple timepoints for MS analysis of the levels of protein of interest in the samples to generate a pharmacokinetic profile [Graphical Abstract, and p 3, para 2].
In view of Schaller, it would have been prima facie obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of Li and Haberger generating a pharmacokinetic profile of the protein of interest administered to a subject over time, as taught by Schaller, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of Li and Haberger because Schaller teaches drug development of biologics such as bispecific antibodies requires formidable characterization alongside preclinical efficacy such as pharmacokinetic profiles. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Schaller relate to methods of analyzing samples comprising antibodies via MS.
Regarding claim 37, the term “biotransformation product” is not specifically defined by the instant specification, but [para 0069] gives examples of a biotransformation product comprising a post-translational modification, truncation, aggregate, fragment, degradation product, or combination thereof, and [para 0136] states that biotransformation products may have modified properties compared to a pre-administration therapeutic. As Li and Schaller both teach methods of detecting fragments of captured antibodies [p 8, final paragraph of Schaller; p 1267, col 1, para 1 of Li], the combined method of Li, Haberger and Schaller is considered to correspond to the detection of a biotransformation product.
Regarding claim 38, Schaller teaches the method can be used to simultaneously detect free and cell-bound drug in order to infer that decay of protein was not simply to do binding of blood cells [p 11, para 1], and as the antibody of Schaller targeted two receptors, the detection of cell-bound drug is considered to correspond the detection of an interacting protein that interacts with a protein of interest.
Therefore, the invention of claims 36-38 would have been obvious to one of ordinary skill in the art before the effective filing date.
Double Patenting
The nonstatutory double patenting rejection is based on a judicially created doctrine grounded in public policy (a policy reflected in the statute) so as to prevent the unjustified or improper timewise extension of the “right to exclude” granted by a patent and to prevent possible harassment by multiple assignees. A nonstatutory double patenting rejection is appropriate where the conflicting claims are not identical, but at least one examined application claim is not patentably distinct from the reference claim(s) because the examined application claim is either anticipated by, or would have been obvious over, the reference claim(s). See, e.g., In re Berg, 140 F.3d 1428, 46 USPQ2d 1226 (Fed. Cir. 1998); In re Goodman, 11 F.3d 1046, 29 USPQ2d 2010 (Fed. Cir. 1993); In re Longi, 759 F.2d 887, 225 USPQ 645 (Fed. Cir. 1985); In re Van Ornum, 686 F.2d 937, 214 USPQ 761 (CCPA 1982); In re Vogel, 422 F.2d 438, 164 USPQ 619 (CCPA 1970); In re Thorington, 418 F.2d 528, 163 USPQ 644 (CCPA 1969).
A timely filed terminal disclaimer in compliance with 37 CFR 1.321(c) or 1.321(d) may be used to overcome an actual or provisional rejection based on nonstatutory double patenting provided the reference application or patent either is shown to be commonly owned with the examined application, or claims an invention made as a result of activities undertaken within the scope of a joint research agreement. See MPEP § 717.02 for applications subject to examination under the first inventor to file provisions of the AIA as explained in MPEP § 2159. See MPEP § 2146 et seq. for applications not subject to examination under the first inventor to file provisions of the AIA . A terminal disclaimer must be signed in compliance with 37 CFR 1.321(b).
The filing of a terminal disclaimer by itself is not a complete reply to a nonstatutory double patenting (NSDP) rejection. A complete reply requires that the terminal disclaimer be accompanied by a reply requesting reconsideration of the prior Office action. Even where the NSDP rejection is provisional the reply must be complete. See MPEP § 804, subsection I.B.1. For a reply to a non-final Office action, see 37 CFR 1.111(a). For a reply to final Office action, see 37 CFR 1.113(c). A request for reconsideration while not provided for in 37 CFR 1.113(c) may be filed after final for consideration. See MPEP §§ 706.07(e) and 714.13.
The USPTO Internet website contains terminal disclaimer forms which may be used. Please visit www.uspto.gov/patent/patents-forms. The actual filing date of the application in which the form is filed determines what form (e.g., PTO/SB/25, PTO/SB/26, PTO/AIA /25, or PTO/AIA /26) should be used. A web-based eTerminal Disclaimer may be filled out completely online using web-screens. An eTerminal Disclaimer that meets all requirements is auto-processed and approved immediately upon submission. For more information about eTerminal Disclaimers, refer to www.uspto.gov/patents/apply/applying-online/eterminal-disclaimer.
A. Claims 1-12, 16-21 and 24-26 are rejected on the ground of nonstatutory double patenting over claim 1 of U.S. Patent No. 12,153,053 (cited on the attached form PTO-892; herein “patent”) in view of Li and Haberger.
Regarding instant claim 1, claim 1 of the patent recites a method for identifying at least one host cell protein from a sample comprising subjecting a sample to a chromatography and performing mass spectrometry analysis on an elution.
The claim of the patent does not recite subjecting the enriched protein of interest or a fragment thereof to SEC under native conditions to form an SEC eluate and contacting the SEC eluate to a denaturing solution to form a denatured eluate.
Li relates to general LC-MS/MS to quantify therapeutic monoclonal antibodies (mAbs) [title].
Regarding instant claim 1, Li discloses a method of quantifying mAbs using tandem liquid chromatographic mass spectrometry (LC-MS/MS) [abstract] and that LC-MS/MS methods can be applied to mAbs for the bioanalysis of preclinical samples, wherein mAbs are enzymatically digested to generate a signature peptide for MS quantification [p 1267, col 1, para 2]. Li further discloses multidimensional chromatography can be used to separate mAb signature peptides from the peptides of the endogenous matrix at the expense of time, and that the use of immunocapture to clean up and concentrate a sample overcome such drawbacks and enhance sensitivity [p 1267, col 1, para 2].
Specifically, Li discloses the addition of the biotinylated human Fc mAb clone 35 (b-Ab35) to streptavidin magnetic beads to produce b-Ab35-coated magnetic beads, followed by the addition of a sample containing alpha-DA-G2, wherein the sample and beads were incubated for 1 h at ambient temperature, washed with 0.01% Tween 20 in PBS, and eluted with a methanol/formic acid mixture, and the resulting supernatant was denatured in the presence of a buffer comprising 8M urea and 20 mM DTT before being subjected to LC-MS/MS analysis [p 1268, col 1, final paragraph; col 2, final paragraph to p 1269, col 1, para 1]. These discloses of Li correspond to steps (i), (ii) and (v), and the subjecting of eluate to a denaturing solution of step (iv) of claim 1.
Haberger relates to characterization of biotherapeutic proteins by SEC coupled to native MS [title], and discusses that SEC is the method of choice for routine product testing to detect and quantify high-molecular aggregates and to determine the content of low molecular forms such as fragments, but does not enable determination of molecular mass of an analyte [p 331, col 1, beginning at final paragraph].
Regarding instant claim 1, Haberger discloses a method of analyzing CrossMAb samples that are in-house bispecific antibodies [p 331, col 2, final paragraph] comprising SEC of CrossMAb samples with a mobile phase of KH2PO4 and KCl and pH 7 [p 338, col 2, para 2], followed by analysis of the eluate via ESI-UV/MS [Figure 5], wherein the SEC protocol was able to resolve the CrossMAb size variants [p 336, col 1, final paragraph, and Figure 5]. The disclosures of Haberger are considered to correspond to subjecting a sample to SEC under native conditions to form an SEC eluate as recited in part (iii) of claim 1.
In view of Li and Haberger, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the claims of the patent by using SEC on the sample before MS analysis, as disclosed by Li and Haberger, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the claims of the patent because Li discloses the a method for identifying antibodies in a sample via affinity chromatography and MS, Haberger discloses SEC can detect and quantify high-molecular aggregates and to determine the content of low molecular forms such as fragments, and Haberger discloses a method resolves size variants of mAbs before MS analysis. One of ordinary skill in the art would have had a reasonable expectation of success because the patent, Li and Haberger relate to methods of identifying proteins via MS.
Regarding instant claim 2, Li discloses the method to analyze alpha-DA-G2 as described above, which is an isotype of the human antidinitrophenol IgG2 mAb [p 1268, col 1, para 2], which is considered to correspond to a drug product.
Regarding instant claim 3, Li discloses the method was carried out on alpha-DA-G2 prepared in cynomolgus monkey serum [p 1271, col 2, final paragraph].
Regarding instant claims 4-5, Li discloses the method carried out on the mAb alpha-DA-G2 as discussed above [p 1268, col 1, final paragraph; col 2, final paragraph to p 1269, col 1, para 1], which is an antibody. As the instant specification does not specifically define a therapeutic antibody, but states in [para 0140] “an anti-human Fc antibody will preferentially bind to the Fc domain of any human antibody, such as, for example, a therapeutic antibody”, and the method of Li uses a biotinylated human Fc mAb clone 35 (b-Ab35) to pull down and analyze the mAb alpha-DA-G2, the alpha-DA-G2 of Li is considered to correspond to a therapeutic antibody.
Regarding instant claims 6-9, the solid surface of the method of Li consists of streptavidin magnetic beads coated with biotinylated human Fc mAb clone 35 (b-Ab35) as discussed in the rejection of claim 1 above.
Regarding instant claim 10, the method of Li comprises a washing step as discussed in the rejection of claim 1 above.
Regarding instant claims 11-12, Li discloses LC-MS/MS methods can be applied to mAbs for the bioanalysis of preclinical samples, wherein mAbs are enzymatically digested, usually with trypsin, to generate a signature peptide for MS quantification [p 1267, col 1, para 2].
Regarding instant claim 16, Haberger discloses the use of an SEC system directly coupled to a mass spectrometer [p 338, col 1, para 3].
Regarding instant claims 17-18, Haberger discloses the use of potassium phosphate as a the SEC mobile phase as discussed above in the rejection of claim 1, and also discloses the use of 25-100 mM ammonium acetate buffer as the SEC mobile phase [p 333, col 2, para 1], and MPEP 2144.05.I states in the case where the claimed ranges overlap or lie inside ranges disclosed by the prior art, a prima facie case of obviousness exists.
Regarding instant claims 19-20, Li discloses the analysis of captured fragments via LC-MS/MS with a mobile phase of 0.1% formic acid in acetonitrile/water, wherein the ratios of acetonitrile/water in mobile phases A and B are 5/95 and 95/5 respectively, and a gradient profile was carried out from 0% to 95% mobile phase B over time [p 1268, col 2, para 3], which is considered to correspond to a denaturing solution from 40% to 80% acetonitrile.
Regarding instant claim 21, Li discloses the use of 3% formic acid on samples to elute from the magnetic beads followed by the transfer of eluate in supernatant to a well plate to dry [p 1269, col 1, para 1]. As the supernatant is understood to contain the eluting solution of 3% formic acid and the eluate, the disclosures of Li are considered to correspond to contacting eluate with a denaturing solution comprising 3% formic acid.
Regarding instant claim 24, Haberger discloses an SEC flow rate of 0.2 ml/min [p 338, col 1, para 3].
Regarding instant claim 25, Haberger discloses an SEC flow rate of 0.2 ml/min [p 338, col 1, para 3] that is coupled directly to MS analysis while Li discloses MS analysis with a mobile phase of 0.1% formic acid in acetonitrile/water that is considered a denaturing solution with a flow rate of 0.6 ml/min [p 1268, col 2, para 3]. It would be obvious to one of ordinary skill in the art in practicing the combined method of Haberger and Li to match the flow rates of the coupled protocols of SEC and MS analysis and maintain a mobile phase of 0.1% formic acid in acetonitrile/water with a flowrate of 0.2 ml/min, as MPEP 2144.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation.
Regarding instant claim 26, Haberger discloses the use of an electrospray ionization mass spectrometer (ESI/MS) [p 338, col 1, para 3].
Claims 13 and 15 are rejected on the ground of nonstatutory double patenting over claim 1 of U.S. Patent No. 12,153,053 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of PR.
The claims of the patent and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the enzyme IdeS or a variant thereof.
PR relates to IdeS as an immunoglobulin-degrading cysteine proteinase of Streptococcus pyogenes [title].
Regarding instant claim 13, PR discloses S. pyogenes expresses proteolytic activity towards IgG via the activity of the cysteine proteinase enzyme IdeS, which cleaves the heavy chain of IgG to form two stable Fab fragments and one Fc fragment [p 52, col 2, paras 2-3].
In view of PR, it would have been obvious to one of ordinary skill in the art before the effective filing date to modify the combined method of the patent, Li and Haberger by using IdeS, since the simple substitution of one known element for another results in a predictable result. One of ordinary skill in the art would have recognized that both trypsin and IdeS are digestive enzymes, and as such both are capable of being incorporated into such methods compositions as described by Li. Thus it would have been obvious to one of ordinary skill in the art to replace trypsin with IdeS, as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because Li describes a method of capturing and analyzing mAbs using the digestive enzyme trypsin, and PR describes a digestive enzyme specific for IgGs.
