DETAILED ACTION
Applicant’s amendment and Arguments/Remarks received on 17 July 2026 have been entered. Claims 1-9, 12-13, 17-18, 22-24, 26-27, 29, and 32 were previously pending in the application. Claims 2 and 17 have been cancelled, and no new claims have been added by Applicant.
Claims 1, 3-9, 12-13, 18, 22-24, 26-27, 29, and 32 are currently pending in the application. Claims 1, 24, 26, 27, and 29 are independent claims.
The election of Group I, drawn to a vector for liver and/or splenic phagocyte-specific expression and a cell comprising the vector, remains in effect in the instant application.
The following election of species remains in effect in the instant application:
Phagocytes: a. Macrophage: ii. MRC1+ macrophage;
Phagocyte-specific promoters and/or enhancers: a. MRC1 promoter and/or enhancer.
Claims 26, 27, and 29 remain withdrawn from consideration as being directed to a nonelected invention, there being no allowable generic or linking claim.
Claim 5 remains withdrawn from consideration as being directed to a nonelected species, there being no allowable generic or linking claim.
Claims 1, 3-4, 6-9, 12-13, 18, 22-24, and 32 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/EP2021/084276, filed 03 December 2021, which claims priority to United Kingdom Application 2019108.6, filed 03 December 2020. Filing of a certified copy of the United Kingdom Application 2019108.6, filed 03 December 2020 is acknowledged.
Thus, the earliest possible priority for the instant application is 03 December 2020.
37 CFR 1.821-1.825
Applicant has not amended the specification to recite SEQ ID NOs for all of the amino acid and/or nucleotide sequences of specification page 81 lines 23-29 wherein the 2) linker sequence, 4) linker sequence, and 6) termination sequence; specification page 84 lines 22-23; and specification pages 84 line 24, page 85 lines 27 and 31, page 86 lines 3, 13, and 28, and page 87 line 21 as identified in the prior action, and so has not addressed the identified issue of noncompliance with the requirements of 37 CFR 1.821 through 1.825. Specifically, the amino acid 2) linker sequence (AGQLM) and 6) termination sequence (TRAS) recited on page 81 lines 24 and 29 of the specification filed 01 June 2023, respectively, are not accompanied by SEQ ID NOs as required by 37 CFR 1.821 through 1.825. Note that page 81 lines 24 and 29 of the specification filed 01 June 2023 corresponds to page 82 lines 17 and 22, respectively, of the specification amendment filed 14 February 2024, which similarly lacks SEQ ID NOs: for the sequence AGQLM and TRAS recited for the linker and termination sequences. As such, the specification filed 17 July 2026 has not been entered.
Therefore, this application still fails to comply with the requirements of 37 CFR 1.821 through 1.825 for the reason(s) set forth below and on the Notice to Comply With Requirements For Patent Applications Containing Nucleotide Sequence And/Or Amino Acid Sequence Disclosures which is attached to this communication.
If the unidentified sequences of specification page 81 lines 23-29 wherein the 2) linker sequence, 4) linker sequence, and 6) termination sequence; specification page 84 lines 22-23; and specification pages 84 line 24, page 85 lines 27 and 31, page 86 lines 3, 13, and 28, and page 87 line 21 are included in the submitted sequence listing, Applicant must amend the claims and specification and/or drawings to comply with the sequence identification requirements. Alternatively, if the unidentified sequences of specification page 81 lines 23-29 wherein the 2) linker sequence, 4) linker sequence, and 6) termination sequence; specification page 84 lines 22-23; and specification pages 84 line 24, page 85 lines 27 and 31, page 86 lines 3, 13, and 28, and page 87 line 21 are not included in the presently submitted sequence listing, Applicant must submit an updated sequence listing in compliance with 37 CFR 1.821(c)-(d) and 37 CFR 1.825(b). See also the attached Notice to Comply.
APPLICANT IS GIVEN A THREE MONTH EXTENDABLE PERIOD WITHIN WHICH TO COMPLY WITH THE SEQUENCE RULES, 37 CFR 1.821-1.825. Failure to comply with these requirements will result in ABANDONMENT of this application under 37 CFR 1.821 (g). Extension of time may be obtained by filing a petition accompanied by the extension fee under the provisions of 37 CFR 1.136. In no case may an applicant extend the period for response beyond the six month statutory period. Applicant is requested to return a copy of the attached Notice to Comply with the response.
Required response – Applicant must provide:
A substitute specification in compliance with 37 CFR 1.52, 1.121(b)(3) and 1.125 inserting the required sequence identifiers, including into the Brief Description of the Drawings, consisting of:
A copy of the previously-submitted specification, with deletions shown with strikethrough or brackets and insertions shown with underlining (marked-up version);
A copy of the amended specification without markings (clean version); and
A statement that the substitute specification contains no new matter.
Applicant argues that the alleged informality is believed to have been addressed in a substitute specification that Applicant filed on 14 February 2024 (second preliminary amendment). However, this is not agreed.
