DETAILED ACTION
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
The amendment filed August 22, 2025, has been received and entered.
Claims 2-4, 13-15, and 17 are canceled.
Claims 1, 5-12, 16, 18, and 19 are pending. Claims 9-12, 18, and 19 are withdrawn.
Claims 1, 5-8, and 16 are examined on the merits.
Response to Amendment
It is noted that the amendment filed August 22, 2025, does not include all markings to indicate the changes that have been made relative to the immediate prior version of claim 1. In particular, the amendment fails to include a strike-through of the recitation “Interleukin 12 (IL-12), Interleukin 15” and the parentheses around the recitation “IL-15” in line 10 of the prior version of claim 1. Additionally, the amendment fails to include a strike-through of the recitation “further” in line 11 of the prior version of the claim.
As indicated in MPEP 714(II)(C), all claims being currently amended must be presented with markings to indicate the changes that have been made relative to the immediate prior version. It is a requirement set forth in 37 C.F.R. 1.121 (see 37 C.F.R. 1.121(c)(2)). MPEP 714(II)(F) states, “If an amendment submitted on or after July 30, 2003, fails to comply with 37 CFR 1.121 (as revised on June 30, 2003), the Office will notify applicant by a Notice of Non-Compliant Amendment, Form PTOL-324, that the amendment fails to comply with the requirements of 37 CFR 1.121 and identify: (1) which section of the amendment is non-compliant (e.g., the amendments to the claims section); (2) items that are required for compliance (e.g., a claim listing in compliance with 37 CFR 1.121(c) ); and (3) the reasons why the section of the amendment fails to comply with 37 CFR 1.121 (e.g., the status identifiers are missing)” (emphasis added). If a subsequent amendment fails to include all markings to indicate the changes, then a Notice of Non-Compliant Amendment will be mailed to Applicant.
Claim Objections
Claims 1, 5-8, and 16 are objected to because of the following informalities:
Claim 1 is objected to because it recites “and” between two commas before the last step (i.e. line 9). The comma after the recitation “and” should be deleted.
Additionally, claim 1 is objected to because the full name corresponding to the abbreviation “IL-15” in line 8 is not recited. Also, claim 1 is objected to because the full name corresponding to the abbreviation “IL-12” in line 10 is not recited. At the first instance an abbreviation appears in the claims, the full name corresponding to the abbreviation should be recited.
Since claim 1 is objected to, then its dependent claims, claims 5-8 and 16, are objected to.
Appropriate correction is required.
Notice Re: Prior Art Available Under Both Pre-AIA and 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.
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 1, 5-8, and 16 are rejected under 35 U.S.C. 103 as being unpatentable over Murphy (US 2018/0008637. Previously cited) in view of Roeven March 2014 (Bone Marrow Transplantation. March 2014. 49(Suppl. 1): ppS50. Previously cited), and Spanholtz `118 (WO 2013/119118. Previously cited).
Murphy discloses methods of producing natural killer (NK) cells from a population of hematopoietic stem or progenitor cells using a three-stage expansion and differentiation method with media comprising stem cell mobilizing factors (abstract; paragraph [0002]). In certain embodiments, the hematopoietic cells are CD34+ cells (paragraph [0300]), and Murphy refers to CD34+ stem cells or progenitors cells as an example of the hematopoietic stem cells or progenitor cells when setting forth the steps of the method (paragraph [0312]). The method comprises cell expansion, during which a plurality of hematopoietic cells within the hematopoietic cell population differentiate into NK cells (paragraph [0312]). The NK cells are CD56+, CD3-, and at least 70% of the natural killer cells are viable with certain embodiments (paragraph [0312]).
In one embodiment, the method comprises:
culturing hematopoietic stem cells or progenitor cells, e.g., CD34+ stem cells or progenitors cells, in a first medium comprising a stem cell mobilizing agent and thrombopoietin (Tpo) to produce a first population of cells, wherein the first medium further comprises one or more of a group of 7 components that include Flt-3L, SCF, and IL-7 (paragraph [0317};
subsequently culturing the cells in a second medium comprising a stem cell mobilizing agent and interleukin-15 (IL-15) and lacking Tpo to produce a second population of cells, wherein the second medium further comprises one or more of 7 components that include Flt-3[L], SCF, and IL-7 (paragraph [0318]);
subsequently culturing the cells in a third medium comprising IL-2 and IL-15, and lacking a stem cell mobilizing agent and LMWH (low-molecular weight heparin; see page 1, paragraph [0009]) to produce a third population of cells, e.g., natural killer cells, wherein the third medium further comprises one or more of 5 components that include SCF and IL-7 (paragraph [0319]).
