DETAILED ACTION
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. Applicant’s Request for Continued Examination, Amendment, and Arguments/Remarks received on 07 May 2026 have been entered. Claims 1-20 were previously pending in the application. New claim 22 has have been added by Applicant. Claims 1-20 and 22 are currently pending in the application. Claims 1, 15, 20, and 22 are independent claims.
The following election of species remains in effect in the instant application:
1) retinal tissue unit shapes: a. hollow ovoid,
2) differentiated living human retinal cells other than retinal epithelium cells: a. rods,
3) retinal diseases: a. age-related macular degeneration.
No claims are withdrawn from examination. Claims 1-20 and 22 are currently pending and under examination in the instant application. An action on the merits follows. The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA . The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
Priority
The present application is a 35 U.S.C. 371 national stage filing of International Application No. PCT/EP2020072567, filed 12 August 2020, which claims priority to FR1909155, filed 12 August 2019.
Filing of a certified untranslated copy of the FR1909155, filed 11 February 2022, is acknowledged.
Should applicant desire to obtain the benefit of foreign priority under 35 U.S.C. 119(a)-(d) prior to declaration of an interference, a certified English translation of the foreign application must be submitted in reply to this action. 37 CFR 41.154(b) and 41.202(e). Failure to provide a certified translation may result in no benefit being accorded for the non-English application.
Thus, the earliest possible priority for the instant application is 12 August 2019.
Election of Species
Applicant argues that “hollow ovoid” is not disclosed or suggested by Singh et al., and as such, Examiner has expanded a search beyond the elected species which requires a re-joinder of non-elected species. However, this is not agreed.
As discussed in the prior actions, the term “ovoid” has been afforded its broadest reasonable interpretation to refer to any 3D structure which is generally round but not a perfect sphere. The retinal tissue organoids of Singh (2015) are three-dimensional structures which are generally round, but not perfectly sphere [Figures 2-7, 9, S1-S2, S4-S10], and as such, the retinal tissue organoids of Singh (2015) are in the form of hollow ovoids.
As discussed in the prior actions, the terms “hollow” and “inner cavity” in claim 1 and 22 have been afforded their broadest reasonable interpretations such that “hollow” refers to a cell-free space interior to the unit, and “inner cavity” refers to a space that is at least partially surrounded by the RPE cell layer. The specification does not provide a limiting definition of “organized around” or “an inner cavity” which would narrow the interpretation of these terms to indicate only a cell layer which fully encompasses an inner space in 3 dimensions such that the cell layer encompasses all surfaces of the cavity.
Accordingly, by searching and applying art within the broadest reasonable interpretation of the terms “hollow” and “ovoid”, Examiner has not expanded the search to encompass non-elected species. As such, the election of species is being maintained.
Claim Objections
Previously presented claim 15 is newly objected to because of the following informalities: claim 15 recites, “if the cells introduced into the microcompartment are the stem cells inducing cell differentiation within the cellular compartment”, which is missing a comma (“,”) after “stem cells”. Appropriate correction is required.
Claim Rejections - 35 USC § 112(b)
Previously presented and new claims 15-19 and 22 are newly rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention. Claims 16-19 are included in this rejection due to their dependence on claim 15.
New independent claim 22 recites “obtained from a pluripotent stem cell” in lines 1-2, which is indefinite because it is unclear in what way the hollow three-dimensional retinal tissue unit is obtained from a pluripotent stem cell in that pluripotent stem cells do not comprise hollow three-dimensional retinal tissue units. As such, the metes and bounds of the claim cannot be determined.
Previously presented independent claim 15 has multiple issues of indefiniteness. Claim 15 recites, “or added” in line 5, which is indefinite because it is unclear whether “or added” is meant to be an active method step of the claimed method.
Claim 15 additionally recites, “a cellular microcompartment comprising” in line 3 … “at least extracellular matrix elements” in lines 5 and “at least differentiated retinal pigment epithelium cells” in line 6, which is indefinite because it is recitation of “comprising… at least” is redundant and it is unclear what is meant to be encompassed by the recitation of “at least” after recitation that the composition is “comprising” the listed components.
As such, the metes and bounds of the claim cannot be determined.
Claim Rejections - 35 USC § 112(a)- Scope of Enablement
The following is a quotation of the first paragraph of 35 U.S.C. 112(a):
(a) IN GENERAL.—The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
The following is a quotation of the first paragraph of pre-AIA 35 U.S.C. 112:
The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor of carrying out his invention.