Although the substitute specification filed 14 February 2024 includes SEQ ID NOs: for the sequences identified on page 81 lines 23-29 of 4) linker sequence; specification page 84 lines 22-23; and specification pages 84 line 24, page 85 lines 27 and 31, page 86 lines 3, 13, and 28, and page 87 line 21 of the originally filed specification; neither the substitute specification filed 14 February 2024 nor the specification filed 17 July 2026 recite SEQ ID NOs: for the sequences of page 81 lines 24 and 29 corresponding to 2) linker sequence AGQLM and 6) termination sequence TRAS (page 82 lines 17 and 22 of the substitute specification filed 14 February 2024). As such, Applicant’s arguments do not overcome the issue of noncompliance with the requirements of 37 CFR 1.821 through 1.825.
Specification
The objection to the specification of the disclosure for the use of trade names and/or marks used in commerce without the accompanying generic terminology and/or proper symbol indicating use in commerce such as ™, SM , or ® following the terms is maintained due to the amendment to the specification of the disclosure filed 17 July 2026 having not been entered. As described above, the specification amendment filed 17 July 2026 still lacks SEQ ID NOs: for recited amino acid sequences.
Note that the specification has not been inspected sufficiently to identify all instances of trade names and/or marks used in commerce. It is Applicant’s responsibility to ensure complete compliance.
Claim Rejections - 35 USC § 112(b)
The rejection of amended and cancelled claims 1-4, 6-9, 12-13, 17-18, 22-24, and 32 under 35 U.S.C. 112(b) as failing to particularly point out and distinctly claim the subject matter which the inventor(s) regards as the invention for multiple issues of indefiniteness is withdrawn in view of Applicant’s amendments to the claims.
**The following new rejection is necessitated by Applicant’s amendments to the claims.
Amended claims 3-4 and 8 are 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 3 and 4 each recites the limitation "the phagocyte" in line 1. There is insufficient antecedent basis for this limitation in the claim. Claims 3 and 4 are each dependent on independent amended claim 1, which now recites “liver and splenic phagocyte-specific expression” in lines 1-2, “a phagocyte-specific promoter and/or enhancer” in line 5, and “liver phagocytes” in line 7. Therefore, it is unclear which phagocytes “the phagocyte” recited in each of claim 3 and 4 are referring.
Claim 3 additionally recites, “wherein the phagocyte is a liver or splenic phagocyte” in lines 1-2. As described above, it is unclear which phagocyte this limitation in claim 3 is references. To the extent that the phagocyte of claim 3 line 1 is referencing the “liver and splenic phagocyte-specific expression” recited in independent claim 1 lines 1-2, the limitation in claim 3 contradicts with the limitation in claim 1 by reciting liver “or” splenic wherein claim 1 requires liver “and” splenic. Similarly, to the extent that “the phagocyte” of claim 3 is referring to the “liver phagocyte” recited in claim 1 line 7, then the limitation of claim 3 conflicts with the limitation of claim 1 which requires the phagocytes to be specifically liver phagocytes.
As such, the metes and bounds of the claim cannot be determined.
Amended claim 8 recites, “wherein the one or more miRNA target sequence suppresses transgene expression in splenic phagocytes” in lines 1-4, which is indefinite because suppression of expression in splenic phagocytes conflicts with the limitation of claim 1, upon which claim 8 depends, which requires “A viral vector for liver and splenic phagocyte-specific expression”. As such, the metes and bounds of the claim cannot be determined.
Claim Rejections - 35 USC § 112(a)- Written Description
The rejection of amended and cancelled claims 1, 2, 6, and 7 under 35 U.S.C. 112(a) for reciting “a fragment thereof” is withdrawn in view of Applicant’s amendments to the claims such that claims 6 and 7 no longer recite “a fragment thereof”.
Claim Rejections - 35 USC § 102
The rejection of amended and cancelled claims 1-4, 6-9, 24, and 32 under 35 U.S.C. 102(a)(1) as being anticipated by Kambara et al. [2015, The American Journal of Pathology, 185(1), 162-171, IDS], as evidenced by Moseman et al. [2013, The Journal of Immunology, 191(11), 5615-5624]; is withdrawn in view of Applicant’s cancellation of claim 2 and amendments to the claims such that claim 1 now recites “A viral vector”.
Claim Rejections - 35 USC § 103
The rejection of amended and cancelled 1-4, 6-9, 12-13, 17-18, 22-24, and 32 under 35 U.S.C. 103 as being unpatentable over He & Marneros [2014, Journal of Biological Chemistry, 289(12), 8019-8028]; in view of Kambara et al. [2015, The American Journal of Pathology, 185(1), 162-171, IDS]; Zhang et al. [2016, Biomaterials, 84, 1-12]; Zhu et al. [2015, Oncology Reports, 34, 2643-2655]; Geisler & Fechner [2016, World Journal of Experimental Medicine, 6(2), 37-54]; and De Palma et al. [2008, Cancer Cell, 14,299-311, IDS]; is withdrawn in view of Applicant’s claims which now recite “wherein the one or more miRNA target sequence … suppresses transgene expression in … liver sinusoidal endothelial cells” in claim 1.