In one embodiment, the stem cell mobilizing compound is an aryl hydrocarbon receptor inhibitor, e.g., an aryl hydrocarbon receptor antagonist (paragraph [0368]). Thus Murphy teaches an embodiment of the above method in which the first medium and the second medium each comprise an aryl hydrocarbon receptor antagonist, and the third medium lacks an aryl hydrocarbon receptor antagonist; these meet limitations of the claimed expansion medium, first differentiation medium, and second differentiation medium, respectively.
In performing the method of Murphy, it would have been prima facie obvious to the skilled artisan to include any combination of the additional components (7 components for the first and second mediums; 5 components for the third medium) in the three mediums since Murphy teaches “one or more of” the list of additional components for inclusion in each of the three mediums (paragraphs [0317]-[0319]). Therefore, it would have been prima facie obvious to select Flt-3L, SCF, and IL-7 as the additional components of the first and second mediums of Murphy for the predictable result of culturing the cells to produce a first and second populations of cells in the respective steps. Further still, it would have been prima facie obvious to select SCF and IL-7 as the additional components of the third medium of Murphy for the predictable result of culturing the cells to finally produce a third population of cells, specifically of NK cells. The first medium rendered obvious by Murphy meets limitations of the claimed ‘expansion medium,’ the second medium rendered obvious by Murphy meets limitations of the claimed ‘first differentiation medium,’ and the third medium rendered obvious by Murphy meets limitations of the claimed ‘second differentiation medium.’
Additionally, since glycosaminoglycans are not disclosed for inclusion in the three mediums of Murphy, then Murphy meets the claimed proviso of mediums that do not comprise a glycosaminoglycan.
In sum, Murphy renders obvious a method meeting limitations of the claimed invention since Murphy renders obvious a method for the ex vivo production of a population of NK cells from CD34+ cells comprising culturing CD34+ cells in a first medium (meeting limitations of the claimed ‘expansion medium’) and subsequently culturing the cultured CD34+ cells in a second medium (meeting limitations of the claimed ‘first differentiation medium’ which is directed to a ‘differentiation medium’), comprising:
culturing in the first medium rendered obvious by Murphy comprising IL-7, SCF, TPO, Flt3L, and an aryl hydrocarbon receptor antagonist (meets limitations of the claimed ‘expansion medium’);
further culturing the expanded CD34+ cells in the second medium rendered obvious by Murphy comprising IL-7, SCF, Flt3L, IL-15, and an aryl hydrocarbon receptor antagonist (meets limitations of the claimed ‘first differentiation medium’); and
culturing in the third medium rendered obvious by Murphy comprising IL-7, SCF, and IL-15, and does not comprise an aryl hydrocarbon receptor antagonist (meets limitations of the claimed ‘second differentiation medium’),
with the proviso that the first, second, and third mediums do not comprise a glycosaminoglycan.
Murphy differs from the claimed invention in that Murphy does not expressly disclose:
the first, second, and third mediums do not contain G-CSF, GM-CSF, and IL-6, and the third medium comprises Interleukin 12 (IL-12); and
the NK cells are ‘highly functional’ which is defined as having at least 10% CD-56 positive cells being capable of secreting IFN-gamma upon stimulation with K562 target cells, and at least 1E+9 highly functional NK cells are produced from a single donor.
Regarding difference (1) (the first, second, and third mediums do not contain G-CSF, GM-CSF, and IL-6, and the third medium comprises Interleukin 12 (IL-12)):
Roeven March 2014 discloses ex vivo generation of NK cells from bone marrow (BM) or mobilized peripheral blood (PB) derived stem cells (“Introduction” paragraph). The culture process comprises expanding CD34+ cells in a medium containing SCF, Flt3L, TPO, IL-7, IL-15, and StemReginin1 (SR1) (“Materials (or patients) and Methods” paragraph). It is noted that Murphy recognizes SR1 as an aryl hydrocarbon receptor inhibitor (paragraph [0022]). Subsequently, the expanded cells were differentiated into NK cells using IL-15 (“Materials (or patients) and Methods” paragraph).
