`DETAILED ACTION
Notice of Pre-AIA or AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
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.
Status of the Claims
Claims 3-11 have been amended. Claims 12, 14, 20, and 22 have been canceled. Claims 1-11, 13, 15-19, 21, and 23 are pending and examined herein.
Priority
This application, 18/715,567, filed 05/31/2024, is a 371 of PCT/EP22/84435 filed on 12/05/2022, and claims benefit of UNITED KINGDOM OF GREAT BRITAIN AND NORTHERN IRELAND application GB2117557.5 filed on 12/03/2021. This priority is acknowledged and the claims examined herein are treated as having an effective filing date of 12/03/2021.
Information Disclosure Statement
The Information Disclosure Statements filed on 05/03/2024 are acknowledged and have been considered.
Claim Rejections - 35 USC § 112
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.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claims 19, 21, and 23 are rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention. Claims 19 and 23 recite “..relative to a control, such as the glycoprotein and/or glycan profile of the protein corona from a sample…”, and “…formed around particles, such as silica particles…” respectively. The recitation of “such as” represents exemplary language that renders the claim indefinite because it is unclear whether the limitations following the phrase are part of the claimed invention, or represent a description of examples or preferences which can lead to confusion over the intended scope of the claim. See MPEP § 2173.05(d).
Claim 21 recites “…the method comprising the detection of one or more lung cancer biomarkers in a sample from a subject, wherein the biomarkers are selected from fibrinogen- derived glycan FA2G2S1 and FA3G3S2; wherein a decrease in the level FA2G2S1 and FA3G3S2 is indicative of lung cancer…”. The language recites “one or more” cancer biomarkers but then recites “FA2G2S1 and FA3G3S2”. Based on the claim language it is unclear if the lung cancer diagnosis can be made by selecting and showing a decrease in one of FA2G2S1 or FA3G3S2, or whether a decrease in both biomarkers is necessary to be measured and make the diagnosis. Therefore, the claim is indefinite.
Appropriate correction is required.
Claim Rejections - 35 USC § 101
35 U.S.C. 101 reads as follows:
Whoever invents or discovers any new and useful process, machine, manufacture, or composition of matter, or any new and useful improvement thereof, may obtain a patent therefor, subject to the conditions and requirements of this title.
Claims 16-19 and 21 are rejected under 35 U.S.C. 101 because the claimed invention is directed to a judicial exception without significantly more.
Claims 16 and 18 recite “A method of diagnosis of a disease or condition of a subject, the method comprising the enrichment of glycoproteins…”. Claim 17 recites a “A method of monitoring the progression or remission of a disease or condition in a subject, the method comprising the enrichment of glycoproteins…”. Claim 19 recites “A method of screening for biomarkers of a disease or condition, the method comprising the comparison of the glycoprotein…”. Claim 21 recites “A method of diagnosis for lung cancer, the method comprising the detection of one or more lung cancer biomarkers in a sample from a subject…”.
The claims are directed to judicial exceptions, mainly they are abstract ideas, specifically, mental processes that can be performed in the human mind, and/or are merely observing naturally occurring correlations (laws of nature/natural correlation). These judicial exceptions are not integrated into a practical application because there is no practical application recited in the claims such as performing a treatment in a way that is particular, and not merely instructions to "apply" the exception in a generic way. The claims do not include additional elements that are sufficient to amount to significantly more than the judicial exception because the additional steps amount to mere data gathering that does not go beyond well-understood, routine, and conventional activity; as detailed below.
Step 1 – Whether a claim is to a statutory category - YES
The instantly claimed invention is directed to methods of diagnosing, monitoring, and screening biomarkers for disease, which in some cases is cancer. The methods use enrichment/measurement of glycoproteins in a sample to make the determination. Therefore, the instantly claimed invention falls into one of the four statutory categories.
Step 2A Prong 1 – Whether the claim is directed to a judicial exception (i.e. Does the claim recite an abstract idea, law of nature, or natural phenomenon?) - YES
Claims 16-19 and 21 recite the following steps which fall under mental processes grouping of abstract ideas and/or laws of nature/natural correlation:
Claim 16-19 and 21 disclose methods of diagnosing, monitoring, and screening biomarkers for disease by the enrichment of glycoproteins, then the glycoprotein and/or glycan profile of the protein corona is “determined”, and the profile is used to “indicate” disease presence (claims 16, 18, and 21), disease progression/remission (claim 17), or disease biomarker validity (claim 19). Claim 19 additionally recites the “comparison” of the glycoprotein and/or glycan profile of the protein corona from a sample of one or more subjects afflicted with the disease or condition relative to a control. Claim 21 additionally recites the detection of specific glycoproteins biomarkers (FA2G2S1 and FA3G3S2) to diagnosis lung cancer.
These limitations recite a law of nature which is a judicial exception, because it is merely observing the correlation between a naturally occurring biomarkers (glycoprotein and/or glycan profile of the protein corona from a sample; FA2G2S1 and FA3G3S2) and its relationship to a disease/disease state (disease/cancer/lung cancer), and using these levels to make a determination of a subjects’ disease state, or determination of biomarker validity, which are abstract ideas, specifically, abstract mental processes. Additionally, some claims recite comparison steps. In claim 17, a subsequent subject sample is compared to the first sample to determine progression/remission, and in claim 19, a comparison is made between the sample from a diseased subject and a control. These comparison steps also represent abstract ideas, specifically, abstract mental processes.
