The Examiner for this application has changed. Please forward all future correspondence to Examiner Kimberly Aron, Art Unit 1633. Additional Contact information can be found at the end of this Action.
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 .
Applicant’s amendments to the claims dated 12/22/25 are acknowledged. Claims 1, 3-7 and 9-36 are pending. Claim 19 is amended.
Election/Restrictions
Applicant’s election without traverse of Group I in the reply filed on 12/22/2025 is acknowledged.
Claims 33-36 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim.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 12/22/25.
The requirement is still deemed proper and is therefore made FINAL.
Prosecution on the merits commences for claims 1, 3-7 and 9-32.
PRIORITY
The instant application, filed 02/28/2023, claims priority to US Provisional Application Nos: 63/448,655 filed 02/27/2023, 63/436,854 filed 01/03/2023, 63/417,873 filed 10/20/2022, 63/411,899 filed 09/30/2022, and 63/315,897 filed 03/02/2022. Thus, the earliest possible priority for the instant application is 03/02/2022.
Information Disclosure Statement
The listing of references in the specification is not a proper information disclosure statement. 37 CFR 1.98(b) requires a list of all patents, publications, or other information submitted for consideration by the Office, and MPEP § 609.04(a) states, "the list may not be incorporated into the specification but must be submitted in a separate paper." Therefore, unless the references have been cited by the examiner on form PTO-892, they have not been considered.
Specification
The disclosure is objected to because it contains an embedded hyperlink and/or other form of browser-executable code. Applicant is required to delete the embedded hyperlink and/or other form of browser-executable code; references to websites should be limited to the top-level domain name without any prefix such as http:// or other browser-executable code. See MPEP § 608.01.
See, paragraphs [0150] and [0268] of the published specification.
Appropriate correction is required.
CLAIMS
Independent claims 1, 9, 12 and 19 are broadly directed to methods of culturing cells and producing an anti-IL-4Ra antibody, including Dupilumab, wherein the viable cell density (VCD) is adjusted.
Claims 1, 12 and 19 require wherein the initial viable cell density is adjusted to specific concentrations, and wherein the cells are cultured in a seed train.
Claims 9 and 12 require wherein viable cell density is measured by establishing a correlation equation of online capacitance and offline viable cell density of cells, (or using a previously generated correlation) and then measuring inline capacitance of cells, and predicting VSD from the established correlation equation.
Claim 19 requires establishing a correlation equation of a first property of a first cell culture measured online with a second property of the first cell culture measured offline, measuring online the first property in a second cell culture, and predicting the measure of the second property of the second culture from the correlation equation established by the first culture.
Claim Objections
Claims 1, 3-4, 7, 14-15, 18, 27 and 29 are objected to because of the following informalities:
Claim 1 should be amended to recite “the initial viable cell density (VCD)” in line 3.
Each of claims 3, 7, 14 and 18 comprise a typographical error within the recited series of percentages, each reciting “14%, 15%, 1%, 17%, 18%” in line 2. The claims should be amended to recite “16%.”
Each of claims 4, 15 and 27 comprise a typographical error, and is missing an x after 1.6 in line 2. The claims should be amended to recite “1.6x.”
Each of claims 7, 18 and 29 should be amended to recite “19%, or 20% in line 2.
Claim 7 should be amended from “resulted in” to “results in” at line 1.
Claim 7 should be amended to recite “increase in final titer (g/L) of Dupilumab.”
Claim 18 should be amended from “resulted in” to “results in” at line 1.
Claim 18 should be amended to recite “increase in final titer (g/L) of Dupilumab.”
Claim 29 should be amended from “resulted in” to “results in” at line 1.
Claim 29 should be amended to recite “increase in final titer (g/L) of Dupilumab.”
Appropriate correction is required.
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 1, 3-7 and 9-32 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.
Claim 1 requires, “wherein the initial VCD is adjusted between 3.5 x 105 to 5.43 x 105 cells/mL in each vessel or bioreactor.” It is unclear whether the claim is attempting to define the initial VCD for each vessel as between 3.5 x 105 to 5.43 x 105 cells/mL (wherein 3.5 x 105 to 5.43 x 105 cells/mL represents the lowest concentration of cells in each vessel), or indicate that the initial VCD is grown to between 3.5 x 105 to 5.43 x 105 cells/mL (wherein 3.5 x 105 to 5.43 x 105 cells/mL represents the highest concentration of cells in each vessel)?
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The same rejection is made for claims 12 and 19.