Regarding instant claim 15, as the combined method of the patent, Li, Haberger and PR uses the IdeS enzyme to digest an immobilized mAb as described above, and PR discloses the activity of IdeS produces Fab and Fc fragments [p 52, col 2, para 3], the combined disclosures of Li, Haberger and PR are considered to satisfy the limitations of claim 15.
Claim 14 is rejected on the ground of nonstatutory double patenting over claim 1 of U.S. Patent No. 12,153,053 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Rosli.
The claims of the patent and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of centrifugation during elution.
Rosli relates to the recovery of biotinylated proteins from streptavidin-based affinity chromatography resins [title], and discusses the release of biotinylated proteins from streptavidin based reagents is a major problem due to the stability of the complex [abstract].
Regarding instant claim 14, Rosli discloses a protocol for the release of biotinylated proteins from streptavidin Sepharose comprising the use of a centrifuge [p 91, para 2 and p 94, Section 3.3].
In view of Rosli, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the patent, Li and Haberger by using centrifugation in the elution step, as disclosed by Rosli, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the patent, Li and Haberger because Rosli discloses the release of biotinylated proteins from streptavidin is a major problem in streptavidin affinity methods and therefore discloses a quantitative release strategy using centrifugation. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Rosli relate to methods comprising streptavidin affinity capture and release.
Claim 22 is rejected on the ground of nonstatutory double patenting over claim 1 of U.S. Patent No. 12,153,053 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Donnelly.
The claims of the patent and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite a denaturing solution comprising about 60% acetonitrile and about 4% formic acid.
Donnelly reviews best practices and benchmarks for intact protein analysis for top-down MS [title], and discusses robust protocols for mass analysis of intact proteins such as antibodies from mixtures of varying complexity [abstract].
Regarding instant claim 22, Donnelly discloses denaturing direct-infusion MS comprising the treatment of samples to 49.95% acetonitrile, 49.95% water and 0.1% formic acid, wherein a 60:35:5 ratio of HPLC grade methanol:water:acetic acid may be used as an alternative resulting in improved signal to noise ratios, and discusses how the use of organic solvents and acids results in efficient ionization [p 591, col 1, para 3]. Therefore Donnelly discloses the result effective variable of organic solvent and acid ratio according to MPEP 2144.05.II.B which effects signal to noise ratio in MS, and thereby quality of the MS analysis. While Donnelly does not disclose the 60% acetonitrile and 4% formic acid recited in the claim, MPEP 2144.05.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. Therefore one of ordinary skill in the art would be motivated to modify the combined method of Li, Haberger and Donnelly to get to the claimed 60% acetonitrile and 4% formic acid because Donnelly discloses the result effective variable of organic solvent and acid ratios, and gives an example of using an increased amount of organic solvent and acid can result in improved signal to noise ratio and therefore improved MS analysis.
In view of Donnelly, it would have been obvious to one of ordinary skill in the art before the effective filing date to modify the combined method of the patent, Li and Haberger by using a denaturing solution of 60% acetonitrile and 4% formic acid because MPEP 2144.05.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. One of ordinary skill in the art would have been motivated to modify the combined method of the patent, Li and Haberger because Donnelly discloses organic solvent and acid ratios are a result effective variable according to MPEP 2144.05.II.B which effects signal to noise ratio in MS, and thereby quality of the MS analysis. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Donnelly relate to methods of MS analysis comprising treatment of protein samples with acetonitrile and formic acid.
Claim 23 is rejected on the ground of nonstatutory double patenting over claim 1 of U.S. Patent No. 12,153,053 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Wong.
The claims of the patent and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of a T-mixer.
Wong relates to a T-mixer as a rapid mixing micromixer [title], and discusses that micromixers are essential components to microfluidic systems because some biochemical analysis require two reagents to be completely mixed before a reaction has proceeded considerably, and can require sub-millisecond mixing times, and therefore micromixers must be capable of mixing two reagents within the shortest time possible [p 359, col 1, paras 1-2 and col 2, paras 1-2].
Regarding instant claim 23, Wong discloses a T-mixer that can sufficiently cause complete mixing of two solutions within less than a millisecond after the two solutions make contact [abstract].
In view of Wong, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the patent, Li and Haberger by a T-mixer to contact the SEC eluate with denaturing solution, as disclosed by Wong, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the patent, Li and Haberger because Wong discloses micromixers such as T-mixers are essential components of microfluidic systems in order to insure reagents are completely mixed in sub-millisecond times. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Wong both relate to methods requiring the efficient mixing of biochemical reagents for downstream analysis of the product.
Claims 27-30 are rejected on the ground of nonstatutory double patenting over claim 1 of U.S. Patent No. 12,153,053 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Ehkirch.
The claims of the patent and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of a flow splitter.
Ehkirch relates to an online four-dimensional HIC x SEC-IM x MS methodology for characterization of antibody drug conjugates [title] and describes an analytical system that combines multiple chromatography separation methods with MS analysis for a comprehensive and streamlined characterization of native and forced degraded sample conformations [abstract].
Regarding instant claim 27, Ehkirch discloses the multidimensional analysis method above comprising a first chromatographic dimension, a second chromatographic dimension which is SEC, and a flow splitter connecting SEC to MS which divides the flow rate by a factor of 7 prior to entering MS [p 1581, col 1, paras 2-3; Figure 2].
In view of Ehkirch, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the patent, Li and Haberger using a flow splitter between the SEC and MS, as disclosed by Ehkirch, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the patent, Li and Haberger because Ehkirch discloses a multidimensional chromatography-MS analytical method for antibody drug conjugate characterization comprising a flow splitter that provides streamlined characterization of native and forced degraded sample conformations. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Ehkirch relate to methods of analyzing samples comprising antibodies via MS.
Regarding instant claim 28, Haberger discloses the use of a detector to observe absorbance at 280 nm [Figure 1], which is considered to correspond to a UV detector.
Regarding instant claim 29, Ehkirch discloses the use of a flow splitter to divide the flow rate by a factor of 7 before entering MS but downstream of a detector [p 1581, col 1, paras 2-3; Figure 2], therefore indicating the flow to the MS to be a low flow relative to the high flow in the detector.
Regarding instant claim 30, Ehkirch discloses reducing the flow rate by a factor of 7 from an inlet flow rate of 100 µL/min [p 1581, col 1, para 3], which is considered to correspond to a low flow of less than 20 µL/min.
Claims 36-38 are rejected on the ground of nonstatutory double patenting over claim 1 of U.S. Patent No. 12,153,053 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Schaller.
The claims of the patent and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the production of a pharmacokinetic profile.
Schaller relates to pharmacokinetic analysis of a bispecific antibody in plasma and whole blood using MS [title], and discusses that drug development of biologics such as bispecific antibodies requires formidable characterization alongside preclinical efficacy [p 3, para 1].
Regarding instant claim 36, Schaller discloses the administration of a drug comprising a protein of interest to a subject (EGFRvIII:CD3 bi-scFv), and the collection of blood at multiple timepoints for MS analysis of the levels of protein of interest in the samples to generate a pharmacokinetic profile [Graphical Abstract, and p 3, para 2].
In view of Schaller, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the patent, Li and Haberger generating a pharmacokinetic profile of the protein of interest administered to a subject over time, as disclosed by Schaller, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the patent, Li and Haberger because Schaller discloses drug development of biologics such as bispecific antibodies requires formidable characterization alongside preclinical efficacy such as pharmacokinetic profiles. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Schaller relate to methods of analyzing samples comprising antibodies via MS.
Regarding instant claim 37, the term “biotransformation product” is not specifically defined by the instant specification, but [para 0069] gives examples of a biotransformation product comprising a post-translational modification, truncation, aggregate, fragment, degradation product, or combination thereof, and [para 0136] states that biotransformation products may have modified properties compared to a pre-administration therapeutic. As Li and Schaller disclose methods of detecting fragments of captured antibodies [p 8, final paragraph of Schaller; p 1267, col 1, para 1 of Li], the combined method of Li, Haberger and Schaller is considered to correspond to the detection of a biotransformation product.
Regarding instant claim 38, Schaller discloses the method can be used to simultaneously detect free and cell-bound drug in order to infer that decay of protein was not simply to do binding of blood cells [p 11, para 1], and as the antibody of Schaller targeted two receptors, the detection of cell-bound drug is considered to correspond the detection of an interacting protein that interacts with a protein of interest.
B. Claims 1-12, 16-21 and 24-26 are provisionally rejected on the ground of nonstatutory double patenting over claim 29 of co-pending Application No. 18/114,047 (herein “reference application”) in view of Li and Haberger.
Regarding instant claim 1, claim 29 of the reference application recites a method for identifying a host cell protein in a sample comprising subjecting the sample to SEC, digesting peptides to SEC fractions and subjecting peptide digests to MS.
The claim of the reference application does not recite subjecting the enriched protein of interest or a fragment thereof to SEC under native conditions to form an SEC eluate; contacting the SEC eluate to a denaturing solution to form a denatured eluate; and subjecting the denatured eluate to MS analysis to identify, quantify, and/or characterize the protein of interest.
Li relates to general LC-MS/MS to quantify therapeutic monoclonal antibodies (mAbs) [title].
Regarding instant claim 1, Li discloses a method of quantifying mAbs using tandem liquid chromatographic mass spectrometry (LC-MS/MS) [abstract] and that LC-MS/MS methods can be applied to mAbs for the bioanalysis of preclinical samples, wherein mAbs are enzymatically digested to generate a signature peptide for MS quantification [p 1267, col 1, para 2]. Li further discloses multidimensional chromatography can be used to separate mAb signature peptides from the peptides of the endogenous matrix at the expense of time, and that the use of immunocapture to clean up and concentrate a sample overcome such drawbacks and enhance sensitivity [p 1267, col 1, para 2].
Specifically, Li discloses the addition of the biotinylated human Fc mAb clone 35 (b-Ab35) to streptavidin magnetic beads to produce b-Ab35-coated magnetic beads, followed by the addition of a sample containing alpha-DA-G2, wherein the sample and beads were incubated for 1 h at ambient temperature, washed with 0.01% Tween 20 in PBS, and eluted with a methanol/formic acid mixture, and the resulting supernatant was denatured in the presence of a buffer comprising 8M urea and 20 mM DTT before being subjected to LC-MS/MS analysis [p 1268, col 1, final paragraph; col 2, final paragraph to p 1269, col 1, para 1]. These discloses of Li correspond to steps (i), (ii) and (v), and the subjecting of eluate to a denaturing solution of step (iv) of claim 1.
Haberger relates to characterization of biotherapeutic proteins by SEC coupled to native MS [title], and discusses that SEC is the method of choice for routine product testing to detect and quantify high-molecular aggregates and to determine the content of low molecular forms such as fragments, but does not enable determination of molecular mass of an analyte [p 331, col 1, beginning at final paragraph].
Regarding instant claim 1, Haberger discloses a method of analyzing CrossMAb samples that are in-house bispecific antibodies [p 331, col 2, final paragraph] comprising SEC of CrossMAb samples with a mobile phase of KH2PO4 and KCl and pH 7 [p 338, col 2, para 2], followed by analysis of the eluate via ESI-UV/MS [Figure 5], wherein the SEC protocol was able to resolve the CrossMAb size variants [p 336, col 1, final paragraph, and Figure 5]. The disclosures of Haberger are considered to correspond to subjecting a sample to SEC under native conditions to form an SEC eluate as recited in part (iii) of claim 1.
In view of Li and Haberger, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the claims of the reference application by using SEC on the sample before MS analysis, as disclosed by Li and Haberger, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the claims of the patent because Li discloses the a method for identifying antibodies in a sample via affinity chromatography and MS, Haberger discloses SEC can detect and quantify high-molecular aggregates and to determine the content of low molecular forms such as fragments, and Haberger discloses a method resolves size variants of mAbs before MS analysis. One of ordinary skill in the art would have had a reasonable expectation of success because the reference application, Li and Haberger relate to methods of identifying proteins via MS.
Regarding instant claim 2, Li discloses the method to analyze alpha-DA-G2 as described above, which is an isotype of the human antidinitrophenol IgG2 mAb [p 1268, col 1, para 2], which is considered to correspond to a drug product.
Regarding instant claim 3, Li discloses the method was carried out on alpha-DA-G2 prepared in cynomolgus monkey serum [p 1271, col 2, final paragraph].
Regarding instant claims 4-5, Li discloses the method carried out on the mAb alpha-DA-G2 as discussed above [p 1268, col 1, final paragraph; col 2, final paragraph to p 1269, col 1, para 1], which is an antibody. As the instant specification does not specifically define a therapeutic antibody, but states in [para 0140] “an anti-human Fc antibody will preferentially bind to the Fc domain of any human antibody, such as, for example, a therapeutic antibody”, and the method of Li uses a biotinylated human Fc mAb clone 35 (b-Ab35) to pull down and analyze the mAb alpha-DA-G2, the alpha-DA-G2 of Li is considered to correspond to a therapeutic antibody.