The “determined/”indicate” steps can be regarded as a law of nature, namely, the naturally occurring correlation between glycoprotein profiles and disease state. Regarding the identification of a correlation between the presence of a biomarker in a bodily sample and disease state the courts have held similar claims to be laws of nature and/or natural phenomena, as in Cleveland Clinic Foundation v. True Health Diagnostics, LLC, 859 F.3d 1352, 1361, 123 USPQ2d 1081, 1087 (Fed. Cir. 2017) which involved claims to simply instruct a user to apply a natural law, by correlating naturally occurring enzyme levels with disease risk. In Mayo, the Supreme Court found that a claim was directed to a natural law, where the claim required administering a drug and determining the levels of a metabolite following administration, where the level of metabolite was indicative of a need to increase or decrease the dosage of the drug. See Mayo Collaborative Services V. Prometheus Labs., Inc., 566 U.S. 66, 74 (2012). The instant claims are similar to those in Mayo as they involve a "relation itself [which] exists in principle apart from any human action" (id. at 77). The instant claims do not recite administering an immunotherapy.
Regarding the steps reciting a “comparison” of glycoprotein profiles to previous samples or control samples, the courts have held similar claims to be abstract mental processes, as in University of Utah Research Foundation v. Ambry Genetics, 774 F.3d 755, 763, 113 USPQ2d 1241, 1246 (Fed. Cir. 2014) which involved claims to "comparing BRCA sequences and determining the existence of alterations," where the claims cover any way of comparing BRCA sequences such that the comparison steps can practically be performed in the human mind. The claims are also similar to that in Classen Immunotherapies, Inc. v. Biogen IDEC, 659 F.3d 1057, 1067, 100 USPQ2d 1492, 1500 (Fed. Cir. 2011), which involved a claim to “collecting and comparing known information” (both of these court cases are discussed in MPEP 2106.04(a)(2) (II)(A)). The step of “determined/”indicate” also constitutes an abstract mental process, involving assessing the comparison of glycoprotein profiles of a sample, and then making an evaluation or judgment as to the severity/state of a test subjects’ particular disease. The “comparison” and “determined/”indicate” steps could be performed in the human mind, or by a human using pen and paper, insofar as it reads on comparing levels and drawing conclusions from this about the health status of a subject.
Thus, claims 16-19 and 21 fall into a judicial exception.
Step 2A: Prong 2 - Does the claim recite additional elements that integrate the judicial exception into a practical application? The Step 2A, Prong 2 analysis requires identifying whether there are any additional elements recited in the claim beyond the judicial exception(s), and evaluating those additional elements to determine whether they integrate the exception into a practical application of the exception.
Claims 16-19 and 21 do not recite any additional element that integrate the exception into a practical application of the exception. The additional step in claim 18 of administering a treatment for the disease or condition, optionally wherein the treatment is treatment for cancer, is insufficient to integrate the exception into a practical application because the purpose is merely to obtain data and/or merely instructions to "apply" the exception in a generic way. Claim 18 does not recite a particular treatment, and as MPEP 2106.04(d)(2) states, the treatment or prophylaxis limitation must be “particular,” i.e., specifically identified so that it does not encompass all applications of the judicial exception(s), for it to integrate the judicial exception.
As in In re Grams, 888 F.2d 835, 839-40; 12 USPQ2d 1824, 1827-28 (Fed. Cir. 1989), such activity involving performing clinical tests on individuals constitutes mere data gathering, and does not go beyond insignificant extra-solution activity. See MPEP §§ MPEP 2106.04(d)(I) and 2106.05(g). There are no subsequent steps recited after the “determined/indicate” or “comparison” steps that would practically apply the method depending on the results of the measurements, e.g., specific treatment or other process steps that are performed after the test subject has been diagnosed with a disease.
In particular, of the claims indicated in the rejection heading, none of the additionally recited limitations amount to an additional element or combination of elements that apply, rely on, or use the judicial exceptions in a manner that impose meaningful limit on the judicial exceptions.
Step 2B; Whether the additional elements contribute an “inventive concept”. In the second step it is determined whether the claimed subject matter includes additional elements that amount to significantly more than the judicial exception. See MPEP 2106.05.
Briefly, claims 16-19 and 21 do not include additional elements that are sufficient to amount to significantly more than the judicial exception because of the following reasons. Simply appending well-understood, routine, conventional activities previously known to the industry, specified at a high level of generality, to the judicial exception, has been found to be insufficient to add “significantly more” (MPEP 2106.05(I)(A)).
