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 .
DETAILED ACTION
Continued Examination Under 37 CFR 1.114
A request for continued examination under 37 CFR 1.114, including the fee set forth in 37 CFR 1.17(e), was filed in this application after final rejection. Since this application is eligible for continued examination under 37 CFR 1.114, and the fee set forth in 37 CFR 1.17(e) has been timely paid, the finality of the previous Office action has been withdrawn pursuant to 37 CFR 1.114. Applicant's submission filed on 11/19/2025 has been entered.
Applicant' s amendment and response filed on 11/19/2025 has been received and entered into the case.
Amendments
In the reply filed 11/19/2025, Applicant has amended claim 1.
Claim Status
Claims 1, 3-11, 13-17, 19, 25 and 27-28 are pending.
Claims 9-11, 13-17, 19, 25 and 27 have been withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim.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 06/09/2023.
Claims 1, 3-8 and 28 are considered on the merits.
Withdrawn Claim Rejections - 35 USC § 103
The prior rejection of claims 1, 3-5 and 28 under 35 U.S.C. 103 as being unpatentable over Katare et al., (PLoS ONE. 2010; 5(2): e9275. P. 1-5) in view of Carrier et al., (Biotechnol Bioeng. 2002;78(6):617-25) is withdrawn in light of Applicant’s amendment to claim 1 to recite new limitation regarding TUNEL assay that is not taught by Katare or Carrier.
The prior rejection of claims 6-8 under 35 U.S.C. 103 as being unpatentable over Katare et al., (PLoS ONE. 2010; 5(2): e9275. P. 1-5) in view of Carrier et al., (Biotechnol Bioeng. 2002;78(6):617-25), as applied to claim 1 above, and further in view of Tulloch et al (Circulation Research. 2011; 109: 47-59. Prior art of record) is withdrawn in light of Applicant’s amendment to claim 1 to recite new limitation regarding TUNEL assay that is not taught by Katare or Carrier.
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 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.
Claims 1, 3-5 and 28 are rejected under 35 U.S.C. 103 as being unpatentable over Katare et al., (PLoS ONE. 2010; 5(2): e9275. P. 1-5. Prior art of record) in view of Bursac et al., (Am J Physiol. 1999;277(2):H433-44) and Li et al., (Sci Rep. 2016:6:37114, p. 1-14).
With respect to claim 1, Katare teaches an engineered heart tissue (EHT) by mixing cardiomyocytes isolated from neonatal rats and cultured in a ring-shaped scaffold suspended in culture medium (see e.g., Fig 3B and online supplemental video) and teaches the EHT is subjected to hypoxia serving as an experimental model in studying the acute ischemia-induced changes in vitro (abstract, see e.g., Fig 3B), thus teaches the preamble an in vitro three-dimensional (3D) myocardial infarct organoid suspended in a culture medium.
Katare teaches cardiomyocytes are isolated from neonatal rats and the resulting cell population (50% cardiomyocytes / 50% nonmyocytes) is subjected to EHT generation (p. 1, right col, para “Cell isolation”), thus teaches the organoid comprises cardiomyocytes and non-myocytes.
Katare teaches the organoid subjected to hypoxia has down-regulation of cell survival proteins Akt and Bcl-2 (p. 3, right col, para 1, see Fig 4), indicating the organoid has reduced cell survival (i.e., increased apoptosis) due to lack of oxygen (i.e., hypoxia).
Katare teaches the organoids have spontaneous contracting (see abstract), and when subjected to hypoxia, EHT demonstrated conduction defects and the synchronous conduction was lost (abstract, p. 3, right col, para 1, see Fig 3B (i) and legend), thus teaches the 3D organoid beats spontaneously in an asynchronous manner.
Katare teaches in Figure 3B (i) that the exemplary organoid subjected to hypoxia has two beats with a duration of about 740 ms in between (~4 divisions x 185 ms/division, see the 1st row for the two beats and see the bottom right of the 3rd row in Fig 3B (i) for the unit 185 ms/div), thus teaches the organoid has (b) a beat rate of about 0 to 90 beats per minute (60 s / 740 ms = 81 beats per minute).