**The following new rejection is necessitated by Applicant’s amendments to the claims.
Amended claims 1, 3-4, 6-9, 12-13, 18, 22-24, and 32 are newly rejected under 35 U.S.C. 103 as being unpatentable over He & Marneros [2014, Journal of Biological Chemistry, 289(12), 8019-8028, cited in a prior action]; in view of Kambara et al. [2015, The American Journal of Pathology, 185(1), 162-171, IDS, cited in a prior action]; Zhang et al. [2016, Biomaterials, 84, 1-12, cited in a prior action]; Zhu et al. [2015, Oncology Reports, 34, 2643-2655, cited in a prior action]; Geisler & Fechner [2016, World Journal of Experimental Medicine, 6(2), 37-54, cited in a prior action]; and Oda et al. [2018, The American Journal of Pathology, 188(4), 916-928, IDS]; De Palma et al. [2008, Cancer Cell, 14,299-311, IDS, cited in a prior action]; Merlin & Follenzi [2019, Molecular Therapy: Methods & Clinical Development, 12, 223-232, IDS]; and Laffont et al. [2016, Cellular Haemostasis and Platelets, 115, 311-323].
Regarding claim 1, He teaches a vector for MCR1+ macrophage specific expression, wherein the vector comprises a transgene (i.e., luciferase) operably linked to an expression control sequence, wherein the expression control sequence is the human MRC1 promoter [column 4 ¶ 2-3, column 6 ¶ 5, Supplemental Figure S1A].
He does not teach the intended use wherein the vector is for liver or splenic-specific expression.
However, Kambara teaches that the MRC1 promoter drives transgene expression in MCR1+ M2 macrophages within the liver and spleen [column 3 ¶ 3, column 8 ¶ 3, Figure 1C]. Kambara also teaches that M2 macrophages upregulate MRC1 and are able to promote tissue remodeling, angiogenesis, and tumor development [column 2 ¶ 1].
Additionally, Zhang teaches that during cancer development and treatment, such as in hepatocellular carcinoma (HCC), tumors are infiltrated by tumor-associated macrophages (TAMs), which are recruited in the monocyte precursor form from the peripheral blood and stimulate tumor progression by facilitating angiogenesis, invasion, and metastasis, and suppressing antitumor immunity [column 2 ¶ 2]. Zhang further teaches that TAMs are typically non-inflammatory M2-type macrophages which specifically express CD206/MRC1 [column 3 ¶ 2]. Zhang also teaches targeting CD206/MCR1-expressing TAMs to treat tumors resistant to conventional therapy, wherein targeting the TAMs inhibited tumor growth and lung metastasis [abstract, column 3 ¶ 4].
Further, Zhu teaches specifically targeting macrophages in the liver, such that CD206/MRC1 is highly expressed in human hepatocellular carcinoma (HCC) tissues compared to the level in peri-carcinoma tissue, and that the macrophage blocker GdCl3 suppresses HCC progression by downregulating the expression of CD206 in M2 TAMs, thereby reducing the invasiveness of HCC cells [abstract, column 2 ¶ 2, column 14 ¶ 1, column 15 ¶ 1, column 19 ¶ 2]. Zhu also teaches that the reduction of invasiveness is possibly by affecting the secretion of cytokines from the M2 macrophages [column 19 ¶ 2].
Therefore, given the teachings of Kambara that the M2 macrophage-specific MRC1 promoter drives transgene expression in liver and spleen, the teachings of Zhang that targeting CD206/MCR1+ macrophages inhibits tumor growth and metastasis, and the teachings of Zhu that targeting CD206/MCR1+ macrophages in HCC reduces the invasiveness of HCC cells by affecting the secretion of cytokines from the macrophages, an ordinarily skilled artisan at the time of filing the instant application would have been motivated to target TAMs in an HCC tumor to modulate TAM secretion and reduce tumor growth, invasiveness, and metastasis.
He does not teach wherein the one or more expression control sequences comprises one or more miRNA target sequence, wherein the one or more miRNA target sequence suppresses transgene expression in cells other than liver phagocytes and suppresses transgene expression in hepatocytes and liver sinusoidal endothelial cells (LSECs).
Geisler teaches that miRNA-dependent post-transcriptional suppression of transgene expression has been emerging as powerful new technology to increase the specificity of vector-mediated transgene expression [column 2 ¶ 1]. Geisler additionally teaches that to control exogenous transgene expression, tandem repeats of artificial miRNA target sites are usually incorporated into the 3’ UTR of the transgene expression cassette, leading to subsequent degradation of transgene mRNA in cells expressing the corresponding miRNA, wherein the targeting strategy, which was first shown for lentiviral vectors in antigen presenting cells, has now been used for tissue-specific expression of vector-encoded therapeutic transgenes to reduce immune response against the transgene, to control virus tropism for oncolytic virotherapy, to increase safety of live attenuated virus vaccines, and to identify and select cell subsets for pluripotent stem cell therapies [column 12 ¶ 1].
Geisler further teaches that miR-122 is a miRNA almost exclusively expressed in liver tissue and represents the best candidate for a specific hepatocyte-de-targeting, such that successful miR-122-mediated suppression of transgene expression in liver was found in several studies [column 6 ¶ 2, Table 1]. Geisler teaches that miR-122 has specifically been used in a metastatic HCC model to de-target hepatocytes for expression of an AAV8-delivered HSV thymidine kinase and luciferase transgenes [Table 1].
Geisler also teaches that miR-126 has been successfully used to de-target lentivirally delivered Interferon-α (IFN-α) away from hematopoietic stem and progenitor cells to target tumor-infiltrating macrophages, thereby inhibiting breast cancer progression [column 16 ¶ 2-column 17 ¶ 1, Table 1].