The additional steps recited above do not add a meaningful limitation to the instant method as they would have been routinely used by those of ordinary skill in the art as supported by Soddu et al. (2020). “Identification of physicochemical properties that modulate nanoparticle aggregation in blood”. Beilstein journal of nanotechnology, 11(1), 550-567. (published 03 April 2020), (herein referred to as Soddu), Kurtz-Chalot et al. (2017). “Impact of silica nanoparticle surface chemistry on protein corona formation and consequential interactions with biological cells”. Materials Science and Engineering: C, 75, 16-24, (herein referred to as Kurtz-Chalot), Ruhaak et al. (2013). “Enrichment strategies in glycomics‐based lung cancer biomarker development”. PROTEOMICS–Clinical Applications, 7(9-10), 664-676, (herein referred to as Ruhaak), Hadjidemetriou et al. (2021). “Nanoparticle-enabled enrichment of longitudinal blood proteomic fingerprints in Alzheimer’s disease”. ACS nano, 15(4), 7357, (herein referred to as Hadjidemetriou), Du et al. (US 10,782,297 B2), (herein referred to as Du), and Kavanagh et al. (2021). “N-Linked glycosylation profiles of therapeutic induced senescent (TIS) triple negative breast cancer cells (TNBC) and their extracellular vesicle (EV) progeny”. Molecular Omics, 17(1), 72-85, (herein referred to as Kavanagh).
Soddu teaches a study in which a set of six silica and carbon nanoparticles (NPs) of known size and morphology was used to evaluate the effect of the size and surface properties on the protein corona composition, platelet activation and aggregation (page 552, column 1, 1st paragraph). Soddu teaches that substantial differences were found in the composition of the protein corona depending on the chemical nature of the nanoparticles, while the surface curvature was found to play a minor role (abstract). Soddu teaches that blood plasma was collected from patients and diluted in PBS in order to obtain solutions with increasing protein, then carbon nanoparticles (CNPs) and silica nanoparticles (SNPs) were then added to the solution and were incubated for 1 h at 37 °C under agitation (page 553, column 1, 6th paragraph). Soddu teaches that the corona composition between the two materials has some similarities when incubated at 10% plasma; however, it becomes highly specific to the NP surface properties at higher concentrations as confirmed by the significant difference in the corona composition (page 556, column 2, 1st paragraph).
Kurtz-Chalot teaches a study investigating the in vitro the influence of NP surface functionalization on their cellular uptake and the biological response induced (abstract). Kurtz-Chalot teaches that 50nm fluorescent silica nanoparticle (NP) were functionalized either with amine or carboxylic groups, in presence or in absence of polyethylene glycol (abstract). Kurtz-Chalot teaches that NP were incubated with macrophages, cellular uptake and cellular response were assessed in terms of cytotoxicity, pro inflammatory response and oxidative stress, and that the NP protein corona was also characterized by protein mass spectroscopy (abstract). Kurtz-Chalot teaches that consequently, NP surface functionalization by modifying surface charge is an efficient and easy way to drive cellular uptake, and that the modulation of the NP physico-chemical features is important not only for the enhancement of cellular uptake but also for the induced biological response (and the potential cytotoxicity) (page 16, column 2, 1st paragraph).
Ruhaak teaches two methods for protein enrichment; affinity capturing of IgG and enrichment of medium abundance proteins, thus allowing us to determine which method yields the best candidate glycan biomarkers for lung cancer (abtract-Purpose). Ruhaak teaches the use of whole plasma N-glycan analysis to further investigate the potential of glycans in blood plasma as diagnostic biomarkers for lung cancer (page 672, column 1, 1st full paragraph). Ruhaak teaches that N-glycans isolated from plasma samples from 20 cases of lung adenocarcinoma and 20 matched controls were analyzed using nLC-PGC-chip-TOF, and that N-glycan profiles were obtained for five different fractions: total plasma, isolated IgG, IgG-depleted plasma, and the bound and flow-through fractions of protein enrichment (abstract-Experimental design). Ruhaak teaches that the results confirm that the N-glycan profile in plasma of lung cancer patients is different from healthy controls and appears to be dominated by alterations in relatively abundant proteins (abstract – Conclusions).
Hadjidemetriou teaches a method of employing the nanoparticle protein corona as a tool to systematically monitor changes in the plasma proteome with Alzheimer’s disease (AD) progression and to reveal underpinning molecular mechanisms (page 7359, column 2, 1st full paragraph). In this Hadjidemetriou teaches a method of identifying and tracking longitudinal alterations of the blood proteome in a transgenic mouse model of AD using a nanotechnology-enabled approach (7359, column 1, 5th full paragraph). Hadjidemetriou teaches that their data revealed the existence of multiple proteomic signals in blood, indicative of the asymptomatic stages of AD (abstract). Hadjidemetriou teaches that comprehensive analysis of the nanoparticle-recovered blood proteome by label-free liquid chromatography−tandem mass spectrometry resulted in the discovery of AD-monitoring signatures that could discriminate the asymptomatic phase from amyloidopathy and cognitive deterioration (abstract). Hadjidemetriou teaches the number of differentially abundant proteins discovered at the three time points (2M, 6M, and 12M) by proteomic analysis of (a) plasma control samples and (b) corona samples (Fig. 2).
Du teaches the use of signatures of secretory proteins are used to identify and distinguish lung cancers, and that the biomarker signatures may also be used to separate lung cancers from other inflammatory diseases, monitor progression, or assess treatment efficacy (abstract). Du teaches that the lung cancer-associated biomarkers are proteins or mRNA selected from the group consisting of: chitinase 3-like 1 (CHI3L1); transthyretin (TTR); fibrinogen beta polypeptide (FGb); fibrinogen-like protein 1 (FGL1); guanylate cyclase activator 2A (GUCA2A); delta-like 1 homolog (DLK1); glucose transporter 3 (GLUT3); cerebellin 1 (CBLN1); elastase 1, pancreatic (ELA1); fibrinogen alpha polypeptide (Fga); histidine-rich glycoprotein (HRG) (column 3, lines 10-15). Du also teaches administering chemotherapy to treat lung cancer (column 27, lines 25-27). Du also teaches that treatment with cytotoxic agents is a well-established standard for most advanced cancers and is also increasingly becoming an integrated part of treatment in primary and early cancers (column 9, lines 14-18).