Amended, previously presented, and new claims 1-20 and 22 are newly rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, because the specification, while being enabling for:
A hollow three-dimensional retinal tissue unit produced within a hydrogel capsule in tissue culture and comprising:
at least one layer of living in vitro differentiated human retinal pigment epithelium cells (RPEs) organized around an inner cavity,
wherein the basal side of each living in vitro differentiated human RPE points outwards and the apical side of each living in vitro differentiated human RPE points towards the inner cavity,
wherein the living in vitro differentiated human RPEs are differentiated in vitro from human induced pluripotent stem cells (hiPSCs) or from human embryonic stem cells (hESCs),
wherein the retinal tissue unit comprises a layer of extracellular matrix on the basal side of the at least one layer of living in vitro differentiated RPEs; and
wherein the three-dimensional retinal tissue unit is produced within an alginate hydrogel capsule, wherein the alginate hydrogel capsule comprises an inner layer of extracellular matrix adjacent to the inner surface of the alginate hydrogel capsule;
-and-
A method for preparing the retinal tissue unit of claim 1, comprising the steps of:
producing a cellular microcompartment comprising an alginate hydrogel capsule,
wherein the alginate hydrogel capsule is produced by:
i. obtaining human cells,
wherein the human cells are induced pluripotent stem cells (hiPSCs), human embryonic stem cells (hESCs) or in vitro differentiated retinal pigment epithelium cells (RPEs), and
wherein the in vitro differentiated RPEs are differentiated in vitro from the hiPSCs or the hESCs;
ii. mixing the human cells with extracellular matrix elements to produce a cell mixture,
and
iii. encapsulating the cell mixture in an alginate hydrogel layer to produce the alginate hydrogel capsule;
b) if the human cells mixed with the extracellular matrix are the hiPSCs or the hESCs, then inducing cell differentiation of the hiPSCs or the hESCs within the cellular microcompartment to obtain one or more differentiated cells,
wherein the one or more differentiated cells comprise the in vitro differentiated RPEs;
c) culturing the in vitro differentiated RPEs or the one or more differentiated cells comprising the in vitro differentiated RPEs within the hydrogel capsule to obtain a hollow three-dimensional retinal tissue unit encapsulated within the hydrogel capsule;
d) separating the hollow three-dimensional retinal tissue unit from the hydrogel capsules; and
e) recovering the in vitro differentiated RPEs or the one or more differentiated cells comprising the in vitro differentiated RPEs in the form of the hollow three-dimensional retinal tissue unit,
wherein at least one layer of the in vitro differentiated RPEs are organized around an inner cavity,
wherein the basal side of each in vitro differentiated RPE points outwards and the apical side of each in vitro differentiated RPE points towards the inner cavity;
does not reasonably provide enablement for:
A hollow three-dimensional retinal tissue unit produced within any hydrogel capsule in tissue culture and comprising, organized around an inner cavity, at least one layer of any differentiated living human retinal pigment epithelium cells, with the basal side of each retinal pigment epithelium cell pointing outwards and the apical side pointing towards the inner cavity;
wherein the retinal tissue unit lacks a layer of extracellular matrix on the basal side of the at least one layer of living in vitro differentiated RPEs; and
wherein the three-dimensional retinal tissue unit is produced within a hydrogel capsule lacking an inner layer of extracellular matrix adjacent to the inner surface of the hydrogel capsule;
-nor-
A method for preparing the retinal tissue unit of claim 1, comprising the steps of:
a) producing a cellular microcompartment comprising, within any hydrogel capsule:
i. at least extracellular matrix elements, secreted by cells or added, and
ii. any stem cells or at least any differentiated retinal pigment epithelium cells,
b) if the cells introduced into the microcompartment are the stem cells, inducing cell differentiation within the cellular microcompartment, so as to obtain at least retinal pigment epithelium cells,
c) removing the hydrogel capsules, and
d) recovering the retinal pigment epithelium cells and any other retinal cells in the form of a hollow three-dimensional retinal tissue unit,
without mixing the human cells with extracellular matrix elements to produce a cell mixture and encapsulating the cell mixture in an alginate hydrogel layer to produce the alginate hydrogel capsule; and
without culturing the differentiated RPEs within the hydrogel capsule to obtain a hollow three-dimensional retinal tissue unit encapsulated within the hydrogel capsule,
wherein at least one layer of the differentiated RPEs are organized around an inner cavity, and
wherein the basal side of each differentiated RPE points outwards and the apical side of each in vitro differentiated RPE points towards the inner cavity.
The specification does not enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make, use, or practice the invention commensurate in scope with these claims.
This rejection comprises three (3) separate issues: 1) the absence of an enabling disclosure for producing the hollow three-dimensional retinal tissue unit within any hydrogel capsule other than an alginate hydrogel capsule comprising an inner layer of extracellular matrix adjacent to the inner surface of the alginate hydrogel capsule; 2) the absence of an enabling disclosure for producing the hollow three-dimensional retinal tissue unit using any differentiated living human retinal pigment epithelial cells other than living in vitro differentiated human RPEs differentiated in vitro from hiPSCs or hESCs; and 3) the lack of an enabling disclosure for generating any retinal tissue unit other than a retinal tissue unit comprising an extracellular matrix layer present along the basal side of the at least one layer of living in vitro differentiated RPEs.
These issues were identified by the Office after analysis of the disclosure provided by the specification. The Office has analyzed the specification in direct accordance to the factors outlined in In re Wands, namely 1) the nature of the invention, 2) the state of the prior art, 3) the predictability of the art, 4) the amount of direction or guidance present, and 5) the presence or absence of working examples, and presented detailed scientific reasons supported by publications from the prior art for the finding of a lack of enablement for the scope of the instant methods. The Wands analysis and supporting specific evidence are presented below for each of the identified issues.
