DETAILED ACTION
Claims 23-37 are pending and under examination in the instant application. The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA . An action on the merits follows.
Information Disclosure Statement
The two information disclosure statements (IDS) submitted on 1/25/24 are in compliance with the provisions of 37 CFR 1.97 and 1.98. Accordingly, the information disclosure statements have been considered by the examiner and initialed and signed copies of the 1449s are attached to this action.
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.
Claim 24 is 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 24 depends on claim 23 and recites the limitation, “wherein the reliability of generating of iPSCs from the lymphoblastoid cells is 100%”. The limitation is in part confusing as “the reliability of generating of iPSCs” may contain a grammatical error where an extra “of” has been inserted between “generating” and “iPSCs”. Further, as “iPSCs” and “lymphoblastoid cells” is plural, it is unclear whether the applicant is intending to claim that the method is 100% reliable in generating at least one or more iPSCs from one or more lymphoblastoid cells, or whether applicant intends to recite that the method is 100% reliable in generating an iPSC from each and every lymphoblastoid cell. As such, the metes and bounds of the limitation regarding 100% reliability cannot be determined.
In the interests of compact prosecution, the limitation regarding 100% reliability has been interpreted to encompass a 100% reliability in generating at least one iPSC from the population of lymphoblastoid cells present in the starting culture.
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.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claims 23-32, and 36-37 are rejected under 35 U.S.C. 103 as being unpatentable over U.S. Patent Application Publication 2012/0009676 (January 12, 2012), hereafter referred to as Mack, in view of Hong et al. (2009) Nature, Vol. 460(7259), 1132-1135, Rajesh et al. (2011) Blood, Vol. 118(7), 1797-1800, U.S. Patent Application Publication 2016/0145642 (2016), hereafter referred to as Cui et al., with an effective filing date of 7/12/13, and U.S. Patent Application Publication 2012/0196360 (August 2, 2012), hereafter referred to as Okita et al.
Mack teaches methods of generating lymphoid-cell derived induced pluripotent stem cells (iPS cells) comprising providing a quantity of mammalian, preferably human, cells comprising lymphoid cells (LCs), such as peripheral blood mononuclear cells, spleen cells, tumor infiltrating lymphocytes, cells from lymph nodes, and lymphoid progenitor cells, and introducing exogenous episomal genetic elements or exogenous RNA genetic elements that express iPS reprogramming factors into the cells (Mack, abstract, and paragraphs 12, 23, 39, 57, 61, 90-91. 111-112, 198, and 299). More specifically, Mack teaches that the reprogramming factors may comprises one or more of Klf4, c-Myc, Lin-28, and SV40 Large T Antigen, that the reprogramming factors may be encoded in one or more oriP/EBNA-1 derived vectors, and that the delivery of reprogramming factors comprises nucleofection (Mack, paragraphs 23, 198, and 299). Mack teaches that the transfected cells are maintained in the transfection media and culture for at least 24 hrs post transfection and then transitioned to reprogramming media over 1-2 days, are subsequently cultured in reprogramming media for at least 7 days, including 7, 8, 9, 10, 11, 12, 13, 14, and 15 days, followed by culture in a serum-free induction media for at least 10 days, where the induction media is TeSR2 (Mack, paragraphs 12, 39, 57, and 111-112). Note that according to the ranges provided by Mack et al., transition to reprogramming media could occur at least about 3-4 days post-transfection. Mack further teaches that the reprogramming media may comprise at least one small chemical induction molecule such as CHIR99021, PD0325901, HA-100 and/or A-83-01 (Mack, paragraph 23). Mack further teaches their methods of culturing obviate the need for serum or MEF feeder cells (Mack et al., paragraph 113). In one embodiment, Mack et al. teaches that the reprogramming media comprises 1% N-2, 2% B-27, 1% NEAA, .5% Glutamax, 0.5 uM PD0325901, 3 uM CHIR99021, 0.5uM A-83-01, 10 uM HA-100, and 100 ng/ml zebrafish FGF (Mack et al., Table 2). Mack also teaches the generation of cell lines of iPS cells obtained using the disclosed method, and further iPS cell lines derived from the peripheral blood of patients with somatic cell disorders and/or an acquired hematologic disorder (Mack, paragraph 75). While Mack et al. primarily teaches to use their methods for generating iPSCs from hematopoietic progenitor cells present in PBMC, Mack et al. does teach that other populations of cells can be used to generate iPSC, including cells from cell lines, see paragraphs 90-91 of Mack et al. Mack et al. also states that their methods can be used to generate iPSCs from CD34 negative cells. See for example paragraph 61 of Mack et al. which states, “ [i]n certain embodiments, certain human hematopoietic progenitor cells may not express CD34, but these cells may nonetheless be converted into iPS cells via the methods disclosed herein” (Mack et al., paragraph 61).