Spanholtz `118 discloses a method for producing natural killer (NK) cells comprising (i) providing a sample of human CD34 positive cells; (ii) expanding said CD34 positive cells ex vivo; and (iii) culturing CD34 positive cells obtained in step ii ex vivo in an NK-cell differentiation medium, wherein the NK-differentiation medium comprises IL-12 (page 2, line 26 through page 3, line 4). Spanholtz `118 discloses that IL-12 modulates ex vivo NK cell differentiation (abstract) and provides the finished NK cell product with new and/or enhanced properties (page 2, third paragraph). In particular, Spanholtz `118 discloses an example of the NK-cell differentiation medium comprising IL-7, IL-15, SCF, and FLT-3L (page 8, lines 25-28). Further still, that example of the NK-cell differentiation medium further comprises IL-12 for the purpose of the Spanholtz `118 invention (page 9, lines 1-3).
As discussed above, it would have been prima facie obvious to select Flt-3L, SCF, and IL-7 for inclusion in the first medium of Murphy which meets limitations the claimed ‘expansion medium.’ In making that selection, then the first medium of Murphy does not comprise G-CSF, GM-CSF, and IL-6. Moreover, before the effective filing date of the claimed invention, it would have been obvious to select Flt-3L, SCF, and IL-7 as the additional components, excluding IL-6, G-CSF, and GM-CSF as additional components, in the first medium of Murphy because Roeven March 2014 teaches a medium that contains the same components of the first medium rendered obvious by Murphy (SCF, Flt-3L, TPO, IL-7, IL-15, an aryl hydrocarbon receptor antagonist) for initial culturing of CD34+ cells for the purpose of obtaining NK cells after differentiation, while not requiring IL-6, G-CSF, and GM-CSF. There would have been a reasonable expectation in producing NK cells by the first medium rendered obvious by Murphy that does not include IL-6, G-CSF, and GM-CSF since Roeven March 2014 demonstrates that such a medium is suitable for initial culturing of CD34+ cells for NK cells production.
Additionally, as discussed above, it would have been prima facie obvious to select Flt-3L, SCF, and IL-7 for inclusion in the second medium of Murphy (meeting limitations of the claimed ‘first differentiation medium’) and selecting SCF and IL-7 for inclusion in the third medium of Murphy (meeting limitations of the claimed ‘second differentiation medium’). In making those selections, the second and third mediums of Murphy do not comprise G-CSF, GM-CSF, and IL-6. Moreover, before the effective filing date of the claimed invention, it would have been obvious to select Flt-3L, SCF, and IL-7 as the additional components, excluding IL-6, G-CSF, and GM-CSF as additional components, in the second medium of Murphy, as well as selecting SCF and IL-7 as the additional components, excluding IL-6, G-CSF, and GM-CSF as additional components, in the third medium of Murphy because Spanholtz `118 teaches an NK-cell differentiation medium comprising IL-7, IL-15, SCF, and FLT-3L for a process in which CD34+ cells are differentiated into NK cells, while not requiring IL-6, G-CSF, and GM-CSF. There would have been a reasonable expectation in producing NK cells using the second and third mediums rendered obvious by Murphy that do not include IL-6, G-CSF, and GM-CSF since Spanholtz `118 teaches that a medium comprising IL-7, IL-15, SCF, and FLT-3 (some of the same components as Murphy’s second and third mediums) that does not include IL-6, G-CSF, and GM-CSF is suitable for the differentiation of CD34+ cells into NK cells.
Further still, before the effective filing date of the claimed invention, it would have been obvious to include IL-12 in the third medium when performing the method of Murphy. One of ordinary skill in the art would have been motivated to do this because IL-12 modulates ex vivo NK cell differentiation and provides an NK cell product with new and/or enhanced properties. There would have been a reasonable expectation of producing NK cells as sought by Murphy since Spanholtz `118 teaches the inclusion of IL-12 in culture medium for differentiation of CD34+ cells into NK cells.