As a first issue (1), the specification does not provide an enabling disclosure for producing the hollow three-dimensional retinal tissue unit within any hydrogel capsule other than an alginate hydrogel capsule comprising an inner layer of extracellular matrix adjacent to the inner surface of the alginate hydrogel capsule, wherein the ECM has been added to the capsule by mixing with the cells prior to encapsulation of the cells with the hydrogel, and wherein the cells are cultured in the hydrogel to produce the retinal tissue.
The broadest independent claims, claims 1 and 22, recite a hollow three-dimensional (3D) retinal tissue unit comprising, organized around an inner cavity, at least one layer of differentiated living human retinal pigment epithelium cells, with the basal side of each RPE cell pointing outwards and the apical side pointing towards the inner cavity. Additionally, claim 1 recites that the retinal tissue units are produced within a hydrogel capsule in tissue culture. Claim 22 recites that the retinal tissue unit is obtained from a pluripotent stem cell. Claims 2-20 depend on and/or encompass independent claim 1.
Claim 15 recite a method of making the retinal tissue unit of claim 1 comprising producing a cellular microcompartment comprising, within a hydrogel capsule, at least extracellular matrix elements, secreted by cells or added, and stem cells or at least differentiated RPE cells, if the cells introduced into the microcompartment are the stem cells, inducing cell differentiation within the cellular microcompartment, so as to obtain at least RPE cells, removing the hydrogel capsules, and recovering the RPE cells and any other retinal cells in the form of a hollow 3D retinal tissue unit.
The specification discloses generic hydrogel capsules [page 6, 7, 14, 22, 23, Figure 3] and also teaches the use of alginate hydrogel capsules [page 8, Figure 5-7-9]. The specification also teaches that the “used hydrogel is preferably biocompatible, i.e., not toxic to cells. The hydrogel capsule must allow the diffusion of oxygen and of nutrients to feed the cells contained in the microcompartment and allow their survival. According to one embodiment, the capsule comprises alginate. It can be formed exclusively of alginate. In particular, the alginate may be a sodium alginate, composed of 80% α-L-guluronate and 20% β-D-mannuronate, with an average molecular weight of 100 to 400 kDa and a total concentration between 0.5 and 5% by weight. The hydrogel capsule makes it possible to protect the cells from the external environment, to limit the uncontrolled proliferation of the cells, and allows for controlled differentiation of the cells into retinal cells, at least into retinal pigment epithelium cells. A capsule very preferably surrounds a single tissue unit according to the invention and each tissue unit is surrounded by a single hydrogel capsule.” And “removal [of the capsule] can be achieved using… an enzyme such as alginate lyase if the hydrogel comprises alginate” [pages 18-19]. The specification as filed does not teach any alternative to alginate for the composition of the hydrogel capsule.
Additionally, the specification teaches generic inclusion of an extracellular matrix (ECM) within the capsule [page 13, Figure 2]. The specification also teaches wherein the extracellular matrix layer can be formed by the cellular matrix secreted by RPE cells and/or by ECM added at the time of preparation of the cell unit, wherein preferably at least part of the extracellular matrix being provided in addition to the ECM naturally secreted by the cells [page 11, 17]. The specification further teaches that when the ECM is present, the retinal cells organized in three-dimensions around the inner cavity advantageously already interact with an ECM, which facilitates their implantation at the retina [page 12]. The specification also teaches methods of producing the hydrogel capsule which only comprise mixing the pluripotent stem cells with an extracellular matrix prior to encapsulating the mixture in a hydrogel layer [page 20, 26].
The specification additionally teaches and that cell polarization can be obtained by depositing a matrix layer on the inner face of alginate capsules [page 8, Figure 9], such that:
“Advantageously, the total or partial encapsulation in the hydrogel and the provision of ECM combined is a means capable of allowing the polarization of the retinal pigment epithelium cells. Indeed, the polarization of said cells can be obtained by depositing a layer of matrix on the inner face of the hydrogel capsules which positions the basal side of the cells, the tissue organizes itself around the cavity following this indication of polarity (as illustrated in figure 9, which shows that an extracellular matrix layer anchored to the alginate shell induces polarization of the cells as evidenced by the flattening of the tissue against the alginate due to the high tensile strength of the gel dictating the shape of the tissue).” [page 18].
Therefore, the specification teaches the criticality of including exogenous extracellular matrix within the alginate hydrogel capsule and provides no examples of producing retinal tissue units without the addition of extracellular matrix into the cell mixture prior to encapsidation within alginate hydrogels.