Therefore, given the teachings of Geisler of the cell-specific de-targeting using miR-122 and/or miR-126, an ordinarily skilled artisan at the time of filing the instant application would have been motivated to include target sites for miRNAs, in a transgene expression vector to reduce expression in non-targeted cells, and particularly to use miR-122 and/or miR-126 target sequences to reduce hepatocyte and HSC expression of transgenes targeting tumor infiltrating macrophages in HCC.
Further, Oda teaches wherein miR-122 (i.e., miR-122-5p) is strongly expressed in hepatocytes and moderately expressed in LSEC and wherein miR-126 (i.e., miR-126-3p) is strongly expressed in LSEC and moderately expressed in hepatocytes [Figure 2]. Therefore, given that LSECs and hepatocytes both comprise miR-122 and miR-126, a viral vector comprising miR-122 and/or miR-126 target sequences to reduce hepatocyte and HSC expression of transgenes targeting tumor infiltrating macrophages in HCC would suppress transgene expression in both hepatocytes and LSECs.
Additionally, Merlin teaches wherein miR-122 represses transgene constructs comprising miR-122 target sequences within hepatocytes and miR-126 represses transgene constructs comprising miR-126 target sequences within LSECs [Figure 1].
He additionally does not teach wherein the vector is a viral vector.
Geisler teaches that miR-126 has been used successfully to de-target lentivirally delivered interferon-α (IFN-α) away from hematopoietic stem and progenitor cells to target IFN-α expression in tumor-infiltrating macrophages, thereby inhibiting breast cancer progression [column 16 ¶ 2-column 17 ¶ 1, Table 1]. De Palma teaches the delivery of the IFN-α transgene to the TEMs in a lentiviral vector (LVs) regulated by the phagocyte-specific Tie2 promoter/enhancer [column 3 ¶ 2, column 20 ¶ 4]. De Palma further teaches that by intravenous injection, LVs preferentially transduce the liver and spleen [column 8 ¶ 2].
Merlin teaches that lentiviral vectors are one of the best tools available in the effort to develop efficient viral vectors for gene therapy, wherein they have several characteristics which make them vary attractive, including 1) an active transport mechanism to translocate the genomic material into the nucleus regardless of cell cycle status; 2) a lack of viral protein expression after transduction; 3) accommodation of expression cassettes of up to 10 kb; 4) a low or absent genotoxicity; and 5) a sustained transgene expression after transduction due to their genome-integration ability [pg 223 col 2 ¶ 5- pg 224 col 1 ¶ 1]. Merlin also teaches wherein pantropic transgene expression can be averted with the use of viral glycoproteins such as the envelope proteins of hepatitis B Virus and the baculovirus GP64, which allow hepatocyte-restricted LV entry [pg 224 col 1 ¶ 3].
Therefore, an ordinarily skilled artisan at the time of filing the instant application would have been motivated to deliver a transgene, such as IFN-α, for macrophage-specific expression in the liver and/or spleen by using a lentiviral vector to achieve preferential transduction of the liver and spleen.
Regarding claims 6-7, He teaches wherein the vector comprises a macrophage-specific MRC1 promoter [column 4 ¶ 2-3, column 6 ¶ 5, Supplemental Figure S1A].
Regarding claim 3-4 and 32, as discussed above, Kambara discloses wherein the MRC1 promoter drives transgene expression in liver and splenic MCR1+ macrophages [column 3 ¶ 3-column 4 ¶ 3, column 8 ¶ 3, Figure 1C].
Regarding claims 8-9, as discussed above, He, Kambara, Zhang, Zhu, Geisler, and Oda teach the limitations of claim 1, including the motivation to include target sites for miRNAs, in a transgene expression vector to reduce expression in non-targeted cells, and particularly to use miR-122 and/or miR-126 target sequences to reduce hepatocyte, LSEC, and HSC expression of transgenes targeting tumor infiltrating macrophages in HCC.
Oda further teaches wherein miR-126-3p and miR-126-5p are moderately expressed in the spleen [Figure 3], but does not teach wherein miR-126 is specifically expressed in splenic phagocytes.
Laffont teaches wherein splenic macrophages take up extracellular platelet-derived microparticles (MPs) comprising functional miR-126-3p which downregulate mRNA targets of miR-126-3p within the macrophages [abstract, pg 311 col 2 ¶ 1]. Therefore, given that splenic macrophages comprise miR-126, a viral vector comprising a miR-126 target sequence to reduce hepatocyte and LSEC expression of a transgene would also suppress transgene expression in splenic macrophages.
Regarding claim 12-13, He, Kambara, Zhang, Zhu, Geisler, and Oda teach the limitations of claim 1, including the motivation to target TAMs in an HCC tumor to modulate TAM secretion and reduce tumor growth, invasiveness, and metastasis. As discussed above, Geisler teaches that miR-126 has been used successfully to de-target lentivirally delivered interferon-α (IFN-α) away from hematopoietic stem and progenitor cells to target IFN-α expression in tumor-infiltrating macrophages, thereby inhibiting breast cancer progression [column 16 ¶ 2-column 17 ¶ 1, Table 1].
Geisler does not teach motivation to specifically deliver IFN-α to tumor-infiltrating macrophages within the liver.