Claim 3 requires “a final Dupilumab titer is about 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 1%, 17%, 18%, 19%, or 20% greater than a titer produced from a cell culture where the initial VCD in said N-5 to N-1 vessel or bioreactor is below 2.5 x10⁵ cells/mL” which is unclear. Is the “a cell culture” part of the seed train of claim 1? Is the claim attempting to compare the titer to the same seed train, but wherein an initial VCD in each vessel or bioreactor is below 2.5 x10⁵ cells/mL? It is unclear how one is able to compare titers produced when it is not clear what methodology is being used to produce the titers from initial VCDs of less than 2.5 x10⁵ cells/mL.
Claim 5 recites the limitation "the final VCD" in line 2. There is insufficient antecedent basis for this limitation in the claim.
Claim 6 recites, wherein there is no substantial difference in peak lactate observed in a final production vessel compared to a final production vessel where the initial VCD vessels or bioreactors of the seed train is below 2.5 x10⁵ cells/mL” which is unclear. Is the “a final production vessel where the initial VCD vessels or bioreactors of the seed train” part of the seed train of claim 1? Is the claim attempting to compare the peak lactate to the same seed train, but wherein an initial VCD in each vessel or bioreactor is below 2.5 x10⁵ cells/mL? It is unclear how one is able to compare peak lactate produced when it is not clear what methodology is being used to produce the lactate from initial VCDs of less than 2.5 x10⁵ cells/mL.
Claim 6 recites, “the initial VCD vessels” in lines 2-3.
Claim 12 is directed to a method of producing Dupilumab by culturing cells wherein an initial viable cell culture has been adjusted to at least 2.5 x 105 cells/mL.
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It is not clear whether the claim, formatted as steps (a)-(d), is attempting to require these steps to be performed in order or whether the steps are simply encompassed by the claimed method. For example, step (a) requires “an initial viable cell density (VCD) in a seed train … is adjusted to at least 2.5 x 105 cells/mL. Step (b) requires measuring VCD by establishing a correlation of capacitance to VSD and step (b) requires adjusting initial VCD in each seen train or bioreactor.
In order to perform the adjustment of VSD in step (a), which is in a seed train vessel or bioreactor, the VSD must necessarily be measured. So, is the VSD of the cells of step (a) measured by the process of steps (b) and (c)? Or is the VSD of step (a) measured otherwise? Also, with regard to step (c), is the step attempting to require that (an? the?) initial VCD in each vessel or bioreactor is adjusted to 2.5 x 105 cells/mL? Or just adjusted to some VCD?
A skilled artisan would not know the metes and bounds of the claimed invention.
Claim 13 recites the limitation "wherein the initial VCD in each vessel or bioreactor of an N-5 to N-1 seed train is adjusted between 3.5 x 105 to 5.43 x 105 cells/mL" is indefinite. Claim 12 recites “initial VCD” of a seed train comprising vessels or bioreactors both in steps (a) (adjusted to at least 2.5 x 105 cells/mL) and (c). Thus, is claim 13 referring back to step (a) or step (c) or claim 12?
A claim is indefinite when it contains words or phrases whose meaning is unclear. In re Packard, 751 F.3d 1307, 1314, 110 USPQ2d 1785, 1789 (Fed. Cir. 2014). The lack of clarity could arise where a claim refers to "said lever" or "the lever," where the claim contains no earlier recitation or limitation of a lever and where it would be unclear as to what element the limitation was making reference. Similarly, if two different levers are recited earlier in the claim, the recitation of "said lever" in the same or subsequent claim would be unclear where it is uncertain which of the two levers was intended. MPEP 2173.05(e).
Claim 12 does not require a “vessel or bioreactor of an N-5 to N-1 seed train.” Thus, the phrasing of claim 13 wherein “the initial VSD in each vessel or bioreactor of an N-5 to N-1 seed train” is unclear. Is the claim attempting to require that the seed train of step (a) is an N-5 to N-1 seed train? The seed train of step (c)? Or is this attempting to add an additional seed train, and that initial seed train is an N-5 to N-1 seed train? The phrase lacks proper antecedent basis.
Claim 14 requires wherein a final Dupilumab titer is greater than “produced from a cell culture where the initial VCD in said seed train is below 2.5 x 105 cells/mL.” Is the claim referring back to a particular step of claim 12 wherein the initial VCD (a) or (c) before adjustment was below 2.5 x 105 cells/mL? Or is the claim attempting to recite “produced from a cell culture where an initial VCD in a seed train is below 2.5 x 105 cells/mL?