Kavanagh teaches the profiling of the N-linked glycans of whole cells, membrane, cytoplasm and extracellular vesicles (EVs) harvested from therapeutic-induced-senescent (TIS) Triple negative breast cancer (TNBC) cells and to compare these to results from non-senescent cells (abstract). Kavanagh teaches that ultra-performance liquid chromatography (UPLC) analysis of exoglycosidase digested N-linked glycans was carried out on TIS compared to non-treated control cells (abstract). Kavanagh teaches that LC-Mass spectrometry (MS) analysis of the N-linked glycans and lectin blotting of samples was carried out to confirm the UPLC results (abstract). Kavanagh also teaches that the HILIC-UPLC and LC-MS results show an increase in the sialylated glycan structure A2G2S1 (peak35) in TIS EVs compared to control EVs, and that there was also a decrease in the sialylated glycan structures FA2G2S1 (peak33+34) and FA3G3S2 (peak47) in TIS EVs compared to control EVs, possibly reflecting enhanced FUCA1 activity in TIS cells (page 78, column 2, 3rd paragraph). Kavanagh also teaches that highly branched and sialylated glycans are associated with cancer, and that N-Glycan signatures may inform on treatment response and overall patient survival (page 73, column 1, 5th paragraph).
For all of these reasons, the claims fail to include additional elements that are sufficient to amount to significantly more than the judicial exception(s).
Therefore, the instantly rejected claims are not drawn to eligible subject matter as they are directed to a law of nature and abstract idea without significantly more. For additional guidance, applicant is directed generally to MPEP § 2106.
Claim Rejections - 35 USC § 102
The following is a quotation of the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
(a)(2) the claimed invention was described in a patent issued under section 151, or in an application for patent published or deemed published under section 122(b), in which the patent or application, as the case may be, names another inventor and was effectively filed before the effective filing date of the claimed invention.
Claims 1, 3, 4, 6-10, and 15 are rejected under 35 U.S.C. 102(a)(1) as being anticipated Soddu et al. (2020). “Identification of physicochemical properties that modulate nanoparticle aggregation in blood”. Beilstein journal of nanotechnology, 11(1), 550-567. (published 03 April 2020), (herein referred to as Soddu).
Regarding claims 1, Soddu teaches a study in which a set of six silica and carbon nanoparticles (NPs) of known size and morphology was used to evaluate the effect of the size and surface properties on the protein corona composition, platelet activation and aggregation (page 552, column 1, 1st paragraph). Soddu teaches that substantial differences were found in the composition of the protein corona depending on the chemical nature of the nanoparticles, while the surface curvature was found to play a minor role (abstract). Soddu teaches that blood plasma was collected from patients and diluted in PBS in order to obtain solutions with increasing protein, then carbon nanoparticles (CNPs) and silica nanoparticles (SNPs) were then added to the solution and were incubated for 1 h at 37 °C under agitation (page 553, column 1, 6th paragraph). Soddu teaches that the corona composition between the two materials has some similarities when incubated at 10% plasma; however, it becomes highly specific to the NP surface properties at higher concentrations as confirmed by the significant difference in the corona composition (page 556, column 2, 1st paragraph). Soddu also teaches that protein grouping (Figure 7) confirmed that the coagulation factors are highly enriched in the corona across all conditions although with different percentage (55–75%) (page 558, column 2, 1st paragraph). Soddu teaches that ApoB100 and histidine-rich glycoprotein were enriched at 10% plasma, but they were displaced by other proteins such as vitronectin and ITIH4 at higher concentrations of plasma (page 560, column 1, 1st paragraph).
Regarding claim 3, Soddu teaches Top 20 most abundant proteins in small silica (SNP-S) and carbon (CNP-S) nanoparticle hard corona samples at three different plasma concentrations (10, 40, 80%) based on the LFQ intensity (Table 4; Figure 4).
Regarding claims 4 and 6, Soddu teaches the use of silica particles (page 553, column 1, 6th paragraph).
Regarding claim 7, Soddu teaches the use of two sets of tailored carbon and silica nanoparticles with three different diameters in the 100–500 nm range with the purpose of investigating the role of surface curvature and chemistry on platelet aggregation, activation and adhesion (abstract; Table 3).
Regarding claim 8, Soddu teaches that blood plasma was collected from patients and diluted in PBS in order to obtain solutions with increasing protein, then carbon nanoparticles (CNPs) and silica nanoparticles (SNPs) were then added to the solution and were incubated for 1 h at 37 °C under agitation (page 553, column 1, 6th paragraph)
Regarding claims 9 and 10, Soddu teaches that after the NPs incubation with human plasma, the nanoparticle–protein corona complexes were pelleted by centrifugation and re-suspended in PBS three times in order to remove the loosely binding coronas as previously described (page 553, column 2, 1st paragraph). Soddu also teaches that after the last washing step, the pelleted samples were suspended in 20 μL PBS and 10 μL of 3X Blue Loading Buffer Reagents (New England biolabs) that contained DTT in a ratio of 1:10 following the manufacturer instructions (page 553, column 2, 1st paragraph).
