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 .
Amendments
In the reply filed 6/10/2026, Applicant has amended Claims 1, 15, 16, and added new claims, Claims 17-19.
Claim 14 is withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a non-elected invention, there being no allowable generic or linking claim.
Claims 1-12, 15-19 are under consideration.
Withdrawn Objection to the Specification
The objection to the disclosure because it contained an embedded hyperlink and/or other form of browser-executable code has been withdrawn due to Applicant’s amendment.
Withdrawn Claim Objections
The objection to Claim 16 has been withdrawn due to Applicant’s amendment.
Withdrawn 35 USC § 112
The prior rejection of Claim 16 under 35 U.S.C. § 112(b) pre-AIA 2nd paragraph as being indefinite is withdrawn in light of Applicant’s amendments of instant claims to describe the mixed culture supernatant.
Declaration under 37 CFR 1.132
The declaration under 37 CFR 1.132 filed by Manabu Itoh on 6/10/2026 is sufficient to overcome the rejection of instant claims based upon 35 U.S.C 102/103 as set forth in the last Office action as Mr. Itoh declares that hepatoblasts to do not support the growth and survival of human liver parenchymal cells.
Withdrawn 35 USC § 102
The prior rejection of Claims 15-16 under 35 U.S.C. 102(a)(1)/(a)(2) as being anticipated by Vallier et al. (WO2020/030822, filed 8/12/2019, published 2/13/2020) is withdrawn in light of Applicant’s amendment of Claim 15 to include the functional limitation that the nonparenchymal cell supports the growth and survival of human hepatic parenchymal cells.
Withdrawn 35 USC § 103
The prior rejection of Claims 1-9 under 35 U.S.C. 103 as being unpatentable over Ortega (US2018/0066233, filed 7/31/2017, published 3/08/2018 see IDS 7/11/2023), in view of Gupta et al., (US2015/0166953, filed 6/27/2013, published 6/18/2015) is withdrawn in light of Applicant’s amendment of Claim 1 to include the functional limitation that the nonparenchymal cell supports the growth and survival of human hepatic parenchymal cells.
The prior rejection of Claims 10-12, and 15-16 under 35 U.S.C. 103 as being unpatentable over Ortega (US2018/0066233, filed 7/31/2017, published 3/08/2018 see IDS 7/11/2023), in view of Gupta et al., (US2015/0166953, filed 6/27/2013, published 6/18/2015), as applied to claim 1, in further view of Vallier et al. (WO2020/030822, filed 8/12/2019, published 2/13/2020) is withdrawn in light of Applicant’s amendment of Claims 1 and 15 to include the functional limitation that the nonparenchymal cell supports the growth and survival of human hepatic parenchymal cells.
New 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 set forth in Graham v. John Deere Co., 383 U.S. 1, 148 USPQ 459 (1966), that are applied 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.
Hepatic stellate cell conditioned media embodiments
Claims 1-3, 5, 7-8, and 17 are rejected under 35 U.S.C. 103 as being unpatentable over Khetani et al., (US2018/0172668, filed 6/23/2026), in view of Akahira et al. (JP2017104057, filed 12/10/2015, published 6/15/2017, see IDS filed 12/27/2024)
In regard to claim 1, Kehtani claims a method for culturing a mixed culture of hepatocytes in a hepatocyte culture medium comprising:
a culture supernatant (i.e., conditioned media) of hepatic stellate cells (HSCs) (see Claim 44 of Kehtani).
In regard to the hepatocytes and HSCs of claim 1, Kehtani teaches they are human hepatocytes such as primary human hepatocytes (PHH) and primary HSCs from one or more donors [0012, 0014, 0016, 0018, 0020, 0022]. Furthermore, the use of human hepatocytes and human HSCs would have been obvious because of the clinical relevance for human liver fibrosis (see more below).