However, Katare is silent on the organoid comprising an apoptotic interior region surrounded by a viable periphery that comprises a region of about 20 µm to about 75 µm from the organoid edge.
Bursac teaches a three-dimensional (3D) in vitro model of engineered cardiac muscle tissue constructs (e.g., title and abstract, equivalent to the engineered heart tissue of Katare). Bursac teaches “Structurally, constructs… contained a 50- to 70-μm-thick outer cardiac tissue-like zone composed of cells that expressed sarcomeric tropomyosin (Fig. 2C) and contained myofilaments, desmosomes, and intercalated disks (Fig. 3D). … The small thickness of the cardiac tissue-like zone in constructs (Fig. 2A)… can be attributed to the low survival rate of metabolically demanding cardiac myocytes located more than 50 µm from a source of gas exchange” (p. 441, right col, para 2). Bursac teaches “at the construct center, cells were sparsely distributed and either elongated, expressing tropomyosin, or round, with pyknotic nuclei and acidophilic cytoplasm (Fig. 2B)” (p. 437, left col., section “Construct morphology”). One of ordinary skill in the art would have immediately expected that those cells that are “round, with pyknotic nuclei and acidophilic cytoplasm” would be apoptotic cells. In summary, Bursac teaches an in vitro 3D engineered heart tissue organoid that comprises an apoptotic interior region due to lack of oxygen (i.e., the region having “low survival rate of metabolically demanding cardiac myocytes located more than 50 µm from a source of gas exchange” and having apoptotic cells that are ”round, with pyknotic nuclei and acidophilic cytoplasm”), surrounded by a viable periphery that comprises a region of about 20 µm to about 75 µm from the organoid edge (i.e., the “50- to 70-μm-thick outer cardiac tissue-like zone composed of cells that expressed sarcomeric tropomyosin”).
Therefore, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to have expected that the 3D myocardial infarct organoid subjected to hypoxia condition of Katare would have comprised an apoptotic interior region due to lack of oxygen surrounded by a viable periphery that comprises a region of about 20 µm to about 75 µm from the organoid edge as suggested by Bursac with a reasonable expectation of success, because Bursac teaches an equivalent cardiac organoid comprises a center region (i.e., an apoptotic interior region) having low survival of cardiac myocytes located more than 50 µm from a source of gas exchange and having apoptotic cells, surrounded by a 50- to 70-μm-thick outer cardiac tissue-like zone (i.e., a viable periphery) (see above) and since Katare teaches the organoid subjected to hypoxia has down-regulation of cell survival proteins (p. 3, right col, para 1, see Fig 4).
However, Katare and Bursac are silent on a TUNEL assay and positive ratio in the apoptotic interior region in an organoid cross-section.
PNG
media_image1.png
608
808
media_image1.png
Greyscale
Nevertheless, as stated supra, Katare teaches the organoid subjected to hypoxia has down-regulation of cell survival proteins Akt and Bcl-2 (p. 3, right col, para 1, see Fig 4). Additionally, Bursac teaches “at the construct center, cells were sparsely distributed and either elongated, expressing tropomyosin, or round, with pyknotic nuclei and acidophilic cytoplasm (Fig. 2B)” (p. 437, right col. Modified Fig 2B is attached, with arrows indicating elongated viable cells and arrow heads indicating apoptotic cells marked by examiner. The apoptotic cells are about 5/15 = ~33%).
In regard to a TUNEL assay, Li uses a model of myocardial ischemia/reperfusion (MI/R) injury to explore a treatment (e.g., abstract) and teaches a TUNEL assay on the cross-section of the MI/R tissue, “Following MI/R, vehicle-treated mice exhibited obvious TUNEL-positive (apoptosis) cells in ischemic myocardium (Fig 3A,B)” and “Furthermore, as shown in Fig. 3C–E, MI/R resulted in a noticeable increase in caspase-3 activity and expression, and the decrease of Bcl-2/Bax ratio” (see p. 2, section “GRS inhibited myocardial apoptosis in MI/R mice”, also see Fig 3A for TUNEL staining (for apoptotic cells) and DAPI staining (for total cells) in a cross-section and Fig 3B for TUNEL positive percentage of about 40% in MI/R tissue).