Mack et al. differs from the instant invention as claimed by not teaching that the reprogramming factors includes a short hairpin RNA targeting p53 (shRNA-p53). However, at the time of filing, Hong et al. teaches that the formation of iPS cells from terminally differentiated somatic cells expressing reprogramming factors is suppressed by p53 expression and that inhibition of p53 expression enhances iPS generation from somatic cells including terminally differentiated lymphocytes (Hong et al., pages 1132-133). Hong et al. teaches that p53 can be effectively inhibited by introduction of a vector comprising a nucleic acid encoding shRNA-p53 and that the introduction of nucleic acids encoding four reprogramming factors and shRNA-p53 to somatic cells resulted in increased generation of iPS cells (Hong et al., Figure 2). More specifically, Hong et al. teaches the successful generation of iPSCs from terminally differentiated p53 negative T lymphocytes- cells which are inherently CD34 negative, using Oct3/4, Sox2, Klf4, and Nanog (Hong et al., pages 2-3). Therefore, in view of the motivation provided by Hong et al. to suppress p53 using a nucleic acid encoding shRNA-p53 in methods of generating iPS cells from somatic cells including lymphoid cells, it would have been prima facie obvious to the skilled artisan at the time of filing to modify the methods of Mack et al. to include the introduction of a vector comprising a nucleic acid encoding shRNA-p53 along with the introduction of vectors encoding reprogramming factors such as Klf4, c-Myc, Lin-28, and SV40 Large T Antigen to a lymphoid cell in order to increase the generation of iPS cells with a reasonable expectation to success.
Mack further differs from the instant invention by not teaching that the lymphoid cells are lymphoblastoid cells derived from a lymphoblastoid cell line. Rajesh et al. supplements Mack et al. and Hong et al. by teaching that large collections of lymphoid cell lines (LCLs) generated by transformation of lymphoid cells with EBV are available internationally to researchers and can be reprogrammed using a cocktail of reprogramming factors and small molecules to generate iPSCs (Rajesh et al., page 1797). The methodology of Rajesh et al. is similar to that of Mack in that Rajesh et al. teaches to transfect lymphoblastoid cell line cells (LCLs) with oriP/EBNA1 episomal vectors encoding Oct4, Sox-2, Klf-4, c-Myc, Lin-28, and SV40 Large T Antigen, culture the transfected LCLs in reprogramming media comprising PD0326901, CHIR99021, HA-100, and A-83-01 for 14 days, followed by culture in an induction media for 14 days (Rajesh et al., see page 1797, and online Supplemental Materials). More specifically, Rajesh et al. teaches that the LCLs undergo nucleofection in RPMI 1640 containing 15% FBS at 37oC and 5% CO2 on Matrigel-coated tissue culture plates, i.e. under feeder free conditions, and then transitioned to reprogramming medium (RM) at about 12 hours post-nucleofection, where the RM consists of DMEM/F12 media supplemented with non-essential amino acids (NEAA), Glutamax, N2, B27, 0.1mM beta-mercaptoethanol, 100ng/mL bFGF, 0.5uM PD0325901, 3uM CHIR99021, 0.5 uM A-83-01, 1000 units/mL hLIF, and 10 uK HA-100 (Rajesh et al., Supplementary Materials page 1). Rajesh et al. further teaches that the cells were fed with fresh media every other day for at least two weeks, and that between days 14-20 the culture medium was transitioned to TeSR-2 Complete medium (Rajesh et al., Supplementary Material, page 1). Rajesh et al. notes that iPSC colonies were detectable at around 20 days (Rajesh et al., Supplementary Material, page 1). Note as well that all culture conditions occurred on the feeder free Matrigel plates. Rajesh et al. further teaches that reprogramming LCLs can offer an unlimited source for patient-specific iPSCs (Rajesh et al., page 1797). Thus, both Hong et al. and Rajesh et al. show the successful generation of iPSCs from mature CD34 negative lymphoid cells, and Rajesh et al. in particular provides specific evidence that LCLs as claimed can be reprogrammed using a technique similar to that disclosed by Mack et al. Therefore, in view of motivation provided by Rajesh et al. to generate lymphoblastoid cell line cell derived