In regard to the hepatocyte culture media of claim 1, Kehtani teaches HSC conditioned media causes a down regulation of hepatocyte CYP3A4 activity, and adds a variety of agents in the hepatocyte culture media to help reduce the negative effects of HSCs on hepatocytes. For example, Kehtani demonstrates that when a TGF-beta inhibitor (SB431542) is combined with the HSC conditioned media to form the hepatocyte culture media, the treatment rescued the CYP3A4 activity of hepatocytes ([0042, 0139], Example 6, see Fig. 19B).
However, Ketani is silent to adding a Wnt signaling agonist and mitogenic growth factor to the hepatocyte culture media.
In regard to the hepatocyte culture media of claim 1, Akahira teaches a method of rescuing CYP3A4 cytochrome p450 activity of primary human hepatocytes in culture by adding Wnt signaling promoters to the hepatocyte culture media (p. 4, Overview, p. 16, [0016], p. 18, Example 1, p. 19, Example 2, see also Claim 1 of Akahira translation). Furthermore, Akahira teaches adding the mitogenic growth factor insulin to the hepatocyte culture media (p. 18, Example 1, [0025], see also Claim 4 of Akahira translation).
Accordingly, it would have been prima facie obvious to one of ordinary skill in the art at the time of filing to practice a method of culturing human hepatocytes in a culture media comprising the supernatant of HSC conditioned media and add one or more agents to rescue hepatocyte CYP3A4 levels as taught by Khetani and add a Wnt agonist and insulin growth factor to the hepatocyte media as taught by Akahira with a reasonable expectation of success. The ordinary skilled artisan would have been motivated to do so as taught by Akahira because the hepatocyte culture media that comprise the Wnt agonists Wnt3a and R-Spondin1 maintained the highest CYP3A4 activity (p. 19, Example 2, [0040] of translation). Furthermore, it would have been obvious to include the mitogenic growth factor insulin not only because it is included in the media of Akahira, but also because Kehtani teaches adding insulin to the co-culture medium [0080] and is part of the “normal hepatocyte maintenance media” [0044].
In regard to claims 2 and 3, as stated supra, Akahira teaches that the hepatocyte culture media comprising the Wnt agonists Wnt3a and R-Spondin1 maintained the highest CYP3A4 activity in hepatocytes.
In regard to claims 5 and 7, as stated supra, Kehtani teaches adding a TGF beta inhibitor in combination with the HSC conditioned media to ameliorate the negative effects of HSC conditioned media on hepatocytes ([0042, 0139], see Figs. 19). Furthermore, MPEP 2144.06 indicates that “it is prima facie obvious to combine two compositions each of which is taught by the prior art to be useful for the same purpose, in order to form a third composition to be used for the very same purpose.... [T]he idea of combining them flows logically from their having been individually taught in the prior art.” In re Kerkhoven, 626 F.2d 846, 850, 205 USPQ 1069, 1072 (CCPA 1980).
In regard to claims 5 and 8, Kehtani teaches the HSC release IL-6 into the conditioned media (see also [0037-0039], Figs. 14-16). Thus, Kehtani teaches the presence of IL-6 in the conditioned media from HSCs.
In regard to claim 17, as stated supra, Kehtani teaches human hepatic stellate cells.
Hence, the claimed invention as a whole was prima facie obvious in the absence of evidence to the contrary.
RESPONSE TO ARGUMENTS
Applicant's arguments filed on 6/10/2026 are acknowledged and have been addressed above.
Claims 10, 12, 15-16, and 18-19 are rejected under 35 U.S.C. 103 as being unpatentable over Khetani et al., (US2018/0172668, filed 6/23/2026), in view of Akahira et al. (JP2017104057, filed 12/10/2015, published 6/15/2017, see IDS filed 12/27/2024), as applied to claim 1, in further view of Kordes et al. (BBRC, 2008, 367:116-123)
As stated supra, Khetani et al. suggest a method for culturing human hepatocytes in a culture media comprising:
the supernatant of human HSC conditioned media,
the Wnt signaling promoters Wnt3a and R-Spondin1, and
the mitogenic growth factor insulin.