Regarding instant claims 6-9, the solid surface of the method of Li consists of streptavidin magnetic beads coated with biotinylated human Fc mAb clone 35 (b-Ab35) as discussed in the rejection of claim 1 above.
Regarding instant claim 10, the method of Li comprises a washing step as discussed in the rejection of claim 1 above.
Regarding instant claims 11-12, Li discloses LC-MS/MS methods can be applied to mAbs for the bioanalysis of preclinical samples, wherein mAbs are enzymatically digested, usually with trypsin, to generate a signature peptide for MS quantification [p 1267, col 1, para 2].
Regarding instant claim 16, Haberger discloses the use of an SEC system directly coupled to a mass spectrometer [p 338, col 1, para 3].
Regarding instant claims 17-18, Haberger discloses the use of potassium phosphate as a the SEC mobile phase as discussed above in the rejection of claim 1, and also discloses the use of 25-100 mM ammonium acetate buffer as the SEC mobile phase [p 333, col 2, para 1], and MPEP 2144.05.I states in the case where the claimed ranges overlap or lie inside ranges disclosed by the prior art, a prima facie case of obviousness exists.
Regarding instant claims 19-20, Li discloses the analysis of captured fragments via LC-MS/MS with a mobile phase of 0.1% formic acid in acetonitrile/water, wherein the ratios of acetonitrile/water in mobile phases A and B are 5/95 and 95/5 respectively, and a gradient profile was carried out from 0% to 95% mobile phase B over time [p 1268, col 2, para 3], which is considered to correspond to a denaturing solution from 40% to 80% acetonitrile.
Regarding instant claim 21, Li discloses the use of 3% formic acid on samples to elute from the magnetic beads followed by the transfer of eluate in supernatant to a well plate to dry [p 1269, col 1, para 1]. As the supernatant is understood to contain the eluting solution of 3% formic acid and the eluate, the disclosures of Li are considered to correspond to contacting eluate with a denaturing solution comprising 3% formic acid.
Regarding instant claim 24, Haberger discloses an SEC flow rate of 0.2 ml/min [p 338, col 1, para 3].
Regarding instant claim 25, Haberger discloses an SEC flow rate of 0.2 ml/min [p 338, col 1, para 3] that is coupled directly to MS analysis while Li discloses MS analysis with a mobile phase of 0.1% formic acid in acetonitrile/water that is considered a denaturing solution with a flow rate of 0.6 ml/min [p 1268, col 2, para 3]. It would be obvious to one of ordinary skill in the art in practicing the combined method of Haberger and Li to match the flow rates of the coupled protocols of SEC and MS analysis and maintain a mobile phase of 0.1% formic acid in acetonitrile/water with a flowrate of 0.2 ml/min, as MPEP 2144.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation.
Regarding instant claim 26, Haberger discloses the use of an electrospray ionization mass spectrometer (ESI/MS) [p 338, col 1, para 3].
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claims 13 and 15 are provisionally rejected on the ground of nonstatutory double patenting over claim 29 of co-pending Application No. 18/114,047 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of PR.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the enzyme IdeS or a variant thereof.
PR relates to IdeS as an immunoglobulin-degrading cysteine proteinase of Streptococcus pyogenes [title].
Regarding instant claim 13, PR discloses S. pyogenes expresses proteolytic activity towards IgG via the activity of the cysteine proteinase enzyme IdeS, which cleaves the heavy chain of IgG to form two stable Fab fragments and one Fc fragment [p 52, col 2, paras 2-3].
In view of PR, it would have been obvious to one of ordinary skill in the art before the effective filing date to modify the combined method of the reference application, Li and Haberger by using IdeS, since the simple substitution of one known element for another results in a predictable result. One of ordinary skill in the art would have recognized that both trypsin and IdeS are digestive enzymes, and as such both are capable of being incorporated into such methods compositions as described by Li. Thus it would have been obvious to one of ordinary skill in the art to replace trypsin with IdeS, as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because Li describes a method of capturing and analyzing mAbs using the digestive enzyme trypsin, and PR describes a digestive enzyme specific for IgGs.
Regarding instant claim 15, as the combined method of the patent, Li, Haberger and PR uses the IdeS enzyme to digest an immobilized mAb as described above, and PR discloses the activity of IdeS produces Fab and Fc fragments [p 52, col 2, para 3], the combined disclosures of Li, Haberger and PR are considered to satisfy the limitations of claim 15.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 14 is provisionally rejected on the ground of nonstatutory double patenting over claim 29 of co-pending Application No. 18/114,047 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Rosli.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of centrifugation during elution.
Rosli relates to the recovery of biotinylated proteins from streptavidin-based affinity chromatography resins [title], and discusses the release of biotinylated proteins from streptavidin based reagents is a major problem due to the stability of the complex [abstract].
Regarding instant claim 14, Rosli discloses a protocol for the release of biotinylated proteins from streptavidin Sepharose comprising the use of a centrifuge [p 91, para 2 and p 94, Section 3.3].
In view of Rosli, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger by using centrifugation in the elution step, as disclosed by Rosli, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Rosli discloses the release of biotinylated proteins from streptavidin is a major problem in streptavidin affinity methods and therefore discloses a quantitative release strategy using centrifugation. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Rosli relate to methods comprising streptavidin affinity capture and release.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 22 is provisionally rejected on the ground of nonstatutory double patenting over claim 29 of co-pending Application No. 18/114,047 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Donnelly.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite a denaturing solution comprising about 60% acetonitrile and about 4% formic acid.
Donnelly reviews best practices and benchmarks for intact protein analysis for top-down MS [title], and discusses robust protocols for mass analysis of intact proteins such as antibodies from mixtures of varying complexity [abstract].
Regarding instant claim 22, Donnelly discloses denaturing direct-infusion MS comprising the treatment of samples to 49.95% acetonitrile, 49.95% water and 0.1% formic acid, wherein a 60:35:5 ratio of HPLC grade methanol:water:acetic acid may be used as an alternative resulting in improved signal to noise ratios, and discusses how the use of organic solvents and acids results in efficient ionization [p 591, col 1, para 3]. Therefore Donnelly discloses the result effective variable of organic solvent and acid ratio according to MPEP 2144.05.II.B which effects signal to noise ratio in MS, and thereby quality of the MS analysis. While Donnelly does not disclose the 60% acetonitrile and 4% formic acid recited in the claim, MPEP 2144.05.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. Therefore one of ordinary skill in the art would be motivated to modify the combined method of Li, Haberger and Donnelly to get to the claimed 60% acetonitrile and 4% formic acid because Donnelly discloses the result effective variable of organic solvent and acid ratios, and gives an example of using an increased amount of organic solvent and acid can result in improved signal to noise ratio and therefore improved MS analysis.
In view of Donnelly, it would have been obvious to one of ordinary skill in the art before the effective filing date to modify the combined method of the reference application, Li and Haberger by using a denaturing solution of 60% acetonitrile and 4% formic acid because MPEP 2144.05.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Donnelly discloses organic solvent and acid ratios are a result effective variable according to MPEP 2144.05.II.B which effects signal to noise ratio in MS, and thereby quality of the MS analysis. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Donnelly relate to methods of MS analysis comprising treatment of protein samples with acetonitrile and formic acid.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 23 is provisionally rejected on the ground of nonstatutory double patenting over claim 29 of co-pending Application No. 18/114,047 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Wong.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of a T-mixer.
Wong relates to a T-mixer as a rapid mixing micromixer [title], and discusses that micromixers are essential components to microfluidic systems because some biochemical analysis require two reagents to be completely mixed before a reaction has proceeded considerably, and can require sub-millisecond mixing times, and therefore micromixers must be capable of mixing two reagents within the shortest time possible [p 359, col 1, paras 1-2 and col 2, paras 1-2].
Regarding instant claim 23, Wong discloses a T-mixer that can sufficiently cause complete mixing of two solutions within less than a millisecond after the two solutions make contact [abstract].
In view of Wong, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger by a T-mixer to contact the SEC eluate with denaturing solution, as disclosed by Wong, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Wong discloses micromixers such as T-mixers are essential components of microfluidic systems in order to insure reagents are completely mixed in sub-millisecond times. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Wong both relate to methods requiring the efficient mixing of biochemical reagents for downstream analysis of the product.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claims 27-30 are provisionally rejected on the ground of nonstatutory double patenting over claim 29 of co-pending Application No. 18/114,047 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Ehkirch.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of a flow splitter.
Ehkirch relates to an online four-dimensional HIC x SEC-IM x MS methodology for characterization of antibody drug conjugates [title] and describes an analytical system that combines multiple chromatography separation methods with MS analysis for a comprehensive and streamlined characterization of native and forced degraded sample conformations [abstract].
Regarding instant claim 27, Ehkirch discloses the multidimensional analysis method above comprising a first chromatographic dimension, a second chromatographic dimension which is SEC, and a flow splitter connecting SEC to MS which divides the flow rate by a factor of 7 prior to entering MS [p 1581, col 1, paras 2-3; Figure 2].
In view of Ehkirch, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger using a flow splitter between the SEC and MS, as disclosed by Ehkirch, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Ehkirch discloses a multidimensional chromatography-MS analytical method for antibody drug conjugate characterization comprising a flow splitter that provides streamlined characterization of native and forced degraded sample conformations. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Ehkirch relate to methods of analyzing samples comprising antibodies via MS.
Regarding instant claim 28, Haberger discloses the use of a detector to observe absorbance at 280 nm [Figure 1], which is considered to correspond to a UV detector.
Regarding instant claim 29, Ehkirch discloses the use of a flow splitter to divide the flow rate by a factor of 7 before entering MS but downstream of a detector [p 1581, col 1, paras 2-3; Figure 2], therefore indicating the flow to the MS to be a low flow relative to the high flow in the detector.
Regarding instant claim 30, Ehkirch discloses reducing the flow rate by a factor of 7 from an inlet flow rate of 100 µL/min [p 1581, col 1, para 3], which is considered to correspond to a low flow of less than 20 µL/min.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claims 36-38 are provisionally rejected on the ground of nonstatutory double patenting over claim 29 of co-pending Application No. 18/114,047 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Schaller.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the production of a pharmacokinetic profile.
Schaller relates to pharmacokinetic analysis of a bispecific antibody in plasma and whole blood using MS [title], and discusses that drug development of biologics such as bispecific antibodies requires formidable characterization alongside preclinical efficacy [p 3, para 1].
Regarding instant claim 36, Schaller discloses the administration of a drug comprising a protein of interest to a subject (EGFRvIII:CD3 bi-scFv), and the collection of blood at multiple timepoints for MS analysis of the levels of protein of interest in the samples to generate a pharmacokinetic profile [Graphical Abstract, and p 3, para 2].
In view of Schaller, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger generating a pharmacokinetic profile of the protein of interest administered to a subject over time, as disclosed by Schaller, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Schaller discloses drug development of biologics such as bispecific antibodies requires formidable characterization alongside preclinical efficacy such as pharmacokinetic profiles. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Schaller relate to methods of analyzing samples comprising antibodies via MS.
Regarding instant claim 37, the term “biotransformation product” is not specifically defined by the instant specification, but [para 0069] gives examples of a biotransformation product comprising a post-translational modification, truncation, aggregate, fragment, degradation product, or combination thereof, and [para 0136] states that biotransformation products may have modified properties compared to a pre-administration therapeutic. As Li and Schaller disclose methods of detecting fragments of captured antibodies [p 8, final paragraph of Schaller; p 1267, col 1, para 1 of Li], the combined method of Li, Haberger and Schaller is considered to correspond to the detection of a biotransformation product.
Regarding instant claim 38, Schaller discloses the method can be used to simultaneously detect free and cell-bound drug in order to infer that decay of protein was not simply to do binding of blood cells [p 11, para 1], and as the antibody of Schaller targeted two receptors, the detection of cell-bound drug is considered to correspond the detection of an interacting protein that interacts with a protein of interest.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
C. Claims 1-12, 16-21 and 24-26 are provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 18/745,346 (herein “reference application”) in view of Li and Haberger.
Regarding instant claim 1, claim 1 of the reference application recites a method for identifying a host cell protein in a sample comprising digesting peptides in the sample and subjecting peptide digests to MS.
The claim of the reference application does not recite contacting a sample including a protein of interest to a solid surface to form an immobilized protein of interest; eluting the immobilized protein of interest of a fragment thereof to form an enriched protein of interest of fragment thereof; and subjecting the enriched protein of interest or a fragment thereof to SEC under native conditions to form an SEC eluate; contacting the SEC eluate to a denaturing solution to form a denatured eluate.