However, De Palma teaches that that type I IFNs, including IFN-α, increase tumor immunogenicity, recruit and activate immune cells within the tumor milieu, and may eventually facilitate tumor rejection [column 3 ¶ 1]. De Palma also teaches delivering tumor-homing proangiogenic, tumor-infiltrating Tie2-expressing monocytes (TEMs) genetically modified to express IFN-α, which achieved substantial antitumor activity in both prevention and intervention trials without measurable toxicity [abstract, column 1 ¶ 1- column 2 ¶ 2, column 3 ¶ 2, SIGNIFICANCE box]. De Palma further teaches that their experiments illustrate the therapeutic potential of gene- and cell-based IFN-α delivery and should allow the development of IFN treatments that more effectively treat cancer [abstract].
Therefore, given the teachings of Geisler of delivering IFN-α to tumor infiltrating macrophages; the teachings of De Palma to modify tumor-infiltrating monocytes to secrete IFN-α to selectively deliver the IFN-α to tumors; and the teaching of Zhang discussed above that tumor-associated macrophages (TAMs) are recruited in the monocyte precursor form from the peripheral blood; an ordinarily skilled artisan at the time of filing the instant application would be motivated to modify tumor infiltrating monocytes/macrophages to express an IFN-α transgene, thereby secreting IFN-α within a tumor as a means to selectively deliver the antitumor IFN-α to tumors to facilitate tumor rejection.
Regarding claims 22, He, Kambara, Zhang, Zhu, Geisler, and Oda teach the limitations of claim 1. He further teaches that MRC1 is a prototypical M2-type marker in human macrophages [column 6 ¶ 5].
Kambara also teaches that Mrc1 is an M2 phenotypic marker [column 3 ¶ 3].
Zhang teaches that M2 macrophages specifically express CD206/MRC1 and using CD206 expression to differentiation M1 and M2 polarized macrophages, wherein only M2 polarized macrophages are stained with an αCD206 antibody probe [column 3 ¶ 2, Figure 1C-E]. Zhang also specifically teaches a FACS for sorting 4T1 tumor cells, wherein less than 5% of cells expressing CD206 are not gated as M2 macrophages [Figure 1D].
Zhu teaches that CD206/MRC1 expression is correlated with the alternatively activated (M2) polarization of macrophages [column 2 ¶ 4].
De Palma teaches that lentivirus preferentially transduces liver and spleen [column 8 ¶ 2].
Additionally, Geisler teaches that cell specificity can be further enhanced by incorporating miRNA target sequences into transgene vectors, including incorporation of target sites for miR-126 to de-target HSCs to facilitate specific targeting of tumor-infiltrating macrophages and miR-122 to de-target hepatocytes in an HCC model [Table 1]. Oda teaches wherein miR-122 and miR-126 are expressed in hepatocytes and LSEC [Figure 2]. Laffont teaches wherein miR-126 is present in splenic macrophages [abstract, pg 311 col 2 ¶ 1].
Therefore, given the teachings of He, Kambara, Zhang, and Zhu that MRC1 expression/ MRC1 promoter activity is specific to M2 macrophages; the teaching of Zhang of less than 5% of cells other than M2 macrophages in a mixed cell population expressing DC206/MRC1; the teaching of De Palma that lentivirus preferentially transduces liver and spleen; and the teachings of Geisler, Oda, and Laffont to incorporate miRNA target sites to de-target hepatocytes, LSECs, splenic macrophages, and HSCs, an ordinarily skilled artisan at the time of filing the instant application would expect the transduced vector to express the transgene specifically in MCR1+ macrophages, such that less than 5% of cells other than phagocytes would express the transgene following transduction.
Regarding claims 23, He, Kambara, Zhang, Zhu, Geisler, and Oda teach the limitations of claim 1.
Kambara also teaches substantially higher expression from the native MRC1 promoter in macrophages of the lung, liver, and spleen compared to the kidney [Figure 1].
Zhang teaches wherein an αCD206 antibody probe stains only liver, kidney, spleen, stomach, and tumor [Figure 3D-E].
Zhu teaches that CD206/MRC1 was highly expressed in the HCC tissues compared to the level in peri-carcinoma tissue [abstract, column 3 ¶ 4, column 6 ¶ 6- column 7 ¶ 1].
De Palma teaches that lentivirus preferentially transduces liver and spleen [column 8 ¶ 2].
Therefore, given the teachings of Kambara that the MRC1 promoter drives expression selectively in lung, liver, and spleen; the teaching of Zhang that an αCD206 antibody probe stains only liver, kidney, spleen, stomach, and tumor; the teaching of Zhu that MRC1 expression is particularly high within HCC tumors; and the teaching of De Palma that lentivirus preferentially transduces liver and spleen; an ordinarily skilled artisan at the time of filing the instant application would expect the transduced vector to express the transgene specifically in liver cells and splenic cells, such that less than 5% of cells of cell types other than liver cells and splenic cells would express the transgene following transduction.
Regarding claim 24, He teaches a cell comprising the MCR1-luc vector [column 4 ¶ 3].