Regarding claim 15, Soddu teaches the enrichment/abundance of fibrinogen, apolipoprotein, and histidine-rich glycoproteins in corona samples at three different plasma concentrations (10, 40, 80%) based on the LFQ intensity (Table 4).
Claim Rejections - 35 USC § 103
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.
Claims 2, 11, and 23 are rejected under 35 U.S.C. 103 as being unpatentable over Soddu, as applied to claims 1, 3, 4, 6-10, and 15 above.
The teachings of Soddu are incorporated herein.
Regarding claims 2 and 23, Soddu teaches all the limitations of claim 1 of the instant application but does not teach that the sample protein concentration to total surface area of the particles is 11 to 330 mg/m2 or 660 to 1760 mg/m2.
It would have been obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to have used a protein concentration to total particle surface area ratio of 11 to 330 mg/m2 or 660 to 1760 mg/ m2 in the method of Soddu as a matter of routine optimization. MPEP 2144.05 (II) (A) states that generally, differences in concentration or temperature will not support the patentability of subject matter encompassed by the prior art unless there is evidence indicating such concentration or temperature is critical. “[W]here 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.” In re Aller, 220 F.2d 454, 456, 105 USPQ 233, 235 (CCPA 1955); see also Peterson, 315 F.3d at 1330, 65 USPQ2d at 1382 (“The normal desire of scientists or artisans to improve upon what is already generally known provides the motivation to determine where in a disclosed set of percentage ranges is the optimum combination of percentages.”).
Soddu teaches that they exposed the same surface area (1.0 × 10−2 m2 - page 553, column 1, 6th paragraph) of silica and carbon NPs of three different sizes to an increasing concentration of human plasma, from 10% to 80%, to mimic the in vitro and in vivo conditions, respectively (page 556, column 2, 1st paragraph). Therefore, Soddu teaches the ratio in terms of the plasma percentage and standardized surface area rather than the specific units and ranges disclosed in claim 2. Soddu teaches that the corona composition between the two materials has some similarities when incubated at 10% plasma; however, it becomes highly specific to the NP surface properties at higher concentrations as confirmed by the significant difference in the corona composition (page 556, column 2, 1st paragraph). Soddu teaches that the nanoparticle hard corona samples contain fibrinogen at the highest abundance in lower plasma concentrations, but the levels decreased at higher concentrations and fibrinogen was displaced by proteins such as histidine-rich glycoprotein, kallikrein B, and plasminogen (Fig. 4; Table 4).
Therefore, Soddu demonstrates that the concentration to surface area ratio is a results-effective variable that can change corona composition and be used to enrich certain proteins by modifying the ratio. Furthermore, the specification of the instantly claimed invention fails to that this magnetization protocol recited produces an unexpected results compared to other ranges; see In re Aller, 220 F.2d 454, 456, 105 USPQ 233, 235 (CCPA 1955) (Claimed process which was performed at a temperature between 40°C and 80°C and an acid concentration between 25% and 70% was held to be prima facie obvious over a reference process which differed from the claims only in that the reference process was performed at a temperature of 100°C and an acid concentration of 10%.). Therefore, the selection of such protein concentration to surface area conditions would be a matter of routine optimization within the level of ordinary skill in the art.
Regarding claim 11, Soddu teaches all the limitations of claim 10 of the instant application but does not teach that the isolated protein corona is washed only once.
Rather than a single wash, Soddu teaches that the nanoparticle–protein corona complexes were pelleted by centrifugation and re-suspended in PBS three times in order to remove the loosely binding coronas. However, it would have been obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to wash the isolated protein corona only once, as routine optimization of the method would have discovered the workable ranges and shown that a single wash step is sufficient to obtain the desired corona purity while minimizing loss. Therefore, using a single wash step would be a matter of routine optimization within the level of ordinary skill in the art depending on the sought after purity and yield.
Claim 5 is rejected under 35 U.S.C. 103 as being unpatentable over Soddu as applied to claims 1, 3, 4, 6-10, and 15 above, in view of Kurtz-Chalot et al. (2017). “Impact of silica nanoparticle surface chemistry on protein corona formation and consequential interactions with biological cells”. Materials Science and Engineering: C, 75, 16-24, (herein referred to as Kurtz-Chalot).
The teachings of Soddu are incorporated herein.
Regarding claim 5, Soddu teaches all the limitations of claim 1 of the instant application but does not teach that the particles are coated with functional groups or a polymeric or silica coating.