The art at the time of filing teaches that cell encapsulation in alginate hydrogel has been extensively investigated for a variety of tissue engineering applications, wherein alginate itself is largely nonadhesive to mammalian cells, but can be modified with adhesion ligands to promote cell attachment [Hunt et al. 2018, Advanced Healthcare Materials, 7, 1800226, 1-31, page 23 col. 2 ¶ 3]. Specifically for tissue engineering of RPE, Hunt (2018) teaches that in order to expand RPE cells for transplantation or in vitro studies, cells are traditionally cultured on tissue culture plastic (TCP), but that TCP culture is often associated with de-differentiation of the RPE cells, wherein the RPE Cells adopt a more fibroblast-like morphology and are less pigmented [page 23 col. 2 ¶ 3]. Hunt (2018) teaches that the suitability of alginate hydrogel to maintain RPE cell viability has been shown [page 24 col. 1 ¶ 1].
Hunt (2018) further teaches that the application of biomaterials to retinal tissue development is yet limited, such that examples of different biomaterials that could be applied to retinal tissue engineering each have associated limitations and benefits [page 10 col 2 ¶ 1, page 10 col 1 ¶ 1, Table 5]. Hunt (2018) teaches that alginate has limited capacity for binding of cells, growth factors, or ECM proteins without modification, exhibits batch-to-batch variability including degradation and mechanical properties [Table 5]. Further, Hunt (2018) teaches that a variety of other biomaterials, such as decellularized tissue, natural polymers of human/animal origin, and synthetic polymers, each have various limitations including possible disease transmission, allergic reactions, limited availability, variability, adverse immunogenic reactions, slow degradation, acidic degradation products, and cytotoxic polymerization methods [Table 5]. Therefore, Hunt (2018) teaches that outcomes from using different biomaterials, such as different hydrogels, are variable such that outcomes with any one biomaterial do not predict outcomes obtained from using a different biomaterial.
The art at the time of filing also teaches that that RPE cells differentiated in culture from human iPSCs and grown on an extracellular matrix will develop in an orientation wherein the basal side of the RPE is attached to the extracellular matrix (ECM) [Khristov et al. 2018, Methods Mol. Biol., 1722, 223-247, page 230 step 7, Figure 1, 5]. Hunt (2018) also teaches polarization of RPE and/or other retinal cells obtained only when using scaffolds that have been treated with an ECM component; for example, PLDLA porous films coated in Col-IV, silk fibroin membrane coated in commercial dECM blend from human placenta, or PLA coated with fibronectin [page 25 col 2 ¶ 1, page 26 col 2 ¶ 2- page 27 col 1 ¶ 1, Table 6-7]. Hunt (2017) teaches that it is increasing being recognized that the extracellular matrix (ECM) is important for the correct development and function of the retina both in vivo and in vitro [Hunt et al. 2017, Acta Biomaterialia, 49, 329-343, page 330 col1 ¶ 4].
Neither the specification nor the art at the time of filing teaches that any hydrogel capsule other than an alginate capsule comprising an inner layer of extracellular matrix adjacent to the inner surface of the alginate can be used to encapsulate human RPE cells and produce a hollow ovoid three-dimensional retinal tissue unit having RPE cells which are polarized such that the basal side of each RPE cell points outward and the apical side of each RPE cell points towards the inner cavity. Additionally, neither the specification nor the art at the time of filing teaches producing a 3D retinal tissue from differentiated RPE cells without culturing in the cells in a matrix to produce the 3D retinal tissue.
Thus, in view of the art recognized unpredictability resulting from various approaches and outcomes employing various biomaterials, including various hydrogel formulations, for the culture and differentiation of retinal cells, including RPE cells, to produce retinal tissues; the limitations of the specification to teaching only alginate hydrogel encapsulation; the limitations in the specification to teaching only alginate hydrogel capsules comprising ECM; the art recognized need to substitute/alter alginate to promoter cellular attachment; the art recognized importance of the ECM for correct development and function of the retina; and the breadth of the claims; the ordinarily skilled artisan at the time of filing the instant application would have considered generating the hollow 3D retinal tissue unit with the organization and polarization as claimed using any hydrogel capsule other than an alginate hydrogel capsule comprising an inner layer of extracellular matrix adjacent to the inner surface of the alginate hydrogel capsule, wherein the ECM was mixed with the cells and thereby added to the capsule rather than merely secreted by the cells, and wherein the differentiated RPE cells were cultured in the hydrogel capsule to generate the 3D tissue, as highly unpredictable. As such, it would have required undue experimentation to practice the scope of Applicant’s invention as claimed.
As a second issue (2), the specification does not provide an enabling disclosure for producing the hollow three-dimensional retinal tissue unit using any differentiated living human retinal pigment epithelial cells other than living in vitro differentiated human RPEs differentiated in vitro from hiPSCs or hESCs.
The broadest independent claims, claims 1 and 22, recite a hollow three-dimensional (3D) retinal tissue unit comprising, organized around an inner cavity, at least one layer of differentiated living human retinal pigment epithelium cells, with the basal side of each RPE cell pointing outwards and the apical side pointing towards the inner cavity. Additionally, claim 1 recites that the retinal tissue units are produced within a hydrogel capsule in tissue culture. Claim 22 recites that the retinal tissue unit is obtained from a pluripotent stem cell. Claims 2-20 depend on and/or encompass independent claim 1.
