DETAILED ACTION
Notice of Pre-AIA or AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Election/Restrictions
Applicant’s election without traverse of Invention I, claims 1-6 and 12-15, drawn to an extracellular matrix-based hydrogel and bioink capable of 3D printing, in the reply filed on May 2, 2025 is acknowledged.
Claims 7-11, drawn to a method of making an extracellular matrix-based hydrogel via Michael addition reaction with a modified collagen compound are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on May 2, 2025.
Applicant’s submission filed on August 28, 2025 has been entered and considered. Rejections and/or objections not reiterated from the previous action mailed May 30, 2025 are hereby withdrawn. The following rejections and/or objections are either newly applied or are reiterated and are the only rejections and/or objections presently applied to the instant application. The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
Status of the Claims
The amended claims filed on August 28, 2025 are acknowledged. Claim 1 has been amended. Claims 5 and 6 have been canceled. Claims 1-4 and 12-15 are examined on the merits.
Priority
Applicant claims foreign priority to Korean patent application KR 10-2021-0005008 filed on January 14, 2021. Receipt is acknowledged of certified copies of papers required by 37 CFR 1.55. Claims 1-4 and 12-15 are entitled to the benefit of Korean patent application KR 10-2021-0005008 and are given a priority date of January 14, 2021.
Information Disclosure Statement
The listing of references in the specification is not a proper information disclosure statement. 37 CFR 1.98(b) requires a list of all patents, publications, or other information submitted for consideration by the Office, and MPEP § 609.04(a) states, "the list may not be incorporated into the specification but must be submitted in a separate paper." Therefore, unless the references have been cited by the examiner on form PTO-892, they have not been considered.
Withdrawn Objection to the Specification
The prior objection to the abstract of the disclosure is withdrawn in light of Applicant’s amended abstract filed 8/28/2025.
Claim Objections
Newly amended claim 1 appears to be a missing a conjunction between the newly added claim limitations “wherein the denatured collagen is selected from methacrylated collagen and acrylated collagen” and “wherein a weight ratio of the extracellular matrix to the denatured collagen contained in the hydrogel is 1: 0.25 to 1: 0.5. For the purposes of examination, it is interpreted that it is Applicant’s intent for both of the newly added claim limitations to be present and that the missing linking word is intended to be “and”. Appropriate correction is required.
Withdrawn Claim Rejections - 35 USC § 103
The prior art rejection of claims 1-6 and 12-15 under 35 U.S.C. 103 as being unpatentable over Kim et al. (2019, Characterization of cornea-specific bioink: transparency, improved in vivo safety, J. of Tissue Engineering, 10) and Bagley et al. (US 20200179563A1) as evidenced by Michal et al. (US 20060233850A1) is withdrawn in light of Applicant’s amendment to claim 1 to recite a modified extracellular matrix-based hydrogel comprising an extracellular matrix-denatured collagen conjugate formed by a Michael addition reaction between the extracellular matrix having an amine group and denatured collagen to which an ethylenically unsaturated bond functional group is introduced to include additional recited limitations “wherein the denatured collagen is selected from methacrylated collagen and acrylated collagen, wherein a weight ratio of the extracellular matrix to the denatured collagen contained in the hydrogel is 1: 0.25 to 1: 0.5”.
Claim Rejections - 35 USC § 103
Claims 1-4 and 12-15 are rejected under 35 U.S.C. 103 as being unpatentable over Kim et al. (2019, Characterization of cornea-specific bioink: transparency, improved in vivo safety, J. of Tissue Engineering, 10, hereafter “Kim”), Bagley et al. (US 20200179563A1, hereafter “Bagley”), and Konuray et al. (2016, Sequential curing of amine-acrylate-methacrylate mixtures based on selective aza-Michael addition followed by radical photopolymerization. European Polymer Journal, 84, 256-267, hereafter “Konuray”) as evidenced by Michal et al. (US 20060233850A1; hereafter “Michal”).
This is a new rejection necessitated by Applicant’s amendment. This rejection shares substantial similarity to the rejection as set forth in the previous office action dated May 30, 2025. Any aspect of Applicant’s traversal that pertains to the rejection as newly set forth with be provided following the new statement of rejection.
With regard to claim 1, Kim teaches an extracellular-matrix based gel bioink comprising corneal-derived decellularized extracellular matrix in which cells can be encapsulated, which is capable of being 3D printed, and is biocompatible (Abstract). Additionally, Kim teaches that collagen hydrogel-based constructs are widely used and well-known in the art (Pg. 2, left column, first full para.). Further, Kim teaches combining cornea-derived decellularized extracellular matrix-based gel with collagen in order to investigate structural integrity (Pg. 6, right column, last full para. And Fig. 3c) and that more concentrated corneal-derived decellularized extracellular matrix-based gels contained collagen fibrils which are more easily crosslinked, leading to more rapid gelation (Pg. 7, right column, last full para.).