Kurtz-Chalot teaches a study investigating the in vitro the influence of NP surface functionalization on their cellular uptake and the biological response induced (abstract). Kurtz-Chalot teaches that 50nm fluorescent silica nanoparticle (NP) were functionalized either with amine or carboxylic groups, in presence or in absence of polyethylene glycol (abstract). Kurtz-Chalot teaches that NP were incubated with macrophages, cellular uptake and cellular response were assessed in terms of cytotoxicity, pro inflammatory response and oxidative stress, and that the NP protein corona was also characterized by protein mass spectroscopy (abstract). Kurtz-Chalot teaches that consequently, NP surface functionalization by modifying surface charge is an efficient and easy way to drive cellular uptake, and that the modulation of the NP physico-chemical features is important not only for the enhancement of cellular uptake but also for the induced biological response (and the potential cytotoxicity) (page 16, column 2, 1st paragraph). Kurtz-Chalot teaches that it is thus crucial to define accurate physico-chemical parameters to take into consideration to manufacture NP with a “safer by design” approach (page 16, column 2, 1st paragraph). Kurtz-Chalot teaches that one easy way to increase NP biocompatibility and enhance their cellular uptake is through surface functionalization, and that amorphous silica is a particularly interesting material because the chemistry of the silanes allows surface modification of silica NP with chemical functional groups like specific antibodies (targeting to cancer cells and drug delivery) or fluorescent labels (tumor labeling) (page 16, column 2, 1st paragraph).
It would have been obvious to person of ordinary skill in the art before the effective filing date of the claimed invention to have modified the particle-based glycoprotein enrichment/measurement method taught by Soddu, to coat the particles with functional groups, as taught by Kurtz-Chalot, because Kurtz-Chalot teaches several advantages of NP surface functionalization, namely that such functionalization’s can easily and efficiently drive cellular uptake, as well as allow for the attachment of specific antibodies or labels that can make the NPs more useful for certain applications. A person of ordinary skill would have had a reasonable expectation of success in making these modifications because Soddu and Kurtz-Chalot are in the same field of endeavor, and both use silicon NPs, and both use protein mass spec to characterize NP protein coronas. Additionally, Kurtz-Chalot provides a detailed method of synthesis of such functionalized NPs.
Claims 13, 16, and 19 are rejected under 35 U.S.C. 103 as being unpatentable over Soddu as applied to claims 1, 3, 4, 6-10, and 15 above, in view of Ruhaak et al. (2013). “Enrichment strategies in glycomics‐based lung cancer biomarker development”. PROTEOMICS–Clinical Applications, 7(9-10), 664-676, (herein referred to as Ruhaak).
The teachings of Soddu are incorporated herein.
Regarding claim 13, 16, and 19, Soddu teaches all the limitations of claim 1 of the instant application but does not teach determining the glycan profile of the enriched glycoproteins, or that the glycoprotein and/or glycan profile is indicative of a disease or condition.
Ruhaak teaches two methods for protein enrichment; affinity capturing of IgG and enrichment of medium abundance proteins, thus allowing us to determine which method yields the best candidate glycan biomarkers for lung cancer (abtract-Purpose). Ruhaak teaches the use of whole plasma N-glycan analysis to further investigate the potential of glycans in blood plasma as diagnostic biomarkers for lung cancer (page 672, column 1, 1st full paragraph). Ruhaak teaches that N-glycans isolated from plasma samples from 20 cases of lung adenocarcinoma and 20 matched controls were analyzed using nLC-PGC-chip-TOF, and that N-glycan profiles were obtained for five different fractions: total plasma, isolated IgG, IgG-depleted plasma, and the bound and flow-through fractions of protein enrichment (abstract-Experimental design). Ruhaak teaches that the results confirm that the N-glycan profile in plasma of lung cancer patients is different from healthy controls and appears to be dominated by alterations in relatively abundant proteins (abstract – Conclusions). Ruhaak teaches that there is a need to identify biomarkers for risk assessment, early detection, and disease monitoring for lung cancer; and that biomarkers may also be provided by protein glycosylation patterns, as glycosylation is one of the most prevalent PTMs (page 664, column 2, 1st paragraph). Ruhaak also teaches that there is increased branching of N-glycans in tumor cells, which has the potential to yield a glycan signature for malignancy and metastasis (page 665, column 1, 1st paragraph).
It would have been obvious to person of ordinary skill in the art before the effective filing date of the claimed invention to have modified the particle-based glycoprotein enrichment/measurement method taught by Soddu, to determine the glycan profile of the enriched glycoprotein, as taught by Ruhaak, to provide a plasma biomarker for lung cancer patients. A person of ordinary skill would have been motivated to make this modification because Ruhaak teaches that there is a need to identify biomarkers for risk assessment, early detection, and disease monitoring for lung cancer, and when would be motivated to identify such biomarkers in order to improve patient outcomes. Furthermore, a person of ordinary skill would have had a reasonable expectation of success in making this modification because: both Soddu and Ruhaak are in the same field of endeavor of protein enrichment methods, both methods use blood plasma samples, and both references use mass spectrometry to quantify proteins.
Claim 17 is rejected under 35 U.S.C. 103 as being unpatentable over Soddu in view of Ruhaak as applied to claims 13, 16, and 19 above, further in view of Hadjidemetriou et al. (2021). “Nanoparticle-enabled enrichment of longitudinal blood proteomic fingerprints in Alzheimer’s disease”. ACS nano, 15(4), 7357, (herein referred to as Hadjidemetriou).
The teachings of Soddu in view of Ruhaak are incorporated herein.