Claim 15 recite a method of making the retinal tissue unit of claim 1 comprising producing a cellular microcompartment comprising, within a hydrogel capsule, at least extracellular matrix elements, secreted by cells or added, and stem cells or at least differentiated RPE cells, if the cells introduced into the microcompartment are the stem cells, inducing cell differentiation within the cellular microcompartment, so as to obtain at least RPE cells, removing the hydrogel capsules, and recovering the RPE cells and any other retinal cells in the form of a hollow 3D retinal tissue unit.
The specification teaches wherein the hollow 3D retinal tissue units generically comprise RPE cells [page 6, 11], and the potential differentiation of RPE cells from generic stem cells or embryonic stem cells [page 20], but only teach the specific use of in vitro differentiated human RPE cells differentiated for hiPSCs [page 27-28, Figure 6-9]. The specification does not teach to use any RPE cells which were isolated from a subject rather than being differentiated in vitro.
The art at the time of filing teaches that the use of human embryonic stem cells (hESCs) and human-induced pluripotent stem cells (hiPSCs) to engineer retinal tissue is of particular interest due to the limited availability of suitable allogenic or autologous tissue [Hunt (2018), abstract]. Additionally, Hunt (2017) teaches the transplantation of retinal tissue and other cell types has been explored to treat retinal disease, wherein the transplantation of adult retinal cells has proved unsuccessful [Hunt (2017), page 330 col 1 ¶ 2]. Hunt (2017) teaches that some success was achieved with human fetal neural retina together with RPE transplanted in to the subretinal space, but that limited availability of fetal tissue for transplantation and the ethical issues associated with its use mean that it is unlikely to be a feasible treatment option for a large number of patients [page 330 col 1 ¶ 2]. Hunt (2017) further teaches that at least immature RPE and other retinal cells can be generated from hESCs or hiPSCs in vitro, and that transplantation of retinal cells derived from hESCs and hiPSCs is considered to be a promising treatment for patients with macular degeneration and inherited retinal disease [page 330 col 1 ¶ 2-3, Figure 1].
Thus, in view of the art-recognized unpredictability of using in vivo differentiated human RPE cells (such as isolated adult RPE cells) for transplantation into subjects to treat retinal disease such as AMD or RP, the art-recognized potential for use of retinal cells derived from hESCs and hiPSCs for transplantation, the lack of teachings in the specification to use any cells other than hESC- or hiPSC-derived retinal cells, the lack of teachings int eh specification of the actual use of any cells other than hiPSC-derived retinal cells, and the breadth of the claims, the ordinarily skilled artisan at the time of filing the instant application would have considered generating the hollow 3D retinal tissue unit with the organization and polarization as claimed using any cells other than living in vitro differentiated human RPEs differentiated in vitro from hiPSCs or hESCs as highly unpredictable. As such, it would have required undue experimentation to practice the scope of Applicant’s invention as claimed.
As a third issue (3), the specification does not provide an enabling disclosure for generating any retinal tissue unit other than a retinal tissue unit comprising an extracellular matrix layer present along the basal side of the at least one layer of living in vitro differentiated RPEs.
The broadest independent claims, claims 1 and 22, recite a hollow three-dimensional (3D) retinal tissue unit comprising, organized around an inner cavity, at least one layer of differentiated living human retinal pigment epithelium cells, with the basal side of each RPE cell pointing outwards and the apical side pointing towards the inner cavity. Additionally, claim 1 recites that the retinal tissue units are produced within a hydrogel capsule in tissue culture. Claim 22 recites that the retinal tissue unit is obtained from a pluripotent stem cell. Claims 2-20 depend on and/or encompass independent claim 1.
Claim 15 recite a method of making the retinal tissue unit of claim 1 comprising producing a cellular microcompartment comprising, within a hydrogel capsule, at least extracellular matrix elements, secreted by cells or added, and stem cells or at least differentiated RPE cells, if the cells introduced into the microcompartment are the stem cells, inducing cell differentiation within the cellular microcompartment, so as to obtain at least RPE cells, removing the hydrogel capsules, and recovering the RPE cells and any other retinal cells in the form of a hollow 3D retinal tissue unit.
As discussed above in detail, the specification teaches generic inclusion of an extracellular matrix (ECM) within the capsule [page 13, Figure 2]. The specification also teaches wherein the extracellular matrix layer can be formed by the cellular matrix secreted by RPE cells and/or by ECM added at the time of preparation of the cell unit, wherein preferably at least part of the extracellular matrix being provided in addition to the ECM naturally secreted by the cells [page 11, 17]. The specification further teaches that when the ECM is present, the retinal cells organized in three-dimensions around the inner cavity advantageously already interact with an ECM, which facilitates their implantation at the retina [page 12]. The specification also teaches methods of producing the hydrogel capsule which only comprise mixing the pluripotent stem cells with an extracellular matrix prior to encapsulating the mixture in a hydrogel layer [page 20, 26].