Given the motivation taught by Kambara, Zhang, and Zhu to target TAMs in an HCC tumor to modulate TAM secretion and reduce tumor growth, invasiveness, and metastasis; the motivation taught by Geisler to include target sites for miRNAs, in a transgene expression vector to reduce expression in non-targeted cells, and particularly to use miR-122 and/or miR-126 target sequences to reduce hepatocyte and HSC expression of transgenes targeting tumor infiltrating macrophages in HCC; the expectation taught by Oda that a viral vector comprising miR-122 and/or miR-126 target sequences would suppress transgene expression in both hepatocytes and LSECs; the motivation taught by Geisler and De Palma to deliver a IFN-α transgene for macrophage-specific expression in the liver and/or spleen by using a lentiviral vector to achieve preferential transduction of the liver and spleen; the expectation taught by Laffont that a viral vector comprising a miR-126 target sequence to reduce hepatocyte and LSEC expression of a transgene would also suppress transgene expression in splenic macrophages; the motivation taught by Geisler, De Palma, and Zhang to modify tumor infiltrating macrophages to express an IFN-α transgene to selectively deliver the antitumor IFN-α to tumors; the expectations taught by He, Kambara, Zhang, Zhu, De Palma, Geisler, Oda, and Laffont that transduction with a lentiviral vector comprising a transgene operably linked to an MCR1 promoter and target sequences for miR-122 and/or miR-126 would result in transgene expression in less than 5% of cells other than phagocytes and less than 5% of cells of cell types other than liver and splenic cells; it would have been prima facie obvious to an ordinarily skilled artisan at the time of filing the instant application to modify the vector of He to replace the luciferase transgene with a transgene encoding IFN-α, to include target sites for miR-122 and/or miR-126 to de-target hepatocytes, LSECs, splenic phagocytes, and/or HSCs, and to package the transgene expression cassette in a lentiviral vector for specific expression in tumor-infiltrating macrophages within the liver and/or spleen to treat a tumor, such as HCC such that 5% or less of non-phagocytes and 5% or less of non-liver and non-splenic cells express the transgene, with a reasonable expectation of success.
Insofar as Applicant’s arguments apply to this new grounds of rejection, Applicant argues that:
the reporter construct of He is a plasmid that comprises the reporter gene luciferase under the control of the MRC1 promoter used to generate a stable cell line to screen for polarization and not to treat HCC, and He does not teach a viral vector, one or more miRNA target sequences, nor specificity for liver and splenic phagocyte expression;
Kambara is silent on miRNA target sequences, does not teach wherein the MRC1 promoter drives transgene expression in MRC1+ M2 macrophages within the liver and spleen, and teaches that MRC1 promoter drives expression in at least the lungs further to the liver and spleen;
Zhang merely teaches testing an antibody probe for tumor-associated macrophages (TAMs) wherein TAMs are possible therapeutic targets for the treatment of cancer, wherein the anti-CD206 antibody conjugate is used to deplete non-organ specific CD206+ TAMs associated with metastatic breast cancer, but does not disclose a viral vector that is specific for transgene expression in liver and splenic phagocytes, and so is not relevant to the problem solved by the instant invention and further teaches away from the instant invention by targeting non-organ specific CD206+ TAMs for depletion;
Zhu is not directed to the same problem as the instant invention and merely teaches that GdCl3 possibly reduces invasiveness of HCC cells by suppression of CD206 expression and promotion of apoptosis of non-organ specific CD206+ TAMs, such that the teachings of Zhu are unrelated to viral vectors specific for transgene expression in liver and splenic phagocytes and only hindsight knowledge would motivate a person of ordinary skill to combine Zhu with He and Kambara;
Geisler does not provide motivation or suggestion to provide a viral vector that is for liver and splenic phagocyte-specific transgene expression nor provide any teaching to the skilled person as to the ability of miRNA target sequences to enable specific expression in liver and splenic phagocytes;
De Palma uses vectors which are not designed for tissue-specific expression and merely transduces HSPCs ex vivo, and further teaches away from the instant invention by suggesting that macrophage progenitors should be modified ex vivo and re-introduced for treatment; and
none of the prior art documents identify the issue of LSEC expression that the inventor successfully ameliorated using miRNA target sequences.
However, this is not agreed.
In response to Applicant’s arguments against the references individually, it is noted that the test for obviousness is not whether the features of a secondary reference may be bodily incorporated into the structure of the primary reference; nor is it that the claimed invention must be expressly suggested in any one or all of the references. Rather, the test is what the combined teachings of the references would have suggested to those of ordinary skill in the art. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981). One cannot show nonobviousness by attacking references individually where the rejections are based on combinations of references. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981); In re Merck & Co., 800 F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986). Further, the Examiner recognizes that obviousness may be established by combining or modifying the teachings of the prior art to produce the claimed invention where there is some teaching, suggestion, or motivation to do so found either in the references themselves or in the knowledge generally available to one of ordinary skill in the art. See In re Fine, 837 F.2d 1071, 5 USPQ2d 1596 (Fed. Cir. 1988), In re Jones, 958 F.2d 347, 21 USPQ2d 1941 (Fed. Cir. 1992), and KSR International Co. v. Teleflex, Inc., 550 U.S. 398, 82 USPQ2d 1385 (2007). In addition, it must be recognized that any judgment on obviousness is in a sense necessarily a reconstruction based upon hindsight reasoning. But so long as it takes into account only knowledge which was within the level of ordinary skill at the time the claimed invention was made, and does not include knowledge gleaned only from the applicant's disclosure, such a reconstruction is proper. See In re McLaughlin, 443 F.2d 1392, 170 USPQ 209 (CCPA 1971).