Claim 15 recites “the initial VCD”, however, like claim 13, it is unclear which “the initial VCD” the claim is referring back to.
Claim 16 recites, “wherein an increased final Dupilumab titer is not dependent on a final VCD in N-5 to N-1 vessels or bioreactors.” The claim is unclear. The claim does not recite an active step, nor clearly articulate that the process results in an increased final Dupilumab, nor clarify how the method comprise an N-5 to N-1 seed train.
Claim 17 recites, “wherein there is no substantial difference in peak lactate observed in a final production vessel compared to a final production vessel or bioreactor where the initial VCD in each N-5 to N-1 vessel or bioreactor is below 2.5 x105 cells/mL.” The claim does not recite an active step, nor clearly articulate that the process results in an increased final production vessel, clarify how the method comprise a N-5 to N-1 seed train.
Claim 18 recites “the seed train” in line 18. The previous claims recite unclear seed train(s). It is not clear which seed train claim 18 is referring back to.
Claim 18 requires the claim results in an increase in final titer, but it is not clear what the final titer is being compared to such that an increase can be determined?
Claim 19 recites,
A method of producing an anti-IL-4Ra antibody, the method comprising:
a) culturing cells capable of expressing an anti-IL-4Ra antibody in a vessel or bioreactor;
b) adjusting an initial viable cell density (VCD) to be 2.5 x 105 cells/mL or greater;
c) using at least one on-line sensor to measure a first property of a first cell culture;
d) using at least one off-line assay to measure a second property of the first cell culture;
e) correlating the measure of the first property of the first cell culture with the measure
of the second property of the first cell culture to determine a correlation equation;
f) using an on-line sensor to measure the first property of a second cell culture;
g) using the measure of the first property of the second cell culture and the correlation
equation to predict at least one measure of the second property of the second cell culture;
h) transferring said cells to another vessel or bioreactor based on the at least one predicted
measure of the second property of the second cell culture; and
i) repeating steps b)-h) along a seed train.
It is unclear whether step (b), “adjusting an initial cell viable density (VCD) to be 2.5 x 105 cells/mL or greater” is adjusting the cells of step (a), the cells necessarily included in the first cell culture of steps (c)-(e), or the cells necessarily included in the second cell culture of steps (f)-(g)?
Step (h) recites “transferring said cells to another vessel or bioreactor based on the at least one predicted measure of the second property of the second cell culture.” It is not clear whether “said cells” of step (h) are cells of the second cell culture, subjected to measurement of the first property of the second cell culture and the correlation equation in step g? Or, are the cells of step h) the same type of cell as those in the second cell culture, but not necessarily those measured in step g)?
In addition, it is unclear what repeating steps b)-h) “along a seed train” means. The claims nor specification nor prior art define the phrase.
Claim 20 recites “wherein said cells are from a cell culture that is the same as the first cell culture.” Claim 19 recites refers to “cells” in steps (a) and (h), but the method inherently requires cells in the first cell culture and the second cell culture, and inherently in the “seed train.” It is unclear which cells the claim is referring back to.
Claim 21 recites, “wherein said cells are from a cell culture that is different from the first cell culture.” It is unclear which cells the claim is referring back to for the same reasons as stated above.
Claim 22 recites, wherein said correlation equation is derived using more than one cell line which is unclear. Claim 19 does not require any “cell lines” or that the correlation equation is derived from a cell culture comprising a cell line. The correlation equation of step (e) only requires “a first cell culture.” In addition, it is not clear how the correlation equation is “derived” from more than one cell line as claim 22 does not contain any additional steps, or how, even with the repetition of steps (b)-(h) how such repetition results in generating the same correlation equation (“the correlation equation) using more than one cell line.
Claim 25 requires wherein at least about 50 percent of variability in said measure of said second property of said first cell culture is due to variance in said measure of said first property of said first cell culture is unclear. The claim does not provide an active step, nor require that there is any variance in the measure of either the first property of the first cell culture or the second property of the first cell culture. In addition, the claim only requires “a measure” (i.e., one measure) of the first property of the first cell culture with “at least one on-line sensor” and “a measure” of the second property of the first cell culture with “at least one off-line assay”.
Claim 31 recites, “wherein said cells are CHO cells.” It is unclear which cells the claim is referring back to for the same reasons as stated above.
Claims 4, 7 and 9-11, 22-24, 26-30, and 32 are included in the rejection because they depend from a rejected claim.