The specification additionally teaches and that cell polarization can be obtained by depositing a matrix layer on the inner face of alginate capsules [page 8, Figure 9], such that:
“Advantageously, the total or partial encapsulation in the hydrogel and the provision of ECM combined is a means capable of allowing the polarization of the retinal pigment epithelium cells. Indeed, the polarization of said cells can be obtained by depositing a layer of matrix on the inner face of the hydrogel capsules which positions the basal side of the cells, the tissue organizes itself around the cavity following this indication of polarity (as illustrated in figure 9, which shows that an extracellular matrix layer anchored to the alginate shell induces polarization of the cells as evidenced by the flattening of the tissue against the alginate due to the high tensile strength of the gel dictating the shape of the tissue).” [page 18].
Therefore, the specification teaches the criticality of including exogenous extracellular matrix within the alginate hydrogel capsule and provides no examples of producing retinal tissue units without the addition of extracellular matrix into the cell mixture prior to encapsidation within alginate hydrogels.
The specification teaches generic language about preferring that the hollow 3D retinal tissue units comprise ECM [page 4, 5], and only demonstrates the production of hollow 3D retinal tissue units which do comprise ECM [page 8, Figure 5-9].
The art at the time of filing also teaches that that RPE cells differentiated in culture from human iPSCs and grown on an extracellular matrix will develop in an orientation wherein the basal side of the RPE is attached to the extracellular matrix (ECM) [Khristov et al. 2018, Methods Mol. Biol., 1722, 223-247, page 230 step 7, Figure 1, 5]. Hunt (2018) also teaches polarization of RPE and/or other retinal cells obtained only when using scaffolds that have been treated with an ECM component; for example, PLDLA porous films coated in Col-IV, silk fibroin membrane coated in commercial dECM blend from human placenta, or PLA coated with fibronectin [page 25 col 2 ¶ 1, page 26 col 2 ¶ 2- page 27 col 1 ¶ 1, Table 6-7]. Hunt (2017) teaches that it is increasing being recognized that the extracellular matrix (ECM) is important for the correct development and function of the retina both in vivo and in vitro [Hunt et al. 2017, Acta Biomaterialia, 49, 329-343, page 330 col1 ¶ 4]. The art at the time of filing further teaches wherein a desirable characteristic of biomaterials for the production of retinal tissue in vitro includes allowing for the normal function of cells such as ECM production, and appropriate degradation time matched to the synthesis of new ECM by cells. Therefore, the art teaches the importance of the retinal tissue cells comprising an ECM. Additionally, the criticality of including the ECM within the biomaterial for production of the retinal tissue indicates the obligatory presence of an ECM upon maturation of the retinal tissue unit.
Neither the specification nor the art at the time of filing teaches that any hydrogel capsule other than an alginate capsule comprising an inner layer of extracellular matrix adjacent to the inner surface of the alginate can be used to encapsulate human RPE cells and produce a hollow ovoid three-dimensional retinal tissue unit having RPE cells which are polarized such that the basal side of each RPE cell points outward and the apical side of each RPE cell points towards the inner cavity.
Thus, in view of the art recognized importance of the ECM for correct development and function of the retina; the limitations in the specification to teaching only alginate hydrogel capsules comprising ECM; the art recognized need to substitute/alter alginate to promoter cellular attachment; the obligatory presence of an ECM in a tissue which has been cultured with the ECM; and the breadth of the claims; the ordinarily skilled artisan at the time of filing the instant application would have considered generating the hollow 3D retinal tissue unit with the organization and polarization as claimed without the tissue unit comprising an extracellular matrix layer adjacent to the basal surface of the RPE cell layer as highly unpredictable. As such, it would have required undue experimentation to practice the scope of Applicant’s invention as claimed.
Claim Rejections - 35 USC § 101
The rejection of amended and previously presented claims 1-3 and 6-8 under 35 U.S.C. 101 as being directed to a product of nature without significantly more is withdrawn over amended and previously presented claims 1-3 and 6-8 and newly applied to new claim 22 in view of Applicant’s amendments to the claim 1 to recite, “A hollow three-dimensional retinal tissue unit produced within a hydrogel capsule in tissue culture” such that the produce-by-process limitation adds sufficient structure to the retinal tissue unit to distinguish it from a naturally occurring eye. Applicant's amendments to the claims and arguments have been fully considered but have not been found persuasive in overcoming the rejection for reasons of record as discussed in detail below.
New claim 22 merely recites, “A hollow three-dimensional retinal tissue unit obtained from a pluripotent stem cell and comprising, organized around an inner cavity, at least one layer of differentiated living human retinal pigment epithelium cells, with the basal side of each retinal pigment epithelium cell pointing outwards and the apical side pointing towards the inner cavity”.
As discussed in the prior actions, the terms “hollow” and “inner cavity” in new claim 22 have been afforded their broadest reasonable interpretations such that “hollow” refers to a cell-free space interior to the unit, and “inner cavity” refers to a space that is at least partially surrounded by the RPE cell layer.