[AltContent: textbox ([img-media_image1.png])]In regard to claims 10 and 15, Kehtani teaches that the method for producing the conditioned media from the HSC dramatically changes the effects on the mixed culture of human hepatocytes (i.e., MPBCs). Specifically, Kehtani teaches that when HSC are cultured under conditions that activate the HSCs and the conditioned media is separated from the pure HSCs and added to the MPBCs this leads to a liver fibrosis model, while when HSCs are culture under conditions that reduced activation of HSC and the conditioned media is separated from the pure HSCs and added to the MPBCs this leads to a hepatocyte culture mimicking a liver closer to a healthy stated ([0028, 0030, 0041, 0138], Figs. 5, 7, 18, see Fig. 14A, adjacent).
In regard to producing the HSC conditioned media as per the HSC culture media of claims 10 and 15, Kehtani teaches adding insulin, which is a mitogenic growth factor, to the HSC culture medium to reduce their activation and lower IL-6 secretion into the conditioned media, and then treating the hepatocytes (i.e., MPBCs) with the HSC conditioned media ([0043-0044, 0140], Example 7, see Figs. 20-21).
However, in regard to producing the HSC conditioned media of claims 10 and 15, Kehtani et al. are silent to adding a Wnt signaling promoter to the HSC culture media when producing the HSC conditioned media.
In regard to claims 10 and 15, Kordes teaches that Wnt signaling maintains the quiescent stage of hepatic stellate cells, and this signal is required to prevent HSC differentiation. Specifically, Kordes demonstrates that HSC cultured in media comprising a Wnt signaling promoter that in inhibits GSK-beta (TWS119), reduces the HSC activation marker alpha-SMA (see Fig. 2E).
Accordingly, it would have been prima facie obvious to one of ordinary skill in the art at the time of filing to practice a method of producing conditioned media from HSCs by including insulin to reduce HSC activation and secretion of IL-6 as taught by Khetani et al. and further include a Wnt signaling promoter to maintain HSC quiescence as taught by Kordes with a reasonable expectation of success. The ordinary skilled artisan would have been motivated to do so because MPEP 2144.06 indicates that “it is prima facie obvious to combine two compositions each of which is taught by the prior art to be useful for the same purpose, in order to form a third composition to be used for the very same purpose.... [T]he idea of combining them flows logically from their having been individually taught in the prior art.” In re Kerkhoven, 626 F.2d 846, 850, 205 USPQ 1069, 1072 (CCPA 1980).
In regard to claim 12, as state supra, Kehtani teaches that while producing the HSC conditioned media, the HSC conditioned medium contains secreted IL-6 ([0037-0039], Figs. 14-16).
In regard to claim 16, as state supra, Kehtani in view of Akahira teach combining the HSC condition media with a Wnt signaling promoter and insulin to form the hepatocyte culture media.
In regard to claims 18-19, as stated supra, Kehtani teaches human hepatic stellate cells.
Hence, the claimed invention as a whole was prima facie obvious in the absence of evidence to the contrary.
RESPONSE TO ARGUMENTS
Applicant's arguments filed on 6/10/2026 are acknowledged and have been addressed above.
Claim 11 is rejected under 35 U.S.C. 103 as being unpatentable over Khetani et al., (US2018/0172668, filed 6/23/2026), in view of Akahira et al. (JP2017104057, filed 12/10/2015, published 6/15/2017, see IDS filed 12/27/2024) and Kordes et al. (BBRC, 2008, 367:116-123), as applied to claims 1 and 10 in further view of Murata (J Hep, 2001, 35:474-481)
As stated supra, Khetani et al. suggests a method for producing condition media from human HSCs that cultured in media to reduce their activation state and lower IL-6 secretion comprising:
a mitogenic growth factor (insulin),
a Wnt signaling promoter (GSK-beta inhibitor), and
collecting the conditioned media for culturing human hepatocytes.
However in regard to claim 11, although Kehatin teaches that the type of attachment and culture dish matrix can affect the activation state of HSCs and conditioned culture therefrom (see Fig. 16A), Khetani is silent to producing the conditioned media from the HSC with a ROCK inhibitor.
Murata teaches a method for producing a conditioned media from HSC with the ROCK inhibitor Y-27632 (p. 475, Materials & Methods, Section 2.4., Preparation of conditioned media).