Claim Rejections - 35 USC § 112
The following is a quotation of the first paragraph of 35 U.S.C. 112(a):
(a) IN GENERAL.—The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
The following is a quotation of the first paragraph of pre-AIA 35 U.S.C. 112:
The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor of carrying out his invention.
Claims 3-7, 14-18, 25, 27- 30 rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, as failing to comply with the written description requirement. The claim(s) contains subject matter which was not described in the specification in such a way as to reasonably convey to one skilled in the relevant art that the inventor or a joint inventor, or for applications subject to pre-AIA 35 U.S.C. 112, the inventor(s), at the time the application was filed, had possession of the claimed invention.
Claims 3-7, 14-18, 25, 27-30 are directed to methods of using a seed train, wherein the claims require specific results:
Claims 3, 14 require wherein a final Dupilumab titer is about 2%....great than a titer produced from a cell culture where the initial VCD in said N-5 to N-1 vessel or bioreactor is below 2.5 x105 cells/mL.
Claims 4, 15, 27 require wherein the initial VCD is about 1.3x greater than an alternative initial VCD in a standard seed train.
Claims 5, 16 require wherein an increased final Dupilumab titer is not dependent on the final VCD in the N-1 seed train vessels or bioreactors.
Claims 6, 17, 28 requires wherein there is no substantial difference in peak lactate observed in a final production vessel compared to a final production vessel where the initial VCD vessels or bioreactors of the seed train is below 2.5 x105 cells/mL.
Claims 7, 18, 29 requires wherein the seed train resulted in a 2% increase in final titer (g/L).
The claims are to a large genus of methods, that require no specific cells, no specific culture methods, fail to define a “standard” seed train. And the specification produces few working examples which are encompassed by the scope of the claims. Thus, there is no structure-function relationship that renders the claims predictable, or indicate that applicant had possession of the claims at the time of the invention.
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.
Claims 9-11 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Schulze et al., Automation of High CHO Cell Density Seed Intensification Via Online Control of the Cell Specific Perfusion Rate and its Impact on the N-stage inoculum Quality, 2021. 335: 65-75, of record (IDS 1/12/24).
With regard to claims 9-11, Schulze discloses methods of culturing CHO cells in a seed train process, from thaw to N-1, and inoculating a production bioreactor (N-stage) to produce a monoclonal antibody (Abstract, page 66, FIG. 1).
Schulze discloses the method comprises establishing a correlation equation using linear regression of online capacitance to viable cell density (VCC therein) by measuring capacitance online (page 66, N-1 Perfusion bioreactor based on rocking motion”) and measuring viable cell density (VCC) offline (page 67 “Offline analytics”; FIG 3). VCC was measured online via inline capacitance during N-1 culture at days including day 5 and 7 (FIG 2). Schulze discloses N-1 culture taken from day 7 is used to inoculate an N-stage bioreactor at an inoculation density of 0.3 x 106 cells/mL and starting volume of 15 mL, resulting in an inoculation volume of 0.05 mL (page 67, “N-stage production bioreactor in multiparallel, automated small-scale bioreactors”).
Thus, Schulze discloses measuring on-line capacitance with a sensor to measure a first capacitance value of a first CHO cell culture; b) measuring viable cell density (VCD) value (VCC therein) of the said first CHO cell culture off line; c) producing a correlation equation of capacitance and VCD (FIG 3); d) measuring on-line capacitance to determine a second capacitance value of a second cell CHO culture (FIG 2, page 67) and using the second capacitance value and said correlation equation to predict the viable cell density value of the second CHO cell culture; and f) adjusting a working volume based on the second viable cell density value to culture the cell (page 67, “N-stage production bioreactor in multiparallel, automated small-scale bioreactors”).
With regard to claim 10, Schulze discloses the same cell lines are used in both the N-1 and N stages (page 66-67, FGI 1).
With regard to claim 11, Schulze discloses the correlation equation is based on linear regression (FIG 3).
Claims 9-11 are rejected under 35 U.S.C. 102(a)(1) and (a)(2) as being anticipated by US Patent Application Publication No. 2020/0255785 to Erlandson.
With regard to claims 9-11, Erlandson discloses methods of culturing CHO cells in a seed train process, from 250 mL, 1L, 3L, one or more 50-L wavebags, to N-1, and inoculating a production bioreactor (N reactor) to produce a monoclonal antibody (Abstract, paragraphs [0004], [0008]-[0012], [0014], [0040], [0043], [0053]-[0059], [0061]-[0063], [0074], [0094], [0096]).