Therefore, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to have expected that the 3D myocardial infarct organoid subjected to hypoxia-reoxygenation to simulate myocardial ischemia-reperfusion (MI/R) in vivo of Katare in view of Bursac (Katare, p. 2, left col, para “Hypoxia-Reoxygenation”) would have had a ratio of a TUNEL-positive area to a DAPI-positive area in the apoptotic interior region in an organoid cross-section ranging from about 0.03 to about 1.0 as suggested by Li with a reasonable expectation of success, because Bursac teaches the construct center comprises apoptotic cells in a percentage of about 33% (see modified Fig 2B above), since Katare teaches the organoid subjected to hypoxia has down-regulation of cell survival proteins Bcl-2 (p. 3, right col, para 1, see Fig 4), and since Li teaches a TUNEL assay on myocardial ischemia/reperfusion (MI/R) injury tissue resulting in a ratio of a TUNEL-positive area to a DAPI-positive area being about 40%, i.e., 0.4, (Fig 3A,B) associated with down-regulation of Bcl-2 (Fig 3E).
With respect to claim 3 directed to the cardiomyocytes comprising primary cardiomyocytes, as stated supra, Katare teaches cardiomyocytes are isolated from neonatal rats, thus teaches the cardiomyocytes comprise primary cardiomyocytes.
With respect to claims 4-5 directed to the ratio of cardiomyocytes to non-myocytes, as stated supra, Katare teaches the resulting cell population is 50% cardiomyocytes / 50% nonmyocytes (p. 1, right col, para “Cell isolation”), thus teaches the cardiomyocytes and non-myocytes are present in a ratio of about 95:5 to about 5:95 in claim 4 and about 60:40 to about 40:60 in claim 5.
With respect to claim 28 directed to a 3D myocardial ischemia-reperfused organoid, Katare teaches the organoid undergoes hypoxia-reoxygenation to simulate myocardial ischemia-reperfusion in vivo (p. 2, left col, para “Hypoxia-Reoxygenation”), thus teaches the organoid is a 3D myocardial ischemia-reperfused organoid.
Hence, the claimed invention as a whole was prima facie obvious to a person of ordinary skill before the effective filing date of the claimed invention in the absence of evidence to the contrary.
Response to Traversal:
Applicant’s arguments filed on 11/19/2025 are acknowledged.
Applicant argues that there also is no teaching or suggestion in Katare of an organoid having an apoptotic interior region that is surrounded by a viable periphery as claimed and that has a TUNEL-positive area to DAPI-positive area ratio as claimed, and that beats spontaneously in an asynchronous manner as claimed, and Carrier fails to correct the deficiencies of Katare (Remarks, p. 9-11).
Applicant’s arguments have been fully considered but become moot because the new ground of rejection does not rely on Carrier for any teaching or matter specifically challenged in the argument. Specifically, as discussed above, Bursac is cited to teach an equivalent cardiac organoid that comprises a center region having low survival of cardiac myocytes located more than 50 µm from a source of gas exchange and having apoptotic cells (i.e., an apoptotic interior region), surrounded by a 50- to 70-μm-thick outer cardiac tissue-like zone (i.e., a viable periphery within the claimed range). Bursac further suggests the construct center comprises apoptotic cells in a percentage of about 33% (see modified Fig 2B above). Li is cited to teach a TUNEL assay on myocardial ischemia/reperfusion (MI/R) injury tissue resulting in a ratio of a TUNEL-positive area to a DAPI-positive area being about 40%, i.e., 0.4, (Fig 3A,B) associated with down-regulation of Bcl-2 (Fig 3E, which is similarly taught in Katare, p. 3, right col, para 1 and Fig 4). Thus, one of ordinary skill in the art would have expected that the 3D myocardial infarct organoid of Katare would have had an apoptotic interior region that is surrounded by a viable periphery as claimed and would have had a TUNEL-positive area to DAPI-positive area ratio as claimed, and would have had spontaneously beating in an asynchronous manner as claimed, as suggested by Bursac and Li.