Kim does not teach a modified hydrogel formed by a Michael addition reaction between an extracellular matrix-based gel and a modified denatured collagen comprising an ethylenically unsaturated bond functional group wherein the denatured collagen is selected from methacrylated collagen and acrylated collagen and wherein a weight ratio of the extracellular matrix to the denatured collagen contained in the hydrogen is 1: 0.25 to 1: 0.5.
Bagley teaches a bioink comprising extracellular matrix molecules, including collagen which may be mixed with synthetic materials and is capable of being 3D printed (Para. [0064], lines 1-6). Additionally, Bagley teaches that the use of denatured collagen, or gelatin, (Para. [0004], line 10) to produce bioinks is known but must be chemically modified, for example methacrylated, in order to be crosslinked (Para. [0006], lines 3-9) which maintains structural integrity. Bagley teaches that the collagen in these bioinks can be crosslinked in order to improve mechanical stability and decrease degradation rate (Para. [0078], lines 6-7) and that this can be done via addition of chemical groups including methacrylate, acrylate, and divinyl sulfone (Para. [0079]) or via addition of a crosslinking agent, e.g. methacrylated gelatin (Para. [0080], lines 3-5). Further, Bagley teaches that bioinks can include additives such as extracellular matrix molecules, decellularized tissue (which may be from any tissue or organ), and may comprise cells (Para. [0083]).
Bagley is silent regarding a Michael addition reaction as being responsible for the crosslinking between extracellular matrix-based collagen and chemically modified components, for example, methacrylated gelatin. However, crosslinking via Michael addition reaction between extracellular matrix based collagen and methacrylated gelatin is evidenced by the teachings of Michal.
Michal teaches a hydrogel bioscaffolding crosslinked by a Michael addition reaction (Para [0011], lines 1-3 wherein the bioscaffolding may be formed of hydrogel components comprising those found in extracellular matrix, e.g. collagen (Para. [0011], lines 6-9). Michal teaches that extracellular matrix derived hydrogel components (for example, hyaluronan) can be modified to include electrophile groups such as acrylate, methacrylate, or vinyl sulfone (Para. [0044], lines 8-10) and that a Michael addition reaction will occur between the hydrogel component having an electrophile (i.e., components which have been modified by addition of acrylate, methacrylate, or vinyl sulfone) and an additional hydrogel component containing a nucleophile (for example, an amine group present in collagen as evidenced by the figure in Para. [0050] of the specification) thus creating a crosslinked hydrogel (Para. [0040], lines 1-9 and 12-14).
Therefore it would have been obvious, to one having ordinary skill in the art, before the effective filing date of the claimed invention, to combine the corneal-derived decellularized extracellular matrix-based hydrogel at taught by Kim with the extracellular matrix derived collagen-based bioink crosslinked by addition of methacrylated gelatin as taught by Bagley. Although Bagley is silent to crosslinking via Michael addition reaction, one having ordinary skill in the art would have recognized, based on the teachings of Michal wherein a hydrogel is formed by a Michael addition reaction which naturally occurs between amine groups and acrylate, methacrylate, or vinyl sulfone groups, that the crosslinking reaction of Bagley, which occurs between extracellular matrix derived collagen and methacrylated gelatin, is due to a Michael addition reaction. One having ordinary skill in the art would have been motivated to combine Kim’s corneal-derived decellularized extracellular matrix-based gel with Bagley’s bioink comprising extracellular matrix-based collagen and a methacrylated gelatin because, as Bagley teaches, crosslinking which occurs between collagen and methacrylated gelatin improves mechanical stability and decreases degradation rate of the resultant bioink which would provide added benefits for use of the bioink in 3D printing and subsequent transplantation. One having ordinary skill in the art would have had a reasonable expectation of success as Kim, Bagley, and Michal teach bioinks and/or bioscaffolds derived from extracellular matrix components for use in tissue regeneration.
While Bagley does teach a mixture of bioink components (e.g. collagen and methacrylated gelatin) in a 1:1 ratio (Para. [0112], line 9), the combined teachings of Kim, Bagley, and Michal do not teach wherein the ratio of the extracellular matrix to the denatured collagen contained in the hydrogel is 1: 0.25 to 1: 0.5.
Konuray teaches an analysis of Michael reaction kinetics between amines and acrylates and/or methacrylates (Abstract) wherein the amine-acrylate ratio indicated for conversion at the gel point was determined to be 0.5 (Pg. 263, 1st Para., lines 1-2).