Li relates to general LC-MS/MS to quantify therapeutic monoclonal antibodies (mAbs) [title].
Regarding instant claim 1, Li discloses a method of quantifying mAbs using tandem liquid chromatographic mass spectrometry (LC-MS/MS) [abstract] and that LC-MS/MS methods can be applied to mAbs for the bioanalysis of preclinical samples, wherein mAbs are enzymatically digested to generate a signature peptide for MS quantification [p 1267, col 1, para 2]. Li further discloses multidimensional chromatography can be used to separate mAb signature peptides from the peptides of the endogenous matrix at the expense of time, and that the use of immunocapture to clean up and concentrate a sample overcome such drawbacks and enhance sensitivity [p 1267, col 1, para 2].
Specifically, Li discloses the addition of the biotinylated human Fc mAb clone 35 (b-Ab35) to streptavidin magnetic beads to produce b-Ab35-coated magnetic beads, followed by the addition of a sample containing alpha-DA-G2, wherein the sample and beads were incubated for 1 h at ambient temperature, washed with 0.01% Tween 20 in PBS, and eluted with a methanol/formic acid mixture, and the resulting supernatant was denatured in the presence of a buffer comprising 8M urea and 20 mM DTT before being subjected to LC-MS/MS analysis [p 1268, col 1, final paragraph; col 2, final paragraph to p 1269, col 1, para 1]. These discloses of Li correspond to steps (i), (ii) and (v), and the subjecting of eluate to a denaturing solution of step (iv) of claim 1.
Haberger relates to characterization of biotherapeutic proteins by SEC coupled to native MS [title], and discusses that SEC is the method of choice for routine product testing to detect and quantify high-molecular aggregates and to determine the content of low molecular forms such as fragments, but does not enable determination of molecular mass of an analyte [p 331, col 1, beginning at final paragraph].
Regarding instant claim 1, Haberger discloses a method of analyzing CrossMAb samples that are in-house bispecific antibodies [p 331, col 2, final paragraph] comprising SEC of CrossMAb samples with a mobile phase of KH2PO4 and KCl and pH 7 [p 338, col 2, para 2], followed by analysis of the eluate via ESI-UV/MS [Figure 5], wherein the SEC protocol was able to resolve the CrossMAb size variants [p 336, col 1, final paragraph, and Figure 5]. The disclosures of Haberger are considered to correspond to subjecting a sample to SEC under native conditions to form an SEC eluate as recited in part (iii) of claim 1.
In view of Li and Haberger, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the claims of the reference application by using SEC on the sample before MS analysis, as disclosed by Li and Haberger, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the claims of the patent because Li discloses the a method for identifying antibodies in a sample via affinity chromatography and MS, Haberger discloses SEC can detect and quantify high-molecular aggregates and to determine the content of low molecular forms such as fragments, and Haberger discloses a method resolves size variants of mAbs before MS analysis. One of ordinary skill in the art would have had a reasonable expectation of success because the reference application, Li and Haberger relate to methods of identifying proteins via MS.
Regarding instant claim 2, Li discloses the method to analyze alpha-DA-G2 as described above, which is an isotype of the human antidinitrophenol IgG2 mAb [p 1268, col 1, para 2], which is considered to correspond to a drug product.
Regarding instant claim 3, Li discloses the method was carried out on alpha-DA-G2 prepared in cynomolgus monkey serum [p 1271, col 2, final paragraph].
Regarding instant claims 4-5, Li discloses the method carried out on the mAb alpha-DA-G2 as discussed above [p 1268, col 1, final paragraph; col 2, final paragraph to p 1269, col 1, para 1], which is an antibody. As the instant specification does not specifically define a therapeutic antibody, but states in [para 0140] “an anti-human Fc antibody will preferentially bind to the Fc domain of any human antibody, such as, for example, a therapeutic antibody”, and the method of Li uses a biotinylated human Fc mAb clone 35 (b-Ab35) to pull down and analyze the mAb alpha-DA-G2, the alpha-DA-G2 of Li is considered to correspond to a therapeutic antibody.
Regarding instant claims 6-9, the solid surface of the method of Li consists of streptavidin magnetic beads coated with biotinylated human Fc mAb clone 35 (b-Ab35) as discussed in the rejection of claim 1 above.
Regarding instant claim 10, the method of Li comprises a washing step as discussed in the rejection of claim 1 above.
Regarding instant claims 11-12, Li discloses LC-MS/MS methods can be applied to mAbs for the bioanalysis of preclinical samples, wherein mAbs are enzymatically digested, usually with trypsin, to generate a signature peptide for MS quantification [p 1267, col 1, para 2].
Regarding instant claim 16, Haberger discloses the use of an SEC system directly coupled to a mass spectrometer [p 338, col 1, para 3].
Regarding instant claims 17-18, Haberger discloses the use of potassium phosphate as a the SEC mobile phase as discussed above in the rejection of claim 1, and also discloses the use of 25-100 mM ammonium acetate buffer as the SEC mobile phase [p 333, col 2, para 1], and MPEP 2144.05.I states in the case where the claimed ranges overlap or lie inside ranges disclosed by the prior art, a prima facie case of obviousness exists.
Regarding instant claims 19-20, Li discloses the analysis of captured fragments via LC-MS/MS with a mobile phase of 0.1% formic acid in acetonitrile/water, wherein the ratios of acetonitrile/water in mobile phases A and B are 5/95 and 95/5 respectively, and a gradient profile was carried out from 0% to 95% mobile phase B over time [p 1268, col 2, para 3], which is considered to correspond to a denaturing solution from 40% to 80% acetonitrile.
Regarding instant claim 21, Li discloses the use of 3% formic acid on samples to elute from the magnetic beads followed by the transfer of eluate in supernatant to a well plate to dry [p 1269, col 1, para 1]. As the supernatant is understood to contain the eluting solution of 3% formic acid and the eluate, the disclosures of Li are considered to correspond to contacting eluate with a denaturing solution comprising 3% formic acid.
Regarding instant claim 24, Haberger discloses an SEC flow rate of 0.2 ml/min [p 338, col 1, para 3].
Regarding instant claim 25, Haberger discloses an SEC flow rate of 0.2 ml/min [p 338, col 1, para 3] that is coupled directly to MS analysis while Li discloses MS analysis with a mobile phase of 0.1% formic acid in acetonitrile/water that is considered a denaturing solution with a flow rate of 0.6 ml/min [p 1268, col 2, para 3]. It would be obvious to one of ordinary skill in the art in practicing the combined method of Haberger and Li to match the flow rates of the coupled protocols of SEC and MS analysis and maintain a mobile phase of 0.1% formic acid in acetonitrile/water with a flowrate of 0.2 ml/min, as MPEP 2144.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation.
Regarding instant claim 26, Haberger discloses the use of an electrospray ionization mass spectrometer (ESI/MS) [p 338, col 1, para 3].
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claims 13 and 15 are provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 18/745,346 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of PR.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the enzyme IdeS or a variant thereof.
PR relates to IdeS as an immunoglobulin-degrading cysteine proteinase of Streptococcus pyogenes [title].
Regarding instant claim 13, PR discloses S. pyogenes expresses proteolytic activity towards IgG via the activity of the cysteine proteinase enzyme IdeS, which cleaves the heavy chain of IgG to form two stable Fab fragments and one Fc fragment [p 52, col 2, paras 2-3].
In view of PR, it would have been obvious to one of ordinary skill in the art before the effective filing date to modify the combined method of the reference application, Li and Haberger by using IdeS, since the simple substitution of one known element for another results in a predictable result. One of ordinary skill in the art would have recognized that both trypsin and IdeS are digestive enzymes, and as such both are capable of being incorporated into such methods compositions as described by Li. Thus it would have been obvious to one of ordinary skill in the art to replace trypsin with IdeS, as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because Li describes a method of capturing and analyzing mAbs using the digestive enzyme trypsin, and PR describes a digestive enzyme specific for IgGs.
Regarding instant claim 15, as the combined method of the patent, Li, Haberger and PR uses the IdeS enzyme to digest an immobilized mAb as described above, and PR discloses the activity of IdeS produces Fab and Fc fragments [p 52, col 2, para 3], the combined disclosures of Li, Haberger and PR are considered to satisfy the limitations of claim 15.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 14 is provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 18/745,346 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Rosli.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of centrifugation during elution.
Rosli relates to the recovery of biotinylated proteins from streptavidin-based affinity chromatography resins [title], and discusses the release of biotinylated proteins from streptavidin based reagents is a major problem due to the stability of the complex [abstract].
Regarding instant claim 14, Rosli discloses a protocol for the release of biotinylated proteins from streptavidin Sepharose comprising the use of a centrifuge [p 91, para 2 and p 94, Section 3.3].
In view of Rosli, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger by using centrifugation in the elution step, as disclosed by Rosli, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Rosli discloses the release of biotinylated proteins from streptavidin is a major problem in streptavidin affinity methods and therefore discloses a quantitative release strategy using centrifugation. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Rosli relate to methods comprising streptavidin affinity capture and release.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 22 is provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 18/745,346 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Donnelly.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite a denaturing solution comprising about 60% acetonitrile and about 4% formic acid.
Donnelly reviews best practices and benchmarks for intact protein analysis for top-down MS [title], and discusses robust protocols for mass analysis of intact proteins such as antibodies from mixtures of varying complexity [abstract].
Regarding instant claim 22, Donnelly discloses denaturing direct-infusion MS comprising the treatment of samples to 49.95% acetonitrile, 49.95% water and 0.1% formic acid, wherein a 60:35:5 ratio of HPLC grade methanol:water:acetic acid may be used as an alternative resulting in improved signal to noise ratios, and discusses how the use of organic solvents and acids results in efficient ionization [p 591, col 1, para 3]. Therefore Donnelly discloses the result effective variable of organic solvent and acid ratio according to MPEP 2144.05.II.B which effects signal to noise ratio in MS, and thereby quality of the MS analysis. While Donnelly does not disclose the 60% acetonitrile and 4% formic acid recited in the claim, MPEP 2144.05.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. Therefore one of ordinary skill in the art would be motivated to modify the combined method of Li, Haberger and Donnelly to get to the claimed 60% acetonitrile and 4% formic acid because Donnelly discloses the result effective variable of organic solvent and acid ratios, and gives an example of using an increased amount of organic solvent and acid can result in improved signal to noise ratio and therefore improved MS analysis.
In view of Donnelly, it would have been obvious to one of ordinary skill in the art before the effective filing date to modify the combined method of the reference application, Li and Haberger by using a denaturing solution of 60% acetonitrile and 4% formic acid because MPEP 2144.05.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Donnelly discloses organic solvent and acid ratios are a result effective variable according to MPEP 2144.05.II.B which effects signal to noise ratio in MS, and thereby quality of the MS analysis. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Donnelly relate to methods of MS analysis comprising treatment of protein samples with acetonitrile and formic acid.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 23 is provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 18/745,346 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Wong.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of a T-mixer.
Wong relates to a T-mixer as a rapid mixing micromixer [title], and discusses that micromixers are essential components to microfluidic systems because some biochemical analysis require two reagents to be completely mixed before a reaction has proceeded considerably, and can require sub-millisecond mixing times, and therefore micromixers must be capable of mixing two reagents within the shortest time possible [p 359, col 1, paras 1-2 and col 2, paras 1-2].
Regarding instant claim 23, Wong discloses a T-mixer that can sufficiently cause complete mixing of two solutions within less than a millisecond after the two solutions make contact [abstract].
In view of Wong, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger by a T-mixer to contact the SEC eluate with denaturing solution, as disclosed by Wong, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Wong discloses micromixers such as T-mixers are essential components of microfluidic systems in order to insure reagents are completely mixed in sub-millisecond times. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Wong both relate to methods requiring the efficient mixing of biochemical reagents for downstream analysis of the product.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claims 27-30 are provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 18/745,346 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Ehkirch.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of a flow splitter.
Ehkirch relates to an online four-dimensional HIC x SEC-IM x MS methodology for characterization of antibody drug conjugates [title] and describes an analytical system that combines multiple chromatography separation methods with MS analysis for a comprehensive and streamlined characterization of native and forced degraded sample conformations [abstract].
Regarding instant claim 27, Ehkirch discloses the multidimensional analysis method above comprising a first chromatographic dimension, a second chromatographic dimension which is SEC, and a flow splitter connecting SEC to MS which divides the flow rate by a factor of 7 prior to entering MS [p 1581, col 1, paras 2-3; Figure 2].
In view of Ehkirch, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger using a flow splitter between the SEC and MS, as disclosed by Ehkirch, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Ehkirch discloses a multidimensional chromatography-MS analytical method for antibody drug conjugate characterization comprising a flow splitter that provides streamlined characterization of native and forced degraded sample conformations. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Ehkirch relate to methods of analyzing samples comprising antibodies via MS.