Claims 6-8 are rejected under 35 U.S.C. 103 as being unpatentable over Katare et al., (PLoS ONE. 2010; 5(2): e9275. P. 1-5. Prior art of record) in view of Bursac et al., (Am J Physiol. 1999;277(2):H433-44) and Li et al., (Sci Rep. 2016:6:37114, p. 1-14), as applied to claim 1 above, and further in view of Tulloch et al (Circulation Research. 2011; 109: 47-59. Prior art of record).
With respect to claim 6 directed to the non-myocytes comprise fibroblasts (FBs), endothelial cells (ECs) and mesenchymal stem cells (MSCs), and claim 7 directed to FBs about 50% to 60%, ECs about 25% to 35% and MSCs about 10% to 20% in the non-myocytes, as stated supra, Katare teaches the source cell population is 50% cardiomyocytes / 50% nonmyocytes (p. 1, right col, para “Cell isolation”).
However, Katare does not specifically teach the non-myocytes comprising fibroblasts, endothelial cells and mesenchymal stem cells, nor teach the proportions of the non-myocytes.
Tulloch teaches a 3D engineered human myocardium organoid (title, abstract). Tulloch teaches in the triculture experiment the engineered heart tissue construct comprises “2 million cardiomyocytes were mixed with 1 million HUVEC and 1 million MSCs or MEFs” (p. 48, right col, para “Cardiac construct generation”), thus teaches the non-myocytes comprise fibroblasts (i.e., MEFs), endothelial cells (i.e., HUVECs) and mesenchymal stem cells (i.e., human marrow stromal cells, MSCs) (p. 48, right col, para “Cell Culture” and “Cardiac construct generation”), i.e., all 3 types of the non-myocytes in claim 6. Tulloch teaches FBs being 50% and ECs being 50%, or MSCs being 50% and EC being 50% (see above, 1 million cells each), thus teaches the non-myocytes are within, or close to, the claimed range in claim 7. Note that the term “about” is “meant to encompass variations of +/- 10%” (specification p. 8).
Tulloch teaches addition of endothelial cells enhances cardiomyocyte proliferation under all stress conditions (14% to 19%), and addition of stromal supporting cells enhances formation of vessel-like structures by about 10-fold (abstract, see Figs 5 and 6). Although Tulloch does not teach a cardiac organoid that comprises all 3 types of non-myocytes in the same organoid, Tulloch does teach “when support cells (MEFs or MSCs) were added into the constructs in triculture with human cardiomyocytes and endothelial cells”, the total number of endothelial structures increase markedly, especially the cord structures (10-fold increase by the addition of MEFs and 8-fold by the addition of MSCs to hCM+EC) (p. 52 last para – p. 53 first para, see Fig 6A-6C and online Figure IV, B).
Therefore, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the 3D myocardial infarct organoid comprising cardiomyocytes and non-myocytes subjected to hypoxia as suggested by Katare in view of Bursac and Li, by choosing and combining fibroblasts, endothelial cells and mesenchymal stem cells so as to comprise all three types of non-myocytes in a defined range of proportions as suggested by Tulloch with a reasonable expectation of success. Since Tulloch teaches addition of endothelial cells enhances cardiomyocyte proliferation (14% to 19%), and addition of support cells (MEFs or MSCs) enhances formation of vessel-like structures by about 10-fold and 8-fold, respectively (abstract, see Figs 5 and 6), one of ordinary skill in the art would have had a reason to choose and combine endothelial cells and supporting fibroblasts and MSCs in the 3D myocardial infarct organoid of Katare in view of Bursac and Li in order to enhance cardiomyocyte proliferation and formation of vessel-like structures to mimic physiological in vivo response to ischemia-induced changes.