Therefore, it would have been obvious before the effective filing date of the claimed invention, to apply the amine-acrylate ratio wherein the conversion at the gel point is 0.5 as taught by Konuray to the extracellular matrix based hydrogel conjugate formed by a Michael addition reaction and comprising extracellular matrix and denatured collagen which has been acrylated or methacrylated with a reasonable expectation of success. A skilled artisan would have been motivated to use the amine-acrylate ratio of 0.5 as taught by Konuray because Konuray teaches a 0.5 amine-acrylate is the ratio at which gelation occurs and one having ordinary skill in the art would have recognized that determination of the optimal ratio needed to produce the desired physical characteristics of the bioink (i.e., gelation) would be beneficial for creating hydrogels that allow for 3D printing and subsequent use in clinical settings while also reducing the cost associated with using more of an acrylated or methacrylated denatured collagen product than required for gelation.
With regard to claims 2-3, Kim et al. teaches that the extracellular matrix is decellularized extracellular matrix derived from the cornea (Abstract).
With regard to claim 4, Bagley et al. teaches that collagen may be modified by chemical groups which permit crosslinking, including methacrylate, acrylate, and divinyl sulfone (Para. [0079], lines 1-7) and that methacrylated gelatin can be used as a crosslinking agent (Para. [0080], line 5).
With regard to claims 12-14, Kim teaches an extracellular based bioink comprising human turbinate-derived mesenchymal stem cells which have been pre-differentiated for keratocyte lineage, which is considered to read on corneal-derived cells, which can be encapsulated at a concentration of 1x106 cells/mL in the extracellular matrix-based gel (Pg. 4, left column, first full para.).
With regard to claim 15, Kim teaches that the corneal-derived extracellular matrix based bioink containing encapsulated cells is capable of being 3D printed and is biocompatible for in-vivo transplantation (Abstract). Further, Kim teaches that their 3D printed corneal-derived extracellular matrix based bioink could be used for “fabrication of patient-specific shaped artificial corneas” (Pg. 11, Conclusion).
Response to Applicant’s Traversal
Applicant’s traversal filed August 28, 2025 is acknowledged and has been fully considered by is not persuasive. Applicant has amended claim 1 to include additional recited limitations “wherein the denatured collagen is selected from methacrylated collagen and acrylated collagen” and “wherein a weight ratio of the extracellular matrix to the denatured collagen contained in the hydrogel is 1: 0.25 to 1: 0.5”. Applicant asserts that the amendment of claim 1 has directed the scope of claim 1 and claims 2-4 and 12-15, which depend from claim 1, to subject matter which is free of the obviousness rejection over Kim and Bagley as evidenced by Michal as applied to claims 1-6 and 12-15 set forth in the office action dated May 30, 2025.
Applicant’s amendment dated August 28, 2025 has canceled claims 5 and 6 which renders any traversal related to these claims moot. Applicant’s amendment of claim 1 has introduced new limitations of the modified extracellular matrix based hydrogel and a new obviousness grounds of rejection of claim 1 and claims 2-4 and 12-15, which depend from claim 1, has been supplied based on the combined teachings of Kim, Bagley, and Konuray as evidenced by Michal.
Applicant asserts on Pg. 6 “The prior art combination relied upon by the Examiner never arrives at this particular chemistry, with this particular pairing of reactants, at this particular non-overlapping ratio range, nor does it teach the unexpected property profile demonstrated in the specification” as relating to “a critical extracellular matrix:denatured collagen ratio of 1:0.25-1:0.5”.
On Pg 7., Applicant asserts that the teachings of Kim are insufficient as Kim does not disclose, teach, or hint at selecting denatured collagen comprising an ethylenically unsaturated functional group which is used in a reaction with amines on the extracellular matrix to create a new conjugate. Additionally, Applicant asserts that the teachings of Bagley are also insufficient as Bagley teaches use of methacrylated gelatin which Applicant asserts is not the same material as methacrylated collagen used in the instantly claimed invention and Applicant’s amendment “confines the denatured collagen to methacrylated collagen or acrylated collagen”. Further, Applicant asserts that Michal only provides generic evidence that electrophiles can undergo Michael addition reactions with nucleophiles present in biological macromolecules but does not teach cornea derived extracellular matrix, methacrylated or acrylated collagen as an electrophile, nor teaches any mass ratio optimization “in the context of maintaining optical clarity and suturability for corneal grafts”.