iPSCs, it would have been prima facie obvious to the skilled artisan at the time of filing to further modify the methods of Mack et al. and Hong et al. by using lymphoblastoid cell line cells as the lymphoid cells used to generate iPSCs with a reasonable expectation of success. Furthermore, based on the teachings of Mack for ranges of culture time for each step in the reprogramming methods including approximately 3 days or more for the time in culture post-transfection and prior to exchange of media to reprogramming media, and culture of the transfected cells in reprogramming media for 15 days, followed by culture in maintenance/induction media for at least 3-5 more days followed by the appearance of iPSC, it would have been prima facie obvious to the skilled artisan at the time of filing to practice the methods of reprogramming lymphoblastoid cell line cells into iPSCs as taught by Mack et al. in view of Hong et al. and Rajesh et al. where the nucleofected lymphoblastoid cells are cultured for about 3 days post nucleofection prior to culturing the media in reprogramming media for about 15 days, following by culture in maintenance/induction media for at least 3-5 more days with a reasonable expectation of success in generating iPSCs. Furthermore, based on the successful generation of iPSCs taught by Rajesh et al., who demonstrates that practice of the instant methods for reprogramming LCLs without shRNA-p53 already exhibited a reprogramming efficiency of 0.01%, the skilled artisan would have had a reasonable expectation that practice of the methods of reprogramming lymphoblastoid cell line cells into iPSCs as taught by Mack et al. in view of Hong et al. and Rajesh et al. would have generated at least 1 iPSC from the culture with 100% reliability.
Further in regards to the elements of the reprogramming media, both Mack et al. and Rajesh et al. as noted above teach that the reprogramming media comprises 0.5 uM PD0325901, 3 uM CHIR99021, 0.5uM A-83-01, 10 uM HA-100, and 100 ng/ml zebrafish FGF, beta-mercaptoethanol, N2, NEAA, B27 and Glutamax (glutamine). Mack et al. further provides specific guidance for using 1% N-2, 2% B-27, 1% NEAA, and .5% Glutamax in the reprogramming media, and Rajesh et al. provides specific guidance for using DMEM/F12 as the base media for the reprogramming media comprising a concentration of 0.1mM beta-mercaptoethanol. While Mack et al. teaches to use .5% Glutamax in the reprogramming media and Rajesh et al. is silent as to the concentration of Glutamax, neither specifically teaches to use 1% Glutamax in the reprogramming media, or provide guidance for the concentration of an antibiotic/antifungal in the reprogramming media. Cui et al. further supplements Mack et al. and Rajesh et al. by teaching that reprogramming media to generate iPSCs which is based on DMEM/F12 can comprise 1% Glutamax and 1% antibiotic/antifungal in addition to 1% N-2, 2% B-27, and 1% NEAA (Cui et al., paragraph 35, 43, and 52). Thus, based on the combined teachings of Mack et al., Rajesh et al., and Cui et al. for the components of reprogramming media capable of generating iPSCs from cells including lymphoid cells and lymphoblastoid cell line cells, it would have been prima facie obvious to the skilled artisan at the time of filing to utilize a reprogramming media based on DMEM/F12 supplemented with 1% Glutamax, 1% antibiotic/antifungal, 1% N-2, 2% B-27, 1% NEAA, 0.1mM beta-mercaptoethanol, 100ng/mL bFGF, 0.5uM PD0325901, 3uM CHIR99021, 0.5 uM A-83-01, 1000 units/mL hLIF, and 10 uK HA-100 in the methods of generating iPSCs from lymphoblastoid cell line cells according to Mack in view of Hong et al. and Rajesh et al. with a reasonable expectation of success. Finally, while Mack teaches the introduction of various reprogramming factors to peripheral blood cells using oriP/EBNA-1 derived vectors, Mack does not specifically teach the introduction of the vectors pEP4 E02S ET2K, pCXLE-hOCT3/4-shp53-F, pCXLE-hSK, and/or pCXLE-hUL as claimed in claims 5 and 18. Okita et al. further supplements Mack by teaching that iPS cells can be derived from various types of human cells, including cells of the blood and immune system by transfecting the cells with episomal vectors including pEP4 E02S