As such, Murphy in view of Roeven March 2014 and Spanholtz `118 renders obvious a first medium that comprises the same materials as the claimed ‘expansion medium,’ a second medium that comprises the same materials as the claimed ‘first differentiation medium,’ and a third medium that comprises the same materials as the claimed ‘second differentiation medium.’ The instant specification states, “An expansion medium according to the invention and a differentiation medium according to the invention comprise a basic medium supplemented with further components as defined herein” (page 7, lines 6-8 of substitute specification filed June 16, 2022). Since Murphy in view of Roeven March 2014 and Spanholtz `118 renders obvious a first medium comprising IL-7, SCF, TPO, Flt3L, and an aryl hydrogen receptor antagonist which are the same materials comprised by the claimed ‘expansion medium’ and identical to the expansion medium of Roeven March 2014, then the first medium rendered obvious by Murphy, Roeven March 2014, and Spanholtz `118 is directed to an ‘expansion medium’ and the culturing in the first medium results in obtaining expanded CD34+ cells (hematopoietic cells which are CD34+ cells). Moreover, there would have been a reasonable expectation of obtaining expanded CD34+ cells from the culturing in the first medium rendered obvious by Murphy in view of Roeven March 2014 and Spanholtz `118 since Roeven March 2014 demonstrates that a medium comprising the same materials is directed to an expansion medium and culturing in that expansion medium results in expanding CD34+ cells.
Likewise, since Murphy in view of Roeven March 2014 and Spanholtz `118 renders obvious a third medium comprising IL-7, SCF, IL-12, and IL-15 which are the same materials comprised by the claimed ‘second differentiation medium’ and comprises some of the same materials as the NK-differentiation medium of Spanholtz `118, then the third medium rendered obvious by Murphy in view of Roeven March 2014 and Spanholtz `118 is directed to a ‘differentiation medium.’
Regarding difference (2) (Murphy does not disclose that the NK cells are ‘highly functional’ which is defined as having at least 10% CD-56 positive cells being capable of secreting IFN-gamma upon stimulation with K562 target cells, and at least 1E+9 highly functional NK cells are produced from a single donor):
Spanholtz `118 teaches that early studies indicated the potency of IL-12 to modulate the differentiation towards a cytotoxic and IFN-gamma producing NK cell (page 29, lines 23-24). In an experiment, the killing efficiency against K562 cells of the IL-12 induced ex vivo generated NK cells was tested (page 26, lines 12-19). See Figure 7 and page 17, lines 10-15. The results in Figure 7 show an enhanced cytotoxic activity of the IL-12 induced ex vivo differentiated NK cells against the K562 cells.
Since Murphy in view of Roeven March 2014 and Spanholtz `118 renders obvious the same steps of the claimed invention, then the produced NK cells necessarily possess the claimed property of being ‘highly functional’ defined as having at least 10% CD56 positive cells being capable of secreting IFN-gamma upon stimulation with K562 target cells, and at least 1E+9 ‘highly functional’ NK cells are necessarily produced from a single donor. Further still, there would have been a reasonable expectation that at least 10% CD56 positive cells are capable of secreting IFN-gamma upon stimulation with K562 target cells because the studies disclosed in Spanholtz `118 found that IL-12 modulates the differentiation towards an IFN-gamma producing NK cell, and that IL-12 induced ex vivo generated NK cells had an enhanced cytotoxic activity against K562 cells. Since IL-12 is included in the third medium of the method rendered by the references, then the claimed IFN-gamma secretion upon stimulation with K562 target cells would have been expected.
Therefore, Murphy in view of Roeven March 2014 and Spanholtz `118 renders obvious instant claim 1.
Regarding instant claim 5, Murphy discloses that in certain embodiments, the stem cell mobilizing compound is StemRegenin-1 (SR-1) (paragraph [0397]), which is an aryl hydrocarbon receptor inhibitor (paragraph [0022]), i.e. aryl hydrocarbon receptor antagonist. SR-1 was used in Examples 1 and 2 as a component of Stage 1 (referring to the step of culturing in a first medium of Murphy) and Stage 2 (referring to the step of culturing in a second medium of Murphy) media of the three-stage method of Murphy (paragraphs [0621]-[0622]; paragraph [0630]). Examples 1 and 2 of Murphy teach culturing CD34+ cells (Example 2 is according to Example 1; see paragraphs [0630] and [0620]). Thus, instant claim 5 (elected species SR1) is rendered obvious.
Regarding instant claim 6, Murphy teaches that the cells are cultured in the first medium for a range of days, including 7, 8, 9, and 10 days, before culturing in the second medium (paragraph [0221]). This is directed to the limitation of instant claim 6 of culturing in the expansion medium for about 7 to 10 days. Additionally, the cells are cultured in the second medium (directed to the claimed ‘first differentiation medium’) for 9, 10, 11, 12, 13, and 14 days, before culturing in the third medium (paragraph [0221]), which meets the limitation of instant claim 6 of culturing in the first differentiation medium from about day 7 to 10 (as would follow according to the teaching regarding the length of time for the first step) to about day 19 to 21 (21-7 days = 14 days maximum; 19-10 days = 9 days minimum). Furthermore, the cells can be cultured in the third medium (directed to the claimed ‘second differentiation medium’) for 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 days, or for more than 30 days (paragraph [0221]), meeting the limitation of instant claim 6 of culturing in the second differentiation medium from about day 19 to 21 (as would follow according to the teachings regarding the length of time for the other steps) for at least about 10 days. Therefore, instant claim 6 is rendered obvious.