Applicant further asserts that the amended ratio of 1:0.25 - 1:0.5 in claim 1 yields the unexpected result of showing a complex modulus that is about 78-fold higher than corneal derived extracellular matrix which is not mixed with a methacrylated denatured collagen and that the data “demonstrate a non-linear, peaked response” which could not have been predicted by a skilled artisan by the combination of Kim and Bagley’s teachings and that, additionally, Michal does not provide a suggestion in the art to use a Michael addition reaction “to covalently graft methacrylated collagen or acrylated collagen to decellularized corneal ECM and to do so at the claimed, critical mass ratio to simultaneously deliver a 78x increase in complex modulus during gelation, maintain shear-thinning and >95% cell viability, and enable suturable, transparent corneal grafts in vivo”.
Applicant’s arguments have been considered, but are not found persuasive.
With regard to Applicant’s assertion regarding the deficiencies in Kim, Bagley, and Michal, MPEP 2145(IV) states:
“One cannot show nonobviousness by attacking references individually where the rejections are based on combinations of references. In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981); In re Merck & Co., Inc., 800 F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986). Where a rejection of a claim is based on two or more references, a reply that is limited to what a subset of the applied references teaches or fails to teach, or that fails to address the combined teaching of the applied references may be considered to be an argument that attacks the reference(s) individually.”
Kim teaches an extracellular-matrix based gel bioink comprising corneal-derived decellularized extracellular matrix (Abstract) which can be combined with collagen in order increase structural integrity (Pg. 6, right column, last full para. And Fig. 3c) and wherein increased collagen fibril concentration in corneal-derived decellularized extracellular matrix-based gels leads to more rapid gelation (Pg. 7, right column, last full para.). Kim also teaches that collagen hydrogel based constructs are well-known and widely used in the art (Pg. 2, left column, first full para.). Bagley teaches a 3D printable bioink comprising extracellular matrix molecules, including the biomacromolecule collagen, which can be crosslinked of via addition of methacrylate or acrylate in order to improve mechanical stability and decrease degradation rate (Para. [0078], lines 6-7). In a specific embodiment Bagley teaches use of gelatin, which Bagley defines as denatured collagen (Para. [0053]) which has been methacrylated (Para. [0080], lines 3-5). Thus, these elements were known in the art at the time of the instant application and one having ordinary skill in the art would have been motivated to combine these element because Bagley teaches use of methacrylated and/or acrylated collagen-based molecules in extracellular matrix based hydrogel constructs provide the benefit of increasing mechanical stability and decreasing degradation of the hydrogel which one having ordinary skill in the art would recognize could easily be applied to Kim’s corneal-derived extracellular matrix based hydrogel which can be combined with collagen and provide the benefit increasing mechanical stability and decreasing degradation. Further, Michal is provided as evidence that Michael addition reactions in hydrogel production are well-known in the art and that a Michal addition reaction will occur between amine groups, which Applicant’s specification indicate are present in the collagen found in extracellular matrix, and collagen-based biomacromolecules which have been methacrylated or acrylated in order to form a crosslinked hydrogel product. As Michal evidences this is a naturally occurring crosslinking reaction between amine groups (which are present in collagen) and acryl or methacryl groups, there is every reason to believe that one having ordinary skill in the art would recognize that a Michael addition reaction would occur in hydrogel as taught by Bagley. Thus, a Michael addition reaction does not need to be used to link methacryl or acryl groups to extracellular matrix components as Applicant asserts on pg. 8, but would occur given Applicant’s hydrogel components as claimed. Accordingly, Applicant’s traversal is unpersuasive.
Additionally, Applicant asserts that the teachings of Bagley are insufficient as Applicant’s amendment is drawn to methacrylated collagen or acrylated collagen wherein Bagley teaches methacrylated gelatin. Applicant’s amendment to claim 1 recites “wherein the denatured collagen is selected from methacrylated collagen and acrylated collagen” and Applicant’s specification further defines “The methacrylated collagen is a denatured collagen in which a methacryl group is linked… and the acrylated collagen is a denatured collagen in which an acryl group is linked…” in Para. [0010]. While Bagley does teach that collagen can may be modified with methacrylate or acrylate groups at Para. [0079], Bagley also teaches that gelatin is defined as collagen which is denatured (Para. [0053]). As Applicant has defined methacrylated collagen as denatured collagen comprising a methacryl group, Bagley’s teachings of denatured collagen (i.e., gelatin) which has been methacrylated is considered to be a reasonable interpretation of Applicant’s claimed denatured collagen wherein methacrylated collagen is defined as a denatured collagen in which a methacryl group is linked. Accordingly, Applicant’s traversal is unpersuasive.