Furthermore, MPEP 2144.06 states "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.". Thus, combining both supporting cells, i.e., fibroblasts and MSCs, of Tulloch is prima facie obvious.
Moreover, regarding the range of proportions of the non-myocytes, notably, in the case where the claimed ranges "overlap or lie inside ranges disclosed by the prior art" a prima facie case of obviousness exists. In re Wertheim, 541 F.2d 257, 191 USPQ 90 (CCPA 1976); In re Woodruff, 919 F.2d 1575, 16 USPQ2d 1934 (Fed. Cir. 1990). It is a routine procedure to optimize component amounts to arrive at an optimal product that is superior for its intended use, since it has been held where the general conditions of a claim are disclosed in the prior art, discovering the optimum or workable ranges involves only routine skill in the art. See M.P.E.P. §2144.05.
With respect to claim 8 directed to the cardiomyocytes and/or non-myocytes being from a human, Katare teaches cardiomyocytes are isolated from neonatal rats (p. 1, right col, para “Cell isolation”), and teaches the organoid is used to screen the pharmacological compounds for treatment of myocardial injury (p. 4, right col).
However, Katare does not teach the cardiomyocytes and/or non-myocytes are from a human.
Tulloch teaches the cardiomyocytes are derived from a human ESC H7 line, and the non-myocytes ECs are human umbilical vein endothelial cells (HUVECs) and the MSCs are human marrow stromal cells (p. 48, right col, para “Cell Culture”), thus teaches the cardiomyocytes and non-myocytes are from a human.
Therefore, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the 3D myocardial infarct organoid comprising cardiomyocytes and non-myocytes from rats suggested by Katare in view of Bursac and Li, by substituting with cardiomyocytes and non-myocytes derived from a human as taught by Tulloch with a reasonable expectation of success. Since Katare teaches the organoid is used to screen the pharmacological compounds for treatment of myocardial injury (p. 4, right col), and since Tulloch teaches “pluripotent stem cells such as human embryonic stem cells (hESCs) or induced pluripotent stem cells (iPSCs) can now be used to generate large-scale cultures of human cardiomyocytes” (p. 47, right col), one of ordinary skill in the art would have had a reason to make this substitution in order to obtain an organoid comprising cardiomyocytes and non-myocytes derived from a human that has better clinical relevance for studies and drug screening applications.
Hence, the claimed invention as a whole was prima facie obvious to a person of ordinary skill before the effective filing date of the claimed invention in the absence of evidence to the contrary.
Response to Traversal:
Applicant’s arguments filed on 11/19/2025 are acknowledged and have been discussed above.
Conclusion
No claims are allowed.
Examiner Contact Information
Any inquiry concerning this communication or earlier communications from the examiner should be directed to Jianjian Zhu whose telephone number is (571)272-0956. The examiner can normally be reached M - F 8:30AM - 4PM (EST).
Examiner interviews are available via telephone, in-person, and video conferencing using a USPTO supplied web-based collaboration tool. To schedule an interview, applicant is encouraged to use the USPTO Automated Interview Request (AIR) at http://www.uspto.gov/interviewpractice.
If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, James Douglas (Doug) Schultz can be reached on (571) 272-0763. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
Information regarding the status of published or unpublished applications may be obtained from Patent Center. Unpublished application information in Patent Center is available to registered users. To file and manage patent submissions in Patent Center, visit: https://patentcenter.uspto.gov. Visit https://www.uspto.gov/patents/apply/patent-center for more information about Patent Center and https://www.uspto.gov/patents/docx for information about filing in DOCX format. For additional questions, contact the Electronic Business Center (EBC) at 866-217-9197 (toll-free). If you would like assistance from a USPTO Customer Service Representative, call 800-786-9199 (IN USA OR CANADA) or 571-272-1000.
/JIANJIAN ZHU/Examiner, Art Unit 1631
Prosecution Insights