Regarding instant claim 7, Murphy in view of Roeven March 2014 and Spanholtz `118 differs from the claimed invention in that they do not expressly disclose that the at least about 1E+9 highly functional NK cells produced from a single donor (see rejection of instant claim 1) are ‘highly pure’ CD56-positive, Perforin-positive and Eomesodermin (EOMES)-positive NK cells, wherein ‘highly pure’ is defined as the amount of CD3-positive cells in the population of NK cells being at most about 0.1%. However, Murphy discloses that the NK cells produced by their method are CD3-negative (paragraph [0009]). Moreover, Murphy teaches that the NK cell population produced by their method comprises no less than 99% CD3-CD56+ cells (paragraph [0500]). Since Murphy in view of Roeven March 2014 and Spanholtz `118 renders obvious the same steps of the claimed invention, then the same effects, including obtaining NK cells that are CD56-positive, Perforin-positive and EOMES-positive and obtaining the same percentage of cells that are differentiated into NK cells, would have necessarily occurred. Therefore, the percentage of cells that are not differentiated into NK cells would have necessarily occurred as claimed, specifically the amount of CD3-positive cells (i.e. cells that are not NK cells) being in the range of ‘at most about 0.1%’ recited in instant claim 7, particularly in view of the fact that Murphy teaches the embodiment of no less than 99% of the NK cell population are CD3-negative (thus cells that are not CD3-negative, i.e. CD3-positive cells, would be 1% or less). Furthermore, Spanholtz `118 teaches that the acquisition of cytotoxic and IFN-gamma producing NK cell functions by IL-12 was correlated with induced expression of the perforin gene (page 5, lines 14-17). Since IL-12 is included in the third medium of the method rendered obvious by Murphy in view of Roeven March 2014 and Spanholtz `118, then there would have been a reasonable expectation that the produced NK cells are Perforin-positive. Therefore, instant claim 7 is rendered obvious.
Regarding instant claim 8, see Figure 1A of Murphy, which shows the fold expansion for various culturing conditions, including ‘NK cell expansion’ for previous NK expansion media resulting in about 1000 fold expansion as compared to Murphy’s three-stage method using SR1 (an aryl hydrogen receptor antagonist) at 1 µM which resulted in 10,000 fold expansion (paragraph [0278]). The greater fold expansion using SR1 is directed to a difference falling in the range of ‘at least about two-fold higher’ of instant claim 8. In including SR1 as the aryl hydrogen receptor antagonist of the method rendered obvious by Murphy in view of Roeven March 2014 and Spanholtz `118, then the same difference in mean overall expansion as compared to when no aryl hydrocarbon receptor antagonist is used, would have been expected. Therefore, instant claim 8 is rendered obvious.
Regarding instant claim 16, Murphy discloses that the hematopoietic cells can be obtained from tissue sources such as bone marrow and peripheral blood (paragraph [0296]). In another embodiment, the hematopoietic cells are obtained from umbilical cord blood (paragraph [0297]). Moreover, Murphy discloses in Example 3 cultivating UCB (umbilical cord blood)) CD34+ cells to produce NK cells (paragraph [0632]). Therefore, instant claim 16 is rendered obvious.
Response to Arguments
Applicant’s arguments, filed August 22, 2025, with respect to the objections to claims 1, 5-8, and 16, the rejections under 35 U.S.C. 112(b) of claims 1, 5-8, and 16, the rejection under 35 U.S.C. 112(a) of claims 1, 5-8, and 16, and the rejection under 35 U.S.C. 103 of claims 1, 5-8, and 16 as being unpatentable over Murphy in view of Roeven March 2014, Spanholtz `118, and Spanholtz `539, have been fully considered and are persuasive. In particular, the claim objections have been overcome by the amendment to claim 1. The rejections under 35 U.S.C. 112(b) have been overcome by the amendments to claims 1 and 6. The rejection under 35 U.S.C. 103 has been overcome by the amendment to claim 1 since it removes the inclusion of TPO and IL-12 from the first differentiation medium. Spanholtz `539 was cited in the last Office Action to render obvious including TPO in the second medium of Murphy (which meets limitations of the claimed ‘first differentiation medium’). Therefore, these objections and rejections have been withdrawn.