Erlandson discloses the method comprises establishing a correlation equation using linear regression of online capacitance to viable cell density (VCD therein) by measuring capacitance online and measuring viable cell density (VCD) offline (paragraphs [0017], [0065], [0091]-[0092], [0095]-[0096], [0104], FIGs 3A, 3B). VCD was measured online via inline capacitance during N-1 culture (paragraphs [0018], [0091]-[0092], [0095]-[0096]). Erlandson discloses when the VCD from N-1 culture reaches a specific concentration, cells are transferred from the N-1 bioreactor to the production bioreactor, and the N-1 bioreactor is reseeded (paragraphs [0019]-[0020], [0068], [0070], [0091]-[0092], [0095]-[0096]). Erlandson discloses the N-1 bioreactors have an initial viable seeding density of 1-2 x 106 cells/mL (10-20 x 105) or 1-1.5 x 106 (10-15 x 105 cells/mL) (paragraphs [0095], [0096], [0098]).
Thus, Erlandson discloses measuring on-line capacitance with a sensor to measure a first capacitance value of a first CHO cell culture; b) measuring viable cell density (VCD) value of the said first CHO cell culture off line; c) producing a correlation equation of capacitance and VCD (paragraph [0104], FIGs 3A, 3B); d) measuring on-line capacitance to determine a second capacitance value of a second cell CHO culture and using the second capacitance value and said correlation equation to predict the viable cell density value of the second CHO cell culture (paragraph [0068]; and f) adjusting a viable cell density based on the second viable cell density value to culture the cell (paragraphs [0068], [0070]).
With regard to claim 10, Erlandson discloses the same cell lines are used in both the N-1 and N production bioreactors (Example 1, paragraphs [0094]-[0098]).
With regard to claim 11, Erlandson discloses the correlation equation is based on linear regression (paragraph [0017], FIGs 3A, 3B).
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.
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.
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Claim 1 is directed to methods of producing Dupilumab in a seed train:
Claim 1 is rejected under 35 U.S.C. 103 as being unpatentable over WO2016/196315 to Moretto, of record (IDS 1/12/24) further in view of US Patent Application No. 2016/0244726 to Lawrence and US Patent Application Publication No. 2010/0120093 to von Fircks.
With regard to claim 1, Morretto discloses methods of culturing recombinant cells to produce biotherapeutic proteins, including Dupilumab, wherein the method comprises culturing the cells in a culture system that monitors and/or controls the culture process to prevent or correct cell culture process deviations in order to increase yield and/or quality (Abstract, page 2, lines 1-8, page 3, lines 5-14, page 4, lines 7-18; page 6, line 13- page 7, line 13, page 40, line 13). Morretto discloses the methods comprise culturing cells in a bioreactor managed through a feedback control automation system (page 4, lines 7-10, page 5, lines 11-12, page 6, line 26 – page 7, line 20; page 35, line 1 – page 37, line 6; FIG. 10). The feedback control automation system monitors one or more of the cell culture process parameters, including culture component levels in a culture medium, evaluates the one or more of the cell culture process parameters against pre-set or predictive threshold levels, and adjusts the cell culture process in order to address and/or correct any process deviations (page 13, lines 1-32, page 16, line 19 – page 18, line 9, page 29, line 25 – page 23, lines 15-23, page 33, line 6 – page 36, line 4, page 37, line 9 – page 38, line 22).
Morretto discloses the recombinant cells are expanded in a seed train, including multiple shake flasks, or expanding high density working cell bank vials in N-2 and N-1 vessels, before being transferred to a production bioreactor (page 51, lines 25-32; page 58, line 27 – page 59, line 22, Table 3, page 66, lines 20-23). Morretto discloses cell cultures are initiated with densities comprising approximately 0.5 x 106 (5 x 105) cells/mL as disclosed on FIG. 2B, FIG. 6D, FIG. 7B, FIG. 17A, and FIG. 18C, although not specific values are disclosed.
However, Morretto does not disclose wherein the seed train comprises N-5 to N-1 vessels or bioreactors, or wherein the initial viable cell density in each vessel or bioreactor is adjusted [to] between 3.5 x 105 to 5.43 x 105 cells/mL, as required by instant claim 1.