Regarding instant claim 28, Haberger discloses the use of a detector to observe absorbance at 280 nm [Figure 1], which is considered to correspond to a UV detector.
Regarding instant claim 29, Ehkirch discloses the use of a flow splitter to divide the flow rate by a factor of 7 before entering MS but downstream of a detector [p 1581, col 1, paras 2-3; Figure 2], therefore indicating the flow to the MS to be a low flow relative to the high flow in the detector.
Regarding instant claim 30, Ehkirch discloses reducing the flow rate by a factor of 7 from an inlet flow rate of 100 µL/min [p 1581, col 1, para 3], which is considered to correspond to a low flow of less than 20 µL/min.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claims 36-38 are provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 18/745,346 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Schaller.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the production of a pharmacokinetic profile.
Schaller relates to pharmacokinetic analysis of a bispecific antibody in plasma and whole blood using MS [title], and discusses that drug development of biologics such as bispecific antibodies requires formidable characterization alongside preclinical efficacy [p 3, para 1].
Regarding instant claim 36, Schaller discloses the administration of a drug comprising a protein of interest to a subject (EGFRvIII:CD3 bi-scFv), and the collection of blood at multiple timepoints for MS analysis of the levels of protein of interest in the samples to generate a pharmacokinetic profile [Graphical Abstract, and p 3, para 2].
In view of Schaller, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger generating a pharmacokinetic profile of the protein of interest administered to a subject over time, as disclosed by Schaller, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Schaller discloses drug development of biologics such as bispecific antibodies requires formidable characterization alongside preclinical efficacy such as pharmacokinetic profiles. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Schaller relate to methods of analyzing samples comprising antibodies via MS.
Regarding instant claim 37, the term “biotransformation product” is not specifically defined by the instant specification, but [para 0069] gives examples of a biotransformation product comprising a post-translational modification, truncation, aggregate, fragment, degradation product, or combination thereof, and [para 0136] states that biotransformation products may have modified properties compared to a pre-administration therapeutic. As Li and Schaller disclose methods of detecting fragments of captured antibodies [p 8, final paragraph of Schaller; p 1267, col 1, para 1 of Li], the combined method of Li, Haberger and Schaller is considered to correspond to the detection of a biotransformation product.
Regarding instant claim 38, Schaller discloses the method can be used to simultaneously detect free and cell-bound drug in order to infer that decay of protein was not simply to do binding of blood cells [p 11, para 1], and as the antibody of Schaller targeted two receptors, the detection of cell-bound drug is considered to correspond the detection of an interacting protein that interacts with a protein of interest.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
D. Claims 1-12, 16-21 and 24-26 are provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 18/916,242 (herein “reference application”) in view of Li and Haberger.
Regarding instant claim 1, claim 1 of the reference application recites a method for identifying a host cell protein in a sample comprising digesting peptides in a sample, subjecting the digested peptides to chromatography and subjecting chromatography eluate to MS.
The claim of the reference application does not recite subjecting the enriched protein of interest or a fragment thereof to SEC under native conditions to form an SEC eluate; and contacting the SEC eluate to a denaturing solution to form a denatured eluate.
Li relates to general LC-MS/MS to quantify therapeutic monoclonal antibodies (mAbs) [title].
Regarding instant claim 1, Li discloses a method of quantifying mAbs using tandem liquid chromatographic mass spectrometry (LC-MS/MS) [abstract] and that LC-MS/MS methods can be applied to mAbs for the bioanalysis of preclinical samples, wherein mAbs are enzymatically digested to generate a signature peptide for MS quantification [p 1267, col 1, para 2]. Li further discloses multidimensional chromatography can be used to separate mAb signature peptides from the peptides of the endogenous matrix at the expense of time, and that the use of immunocapture to clean up and concentrate a sample overcome such drawbacks and enhance sensitivity [p 1267, col 1, para 2].
Specifically, Li discloses the addition of the biotinylated human Fc mAb clone 35 (b-Ab35) to streptavidin magnetic beads to produce b-Ab35-coated magnetic beads, followed by the addition of a sample containing alpha-DA-G2, wherein the sample and beads were incubated for 1 h at ambient temperature, washed with 0.01% Tween 20 in PBS, and eluted with a methanol/formic acid mixture, and the resulting supernatant was denatured in the presence of a buffer comprising 8M urea and 20 mM DTT before being subjected to LC-MS/MS analysis [p 1268, col 1, final paragraph; col 2, final paragraph to p 1269, col 1, para 1]. These discloses of Li correspond to steps (i), (ii) and (v), and the subjecting of eluate to a denaturing solution of step (iv) of claim 1.
Haberger relates to characterization of biotherapeutic proteins by SEC coupled to native MS [title], and discusses that SEC is the method of choice for routine product testing to detect and quantify high-molecular aggregates and to determine the content of low molecular forms such as fragments, but does not enable determination of molecular mass of an analyte [p 331, col 1, beginning at final paragraph].
Regarding instant claim 1, Haberger discloses a method of analyzing CrossMAb samples that are in-house bispecific antibodies [p 331, col 2, final paragraph] comprising SEC of CrossMAb samples with a mobile phase of KH2PO4 and KCl and pH 7 [p 338, col 2, para 2], followed by analysis of the eluate via ESI-UV/MS [Figure 5], wherein the SEC protocol was able to resolve the CrossMAb size variants [p 336, col 1, final paragraph, and Figure 5]. The disclosures of Haberger are considered to correspond to subjecting a sample to SEC under native conditions to form an SEC eluate as recited in part (iii) of claim 1.
In view of Li and Haberger, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the claims of the reference application by using SEC on the sample before MS analysis, as disclosed by Li and Haberger, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the claims of the patent because Li discloses the a method for identifying antibodies in a sample via affinity chromatography and MS, Haberger discloses SEC can detect and quantify high-molecular aggregates and to determine the content of low molecular forms such as fragments, and Haberger discloses a method resolves size variants of mAbs before MS analysis. One of ordinary skill in the art would have had a reasonable expectation of success because the reference application, Li and Haberger relate to methods of identifying proteins via MS.
Regarding instant claim 2, Li discloses the method to analyze alpha-DA-G2 as described above, which is an isotype of the human antidinitrophenol IgG2 mAb [p 1268, col 1, para 2], which is considered to correspond to a drug product.
Regarding instant claim 3, Li discloses the method was carried out on alpha-DA-G2 prepared in cynomolgus monkey serum [p 1271, col 2, final paragraph].
Regarding instant claims 4-5, Li discloses the method carried out on the mAb alpha-DA-G2 as discussed above [p 1268, col 1, final paragraph; col 2, final paragraph to p 1269, col 1, para 1], which is an antibody. As the instant specification does not specifically define a therapeutic antibody, but states in [para 0140] “an anti-human Fc antibody will preferentially bind to the Fc domain of any human antibody, such as, for example, a therapeutic antibody”, and the method of Li uses a biotinylated human Fc mAb clone 35 (b-Ab35) to pull down and analyze the mAb alpha-DA-G2, the alpha-DA-G2 of Li is considered to correspond to a therapeutic antibody.
Regarding instant claims 6-9, the solid surface of the method of Li consists of streptavidin magnetic beads coated with biotinylated human Fc mAb clone 35 (b-Ab35) as discussed in the rejection of claim 1 above.
Regarding instant claim 10, the method of Li comprises a washing step as discussed in the rejection of claim 1 above.
Regarding instant claims 11-12, Li discloses LC-MS/MS methods can be applied to mAbs for the bioanalysis of preclinical samples, wherein mAbs are enzymatically digested, usually with trypsin, to generate a signature peptide for MS quantification [p 1267, col 1, para 2].
Regarding instant claim 16, Haberger discloses the use of an SEC system directly coupled to a mass spectrometer [p 338, col 1, para 3].
Regarding instant claims 17-18, Haberger discloses the use of potassium phosphate as a the SEC mobile phase as discussed above in the rejection of claim 1, and also discloses the use of 25-100 mM ammonium acetate buffer as the SEC mobile phase [p 333, col 2, para 1], and MPEP 2144.05.I states in the case where the claimed ranges overlap or lie inside ranges disclosed by the prior art, a prima facie case of obviousness exists.
Regarding instant claims 19-20, Li discloses the analysis of captured fragments via LC-MS/MS with a mobile phase of 0.1% formic acid in acetonitrile/water, wherein the ratios of acetonitrile/water in mobile phases A and B are 5/95 and 95/5 respectively, and a gradient profile was carried out from 0% to 95% mobile phase B over time [p 1268, col 2, para 3], which is considered to correspond to a denaturing solution from 40% to 80% acetonitrile.
Regarding instant claim 21, Li discloses the use of 3% formic acid on samples to elute from the magnetic beads followed by the transfer of eluate in supernatant to a well plate to dry [p 1269, col 1, para 1]. As the supernatant is understood to contain the eluting solution of 3% formic acid and the eluate, the disclosures of Li are considered to correspond to contacting eluate with a denaturing solution comprising 3% formic acid.
Regarding instant claim 24, Haberger discloses an SEC flow rate of 0.2 ml/min [p 338, col 1, para 3].
Regarding instant claim 25, Haberger discloses an SEC flow rate of 0.2 ml/min [p 338, col 1, para 3] that is coupled directly to MS analysis while Li discloses MS analysis with a mobile phase of 0.1% formic acid in acetonitrile/water that is considered a denaturing solution with a flow rate of 0.6 ml/min [p 1268, col 2, para 3]. It would be obvious to one of ordinary skill in the art in practicing the combined method of Haberger and Li to match the flow rates of the coupled protocols of SEC and MS analysis and maintain a mobile phase of 0.1% formic acid in acetonitrile/water with a flowrate of 0.2 ml/min, as MPEP 2144.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation.
Regarding instant claim 26, Haberger discloses the use of an electrospray ionization mass spectrometer (ESI/MS) [p 338, col 1, para 3].
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claims 13 and 15 are provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 18/916,242 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of PR.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the enzyme IdeS or a variant thereof.
PR relates to IdeS as an immunoglobulin-degrading cysteine proteinase of Streptococcus pyogenes [title].
Regarding instant claim 13, PR discloses S. pyogenes expresses proteolytic activity towards IgG via the activity of the cysteine proteinase enzyme IdeS, which cleaves the heavy chain of IgG to form two stable Fab fragments and one Fc fragment [p 52, col 2, paras 2-3].
In view of PR, it would have been obvious to one of ordinary skill in the art before the effective filing date to modify the combined method of the reference application, Li and Haberger by using IdeS, since the simple substitution of one known element for another results in a predictable result. One of ordinary skill in the art would have recognized that both trypsin and IdeS are digestive enzymes, and as such both are capable of being incorporated into such methods compositions as described by Li. Thus it would have been obvious to one of ordinary skill in the art to replace trypsin with IdeS, as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because Li describes a method of capturing and analyzing mAbs using the digestive enzyme trypsin, and PR describes a digestive enzyme specific for IgGs.
Regarding instant claim 15, as the combined method of the patent, Li, Haberger and PR uses the IdeS enzyme to digest an immobilized mAb as described above, and PR discloses the activity of IdeS produces Fab and Fc fragments [p 52, col 2, para 3], the combined disclosures of Li, Haberger and PR are considered to satisfy the limitations of claim 15.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 14 is provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 18/916,242 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Rosli.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of centrifugation during elution.
Rosli relates to the recovery of biotinylated proteins from streptavidin-based affinity chromatography resins [title], and discusses the release of biotinylated proteins from streptavidin based reagents is a major problem due to the stability of the complex [abstract].
Regarding instant claim 14, Rosli discloses a protocol for the release of biotinylated proteins from streptavidin Sepharose comprising the use of a centrifuge [p 91, para 2 and p 94, Section 3.3].
In view of Rosli, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger by using centrifugation in the elution step, as disclosed by Rosli, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Rosli discloses the release of biotinylated proteins from streptavidin is a major problem in streptavidin affinity methods and therefore discloses a quantitative release strategy using centrifugation. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Rosli relate to methods comprising streptavidin affinity capture and release.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 22 is provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 18/916,242 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Donnelly.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite a denaturing solution comprising about 60% acetonitrile and about 4% formic acid.
Donnelly reviews best practices and benchmarks for intact protein analysis for top-down MS [title], and discusses robust protocols for mass analysis of intact proteins such as antibodies from mixtures of varying complexity [abstract].