Specifically, regarding Applicant’s argument 1), note that He was not relied on for teaching every element of the claimed invention. He was cited for teaching a vector for MCR1+ macrophage specific expression, wherein the vector comprises a transgene (i.e., luciferase) operably linked to an expression control sequence, wherein the expression control sequence is the human MRC1 promoter [column 4 ¶ 2-3, column 6 ¶ 5, Supplemental Figure S1A]. He was not relied upon to teach a viral vector, one or more miRNA target sequences, nor specificity for liver and splenic phagocyte expression.
As discussed above, lentiviral delivery of transgenes was well-known at the time of filing the instant application, including preferential transduction of liver and splenic-specific, as taught by Geisler, De Palma, and Merlin.
As further discussed above, the MCR1+ promoter was well known at the time of filing to produce phagocyte-specific expression, including M2 macrophages in the liver, as taught by Kambara and Zhu.
Additionally, Geisler, Oda, Merlin, and Laffont were cited for teaching the motivation to de-target non-target cells types by including cell-type specific miRNA target sequences within the expression construct to minimize expression within cell types expressing the corresponding miRNA.
Therefore, as discussed above, given the teachings of the cited art, the combination of a cell type specific promoter, a tissue selective viral vector, and cell-type specific miRNA targeting sequences for targeted delivery of a transgene would have been obvious to an ordinarily skilled artisan at the time of filing the instant application. In fact, Merlin explicitly teaches the several approaches to restrict transgene expression in desired cell types after vector delivery include 1) transcriptional regulation using cell-type-specific promoters; 2) post-transcriptional regulation, or transgene de-targeting, taking advantage of the expression of different miRNAs in certain cell types; and 3) restriction of transduction by means of glycoproteins derived from several viruses which confer LVs to the tropism for the desired cell types [pg 224 col 1 ¶ 4- col 2 l¶ 1, Figure 1].
Additionally, an ordinarily skilled artisan at the time of filing the instant application would have had a reasonable expectation of success not only in obtaining the structure of the claimed viral vector, but also in obtaining the functions of the claimed viral vector for cell-type specific expression of the transgene within macrophages of the liver.
Regarding Applicant’s argument 2), note that Kambara was not cited for teaching miRNA target sequence. Kambara was cited for teaching that the MRC1 promoter drives transgene expression in MCR1+ M2 macrophages within the liver and spleen [column 3 ¶ 3, column 8 ¶ 3, Figure 1C], albeit not exclusively within the liver and spleen in that expression was also observed within the lung and kidney. Kambara also teaches that M2 macrophages upregulate MRC1 and are able to promote tissue remodeling, angiogenesis, and tumor development [column 2 ¶ 1].
Note that the cell type specific expression attributed to the instantly claimed vector is not obtained merely by use of a cell type specific promoter, but it a combination of a cell-type specific promoter, a tissue selective viral vector, and cell-type specific de-targeting using miRNAs. As such, the liver macrophage expression results from the intersection of the three impacting transgene expression. Accordingly, the fact that M2 macrophages in non-liver/spleen organs are capable of expressing the MCR1 promoter is not a teaching away from the instant invention.
Regarding Applicant’s argument 3), Zhang was not cited for teaching a viral vector that is specific for transgene expression in liver and splenic phagocytes. Zhang was cited for teaching that during cancer development and treatment, such as in hepatocellular carcinoma (HCC), tumors are infiltrated by tumor-associated macrophages (TAMs), which are recruited in the monocyte precursor form from the peripheral blood and stimulate tumor progression by facilitating angiogenesis, invasion, and metastasis, and suppressing antitumor immunity [column 2 ¶ 2]. Zhang further teaches that TAMs are typically non-inflammatory M2-type macrophages which specifically express CD206/MRC1 [column 3 ¶ 2]. Zhang also teaches targeting CD206/MCR1-expressing TAMs to treat tumors resistant to conventional therapy, wherein targeting the TAMs inhibited tumor growth and lung metastasis [abstract, column 3 ¶ 4]. As such, Zhang teaches the importance of TAMs for tumor targeting.
Although Zhang does not teach to only target the liver and spleen, Zhang does teach motivation for targeting the liver by teaching a need to treat advanced HCC, among other forms of cancer [col 1 ¶ 2- col 2 ¶ 1].
Note that the instant claims do not require that the macrophage targeting preserve the presence of the macrophages within the tumor, but merely that the vector is able to specifically express a gene within the macrophages. Therefore, teaching to deplete the TAMs is not a teaching away from the instant invention.
Further, Zhang teaches wherein the anti-CD206 antibody primarily stains the liver, spleen, kidney, stomach, and tumor with only minimal (<5%) staining in other organs [Figure 3]. As such, as discussed above for Kambara, the fact that Zhang teaches targeting organs in addition to the liver or spleen does not indicate a teaching away from specifically targeting the liver and spleen.