Regarding claim 17, Soddu teaches all the limitations of claim 1 of the instant application, and Soddu in view of Ruhaak determining the glycan profile of the enriched glycoprotein that is indicative of a disease or condition, but does not teach the enrichment of glycoproteins in a subsequent sample from the subject, wherein the glycoprotein and/or glycan profile of the protein corona is determined and wherein a change in the determined glycoprotein and/or glycan profile between the first sample and subsequent sample is indicative of the progression or remission of the disease or condition.
Hadjidemetriou teaches a method of employing the nanoparticle protein corona as a tool to systematically monitor changes in the plasma proteome with Alzheimer’s disease (AD) progression and to reveal underpinning molecular mechanisms (page 7359, column 2, 1st full paragraph). In this Hadjidemetriou teaches a method of identifying and tracking longitudinal alterations of the blood proteome in a transgenic mouse model of AD using a nanotechnology-enabled approach (7359, column 1, 5th full paragraph). Hadjidemetriou teaches that their data revealed the existence of multiple proteomic signals in blood, indicative of the asymptomatic stages of AD (abstract). Hadjidemetriou teaches that comprehensive analysis of the nanoparticle-recovered blood proteome by label-free liquid chromatography−tandem mass spectrometry resulted in the discovery of AD-monitoring signatures that could discriminate the asymptomatic phase from amyloidopathy and cognitive deterioration (abstract). Hadjidemetriou teaches the number of differentially abundant proteins discovered at the three time points (2M, 6M, and 12M) by proteomic analysis of (a) plasma control samples and (b) corona samples (Fig. 2).
It would have been obvious to person of ordinary skill in the art before the effective filing date of the claimed invention to have modified the particle-based glycoprotein enrichment/measurement method taught by Soddu in view of Ruhaak, to take samples at multiple timepoints to determine a change in the glycoprotein and/or glycan profile that is indicative of the progression or remission of the disease or condition, as taught by Hadjidemetriou, as it would be “obvious to try”. Soddu in view of Ruhaak already teaches determining the glycan profile of the enriched glycoprotein that is indicative of a disease or condition, and the use of repeated/longitudinal sampling to monitor disease state is a practice that is well-known, routine, and conventional in the art with regards to prognosis/diagnosis. One of ordinary skill in the art would also recognize that repeated sampling to monitor disease can be more accurate than a single sampling point due to establishment of in individuals “baseline” level of the marker. A person of ordinary skill would have been motivated to make this modification in order to improve the prognostic value of a biomarker with regards to a disease in order to improve patient outcomes. Furthermore, a person of ordinary skill would have had a reasonable expectation of success in making this modification because: both Soddu, Ruhaak, and Hadjidemetriou are in the same field of endeavor of protein enrichment methods, all three references methods use blood plasma samples, and all three references use mass spectrometry to quantify proteins, and the methods of Soddu and Hadjidemetriou both use nanoparticle-based proteomics enrichment methods. Furthermore, the incorporation of longitudinal sampling into the method of Soddu in view of Ruhaak would merely require the repetition of steps already found in their methods, not physical incorporation.
Claim 18 is rejected under 35 U.S.C. 103 as being unpatentable over Soddu in view of Ruhaak as applied to claims 13, 16, and 19 above, further in view of Du et al. (US 10,782,297 B2), (herein referred to as Du).
The teachings of Soddu in view of Ruhaak are incorporated herein.
Regarding claim 18, Soddu in view of Ruhaak teaches all the limitations of claim 16 of the instant application but does not teach administering a treatment for the disease or condition, which is optionally treatment for cancer.
Du teaches the use of signatures of secretory proteins are used to identify and distinguish lung cancers, and that the biomarker signatures may also be used to separate lung cancers from other inflammatory diseases, monitor progression, or assess treatment efficacy (abstract). Du teaches that the lung cancer-associated biomarkers are proteins or mRNA selected from the group consisting of: chitinase 3-like 1 (CHI3L1); transthyretin (TTR); fibrinogen beta polypeptide (FGb); fibrinogen-like protein 1 (FGL1); guanylate cyclase activator 2A (GUCA2A); delta-like 1 homolog (DLK1); glucose transporter 3 (GLUT3); cerebellin 1 (CBLN1); elastase 1, pancreatic (ELA1); fibrinogen alpha polypeptide (Fga); histidine-rich glycoprotein (HRG) (column 3, lines 10-15). Du also teaches administering chemotherapy to treat lung cancer (column 27, lines 25-27). Du also teaches that treatment with cytotoxic agents is a well-established standard for most advanced cancers and is also increasingly becoming an integrated part of treatment in primary and early cancers (column 9, lines 14-18).
It would have been obvious to person of ordinary skill in the art before the effective filing date of the claimed invention to have modified the particle-based glycoprotein enrichment/measurement method taught by Soddu in view of Ruhaak, to administer a treatment for the disease or condition, as taught by Du, as it would be “obvious to try”. It would be obvious to try as Du teaches that treatment with cytotoxic agents is a well-established standard for most advanced cancers and is also increasingly becoming an integrated part of treatment in primary and early cancers. A person of ordinary skill would have had a reasonable expectation of success in making this modification because Soddu in view of Ruhaak already teaches determining the glycan profile of the enriched glycoprotein that is indicative of a disease or condition, and Ruhaak specifically teaches lung cancer as the disease that the biomarkers are being measured for, analogous to the disease in Du. Furthermore, treatment administration for a disease state is a practice that is well-known, routine, and conventional in the art, especially with regards to cancer.