However, upon further consideration, a new ground(s) of rejection is made under 35 U.S.C. 103 in view of the previously cited references Murphy, Roeven March 2014, and Spanholtz `118, as necessitated by the amendments to the claims.
To the extent Applicant’s arguments are directed to the new ground of rejection under 35 U.S.C. 103, Applicant’s arguments are unpersuasive. In the last Office Action and repeated above, Roeven March 2014 and Spanholtz `118 are cited to render obvious difference (1): the first, second, and third mediums do not contain G-CSF, GM-CSF, and IL-6, and the third medium comprises IL-12. Applicant disagrees that Roeven March 2014 and Spanholtz `118 make up for difference (1), arguing that the technical effect obtained in the comparative examples 1 and 2 in the instant application could not have been foreseen by one skilled in the art. Applicant points out that in example 1, NK cells are produced in media wherein an aryl hydrocarbon receptor antagonist is present together with a glycosaminoglycan, G-CSF, GM-CSF, and IL-6, the resulting NK cells produce 170 pg/ml IFN-gamma upon stimulation with K562 cells (e.g., Figure 5A). Also, Applicant states that in example 2, NK cells are produced in a method (asserted by Applicant as claimed) in a differentiation medium wherein an aryl hydrocarbon antagonist is present while a glycosaminoglycan and G-CSF, GM-CSF, and IL-6 are absent, the resulting NK cells product 700 pg/mL IFN-gamma upon stimulation with K562 cells (e.g., Figure 10D), i.e. a four-fold increase in IFN-gamma production. Applicant argues that the big difference between examples 1 and 2 are the lack of a glycosaminoglycan, G-CSF, GM-CSF, and IL-6 in the media of example 2. It is asserted that the technical effect conferred by the difference is nowhere suggested in the prior art.
However, in example 1 of the specification, the NK cells were generated from CD34+ HSPCs isolated from bone marrow samples collected from healthy donors in the presence of SR1 (directed to an aryl hydrocarbon antagonist), with the activity of the cells investigated after 5 weeks (page 20, lines 32-33; page 29, lines 22-31 of originally filed specification). In example 1, the cells were expanded during 9 or 10 days with a high-dose cytokine cocktail (Expansion cocktail I) consisting of 25 ng/mL IL-7, 25 ng/mL SCF, 25 ng/mL TPO, and 25 ng/ml Flt3L; cultured from day 9 or 10 until day 14 or 15 in a medium in which TPO was replaced by 25 ng/mL IL-15; and cultured, where cell differentiation was initiated, after day 14 or 15 by replacing Expansion cocktail I by a new high-dose cytokine cocktail (Differentiation cocktail) consisting of 20 ng/mL IL-7, 20 ng/mL SCF, 20 ng/mL IL-15, and 1000 U/mL IL-12 (page 30, lines 2-9). Figure 1A shows that 2 µM SR1 was also included in all of the media (page 30, lines 9-11). Example 1 also states that cells were cultured in Cellgro GMP DC medium supplemented with 10% human serum and a low dose cytokine cocktail consisting of G-CSF, GM-CSF, and IL-6 (page 30, lines 11-15).
On the other hand, in example 2 of the specification, the NK cells were generated from CD34+ HSPCs isolated from umbilical cord blood (UCB) in the presence of SR1 (directed to an aryl hydrocarbon antagonist), and a phenotypical analysis was performed after 6 weeks (page 23, lines 22-23; page 45, lines 23-29). CD34+ cells were first cultured for 9 or 10 days with 2 µM SR1, 25 ng/mL IL-7, 25 ng/mL SCF, 25 ng/mL Flt3L, and 25 ng/mL TPO; from day 9 or 10 until day 14 or 15, cultured in a medium in which TPO was replaced by 50 ng/mL IL-15; and cultured from day 14 or 15 in differentiation medium consisting of 20 ng/mL IL-7, 20 ng/mL SCF, 50 ng/mL IL-15, and 0.2 ng/mL IL-12; SR1 was added till day 21 (page 45, line 33 through page 46, line 6). Also, the culture were performed over 5-6 weeks using Cellgro DC medium or NK MACS medium supplemented with 10% and 2% human serum during the expansion and the differentiation phase, respectively (page 45, lines 29-33).