The recited retinal tissue unit is not markedly different from its naturally occurring counterpart because, as claimed, the retinal tissue unit encompasses a naturally occurring human eye. As taught by Singh (2018), a naturally occurring human eye comprises a hollow (e.g., cell-free) 3D retinal tissue unit comprising a layer of differentiated living human RPE cells organizes around an inner cell-free cavity, with the basal side of each RPE cell pointing outwards (e.g., towards Bruch’s membrane) and the apical side of each RPE cells pointing towards the inner cavity (e.g., towards the photoreceptor outer segments) [Singh et al. 2018, Stem Cell Reviews & Reports, 14, 463-483, cited in a prior action, Figure 1]. Singh (2018) teaches that the human eye is a hollow spheroid/ ovoid structure [Figure 1]. Singh (2018) also teaches wherein during development, six different neuronal cell types and a single glial cell type develop in a sequential order from a pool of cycling multipotential retinal progenitors [pp 465 col 1 ¶ 2]. Singh (2018) also teaches that human embryonic stem cells (hESCs) are derived from the inner cell mass of a blastocyst and are considered a limitless source of pluripotent stem cells [pp 467 col 1 ¶ 2]; as such, all differentiated cell types arise from pluripotent stem cells during the natural development of a human, including the cells of the eye.
Therefore, all the features recited in new claim 22 of the instant invention are indistinguishable from a naturally occurring human eye.
This judicial exception is not integrated into a practical application because the claim as written does not recite any additional features other than features of the naturally occurring eye, and as such, fails to integrate the judicial exception into a practical application. The use of the term “unit” is a generic phrase which does not impart any structure, and as such, does not distinguish the claimed retinal tissue unit from its naturally occurring counterpart of a human eye. Merely calling the product of nature by a generic name such as “unit” does not add a meaningful limitation as it is merely a nominal or token extra-solution component of the claim, and is nothing more than an attempt to generally link the product of nature to a particular technological environment.
The claim(s) does/do not include additional elements that are sufficient to amount to significantly more than the judicial exception because the claims do not recite any additional elements. Therefore, the claimed invention is directed to a product of nature without significantly more.
Applicant argues that claim 1 now recites in part “produced within a hydrogel capsule in tissue culture” and therefore this claim does not encompass the human eye.
However, this is agreed in part.
The limitation “produced within a hydrogel capsule in tissue culture” now recited in amended independent claim 1 is a product-by-process limitation, which limits the product only to the extent that the process necessarily requires or results in particular structural features.
Note that “even though product-by-process claims are limited by and defined by the process, determination of patentability is based on the product itself. The patentability of a product does not depend on its method of production. If the product in the product-by-process claim is the same as or obvious from a product of the prior art, the claim is unpatentable even though the prior product was made by a different process.” In re Thorpe, 777 F.2d 695, 698, 227 USPQ 964, 966 (Fed. Cir. 1985) (citations omitted).
In the instant case, the product-by-process limitation recited in claim 1 imparts sufficient structure such that any retinal tissue unit produced “within a hydrogel capsule in tissue culture” will not be able to fully reproduce the complex structure of a natural eye, such as a natural human eye. As such, the limitation overcomes the product of nature rejection over claims 1-20.
However, new claim 22 is not dependent on independent claim 1, and as such, the limitation does not overcome a finding of unpatentability under 35 U.S.C. 101 for new independent claim 22 encompassing a product of nature without significantly more. As such, Applicant’s arguments additionally do not overcome a finding of unpatentability under 35 U.S.C. 101 for new independent claim 22 encompassing a product of nature without significantly more.
Claim Rejections - 35 USC § 102
The rejection of amended and previously presented claims 1, 3-5, 7-9, and 12-14 under 35 U.S.C. 102(a)(1) as being anticipated by Singh et al. [2015, Stem Cell & Development, 24(23), 2778-2795], is withdrawn in view of Applicant’s claims which now recites “A hollow three-dimensional retinal tissue unit produced within a hydrogel capsule in tissue culture” in independent claim 1 and in view of the Declaration of Dr. Maxime Feyeux under 37 CFR 1.132.
Claim Rejections - 35 USC § 103
The rejection of amended and previously presented claims 1-20 under 35 U.S.C. 103 as being unpatentable over Singh et al. (2015, Stem Cell & Development, 24(23), 2778-2795); in view of Feyeux (WO2018096277A1, published 31 May 2018, IDS, English translation from PE2E); Singh et al. (2018, Stem Cell Reviews & Reports, 14, 463-483); Rizzolo et al. (2011, Progress in Retinal & Eye Research, 30, 296-323); and Singh et al. (2019, Stem Cells & Development, 28(17), 1151-1166, published 18 June 2019), is withdrawn in view of Applicant’s claims which now recites “A hollow three-dimensional retinal tissue unit produced within a hydrogel capsule in tissue culture” in independent claim 1 and in view of the Declaration of Dr. Maxime Feyeux under 37 CFR 1.132.
Double Patenting
The rejection of amended and previously presented claims 1-20 on the ground of nonstatutory double patenting as being unpatentable over claims 1-24 of copending Application No. 18/270,931, hereafter referred to as the ‘931 application, in view of Singh et al. (2015, Stem Cell & Development, 24(23), 2778-2795), is withdrawn.