Regarding Applicant’s argument 4), Zhu was not cited for teaching viral vectors for transgene expression in liver and splenic phagocytes, but for teaching the motivation to target liver macrophages in HCC. Zhu was specifically cited for teaching the targeting of macrophages in the liver, such that CD206/MRC1 is highly expressed in human hepatocellular carcinoma (HCC) tissues compared to the level in peri-carcinoma tissue, and that targeting macrophages suppresses HCC progression by downregulating the expression of CD206 in M2 TAMs, thereby reducing the invasiveness of HCC cells [abstract, column 2 ¶ 2, column 14 ¶ 1, column 15 ¶ 1, column 19 ¶ 2]. Zhu also teaches that the reduction of invasiveness is possibly by affecting the secretion of cytokines from the M2 macrophages [column 19 ¶ 2]. The fact that Zhu’s teaching regarding the mechanism for targeting macrophages to reduce invasiveness utilizes the word “possibly” does not change the fact that Zhu teaches the idea of reducing invasiveness by targeting macrophages to reduce the secretion of cytokines. Additionally, the result on a tumor of targeting the liver macrophages for transgene expression is not an instantly claimed limitation either structurally or functionally.
Regarding Applicant’s argument 5), as discussed above, Geisler was cited for teaching the motivation to include miRNA target sequences in a transgene construct for de-targeting expression in non-target cells. In particular, Geisler was cited for teaching that miRNA-dependent post-transcriptional suppression of transgene expression has been emerging as powerful new technology to increase the specificity of vector-mediated transgene expression [column 2 ¶ 1].
Applicant asserts that Geisler does not explicitly teach the specific cell types of liver and spleen macrophages to which expression is limited. However, Geisler does teach the specific targeting of HCC cells accompanied by the de-targeting of hepatocytes as well as targeting tumor-infiltrating macrophages accompanied by de-targeting of HSPCs [Table 1].
Geisler was cited for teaching the subtractive component of the liver macrophage targeting, wherein expression is prevented in non-target cells which otherwise would be capable of expressing the cell-type specific promoter and being transduced by the tissue-selected viral vector.
Additionally, Geisler teaches the use of lentiviral vectors for delivering gene therapy constructs comprising miRNA targeting sequences for de-targeting, wherein the targeting strategy was first shown for lentiviral vectors in antigen presenting cells[column 12 ¶ 1]. Geisler was also cited for teaching that the targeting strategy has now been used for tissue-specific expression of vector-encoded therapeutic transgenes to reduce immune response against the transgene, to control virus tropism for oncolytic virotherapy, to increase safety of live attenuated virus vaccines, and to identify and select cell subsets for pluripotent stem cell therapies [column 12 ¶ 1]. Accordingly, Geisler specifically teaches the use of a viral vector to deliver a transgene to a cell type of interest, wherein the transgene construct comprises miRNA target sequences to ensure cell-type specific expression.
Additionally, Geisler was cited for teaching each of miR-122 and miR-126 for promoting cell-type specific expression wherein miR-122 represents the best candidate for a specific hepatocyte-de-targeting [column 6 ¶ 2, Table 1] and wherein miR-126 has been successfully used to de-target lentivirally delivered Interferon-α (IFN-α) away from hematopoietic stem and progenitor cells to target tumor-infiltrating macrophages, thereby inhibiting breast cancer progression [column 16 ¶ 2-column 17 ¶ 1, Table 1].
Regarding Applicant’s argument 6), De Palma was cited for teaching to modify tumor-infiltrating monocytes to secrete IFN-α to selectively deliver the IFN-α to tumors, wherein delivering tumor-homing proangiogenic, tumor-infiltrating Tie2-expressing monocytes (TEMs) genetically modified to express IFN-α achieved substantial antitumor activity in both prevention and intervention trials without measurable toxicity [abstract, column 1 ¶ 1- column 2 ¶ 2, column 3 ¶ 2, SIGNIFICANCE box]. De Palma further teaches that their experiments illustrate the therapeutic potential of gene- and cell-based IFN-α delivery, for example via tumor-infiltrating monocytes/macrophages, and should allow the development of IFN treatments that more effectively treat cancer [abstract, pg 301 col 2 ¶ 1]. De Palma was also cited for teaching the delivery of the IFN-α transgene to the TEMs in a lentiviral vector (LVs) regulated by the phagocyte-specific Tie2 promoter/enhancer [column 3 ¶ 2, column 20 ¶ 4]. De Palma further teaches that by intravenous injection, LVs preferentially transduce the liver and spleen [column 8 ¶ 2].
Applicant asserts that De Palma teaches away from the instant invention by teaching ex vivo transduction of cells and by suggesting that macrophage progenitors should be modified ex vivo and re-introduced for treatment. However, teachings by De Palma to treat ex vivo is not necessarily a teaching away from in vivo treatment at large. De Palma specifically treats mice with intravenous injection of lentivirus and observes the preferential transduction in the liver and spleen. De Palma was specifically looking for effects on glioma, such that transduction of the liver and spleen was not desirable. However, an ordinarily skilled artisan at the time of filing would understand that transduction of the liver and spleen would be desirable when targeting liver and spleen cells.
Regarding Applicant’s argument 7), Oda was cited for teaching wherein miR-122 and miR-126 are expressed within hepatocytes and LSECs [Figure 2], such that an ordinarily skilled artisan would understand than inserting an expression construct comprising miR-122 and/or miR-126 targeting sequences into hepatocytes and/or LSECs would result in suppression of expression of the mRNAs expressed therefrom.
Therefore, Applicant’s arguments do not overcome a finding of obviousness under 35 U.S.C. 103 over He, Kambara, Zhang, Zhu, Geisler, Oda, De Palma, Merlin, and Laffont.
Conclusion
No claim is allowed.
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
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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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