Examples 1 and 2 have a couple of other differences besides the difference pointed out by Applicant (example 1 cultured the cells in a Cellgro GMP DC medium supplemented with a low dose cytokine cocktail consisting of G-CSF, GM-CSF, and IL-6, which is not taught in example 2). First, example 2 states that SR1 was added till day 21 (page 46, line 6), whereas SR1 was present in example 1 throughout the 5 week, i.e. 35 days, time period (page 30, lines 9-11; Figure 1). Also, example 2 required that 50 ng/mL IL-15 and 0.2 ng/mL IL-12 were included in the differentiation medium after day 14 or 15 (page 46, lines 4-6), whereas example 1 required different concentrations of IL-15 and IL-12, specifically 20 ng/mL IL-15 and 1000 U/mL IL-12 (page 30, lines 6-9). Given these differences, then it cannot be concluded that the increase in IFN-gamma production is necessarily because the media of example 2 lack glycosaminoglycan, G-CSF, GM-CSF, and IL-6.
Moreover, contrary to Applicant’s assertion, example 2 of the specification is not directed to the method as claimed. In particular, example 2 states that SR1 (directed to an aryl hydrocarbon receptor antagonist) was added till day 21 (page 46, line 6). The differentiation medium in example 2 after day 14 or 15 meets limitations of the claimed ‘second differentiation medium’ except that SR1 remains in that differentiation medium until day 21. Thus, example 2 fails to meet the claimed limitation of ‘culturing in a second differentiation medium comprising IL-7, SCF, IL-12 and IL-15…with the proviso that…the second differentiation medium does not comprise an aryl hydrocarbon receptor antagonist’ (instant claim 1). Since the evidence of example 2 is not commensurate is scope with the claimed invention, then it is unpersuasive as evidence of unexpected results to overcome the rejection under 35 U.S.C. 103.
The Declaration of Harmen Dolstra pursuant to 37 C.F.R. 1.130(a) was filed August 22, 2025, and has been considered by the Examiner. Applicant refers to the Declaration as a Declaration under 37 C.F.R. 1.132. Applicant asserts that Roeven March 2014 is a reflection of Example 1 of the instant application, and that the complete study was later published by Roeven in 2015 (STEM CELLS AND DEVELOPMENT, Volume 24, Number 24, 2015). This is set forth in paragraph 10 of the Declaration. Paragraph 11 of the Declaration also states that the experiments conducted in Roeven March 2014, example 1 of the instant application, and the Roeven 2015 article, all use heparin and a low dose cytokine cocktail (G-CSF, GM-CSF, and IL-6). Paragraph 12 of the Declaration states that the heparin and low dose cytokines were not mentioned in Roeven March 2014 due to lack of space in the abstract, but they were present in the experiments. Paragraph 13 refers to a lab journal entry dated December 18, 2013, which demonstrates that the protocol used the low dose cytokine cocktail. In view of the Declaration, Applicant asserts that Roeven March 2014 and Spanholtz `118 cannot remedy the deficiencies noted in at least difference point 1) in Murphy. However, in reading Roeven March 2014, the skilled artisan would not have recognized that the media used in the reference included G-CSF, GM-CSF, and IL-6. As pointed out by MPEP 2123(I), “A reference may be relied upon for all that it would have reasonably suggested to one having ordinary skill in the art, including nonpreferred embodiments.” Since G-CSF, GM-CSF, and IL-6 are not disclosed in Roeven March 2014, before the effective filing date of the claimed invention, the person of ordinary skill in the art would not have recognized that they are necessary for the ex vivo generation of NK cells. As such, Applicant’s arguments based on the Declaration are unpersuasive.
Additionally, Applicant asserts that it is not correct, when assessing obviousness to recreate the subject matter of a claim including its limitations, to pick elements from the art to derive the claimed subject matter and then conclude that a technical effect would be inherent to such subject matter. Applicant cites Ex parte Vigano (Appeal No. 2010-007666, March 13, 2012), to support this. However, this Board decision is not precedential and thus Applicant’s argument is not considered persuasive.
Conclusion
No claims are allowed.
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
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/SUSAN E. FERNANDEZ/Examiner, Art Unit 1651
/DAVID W BERKE-SCHLESSEL/Primary Examiner, Art Unit 1651