With regard to Applicant’s assertion that amended ratio of 1:0.25 - 1:0.5 in claim 1 is not taught by Kim, Bagley, and Michal; a new obviousness grounds of rejection has been supplied incorporating the teachings of Konuray. Konuray teaches wherein the amine-acrylate ratio for conversion at the gel point is 0.5 and therefore would have been obvious to combine with Kim, Bagley, and Michal as discussed in the rejection above. Accordingly, Applicant’s traversal is unpersuasive against the rejection as newly set forth.
With regard to Applicant’s assertion of alleged unexpected results of the critical claimed ratio, the combined teachings of Kim, Bagley, and Michal make obvious extracellular matrix based hydrogels comprising extracellular matrix components (for example, collagen) which can be crosslinked to methacrylated or acrylated biomolecules such as gelatin via Michael addition reactions and that this crosslinking of components is used in order to increase structural stability of the extracellular matrix based hydrogel, one having ordinary skill in the art would expect that when a methacrylated or acrylated denatured collagen was added to extracellular matrix comprising amine groups, a Micheal addition reaction would occur resulting in increased structural stability when compared to a control group which would be reflected as increases in complex modulus as shown in Fig. 2 of the instant specification. Further, Applicant asserts, regarding the ratio, that “under lower loading (≈ 1:0.05), the reaction proceeds insufficiently; under higher loading (≈ 1:0.5), the FT-IR indicates little additional C-C growth relative to the 1: 0.25 condition, evidencing a saturation or diminishing-returns region.” It is noted that Applicant asserts the reaction still proceeds, albeit insufficiently, under lower loading conditions of 1: 0.05 which is outside of the claimed critical range. Applicant’s working examples are to a single ratio of 1:0.25 so it is not possible to evaluate whether ratios which are outside of the claimed critical range, for example 1:0.2, would also generate a similar allegedly unexpected results. Para. [0053] of Applicant’s specification states that use of 1.0% w/v hydrogel (1.0 MACCO) did not increase the number of single bonds as compared to 0.5% w/v hydrogel (0.5 MACCO) and that “These results indicate that the reaction is closest to the maximum in the 0.5 MACCO hydrogel” which corresponds to a 1: 0.25 ratio. Thus, it appears that Applicant’s claimed critical ratio corresponds to the saturation point of the reaction, but that other ratios which are outside of the claimed critical ratio are likely to produce similar allegedly unexpected results.
Additionally, unexpected results are based on objective scientific evidence (See MPEP 2145). Fig. 5(b) of Kim shows a time sweep analysis of various concentrations of corneal-derived extracellular matrix (co-dECM), including 2.0% wt. The 2.0% wt co-dECM of Kim appears to be the same compound as instantly claimed in the working examples and which is indicated as the control condition in the time sweep analysis provided in Applicant’s Fig. 2, right side. Both Kim and the instant specification teach that gelation kinetics were tested at 37°C. However, the time sweep analysis of gelation as instantly claimed for co-dECM (Fig 2, black line) does not appear to indicate the same modulus as what is shown in the prior art of Kim for 2.0% co-dECM (Fig. 5, red line). As these compounds appear to be exactly the same and appear to be tested under the same conditions, one having ordinary skill in the art would expect the gelation to occur with similar results. Thus, the discrepancy between the prior art and the instantly claimed working examples provides some uncertainty regarding Applicant’s alleged unexpected results.
Additionally, MPEP 716.02(d) states:
“Whether the unexpected results are the result of unexpectedly improved results or a property not taught by the prior art, the "objective evidence of nonobviousness must be commensurate in scope with the claims which the evidence is offered to support." In other words, the showing of unexpected results must be reviewed to see if the results occur over the entire claimed range. In re Clemens, 622 F.2d 1029, 1036, 206 USPQ 289, 296 (CCPA 1980)” and in 716.02(d)(II) “To establish unexpected results over a claimed range, applicants should compare a sufficient number of tests both inside and outside the claimed range to show the criticality of the claimed range. In re Hill, 284 F.2d 955, 128 USPQ 197 (CCPA 1960).”
Applicant has supplied evidence of alleged unexpected results at a single ratio, 1: 0.25, and only using methacrylated collagen and therefore has not shown evidence of nonobviousness which is commensurate in scope with the entirety of the range as instantly claimed nor over the entirety of denatured collagen species as claimed. Accordingly, Applicant’s traversal is unpersuasive.
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 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.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to ERIN V PAULUS whose telephone number is (571)272-6301. The examiner can normally be reached Mon-Fri 8 AM-5 PM.
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/ERIN V PAULUS/Examiner, Art Unit 1631
/ARTHUR S LEONARD/Examiner, Art Unit 1631