Accordingly, it would have been prima facie obvious to one of ordinary skill in the art at the time of filing to practice a method of producing conditioned media from HSC as taught by Khetani and combine the ROCK inhibitor Y-27632 as taught by Murata with a reasonable expectation of success. The ordinary skilled artisan would have been motivated to do so as taught by Murata because the presence of Y-27632 reduces the activation of HSC (Abstract, p. 477, Discussion, 1st para.), thereby reducing the fibrotic effects of the conditioned media produced on hepatocytes. Furthermore, MPEP 2144.06 indicates that “it is prima facie obvious to combine two compositions each of which is taught by the prior art to be useful for the same purpose, in order to form a third composition to be used for the very same purpose.... [T]he idea of combining them flows logically from their having been individually taught in the prior art.” In re Kerkhoven, 626 F.2d 846, 850, 205 USPQ 1069, 1072 (CCPA 1980).
Hence, the claimed invention as a whole was prima facie obvious in the absence of evidence to the contrary.
RESPONSE TO ARGUMENTS
Applicant's arguments filed on 6/10/2026 are acknowledged and have been addressed above.
Kupffer cell conditioned media embodiments
Claims 1, 4-7, 9, and 17 are rejected under 35 U.S.C. 103 as being unpatentable over Clevers et al., (WO 2020/109324, filed 11/26/2019, see IDS filed 7/11/2023), in view of Katsumoto et al., (Hepatology, 1989, 9(3):405-410).
In regard to claims 1, 4-7, and 9, Clevers teaches a method for expanding cultured primary human hepatocytes (PHHs) in a hepatocyte culture medium (Hep-medium) comprising:
A Wnt signaling promoter (CHIR99021 and R-spondin1 conditioned medium),
A mitogenic factor (EGF and FGF),
Nicotinamide and the antioxidant N-acetylcysteine,
A ROCK inhibitor (Y-27632), and
A TGF-beta signaling inhibitor (A83-01)
(p. 21, “The hepatocyte culture medium (Hep-medium) of the invention” to p. 23 Table 2, “Hep-Medium (Human)” column, see also Fig. 12A, “PHH”).
However, Clevers is silent with respect to including the conditioned media of Kupffer cells in the hepatocyte expansion media.
In regard to claims 1 and 17, Katsumoto teaches a method of expanding primary hepatocytes in a culture media comprising the supernatant of conditioned media from Kupffer cells (Abstract, see Figs. 2,3 and 5).
Accordingly, it would have been prima facie obvious to one of ordinary skill in the art at the time of filing to practice the culture method of expanding primary human hepatocytes comprising a R-spondin conditioned media as taught by Clevers, and combine a Kupffer cell conditioned media as taught by Katsumoto with a reasonable expectation of success. The ordinary skilled artisan would have been motivated to do so because MPEP 2144.06 indicates that “it is prima facie obvious to combine two compositions each of which is taught by the prior art to be useful for the same purpose, in order to form a third composition to be used for the very same purpose.... [T]he idea of combining them flows logically from their having been individually taught in the prior art.” In re Kerkhoven, 626 F.2d 846, 850, 205 USPQ 1069, 1072 (CCPA 1980). Furthermore, it would have been obvious to use human primary Kupffer cells to avoid xeno-contamination, because Clevers teaches the human primary hepatocytes are eventually used in humans to treat human liver damage (p. 47, Example 8).
Hence, the claimed invention as a whole was prima facie obvious in the absence of evidence to the contrary.
RESPONSE TO ARGUMENTS
Applicant's arguments filed on 6/10/2026 are acknowledged and have been addressed above.
Conclusion
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 extension fee 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 date of this final action.
No claims are allowed.
Examiner Contact Information
Any inquiry concerning this communication or earlier communications from the examiner should be directed to ARTHUR S LEONARD whose telephone number is (571)270-3073. The examiner can normally be reached on Mon-Fri 9am-5pm.
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/ARTHUR S LEONARD/Examiner, Art Unit 1631
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