Amended, previously presented, and new claims 1-20 and 22 are newly rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1-24 of copending Application No. 18/270,931, hereafter referred to as the ‘931 application, in view of Khristov et al. [2018, Methods Mol. Biol., 1722, 223-247].
The ‘931 application claims recite a 3D microcompartment of ovoid shape comprising an external hydrogel layer defining an internal part, said internal part comprising at least an ECM and at least two cysts, each cyst being formed by at least one layer of human or animal cells, organized three-dimensionally around a lumen, the smallest radius of the internal part being at least 100 µm (claim 1). Claim 3 further limits the cells to include RPE cells.
As such, the ‘931 application claims encompass the claims of the instant application by reciting a cyst formed of at least one layer of human RPE cells organized three-dimensionally around a lumen (e.g., a hollow 3D retinal tissue unit comprising at least one layer of hRPE cells organized around an inner cavity) and having an ECM around the retinal tissue/cyst which is encapsulated in a hydrogel capsule. Although the co-pending ‘931 application claims do not recite that the basal side of the each RPE cell is pointing outwards, and the apical side of each RPE cell is pointing towards the inner cavity, Khristov (2018) teaches that RPE cells differentiated in culture from human PSCs and grown on an extracellular matrix will develop in an orientation wherein the basal side of the RPE is attached to the extracellular matrix (ECM) [page 230 step 7, Figure 1, 5]. Therefore, given the teachings of Khristov that hPSC-derived RPE cells grown on an extracellular matrix will polarize such that the basal side is connect to the ECM, an ordinarily skilled artisan at the time of filing would expect that the RPE cells of the ‘931 application, when grown inside of the ECM-coated hydrogel capsule will orient in a direction such that the basal side of the cell is attached to the ECM, thereby orienting so that the basal side is along the outside of RPE layer and the apical side is pointing towards the inner cavity.
The ’931 application claim 5 also recites that the smallest radius is at least 200 um (e.g., between 10 and 1000 um, instant claim 5).
The ‘931 application claim 14 recites a method for preparing a cellular microcompartment according to claim 1, comprising incubating human or animal cells in a culture medium containing at least one cytoprotective factor, mixing the cells with ECM, encapsulating the suspension of cells in a hydrogel layer to form a microcompartment of ovoid shape, culturing the resulting microcompartments in an isotonic sensing buffer, then in a culture medium, and culturing the microcompartments for at least two cell division cycles (amplifying) in a culture medium without a cryoprotective factor, and optionally recovering the resulting cellular microcompartments. Accordingly, the method of the co-pending application is a species of the instant method for preparing a retinal tissue unit comprising the steps of producing a cellular microcompartment comprising, within a hydrogel capsule, at least ECM and differentiated RPE cells, followed by removing the hydrogel capsules to recover the 3D tissue unit.
Therefore, the ‘931 application claims encompass and render obvious the hollow 3D retinal tissue unit of the instant application. Additionally, the ‘931 application claims recite a method which is a species of the method of the instant application, thereby rendering obvious the instant method.
This is a provisional nonstatutory double patenting rejection.
Applicant argues that:
present claims are novel over Singh (2015) and also non-obvious over Singh (2015) even as combined with 4 other references; and
the earliest priority claimed in the co-pending application is after the priority date of the instant application, and since the present claims are novel and non-obvious, the double-patenting rejection must be withdrawn.
However, this is not agreed.
Regarding argument 1), Khristov (2018) was cited for that RPE cells differentiated in culture from human PSCs and grown on an extracellular matrix will develop in an orientation wherein the basal side of the RPE is attached to the extracellular matrix (ECM) [page 230 step 7, Figure 1, 5].
Regarding argument 2), the present claims are currently not allowable. Provisional nonstatutory double patenting rejections do not require that the co-pending application have a prior effective filing date. As set forth in MPEP 804(I)(B)(1), if two (or more) pending applications are filed, in each of which a rejection of one claimed invention over the other on the ground of provisional nonstatutory double patenting (NSDP) is proper, the provisional NSDP rejection will be made in each application (emphasis added). Additionally, a complete response to a nonstatutory double patenting (NDP) rejection is either a reply by applicant showing that the claims subject to the rejection are patentably distinct from the reference claims or the filing of a terminal disclaimer in accordance with 37 CFR 1.321 in the pending application(s) with a reply to the Office action (MPEP 804 (B)(1)). Such a response is required even when the nonstatutory double patenting rejection is provisional.
Therefore, Applicant’s amendments and arguments do not overcome the provisional nonstatutory double rejection over claims 1-24 of copending Application No. 18/270,931, in view of Khristov et al. [2018, Methods Mol. Biol., 1722, 223-247].
Conclusion
No claim is allowed.
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DR. KATIE L. PENNINGTON
Examiner
Art Unit 1634
/KATIE L PENNINGTON/Examiner, Art Unit 1634
Dr. A.M.S. Wehbé
/ANNE MARIE S WEHBE/Primary Examiner, Art Unit 1634
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