Lawrence discloses methods of culturing recombinant cells in a seed train to produce therapeutic proteins (abstract, paragraphs [0002]-[0003], [0010]-[0013], [0020], [0026]). At paragraph [0045], Lawrence discloses:
The phrase "seed culture" or "seed train" (also referred to as inoculum train) as used herein includes the inoculation source of a cell population which is allowed to expand in batch culture, or series of batch cultures, until ready for production scale. The seed train expansion process constitutes the initial growth phase of the cells, or inoculum growth phase, following a thaw of frozen cells. The interval between cell thawing and the accumulation of sufficient cell mass to inoculate a production bioreactor constitutes the seed train expansion phase. The cell mass may be scaled up through several bioreactor stages in seed culture, and the cells are grown in cell culture medium under conditions favorable to the survival, growth and viability of the cell culture. It is understood that the seed train is intended to maximize the exponential growth phase, or achieve the maximal growth rate for the particular cell type being cultured. Therefore, passaging of cells from one bioreactor or vessel to another may be one way to achieve maximal growth rate. The precise conditions will vary depending on the cell type, the organism from which the cell was derived, and the nature and character of the expressed polypeptide or protein. A shift to lactate consumption metabolism may occur or be detected in any one of the vessels in a seed train expansion.
Lawrence teaches the series of expansion cultures in a seed train are numbered N-X cultures, wherein “X” is the number of expansion culture steps in the train, and N-1 is the culture that inoculates a production bioreactor (paragraphs [0056], [0058]). Lawrence discloses the number of cultures in a seed train is determined by the skilled artisan, and may comprise starting seed trains at N-8, N-7, N-6, N-5, N-4, N-3, or N-2 cultures, wherein when an expansion culture step reaches a specific growth or cell density target, cells from the culture are transferred, serving to inoculate to the next step (thus, N-5 serves to inoculate N-4, expansion of N-4 serves to inoculate N-3, expansion of N-3 serves to inoculate N-2, etc., increasing the overall seed train cell density (paragraphs [0056]-[0059], [0063], [0065]-[0068]). Figures 3 and 8 of Lawrence show, at least, exemplary seed train processes, comprising seed vessels N-5 (thaw culture) through N-1 (paragraphs [0056]-[0057]). Lawrence discloses the initial viable cell density in a seed train varies, depending on the starting cell line, and can range from 0.5 - 3 x 106 cells/mL (paragraphs [0056]-[0057]).
US Patent Application Publication No. 2010/0120093 to von Fircks discloses methods of expanding recombinant cells in a seed train (paragraphs [0050]-[0053], wherein the expansion cultures are repeated with progressive increases in the size of the seed culture vessels until a minimum cell density is obtained (paragraph [0064]). Von Fircks discloses “Typically, cells are inoculated into a seed culture vessel at an initial density of about 0.2 – 0.4 x 106 cells/mL” (paragraph [0064]).
It would have been obvious to combine the method of producing Dupilumab in a seed train, according to Moretto, further with the disclosures of Lawrence and von Fircks. It would have been obvious to culture cells in a seed train of N-5 to N-1 vessels or bioreactors, as Lawrence discloses the number of expansion cultures in a seed train is determined by a skilled artisan, in culture steps sufficient to product a viable cell density suitable to inoculate a production bioreactor, and exemplifies a seed train of N-5 to N-1 vessels is able to do so. It would have been obvious to utilize adjust the initial viable cell density in each of the seed train vessels because Lawrence discloses initial viable cell densities used to inoculate seed train cultures comprises 0.5 - 3 x 106 cells/mL, and von Fircks discloses an initial density of about 0.2 – 0.4 x 106 cells/mL is a typical density to inoculate seed train vessels. It would have been obvious to utilize known seed train culture parameters in the method of Moretto when producing Dupilumab. Combining prior art elements according to known methods to yield predictable results supports a conclusion of obviousness (MPEP 2143 (I)(A)). In the instant case, the prior art includes each element claimed, although not in a single prior art reference, and that in combination, each element merely performs the same function as it does separately. In addition, In the case where the claimed ranges "overlap or lie inside ranges disclosed by the prior art" a prima facie case of obviousness exists (MPEP 2144.05). A skilled artisan would have recognized using known seed train parameters of culturing recombinant cells expressing a protein would have resulted in the successful production of Dupilumab.
Claims 12-13, 19-24, 26 and 31-32 are rejected under 35 U.S.C. 103 as being unpatentable over WO2016/196315 to Moretto, of record (IDS 1/12/24) in view of WO2020/169432 (WO’432), Metze Monitoring Online Biomass with a Capacitance Sensor During Scale-up of Industrially Relevant CHO Cell Culture Fed-batch Processes in Single-Use Bioreactors. Bioprocess and Biosystems Engineering, 2019. 43(2): pages 193-205, of record, and Kern et al. Model-based Strategy for Cell Culture Seed Train Layout Verified at Lab Scale. Cytotechnology, 2016, of record.