ET2K, pCXLE-hOCT3/4-shp53-F, pCXLE-hSK, and/or pCXLE-hUL (Okita et al., paragraphs 8, 72-80, 101-102, and 154). Okita et al. further teaches that use of these reprogramming vectors efficiently establishes safe human iPS cells suitable for clinical application without integration of the nucleic acid factors into the genome (Okita et al., paragraph 8). Therefore, in view of the teachings of Mack to use oriP/EBNA-1 derived vectors to express reprogramming factors in lymphoid cells to generate iPS cells, and the motivation provided by Okita et al. to generate iPS from cells such as blood and immune cells using the EBNA-1 based vectors pEP4 E02S ET2K, pCXLE-hOCT3/4-shp53-F, pCXLE-hSK, and/or pCXLE-hUL in order to efficiently establish safe human iPS cells suitable for clinical application without integration of the nucleic acid factors into the genome, it would have been prima facie obvious to the skilled artisan at the time of filing to practice the methods of generating iPS cells from lymphoblastoid cell line cells taught by Mack et al. in view of Hong et al., Rajesh et al., and Cui et al., using the pEP4 E02S ET2K, pCXLE-hOCT3/4-shp53-F, pCXLE-hSK, and/or pCXLE-hUL vectors taught by Okita et al. with a reasonable expectation of success.
Claims 33-35 are rejected under 35 U.S.C. 103 as being unpatentable over U.S. Patent Application Publication 2012/0009676 (January 12, 2012), hereafter referred to as Mack, in view of Hong et al. (2009) Nature, Vol. 460(7259), 1132-1135, Rajesh et al. (2011) Blood, Vol. 118(7), 1797-1800, U.S. Patent Application Publication 2016/0145642 (2016), hereafter referred to as Cui et al., and U.S. Patent Application Publication 2012/0196360 (August 2, 2012), hereafter referred to as Okita et al., as applied to claims 23-32, and 36-37 above, and further in view of US Patent 8,257,941 (September 4, 2012), hereafter referred to as Sakurada et al.
As set forth above, Mack in view of Hong et al., Rajesh et al., Cui et al., and Okita et al. provides the teachings and motivation to generate iPSCs from lymphoblastoid cell line cells as claimed in claims 23-32 and 36-37.
While Mack teaches that the iPS cells can be derived from the peripheral blood of patient with a somatic cell disorder, Mack does not specifically teach that the somatic cell disorder comprises a cell mutation or that the mutation is a disease mutation associated with a neurodegenerative disease/disorder or an inflammatory bowel disease/disorder. Sakurada et al. supplements Mack by teaching iPSC derived from patients having disease mutations, and more specifically a neurodegenerative disease where the patient has at least one endogenous allele associated with the neurodegenerative disease, or where the disease mutation is associated with inflammatory bowel disease such as Crohn’s disease (Sakurada et al., columns 1-2 and Table 3). Thus, based on motivation provided by Mack to generate iPSCs from peripheral blood of patient with a somatic disorder, and motivation provided by Sakurada et al. to generate iPS from patients have mutations associated with neurodegenerative disease or an inflammatory bowel disorder, it would have been prima facie obvious to the skilled artisan at the time of filing to generate iPSCs from lymphoblastoid cell line cells derived from the peripheral blood of patients with a neurodegenerative or inflammatory bowel disorder using the methods of Mack in view of Hong, Rajesh, and Okita with a reasonable expectation of success.
No claims are allowed.
Any inquiry concerning this communication from the examiner should be directed to Anne Marie S. Wehbé, Ph.D., whose telephone number is (571) 272-0737. If the examiner is not available, the examiner’s supervisor, Maria Leavitt, can be reached at (571) 272-1085. For all official communications, the technology center fax number is (571) 273-8300. Please note that all official communications and responses sent by fax must be directed to the technology center fax number. For informal, non-official communications only, the examiner’s direct fax number is (571) 273-0737. For any inquiry of a general nature, please call (571) 272-0547.
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Dr. A.M.S. Wehbé
/ANNE MARIE S WEHBE/Primary Examiner, Art Unit 1634