Regarding instant claim 22, Donnelly discloses denaturing direct-infusion MS comprising the treatment of samples to 49.95% acetonitrile, 49.95% water and 0.1% formic acid, wherein a 60:35:5 ratio of HPLC grade methanol:water:acetic acid may be used as an alternative resulting in improved signal to noise ratios, and discusses how the use of organic solvents and acids results in efficient ionization [p 591, col 1, para 3]. Therefore Donnelly discloses the result effective variable of organic solvent and acid ratio according to MPEP 2144.05.II.B which effects signal to noise ratio in MS, and thereby quality of the MS analysis. While Donnelly does not disclose the 60% acetonitrile and 4% formic acid recited in the claim, MPEP 2144.05.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. Therefore one of ordinary skill in the art would be motivated to modify the combined method of Li, Haberger and Donnelly to get to the claimed 60% acetonitrile and 4% formic acid because Donnelly discloses the result effective variable of organic solvent and acid ratios, and gives an example of using an increased amount of organic solvent and acid can result in improved signal to noise ratio and therefore improved MS analysis.
In view of Donnelly, it would have been obvious to one of ordinary skill in the art before the effective filing date to modify the combined method of the reference application, Li and Haberger by using a denaturing solution of 60% acetonitrile and 4% formic acid because MPEP 2144.05.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Donnelly discloses organic solvent and acid ratios are a result effective variable according to MPEP 2144.05.II.B which effects signal to noise ratio in MS, and thereby quality of the MS analysis. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Donnelly relate to methods of MS analysis comprising treatment of protein samples with acetonitrile and formic acid.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 23 is provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 18/916,242 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Wong.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of a T-mixer.
Wong relates to a T-mixer as a rapid mixing micromixer [title], and discusses that micromixers are essential components to microfluidic systems because some biochemical analysis require two reagents to be completely mixed before a reaction has proceeded considerably, and can require sub-millisecond mixing times, and therefore micromixers must be capable of mixing two reagents within the shortest time possible [p 359, col 1, paras 1-2 and col 2, paras 1-2].
Regarding instant claim 23, Wong discloses a T-mixer that can sufficiently cause complete mixing of two solutions within less than a millisecond after the two solutions make contact [abstract].
In view of Wong, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger by a T-mixer to contact the SEC eluate with denaturing solution, as disclosed by Wong, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Wong discloses micromixers such as T-mixers are essential components of microfluidic systems in order to insure reagents are completely mixed in sub-millisecond times. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Wong both relate to methods requiring the efficient mixing of biochemical reagents for downstream analysis of the product.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claims 27-30 are provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 18/916,242 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Ehkirch.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of a flow splitter.
Ehkirch relates to an online four-dimensional HIC x SEC-IM x MS methodology for characterization of antibody drug conjugates [title] and describes an analytical system that combines multiple chromatography separation methods with MS analysis for a comprehensive and streamlined characterization of native and forced degraded sample conformations [abstract].
Regarding instant claim 27, Ehkirch discloses the multidimensional analysis method above comprising a first chromatographic dimension, a second chromatographic dimension which is SEC, and a flow splitter connecting SEC to MS which divides the flow rate by a factor of 7 prior to entering MS [p 1581, col 1, paras 2-3; Figure 2].
In view of Ehkirch, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger using a flow splitter between the SEC and MS, as disclosed by Ehkirch, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Ehkirch discloses a multidimensional chromatography-MS analytical method for antibody drug conjugate characterization comprising a flow splitter that provides streamlined characterization of native and forced degraded sample conformations. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Ehkirch relate to methods of analyzing samples comprising antibodies via MS.
Regarding instant claim 28, Haberger discloses the use of a detector to observe absorbance at 280 nm [Figure 1], which is considered to correspond to a UV detector.
Regarding instant claim 29, Ehkirch discloses the use of a flow splitter to divide the flow rate by a factor of 7 before entering MS but downstream of a detector [p 1581, col 1, paras 2-3; Figure 2], therefore indicating the flow to the MS to be a low flow relative to the high flow in the detector.
Regarding instant claim 30, Ehkirch discloses reducing the flow rate by a factor of 7 from an inlet flow rate of 100 µL/min [p 1581, col 1, para 3], which is considered to correspond to a low flow of less than 20 µL/min.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claims 36-38 are provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 18/916,242 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Schaller.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the production of a pharmacokinetic profile.
Schaller relates to pharmacokinetic analysis of a bispecific antibody in plasma and whole blood using MS [title], and discusses that drug development of biologics such as bispecific antibodies requires formidable characterization alongside preclinical efficacy [p 3, para 1].
Regarding instant claim 36, Schaller discloses the administration of a drug comprising a protein of interest to a subject (EGFRvIII:CD3 bi-scFv), and the collection of blood at multiple timepoints for MS analysis of the levels of protein of interest in the samples to generate a pharmacokinetic profile [Graphical Abstract, and p 3, para 2].
In view of Schaller, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger generating a pharmacokinetic profile of the protein of interest administered to a subject over time, as disclosed by Schaller, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Schaller discloses drug development of biologics such as bispecific antibodies requires formidable characterization alongside preclinical efficacy such as pharmacokinetic profiles. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Schaller relate to methods of analyzing samples comprising antibodies via MS.
Regarding instant claim 37, the term “biotransformation product” is not specifically defined by the instant specification, but [para 0069] gives examples of a biotransformation product comprising a post-translational modification, truncation, aggregate, fragment, degradation product, or combination thereof, and [para 0136] states that biotransformation products may have modified properties compared to a pre-administration therapeutic. As Li and Schaller disclose methods of detecting fragments of captured antibodies [p 8, final paragraph of Schaller; p 1267, col 1, para 1 of Li], the combined method of Li, Haberger and Schaller is considered to correspond to the detection of a biotransformation product.
Regarding instant claim 38, Schaller discloses the method can be used to simultaneously detect free and cell-bound drug in order to infer that decay of protein was not simply to do binding of blood cells [p 11, para 1], and as the antibody of Schaller targeted two receptors, the detection of cell-bound drug is considered to correspond the detection of an interacting protein that interacts with a protein of interest.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
E. Claims 1-12, 16-21 and 24-26 are provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 19/042,366 (herein “reference application”) in view of Li and Haberger.
Regarding instant claim 1, claim 1 of the reference application recites a method for identifying a multimer of interest from an antibody in a sample comprising separating the sample by SEC and analyzing the separated sample by MS.
The claim of the reference application does not recite contacting a sample including a protein of interest to a solid surface to form an immobilized protein of interest; eluting the immobilized protein of interest of a fragment thereof to form an enriched protein of interest of fragment thereof; subjecting the enriched protein of interest or a fragment thereof to SEC under native conditions to form an SEC eluate; and contacting the SEC eluate to a denaturing solution to form a denatured eluate.
Li relates to general LC-MS/MS to quantify therapeutic monoclonal antibodies (mAbs) [title].
Regarding instant claim 1, Li discloses a method of quantifying mAbs using tandem liquid chromatographic mass spectrometry (LC-MS/MS) [abstract] and that LC-MS/MS methods can be applied to mAbs for the bioanalysis of preclinical samples, wherein mAbs are enzymatically digested to generate a signature peptide for MS quantification [p 1267, col 1, para 2]. Li further discloses multidimensional chromatography can be used to separate mAb signature peptides from the peptides of the endogenous matrix at the expense of time, and that the use of immunocapture to clean up and concentrate a sample overcome such drawbacks and enhance sensitivity [p 1267, col 1, para 2].
Specifically, Li discloses the addition of the biotinylated human Fc mAb clone 35 (b-Ab35) to streptavidin magnetic beads to produce b-Ab35-coated magnetic beads, followed by the addition of a sample containing alpha-DA-G2, wherein the sample and beads were incubated for 1 h at ambient temperature, washed with 0.01% Tween 20 in PBS, and eluted with a methanol/formic acid mixture, and the resulting supernatant was denatured in the presence of a buffer comprising 8M urea and 20 mM DTT before being subjected to LC-MS/MS analysis [p 1268, col 1, final paragraph; col 2, final paragraph to p 1269, col 1, para 1]. These discloses of Li correspond to steps (i), (ii) and (v), and the subjecting of eluate to a denaturing solution of step (iv) of claim 1.
Haberger relates to characterization of biotherapeutic proteins by SEC coupled to native MS [title], and discusses that SEC is the method of choice for routine product testing to detect and quantify high-molecular aggregates and to determine the content of low molecular forms such as fragments, but does not enable determination of molecular mass of an analyte [p 331, col 1, beginning at final paragraph].
Regarding instant claim 1, Haberger discloses a method of analyzing CrossMAb samples that are in-house bispecific antibodies [p 331, col 2, final paragraph] comprising SEC of CrossMAb samples with a mobile phase of KH2PO4 and KCl and pH 7 [p 338, col 2, para 2], followed by analysis of the eluate via ESI-UV/MS [Figure 5], wherein the SEC protocol was able to resolve the CrossMAb size variants [p 336, col 1, final paragraph, and Figure 5]. The disclosures of Haberger are considered to correspond to subjecting a sample to SEC under native conditions to form an SEC eluate as recited in part (iii) of claim 1.
In view of Li and Haberger, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the claims of the reference application by using SEC on the sample before MS analysis, as disclosed by Li and Haberger, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the claims of the patent because Li discloses the a method for identifying antibodies in a sample via affinity chromatography and MS, Haberger discloses SEC can detect and quantify high-molecular aggregates and to determine the content of low molecular forms such as fragments, and Haberger discloses a method resolves size variants of mAbs before MS analysis. One of ordinary skill in the art would have had a reasonable expectation of success because the reference application, Li and Haberger relate to methods of identifying proteins via MS.
Regarding instant claim 2, Li discloses the method to analyze alpha-DA-G2 as described above, which is an isotype of the human antidinitrophenol IgG2 mAb [p 1268, col 1, para 2], which is considered to correspond to a drug product.
Regarding instant claim 3, Li discloses the method was carried out on alpha-DA-G2 prepared in cynomolgus monkey serum [p 1271, col 2, final paragraph].
Regarding instant claims 4-5, Li discloses the method carried out on the mAb alpha-DA-G2 as discussed above [p 1268, col 1, final paragraph; col 2, final paragraph to p 1269, col 1, para 1], which is an antibody. As the instant specification does not specifically define a therapeutic antibody, but states in [para 0140] “an anti-human Fc antibody will preferentially bind to the Fc domain of any human antibody, such as, for example, a therapeutic antibody”, and the method of Li uses a biotinylated human Fc mAb clone 35 (b-Ab35) to pull down and analyze the mAb alpha-DA-G2, the alpha-DA-G2 of Li is considered to correspond to a therapeutic antibody.
Regarding instant claims 6-9, the solid surface of the method of Li consists of streptavidin magnetic beads coated with biotinylated human Fc mAb clone 35 (b-Ab35) as discussed in the rejection of claim 1 above.
Regarding instant claim 10, the method of Li comprises a washing step as discussed in the rejection of claim 1 above.
Regarding instant claims 11-12, Li discloses LC-MS/MS methods can be applied to mAbs for the bioanalysis of preclinical samples, wherein mAbs are enzymatically digested, usually with trypsin, to generate a signature peptide for MS quantification [p 1267, col 1, para 2].
Regarding instant claim 16, Haberger discloses the use of an SEC system directly coupled to a mass spectrometer [p 338, col 1, para 3].
Regarding instant claims 17-18, Haberger discloses the use of potassium phosphate as a the SEC mobile phase as discussed above in the rejection of claim 1, and also discloses the use of 25-100 mM ammonium acetate buffer as the SEC mobile phase [p 333, col 2, para 1], and MPEP 2144.05.I states in the case where the claimed ranges overlap or lie inside ranges disclosed by the prior art, a prima facie case of obviousness exists.
Regarding instant claims 19-20, Li discloses the analysis of captured fragments via LC-MS/MS with a mobile phase of 0.1% formic acid in acetonitrile/water, wherein the ratios of acetonitrile/water in mobile phases A and B are 5/95 and 95/5 respectively, and a gradient profile was carried out from 0% to 95% mobile phase B over time [p 1268, col 2, para 3], which is considered to correspond to a denaturing solution from 40% to 80% acetonitrile.
Regarding instant claim 21, Li discloses the use of 3% formic acid on samples to elute from the magnetic beads followed by the transfer of eluate in supernatant to a well plate to dry [p 1269, col 1, para 1]. As the supernatant is understood to contain the eluting solution of 3% formic acid and the eluate, the disclosures of Li are considered to correspond to contacting eluate with a denaturing solution comprising 3% formic acid.
Regarding instant claim 24, Haberger discloses an SEC flow rate of 0.2 ml/min [p 338, col 1, para 3].
Regarding instant claim 25, Haberger discloses an SEC flow rate of 0.2 ml/min [p 338, col 1, para 3] that is coupled directly to MS analysis while Li discloses MS analysis with a mobile phase of 0.1% formic acid in acetonitrile/water that is considered a denaturing solution with a flow rate of 0.6 ml/min [p 1268, col 2, para 3]. It would be obvious to one of ordinary skill in the art in practicing the combined method of Haberger and Li to match the flow rates of the coupled protocols of SEC and MS analysis and maintain a mobile phase of 0.1% formic acid in acetonitrile/water with a flowrate of 0.2 ml/min, as MPEP 2144.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation.