Claim 19 is directed to a method of producing an anti-IL4Ra antibody in a cell culture seed train, wherein the method includes generating a cell culture parameter model by comparing on-line data of a first parameter to the off-line data of a second parameter, correlating the data to create the model, and then verifying the model by generating more on-line data of the first parameter.
Claim 12 requires wherein viable cell density is measured by online capacitance and utilizes a previously generated correlation of inline capacitance to offline VCD, and generating Dupliumab.
With regards to claims 12 and 19, Morretto discloses methods of culturing recombinant cells to produce biotherapeutic proteins, including Dupilumab, wherein the method comprises culturing the cells in a culture system that monitors and/or controls the culture process to prevent or correct cell culture process deviations in order to increase yield and/or quality (Abstract, page 2, lines 1-8, page 3, lines 5-14, page 4, lines 7-18; page 6, line 13- page 7, line 13, page 40, line 13). Morretto discloses the methods comprise culturing cells in a bioreactor managed through a feedback control automation system (page 4, lines 7-10, page 5, lines 11-12, page 6, line 26 – page 7, line 20; page 35, line 1 – page 37, line 6; FIG. 10). The feedback control automation system monitors one or more of the cell culture process parameters, including culture component levels in a culture medium, evaluates the one or more of the cell culture process parameters against pre-set or predictive threshold levels, and adjusts the cell culture process in order to address and/or correct any process deviations (page 13, lines 1-32, page 16, line 19 – page 18, line 9, page 29, line 25 – page 23, lines 15-23, page 33, line 6 – page 36, line 4, page 37, line 9 – page 38, line 22).
Morretto discloses the recombinant cells are expanded in a seed train, including multiple shake flasks, or expanding high density working cell bank vials in N-2 and N-1 vessels, before being transferred to a production bioreactor (page 51, lines 25-32; page 58, line 27 – page 59, line 22, Table 3, page 66, lines 20-23). Morretto discloses cell cultures are initiated with densities comprising approximately 0.5 x 106 (5 x 105) cells/mL as disclosed on FIG. 2B, FIG. 6D, FIG. 7B, FIG. 17A, and FIG. 18C, although not specific values are disclosed.
However, Moretto does not disclose discloses a method (a) culturing cells wherein an initial viable cell density (VCD) in a seed train in vessels or bioreactors is adjusted to a specific cell density adjusted to at least 2.5 x 105; (b) measuring viable cell density by (i) applying an electric field to said cells cultured in a vessel or bioreactor; and (ii) measuring capacitance; and (iii) correlating capacitance to viable cell density; (c) adjusting initial VCD in each seed train vessel or bioreactor.
WO’432 discloses a cell culture bioprocessing device and bioreactor assembly. WO’432 discloses the bioprocess device assembly, and methods of generating an inoculum, comprises multiple pre-culture (inoculation) bioreactors (N-X, N-2, N-1), wherein when the bioreactors are seeded, the device transfers a volume of the inoculum from one bioreactor to the next, thus forming a seed train. WO’432 discloses the number of pre-culture bioreactors can be increased (N-X stage to N-X+1) in order to scale up the process, wherein N-x (is the initial seed culture) to N-1 (last seed culture before production N) (page 5, lines 5-7; page 8, lines 6-13, FIG 3).
WO’432 discloses the bioreactors comprise one or more in-line sensors which monitor and measure at least one parameter relating to the quantity and/or quality of the culture. The system includes one or more in-line sensors that can measure viable cell density (VCD), wherein when a predetermined VCD level or threshold is detected in the bioreactor, the system transfers a volume of inoculum into the next bioreactor (pages 2, 4).
WO’432 discloses the in-line VCD is determined by repeatedly measuring the in-line capacitance of the culture when in perfusion mode with a capacitance sensor, and correlating the capacitance data to a correlation equation determined earlier empirically, via modeling or, is generated in parallel to the culture steps, allowing for real time monitoring and transfer (page 3, page 4, page 5, page 6, lines 13-27, line 25 – page 7, line 11).
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WO’432 discloses the system can be used to monitor and adjust different parameters in the cell culture.
Thus, WO’432 discloses a method (a) culturing cells wherein an initial viable cell density (VCD) in a seed train in vessels or bioreactors is adjusted to a specific cell density; (b) measuring viable cell density by (i) applying an electric field to said cells cultured in a vessel or bioreactor; and (ii) measuring capacitance; and (iii) correlating capacitance to viable cell density; (c) adjusting initial VCD in each seed train vessel or bioreactor.