Claim 21 is rejected under 35 U.S.C. 103 as being unpatentable over Kavanagh et al. (2021). “N-Linked glycosylation profiles of therapeutic induced senescent (TIS) triple negative breast cancer cells (TNBC) and their extracellular vesicle (EV) progeny”. Molecular Omics, 17(1), 72-85, (herein referred to as Kavanagh), in view of Ruhaak et al. (2013). “Enrichment strategies in glycomics‐based lung cancer biomarker development”. PROTEOMICS–Clinical Applications, 7(9-10), 664-676, (herein referred to as Ruhaak).
The teachings of Ruhaak are incorporated herein.
Regarding claim 21, Kavanagh teaches the profiling of the N-linked glycans of whole cells, membrane, cytoplasm and extracellular vesicles (EVs) harvested from therapeutic-induced-senescent (TIS) Triple negative breast cancer (TNBC) cells and to compare these to results from non-senescent cells (abstract). Kavanagh teaches that ultra-performance liquid chromatography (UPLC) analysis of exoglycosidase digested N-linked glycans was carried out on TIS compared to non-treated control cells (abstract). Kavanagh teaches that LC-Mass spectrometry (MS) analysis of the N-linked glycans and lectin blotting of samples was carried out to confirm the UPLC results (abstract). Kavanagh also teaches that the HILIC-UPLC and LC-MS results show an increase in the sialylated glycan structure A2G2S1 (peak35) in TIS EVs compared to control EVs, and that there was also a decrease in the sialylated glycan structures FA2G2S1 (peak33+34) and FA3G3S2 (peak47) in TIS EVs compared to control EVs, possibly reflecting enhanced FUCA1 activity in TIS cells (page 78, column 2, 3rd paragraph). Kavanagh also teaches that highly branched and sialylated glycans are associated with cancer, and that N-Glycan signatures may inform on treatment response and overall patient survival (page 73, column 1, 5th paragraph).
However, Kavanagh does not teach a method of diagnosis for lung cancer, comprising the detection of one or more lung cancer biomarkers in a sample from a subject.
As discussed in the above rejection, Ruhaak teaches the use of whole plasma N-glycan analysis to further investigate the potential of glycans in blood plasma as diagnostic biomarkers for lung cancer (page 672, column 1, 1st full paragraph). Ruhaak teaches that the that the N-glycan profile in plasma of lung cancer patients is different from healthy controls and appears to be dominated by alterations in relatively abundant proteins (abstract – Conclusions). Ruhaak teaches that there is a need to identify biomarkers for risk assessment, early detection, and disease monitoring for lung cancer; and that biomarkers may also be provided by protein glycosylation patterns, as glycosylation is one of the most prevalent PTMs (page 664, column 2, 1st paragraph). Ruhaak also teaches that there is increased branching of N-glycans in tumor cells, which has the potential to yield a glycan signature for malignancy and metastasis (page 665, column 1, 1st paragraph).
It would have been obvious to person of ordinary skill in the art before the effective filing date of the claimed invention to have modified the N-Linked glycosylation profiling method of triple negative breast cancer cells and measurement of FA2G2S1 and FA3G3S2 , as taught by Kavanagh, to use those biomarkers to diagnose lung cancer, as taught by Ruhaak, as it would be “obvious to try”. It would be obvious to try as Kavanagh teaches that sialylated glycan structures FA2G2S1 and FA3G3S2 levels in cancer cells showed a decrease compared to control cells, and that generally, such highly branched and sialylated glycans are associated with cancer. Additionally, Ruhaak teaches that there is increased branching of N-glycans in tumor cells, which has the potential to yield a glycan signature for malignancy and metastasis. Similar to Kavanagh, Ruhaak teaches the measurement of sialylated glycans, just in lung cancer samples, and showed that fewer sialylated glycans in the IgG fraction.
Armed with this knowledge that breast cancer cells show decreased FA2G2S1 and FA3G3S2, and the knowledge that other sialylated glycans are reduced in lung cancer specifically, it would be obvious to modify the method of Kavanaugh to use a decrease in FA2G2S1 and FA3G3S2 as a biomarker for lung cancer diagnosis. A person of ordinary skill would have been motivated to make this modification because Ruhaak teaches that there is a need to identify biomarkers for risk assessment, early detection, and disease monitoring for lung cancer, and one would be motivated to identify such biomarkers in order to improve patient outcomes, and Kavanagh teaches that N-Glycan signatures may inform on treatment response and overall patient survival. Furthermore, a person of ordinary skill would have had a reasonable expectation of success in making this modification because: both Soddu and Ruhaak are in the same field of endeavor glycan profiling for cancer biomarkers, both methods use blood plasma samples, and both references use mass spectrometry to quantify proteins. Furthermore, the use of blood biomarkers to diagnose disease or disease progression is a practice that is well-known, routine, and conventional in the art, especially with regards to cancer.
Conclusion
For all the reasons discussed above, claims 1-11, 13, 15-19, 21, and 23 are rejected and therefore no claims are allowed.
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/ALEXANDER J. HOFFMAN/ Examiner, Art Unit 1677
/BAO-THUY L NGUYEN/Supervisory Patent Examiner, Art Unit 1677 August 31, 2026
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