Regarding instant claim 26, Haberger discloses the use of an electrospray ionization mass spectrometer (ESI/MS) [p 338, col 1, para 3].
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claims 13 and 15 are provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 19/042,366 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of PR.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the enzyme IdeS or a variant thereof.
PR relates to IdeS as an immunoglobulin-degrading cysteine proteinase of Streptococcus pyogenes [title].
Regarding instant claim 13, PR discloses S. pyogenes expresses proteolytic activity towards IgG via the activity of the cysteine proteinase enzyme IdeS, which cleaves the heavy chain of IgG to form two stable Fab fragments and one Fc fragment [p 52, col 2, paras 2-3].
In view of PR, it would have been obvious to one of ordinary skill in the art before the effective filing date to modify the combined method of the reference application, Li and Haberger by using IdeS, since the simple substitution of one known element for another results in a predictable result. One of ordinary skill in the art would have recognized that both trypsin and IdeS are digestive enzymes, and as such both are capable of being incorporated into such methods compositions as described by Li. Thus it would have been obvious to one of ordinary skill in the art to replace trypsin with IdeS, as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because Li describes a method of capturing and analyzing mAbs using the digestive enzyme trypsin, and PR describes a digestive enzyme specific for IgGs.
Regarding instant claim 15, as the combined method of the patent, Li, Haberger and PR uses the IdeS enzyme to digest an immobilized mAb as described above, and PR discloses the activity of IdeS produces Fab and Fc fragments [p 52, col 2, para 3], the combined disclosures of Li, Haberger and PR are considered to satisfy the limitations of claim 15.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 14 is provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 19/042,366 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Rosli.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of centrifugation during elution.
Rosli relates to the recovery of biotinylated proteins from streptavidin-based affinity chromatography resins [title], and discusses the release of biotinylated proteins from streptavidin based reagents is a major problem due to the stability of the complex [abstract].
Regarding instant claim 14, Rosli discloses a protocol for the release of biotinylated proteins from streptavidin Sepharose comprising the use of a centrifuge [p 91, para 2 and p 94, Section 3.3].
In view of Rosli, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger by using centrifugation in the elution step, as disclosed by Rosli, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Rosli discloses the release of biotinylated proteins from streptavidin is a major problem in streptavidin affinity methods and therefore discloses a quantitative release strategy using centrifugation. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Rosli relate to methods comprising streptavidin affinity capture and release.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 22 is provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 19/042,366 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Donnelly.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite a denaturing solution comprising about 60% acetonitrile and about 4% formic acid.
Donnelly reviews best practices and benchmarks for intact protein analysis for top-down MS [title], and discusses robust protocols for mass analysis of intact proteins such as antibodies from mixtures of varying complexity [abstract].
Regarding instant claim 22, Donnelly discloses denaturing direct-infusion MS comprising the treatment of samples to 49.95% acetonitrile, 49.95% water and 0.1% formic acid, wherein a 60:35:5 ratio of HPLC grade methanol:water:acetic acid may be used as an alternative resulting in improved signal to noise ratios, and discusses how the use of organic solvents and acids results in efficient ionization [p 591, col 1, para 3]. Therefore Donnelly discloses the result effective variable of organic solvent and acid ratio according to MPEP 2144.05.II.B which effects signal to noise ratio in MS, and thereby quality of the MS analysis. While Donnelly does not disclose the 60% acetonitrile and 4% formic acid recited in the claim, MPEP 2144.05.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. Therefore one of ordinary skill in the art would be motivated to modify the combined method of Li, Haberger and Donnelly to get to the claimed 60% acetonitrile and 4% formic acid because Donnelly discloses the result effective variable of organic solvent and acid ratios, and gives an example of using an increased amount of organic solvent and acid can result in improved signal to noise ratio and therefore improved MS analysis.
In view of Donnelly, it would have been obvious to one of ordinary skill in the art before the effective filing date to modify the combined method of the reference application, Li and Haberger by using a denaturing solution of 60% acetonitrile and 4% formic acid because MPEP 2144.05.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Donnelly discloses organic solvent and acid ratios are a result effective variable according to MPEP 2144.05.II.B which effects signal to noise ratio in MS, and thereby quality of the MS analysis. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Donnelly relate to methods of MS analysis comprising treatment of protein samples with acetonitrile and formic acid.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 23 is provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 19/042,366 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Wong.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of a T-mixer.
Wong relates to a T-mixer as a rapid mixing micromixer [title], and discusses that micromixers are essential components to microfluidic systems because some biochemical analysis require two reagents to be completely mixed before a reaction has proceeded considerably, and can require sub-millisecond mixing times, and therefore micromixers must be capable of mixing two reagents within the shortest time possible [p 359, col 1, paras 1-2 and col 2, paras 1-2].
Regarding instant claim 23, Wong discloses a T-mixer that can sufficiently cause complete mixing of two solutions within less than a millisecond after the two solutions make contact [abstract].
In view of Wong, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger by a T-mixer to contact the SEC eluate with denaturing solution, as disclosed by Wong, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Wong discloses micromixers such as T-mixers are essential components of microfluidic systems in order to insure reagents are completely mixed in sub-millisecond times. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Wong both relate to methods requiring the efficient mixing of biochemical reagents for downstream analysis of the product.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claims 27-30 are provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 19/042,366 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Ehkirch.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of a flow splitter.
Ehkirch relates to an online four-dimensional HIC x SEC-IM x MS methodology for characterization of antibody drug conjugates [title] and describes an analytical system that combines multiple chromatography separation methods with MS analysis for a comprehensive and streamlined characterization of native and forced degraded sample conformations [abstract].
Regarding instant claim 27, Ehkirch discloses the multidimensional analysis method above comprising a first chromatographic dimension, a second chromatographic dimension which is SEC, and a flow splitter connecting SEC to MS which divides the flow rate by a factor of 7 prior to entering MS [p 1581, col 1, paras 2-3; Figure 2].
In view of Ehkirch, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger using a flow splitter between the SEC and MS, as disclosed by Ehkirch, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Ehkirch discloses a multidimensional chromatography-MS analytical method for antibody drug conjugate characterization comprising a flow splitter that provides streamlined characterization of native and forced degraded sample conformations. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Ehkirch relate to methods of analyzing samples comprising antibodies via MS.
Regarding instant claim 28, Haberger discloses the use of a detector to observe absorbance at 280 nm [Figure 1], which is considered to correspond to a UV detector.
Regarding instant claim 29, Ehkirch discloses the use of a flow splitter to divide the flow rate by a factor of 7 before entering MS but downstream of a detector [p 1581, col 1, paras 2-3; Figure 2], therefore indicating the flow to the MS to be a low flow relative to the high flow in the detector.
Regarding instant claim 30, Ehkirch discloses reducing the flow rate by a factor of 7 from an inlet flow rate of 100 µL/min [p 1581, col 1, para 3], which is considered to correspond to a low flow of less than 20 µL/min.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claims 36-38 are provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 19/042,366 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Schaller.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the production of a pharmacokinetic profile.
Schaller relates to pharmacokinetic analysis of a bispecific antibody in plasma and whole blood using MS [title], and discusses that drug development of biologics such as bispecific antibodies requires formidable characterization alongside preclinical efficacy [p 3, para 1].
Regarding instant claim 36, Schaller discloses the administration of a drug comprising a protein of interest to a subject (EGFRvIII:CD3 bi-scFv), and the collection of blood at multiple timepoints for MS analysis of the levels of protein of interest in the samples to generate a pharmacokinetic profile [Graphical Abstract, and p 3, para 2].
In view of Schaller, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger generating a pharmacokinetic profile of the protein of interest administered to a subject over time, as disclosed by Schaller, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Schaller discloses drug development of biologics such as bispecific antibodies requires formidable characterization alongside preclinical efficacy such as pharmacokinetic profiles. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Schaller relate to methods of analyzing samples comprising antibodies via MS.
Regarding instant claim 37, the term “biotransformation product” is not specifically defined by the instant specification, but [para 0069] gives examples of a biotransformation product comprising a post-translational modification, truncation, aggregate, fragment, degradation product, or combination thereof, and [para 0136] states that biotransformation products may have modified properties compared to a pre-administration therapeutic. As Li and Schaller disclose methods of detecting fragments of captured antibodies [p 8, final paragraph of Schaller; p 1267, col 1, para 1 of Li], the combined method of Li, Haberger and Schaller is considered to correspond to the detection of a biotransformation product.
Regarding instant claim 38, Schaller discloses the method can be used to simultaneously detect free and cell-bound drug in order to infer that decay of protein was not simply to do binding of blood cells [p 11, para 1], and as the antibody of Schaller targeted two receptors, the detection of cell-bound drug is considered to correspond the detection of an interacting protein that interacts with a protein of interest.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
F. Claims 1-12, 16-21 and 24-26 are provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 17/863,332 (herein “reference application”) in view of Li and Haberger.
Regarding instant claim 1, claim 1 of the reference application recites a method for characterizing an antibody in a sample comprising immobilizing the antibody on solid-phase substrate, digesting the antibody, eluting the digested fragment, and subjecting the eluate to strong cation-exchange mass spectrometry.
The claim of the reference application does not recite subjecting the enriched protein of interest or a fragment thereof to SEC under native conditions to form an SEC eluate; and contacting the SEC eluate to a denaturing solution to form a denatured eluate.
Li relates to general LC-MS/MS to quantify therapeutic monoclonal antibodies (mAbs) [title].
Regarding instant claim 1, Li discloses a method of quantifying mAbs using tandem liquid chromatographic mass spectrometry (LC-MS/MS) [abstract] and that LC-MS/MS methods can be applied to mAbs for the bioanalysis of preclinical samples, wherein mAbs are enzymatically digested to generate a signature peptide for MS quantification [p 1267, col 1, para 2]. Li further discloses multidimensional chromatography can be used to separate mAb signature peptides from the peptides of the endogenous matrix at the expense of time, and that the use of immunocapture to clean up and concentrate a sample overcome such drawbacks and enhance sensitivity [p 1267, col 1, para 2].
Specifically, Li discloses the addition of the biotinylated human Fc mAb clone 35 (b-Ab35) to streptavidin magnetic beads to produce b-Ab35-coated magnetic beads, followed by the addition of a sample containing alpha-DA-G2, wherein the sample and beads were incubated for 1 h at ambient temperature, washed with 0.01% Tween 20 in PBS, and eluted with a methanol/formic acid mixture, and the resulting supernatant was denatured in the presence of a buffer comprising 8M urea and 20 mM DTT before being subjected to LC-MS/MS analysis [p 1268, col 1, final paragraph; col 2, final paragraph to p 1269, col 1, para 1]. These discloses of Li correspond to steps (i), (ii) and (v), and the subjecting of eluate to a denaturing solution of step (iv) of claim 1.
Haberger relates to characterization of biotherapeutic proteins by SEC coupled to native MS [title], and discusses that SEC is the method of choice for routine product testing to detect and quantify high-molecular aggregates and to determine the content of low molecular forms such as fragments, but does not enable determination of molecular mass of an analyte [p 331, col 1, beginning at final paragraph].
Regarding instant claim 1, Haberger discloses a method of analyzing CrossMAb samples that are in-house bispecific antibodies [p 331, col 2, final paragraph] comprising SEC of CrossMAb samples with a mobile phase of KH2PO4 and KCl and pH 7 [p 338, col 2, para 2], followed by analysis of the eluate via ESI-UV/MS [Figure 5], wherein the SEC protocol was able to resolve the CrossMAb size variants [p 336, col 1, final paragraph, and Figure 5]. The disclosures of Haberger are considered to correspond to subjecting a sample to SEC under native conditions to form an SEC eluate as recited in part (iii) of claim 1.
In view of Li and Haberger, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the claims of the reference application by using SEC on the sample before MS analysis, as disclosed by Li and Haberger, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the claims of the patent because Li discloses the a method for identifying antibodies in a sample via affinity chromatography and MS, Haberger discloses SEC can detect and quantify high-molecular aggregates and to determine the content of low molecular forms such as fragments, and Haberger discloses a method resolves size variants of mAbs before MS analysis. One of ordinary skill in the art would have had a reasonable expectation of success because the reference application, Li and Haberger relate to methods of identifying proteins via MS.
Regarding instant claim 2, Li discloses the method to analyze alpha-DA-G2 as described above, which is an isotype of the human antidinitrophenol IgG2 mAb [p 1268, col 1, para 2], which is considered to correspond to a drug product.
Regarding instant claim 3, Li discloses the method was carried out on alpha-DA-G2 prepared in cynomolgus monkey serum [p 1271, col 2, final paragraph].