Metze generates a model correlating in-line capacitance to off-line viable cell density (VCD) (VCC therein) of recombinant CHO cells which express an antibody, cultured in a bioreactor in a scale up process (Abstract, page 195). Metze generates the model by culturing CHO cells in a bioreactor with an initial seed density of 3 x 105 cells/mL (paqe 195 “Seed Culture for Process A and Process B”, and Main Culture For Process A” and “Main Process for Process B”).
The model is produced by a) using at least one on-line capacitance probe to measure a first capacitance value of a first cell culture; b) using at least one off-line assay to measure a first viable cell density value of the said first cell culture; c) correlating said first capacitance value with the said first viable cell density value to determine a correlation equation (Page 196 “Online Capacitance Measurements” “Offline Capacitance Measurements,” Page 198, FIGs.1d, 2d; FIG 8a). The model is validated by d) using an on-line capacitance probe to determine a second capacitance value of a second cell culture; e) using said second capacitance value and said correlation equation to predict at least one second viable cell density value of said second cell culture (Page 199, 200-201; FIG 5, FIG 8b). Metze discloses the model is accurate for measuring VCD through exponential growth phase, until peak cell density. Metze discloses the use of capacitance to measure VCD is useful for scale up production, as it appears to be able to provide scale independent models of VCD (page 194, 200, 203). Metze discloses the use of inline capacitance can be used in automated scale up processes (page 202).
Kern discloses a seed train modeling tool (abstract). Kern discloses seed trains require expanding cells in successively larger vessels, from vial to bioreactors until a sufficient quantity of cells are produced to inoculate a production vessel (page 1020). Cell expansion during seed train expansion is monitored via measuring viable cell density from initial cell seeding density to the end of the exponential growth phase, as cell passage into the next stage should be done prior to the cells exiting the exponential growth phase (page 1020, 1023-1024). Kern discloses initial seeding density of CHO cells is 0.3 x 109 cells/L (3.0 x 105 cells/mL), and following expansion and passage, the seeding density is adjusted back to the same density in each vessel (page 1022; Fig 6a).
It would have been obvious to combine the disclosure of Moretto, on methods of generating Dupilumab in an automated feedback system. Further with the disclosures of WO/342, Metze and Kern. A skilled artisan would have been motivated to use the automated system of WO’432 for its accuracy, as WO’432 discloses its system is able to monitor and adjust the conditions between bioreactors in real time, calculates the correct inoculation volume and accurately transfers it to the next vessel, can be performed multiple times, and is scalable (pages 4-8). WO’432 discloses its system can use modeling, and/or can gather measurements to generate the correlation repeatedly (pages 6-7). In addition, a skilled artisan would have been motivated further during seed train expansion of dupilumab, as Metze discloses use of online capacitance to measure VCD, based off of inline capacitance and offline VCD correlation equations, provides accurate measurements that can be used in automated scale up production of antibodies (page 202). Finally, the claimed requirements relating to specific adjustments of initial seed densities in seed train modeling are known in the art, as disclosed by Kern who shows initial seeding densities of CHO cells include 3.0 x 105 cells/mL, and following expansion and passage, the seeding density is adjusted back to the same density in each vessel (page 1022; Fig 6a). A skilled artisan would have had a reasonable expectation of success in practicing the claimed invention as all of the individual portions relating to seed train scale up were known at the time of the invention.
With regard to claim 12, the use of an N-5 to N-1 would have been obvious from WO’432 in light of the art, depending on need.
With regard to claims 20-22, Metze shows the model is accurate for cells from the same culture, using different cell lines (pages 195-200), and then also accurate in cells from a second culture, and use of different cell lines (pages 201-203).
With regard to claims 23-24 Metze discloses the method comprises generating more than one measurement of online capacitance and offline VCC (page 196, FIGURES).
With regard to claim 26, WO’342 discloses the correlation equation is generated using linear regression (page 3, page 4, page 5, page 6, lines 13-27, line 25 – page 7, line 11, FIG 2). Metze discloses the correlation equation is generated using linear regression (pages 197, FIGURES).
With regard to claim 31, Moretto discloses the cells are CHO cells (page 4, 12, 44).
With regard to claim 32, Moretto discloses the antibody is dupilmab (page 40).
Conclusion
No claims are allowed.
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/CHRISTOPHER M BABIC/Supervisory Patent Examiner, Art Unit 1633
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