Regarding instant claims 4-5, Li discloses the method carried out on the mAb alpha-DA-G2 as discussed above [p 1268, col 1, final paragraph; col 2, final paragraph to p 1269, col 1, para 1], which is an antibody. As the instant specification does not specifically define a therapeutic antibody, but states in [para 0140] “an anti-human Fc antibody will preferentially bind to the Fc domain of any human antibody, such as, for example, a therapeutic antibody”, and the method of Li uses a biotinylated human Fc mAb clone 35 (b-Ab35) to pull down and analyze the mAb alpha-DA-G2, the alpha-DA-G2 of Li is considered to correspond to a therapeutic antibody.
Regarding instant claims 6-9, the solid surface of the method of Li consists of streptavidin magnetic beads coated with biotinylated human Fc mAb clone 35 (b-Ab35) as discussed in the rejection of claim 1 above.
Regarding instant claim 10, the method of Li comprises a washing step as discussed in the rejection of claim 1 above.
Regarding instant claims 11-12, Li discloses LC-MS/MS methods can be applied to mAbs for the bioanalysis of preclinical samples, wherein mAbs are enzymatically digested, usually with trypsin, to generate a signature peptide for MS quantification [p 1267, col 1, para 2].
Regarding instant claim 16, Haberger discloses the use of an SEC system directly coupled to a mass spectrometer [p 338, col 1, para 3].
Regarding instant claims 17-18, Haberger discloses the use of potassium phosphate as a the SEC mobile phase as discussed above in the rejection of claim 1, and also discloses the use of 25-100 mM ammonium acetate buffer as the SEC mobile phase [p 333, col 2, para 1], and MPEP 2144.05.I states in the case where the claimed ranges overlap or lie inside ranges disclosed by the prior art, a prima facie case of obviousness exists.
Regarding instant claims 19-20, Li discloses the analysis of captured fragments via LC-MS/MS with a mobile phase of 0.1% formic acid in acetonitrile/water, wherein the ratios of acetonitrile/water in mobile phases A and B are 5/95 and 95/5 respectively, and a gradient profile was carried out from 0% to 95% mobile phase B over time [p 1268, col 2, para 3], which is considered to correspond to a denaturing solution from 40% to 80% acetonitrile.
Regarding instant claim 21, Li discloses the use of 3% formic acid on samples to elute from the magnetic beads followed by the transfer of eluate in supernatant to a well plate to dry [p 1269, col 1, para 1]. As the supernatant is understood to contain the eluting solution of 3% formic acid and the eluate, the disclosures of Li are considered to correspond to contacting eluate with a denaturing solution comprising 3% formic acid.
Regarding instant claim 24, Haberger discloses an SEC flow rate of 0.2 ml/min [p 338, col 1, para 3].
Regarding instant claim 25, Haberger discloses an SEC flow rate of 0.2 ml/min [p 338, col 1, para 3] that is coupled directly to MS analysis while Li discloses MS analysis with a mobile phase of 0.1% formic acid in acetonitrile/water that is considered a denaturing solution with a flow rate of 0.6 ml/min [p 1268, col 2, para 3]. It would be obvious to one of ordinary skill in the art in practicing the combined method of Haberger and Li to match the flow rates of the coupled protocols of SEC and MS analysis and maintain a mobile phase of 0.1% formic acid in acetonitrile/water with a flowrate of 0.2 ml/min, as MPEP 2144.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation.
Regarding instant claim 26, Haberger discloses the use of an electrospray ionization mass spectrometer (ESI/MS) [p 338, col 1, para 3].
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claims 13 and 15 are provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 17/863,332 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of PR.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the enzyme IdeS or a variant thereof.
PR relates to IdeS as an immunoglobulin-degrading cysteine proteinase of Streptococcus pyogenes [title].
Regarding instant claim 13, PR discloses S. pyogenes expresses proteolytic activity towards IgG via the activity of the cysteine proteinase enzyme IdeS, which cleaves the heavy chain of IgG to form two stable Fab fragments and one Fc fragment [p 52, col 2, paras 2-3].
In view of PR, it would have been obvious to one of ordinary skill in the art before the effective filing date to modify the combined method of the reference application, Li and Haberger by using IdeS, since the simple substitution of one known element for another results in a predictable result. One of ordinary skill in the art would have recognized that both trypsin and IdeS are digestive enzymes, and as such both are capable of being incorporated into such methods compositions as described by Li. Thus it would have been obvious to one of ordinary skill in the art to replace trypsin with IdeS, as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because Li describes a method of capturing and analyzing mAbs using the digestive enzyme trypsin, and PR describes a digestive enzyme specific for IgGs.
Regarding instant claim 15, as the combined method of the patent, Li, Haberger and PR uses the IdeS enzyme to digest an immobilized mAb as described above, and PR discloses the activity of IdeS produces Fab and Fc fragments [p 52, col 2, para 3], the combined disclosures of Li, Haberger and PR are considered to satisfy the limitations of claim 15.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 14 is provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 17/863,332 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Rosli.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of centrifugation during elution.
Rosli relates to the recovery of biotinylated proteins from streptavidin-based affinity chromatography resins [title], and discusses the release of biotinylated proteins from streptavidin based reagents is a major problem due to the stability of the complex [abstract].
Regarding instant claim 14, Rosli discloses a protocol for the release of biotinylated proteins from streptavidin Sepharose comprising the use of a centrifuge [p 91, para 2 and p 94, Section 3.3].
In view of Rosli, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger by using centrifugation in the elution step, as disclosed by Rosli, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Rosli discloses the release of biotinylated proteins from streptavidin is a major problem in streptavidin affinity methods and therefore discloses a quantitative release strategy using centrifugation. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Rosli relate to methods comprising streptavidin affinity capture and release.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 22 is provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 17/863,332 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Donnelly.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite a denaturing solution comprising about 60% acetonitrile and about 4% formic acid.
Donnelly reviews best practices and benchmarks for intact protein analysis for top-down MS [title], and discusses robust protocols for mass analysis of intact proteins such as antibodies from mixtures of varying complexity [abstract].
Regarding instant claim 22, Donnelly discloses denaturing direct-infusion MS comprising the treatment of samples to 49.95% acetonitrile, 49.95% water and 0.1% formic acid, wherein a 60:35:5 ratio of HPLC grade methanol:water:acetic acid may be used as an alternative resulting in improved signal to noise ratios, and discusses how the use of organic solvents and acids results in efficient ionization [p 591, col 1, para 3]. Therefore Donnelly discloses the result effective variable of organic solvent and acid ratio according to MPEP 2144.05.II.B which effects signal to noise ratio in MS, and thereby quality of the MS analysis. While Donnelly does not disclose the 60% acetonitrile and 4% formic acid recited in the claim, MPEP 2144.05.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. Therefore one of ordinary skill in the art would be motivated to modify the combined method of Li, Haberger and Donnelly to get to the claimed 60% acetonitrile and 4% formic acid because Donnelly discloses the result effective variable of organic solvent and acid ratios, and gives an example of using an increased amount of organic solvent and acid can result in improved signal to noise ratio and therefore improved MS analysis.
In view of Donnelly, it would have been obvious to one of ordinary skill in the art before the effective filing date to modify the combined method of the reference application, Li and Haberger by using a denaturing solution of 60% acetonitrile and 4% formic acid because MPEP 2144.05.II.A states where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Donnelly discloses organic solvent and acid ratios are a result effective variable according to MPEP 2144.05.II.B which effects signal to noise ratio in MS, and thereby quality of the MS analysis. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Donnelly relate to methods of MS analysis comprising treatment of protein samples with acetonitrile and formic acid.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 23 is provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 17/863,332 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Wong.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of a T-mixer.
Wong relates to a T-mixer as a rapid mixing micromixer [title], and discusses that micromixers are essential components to microfluidic systems because some biochemical analysis require two reagents to be completely mixed before a reaction has proceeded considerably, and can require sub-millisecond mixing times, and therefore micromixers must be capable of mixing two reagents within the shortest time possible [p 359, col 1, paras 1-2 and col 2, paras 1-2].
Regarding instant claim 23, Wong discloses a T-mixer that can sufficiently cause complete mixing of two solutions within less than a millisecond after the two solutions make contact [abstract].
In view of Wong, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger by a T-mixer to contact the SEC eluate with denaturing solution, as disclosed by Wong, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Wong discloses micromixers such as T-mixers are essential components of microfluidic systems in order to insure reagents are completely mixed in sub-millisecond times. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Wong both relate to methods requiring the efficient mixing of biochemical reagents for downstream analysis of the product.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claims 27-30 are provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 17/863,332 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Ehkirch.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the use of a flow splitter.
Ehkirch relates to an online four-dimensional HIC x SEC-IM x MS methodology for characterization of antibody drug conjugates [title] and describes an analytical system that combines multiple chromatography separation methods with MS analysis for a comprehensive and streamlined characterization of native and forced degraded sample conformations [abstract].
Regarding instant claim 27, Ehkirch discloses the multidimensional analysis method above comprising a first chromatographic dimension, a second chromatographic dimension which is SEC, and a flow splitter connecting SEC to MS which divides the flow rate by a factor of 7 prior to entering MS [p 1581, col 1, paras 2-3; Figure 2].
In view of Ehkirch, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger using a flow splitter between the SEC and MS, as disclosed by Ehkirch, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Ehkirch discloses a multidimensional chromatography-MS analytical method for antibody drug conjugate characterization comprising a flow splitter that provides streamlined characterization of native and forced degraded sample conformations. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Ehkirch relate to methods of analyzing samples comprising antibodies via MS.
Regarding instant claim 28, Haberger discloses the use of a detector to observe absorbance at 280 nm [Figure 1], which is considered to correspond to a UV detector.
Regarding instant claim 29, Ehkirch discloses the use of a flow splitter to divide the flow rate by a factor of 7 before entering MS but downstream of a detector [p 1581, col 1, paras 2-3; Figure 2], therefore indicating the flow to the MS to be a low flow relative to the high flow in the detector.
Regarding instant claim 30, Ehkirch discloses reducing the flow rate by a factor of 7 from an inlet flow rate of 100 µL/min [p 1581, col 1, para 3], which is considered to correspond to a low flow of less than 20 µL/min.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claims 36-38 are provisionally rejected on the ground of nonstatutory double patenting over claim 1 of co-pending Application No. 17/863,332 in view of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 above, and further in view of Schaller.
The claims of the reference application and disclosures of Li and Haberger as applied to claims 1-12, 16-21 and 24-26 are described above. The claims of the patent do not recite the production of a pharmacokinetic profile.
Schaller relates to pharmacokinetic analysis of a bispecific antibody in plasma and whole blood using MS [title], and discusses that drug development of biologics such as bispecific antibodies requires formidable characterization alongside preclinical efficacy [p 3, para 1].
Regarding instant claim 36, Schaller discloses the administration of a drug comprising a protein of interest to a subject (EGFRvIII:CD3 bi-scFv), and the collection of blood at multiple timepoints for MS analysis of the levels of protein of interest in the samples to generate a pharmacokinetic profile [Graphical Abstract, and p 3, para 2].
In view of Schaller, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined method of the reference application, Li and Haberger generating a pharmacokinetic profile of the protein of interest administered to a subject over time, as disclosed by Schaller, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined method of the reference application, Li and Haberger because Schaller discloses drug development of biologics such as bispecific antibodies requires formidable characterization alongside preclinical efficacy such as pharmacokinetic profiles. One of ordinary skill in the art would have had a reasonable expectation of success because Li and Schaller relate to methods of analyzing samples comprising antibodies via MS.
Regarding instant claim 37, the term “biotransformation product” is not specifically defined by the instant specification, but [para 0069] gives examples of a biotransformation product comprising a post-translational modification, truncation, aggregate, fragment, degradation product, or combination thereof, and [para 0136] states that biotransformation products may have modified properties compared to a pre-administration therapeutic. As Li and Schaller disclose methods of detecting fragments of captured antibodies [p 8, final paragraph of Schaller; p 1267, col 1, para 1 of Li], the combined method of Li, Haberger and Schaller is considered to correspond to the detection of a biotransformation product.
Regarding instant claim 38, Schaller discloses the method can be used to simultaneously detect free and cell-bound drug in order to infer that decay of protein was not simply to do binding of blood cells [p 11, para 1], and as the antibody of Schaller targeted two receptors, the detection of cell-bound drug is considered to correspond the detection of an interacting protein that interacts with a protein of interest.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Conclusion
Status of the Application:
Claims 1-30, 32, and 34-40 are pending.
Claims 32, 34-35 and 39-40 are withdrawn.
Claims 1-30 and 36-38 are rejected.
No claim is in condition for allowance.
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/JOSEPH R SPANGLER/
Examiner
Art Unit 1656
/David Steadman/Primary Examiner, Art Unit 1656
Prosecution Insights