DETAILED ACTION
Notice of Pre-AIA or AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Continued Examination Under 37 CFR 1.114
A request for continued examination under 37 CFR 1.114, including the fee set forth in 37 CFR 1.17(e), was filed in this application after final rejection. Since this application is eligible for continued examination under 37 CFR 1.114, and the fee set forth in 37 CFR 1.17(e) has been timely paid, the finality of the previous Office action has been withdrawn pursuant to 37 CFR 1.114. Applicant's submission filed on May 26, 2026 has been entered.
Application Status
Applicant’s remarks and amendments to the claims filed May 26, 2026 are acknowledged. Claims 1, 3-7, 10-18, 23, and 25 were cancelled, claims 19-20, 26, and 33 were amended, and claims 36-39 were introduced. Claims 19-22, 24, 26, and 30-39 are pending.
A record of the correspondence between Applicant and Examiner subsequent to the Interview held on April 28, 2026 appears in Appendix I.
Restriction/Election
Claims 30-35 remain withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention. Claims 19-22, 24, 26, and 36-39 are under examination herein.
Priority
Applicant’s priority claims to foreign application CN201910322504.8 and PCT/CN2020/085366 are acknowledged. Claims 19-22, 24, 26, and 36-39 find support in CN201910322504.8, filed April 19, 2019. The effective filing date of all claims currently under examination is April 19, 2019.
Withdrawn Rejections
Applicant’s amendments to the claims are sufficient to overcome the § 102 rejections raised in the prior action over Nakashima. Applicant’s amendments to claims 19 and 20, to require an AAV vector or set of AAV vectors, wherein the tyrosine hydroxylase variant and aromatic L-amino acid decarboxylase are the only “exogenous target genes” in the vector or set of vectors, are sufficient to overcome the § 103 rejections raised over Azzouz, NM_199292.2, Palfi, Nakashima, and Ota as presented in the previous action, and the aforementioned references as evidenced by M76180 and/or in further view of G-Biosciences, Li, or Gray. These rejections are withdrawn, accordingly.
Applicant’s remarks and amendments have been thoroughly reviewed, but are not persuasive to place the claims in condition for allowance for the reasons that follow. Any rejection or objection not reiterated herein has been overcome by amendment.
Claim Rejections - 35 USC § 112(b)
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claims 19-22, 24, 26, and 38-39 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. The rejections that follow are new.
Claims 19 and 20 recite “a tyrosine hydroxylase variant comprising the amino acid sequence set forth in SEQ ID NO: 1, wherein the tyrosine hydroxylase comprises an N-terminal deletion of 90 amino acid residues from SEQ ID NO: 1.” It is not clear how the tyrosine hydroxylase variant can comprise “the amino acid sequence set forth in SEQ ID NO: 1,” i.e., each consecutive residue of SEQ ID NO: 1, and simultaneously comprise 90 fewer N-terminal amino acid residues from SEQ ID NO: 1. It is also not clear whether the phrase “comprises an N-terminal deletion of 90 amino acid residues from SEQ ID NO: 1,” should be interpreted as requiring deletion of exactly 90 N-terminal residues from SEQ ID NO: 1, or alternatively, whether the term “comprises” should be interpreted as allowing for deletion of 90 or more N-terminal residues from SEQ ID NO: 1. Because the scope of tyrosine hydroxylase variants encompassed by the claims are unclear, the structures of the vectors encoding the tyrosine hydroxylase variants are also unclear.
Claims 21-22, 24, 26, and 38-39 are rejected for depending from claims 19 or 20 and failing to remedy the indefiniteness. In the interest of compact prosecution, the claims will be interpreted as requiring a tyrosine hydroxylase variant consisting from N- to C-terminus of the amino acid methionine and amino acids 1-437 of SEQ ID NO: 2 (i.e., amino acids 92-528 of SEQ ID NO: 1), as it is believed that this is Applicant’s preferred embodiment based on the specification and remarks ([0073; [00169]).
Claims 19 and 20 require that “the exogenous target genes encoding the tyrosine hydroxylase variant and the aromatic L-amino acid decarboxylase are the only exogenous target genes in the vector” or “set of vectors.” The term “exogenous target gene” is not explicitly defined in the claim or specification, and neither the claim nor specification make clear what the gene must be “exogenous” relative to, e.g., an AAV vector, a particular cell or organism, etc., or what would satisfies the term “target” given that what would be considered a “target” to one artisan, may not be a “target” to another artisan.
For example, the prior art teaches AAV vectors encoding “GTP cyclohydrolase I (GCH),”, a gene involved in dopamine synthesis (see Figs. 1-2 of Shen et al., 2000, Human Gene Therapy, 11:2509-1519). The specification describes that AAV vectors comprise heterologous sequences, which would be interpreted as sequences derived from a different origin than the AAV vector ([00112]). It is not clear whether the CH1 gene should be considered “exogenous” because I) it is part of a known AAV vector, and therefore, would not necessarily be considered “exogenous” thereto, and because II) it appears to fall within the scope of elements comprised by AAV vectors based on the specification (i.e., heterologous sequences). Furthermore, the skilled artisan may not consider GCH a “target” in particular contexts, e.g., when assessing the consequence of adding further dopamine synthesis genes to an existing AAV vector. The skilled artisan would not be reasonably apprised of the genes which would be considered “exogenous target[s],” and therefore, would not be able to ascertain the structures of the vector or set of vectors which fall within or outside the claim scope.
Claims 21-22, 24, 26, and 38-39 are rejected for depending from claims 19 or 20 and failing to remedy the indefiniteness. In the interest of compact prosecution, the aforementioned phrase will be interpreted as excluding any additional genes involved in dopamine synthesis, as it is believed that this is Applicant’s intention based on the remarks.
Claim Rejections - 35 USC § 101
35 U.S.C. 101 reads as follows:
Whoever invents or discovers any new and useful process, machine, manufacture, or composition of matter, or any new and useful improvement thereof, may obtain a patent therefor, subject to the conditions and requirements of this title.
Claims 36-37 are rejected under 35 U.S.C. 101 because the claimed inventions are directed to a law of nature or natural phenomena without significantly more. The claims recites a “tyrosine hydroxylase variant with an amino acid sequence as set forth in SEQ ID NO: 2,” or “as set forth in SEQ ID NO: 3,” which is interpreted as a tyrosine hydroxylase comprising a sequence difference from any other tyrosine hydroxylase (e.g., an amino acid deletion, substitution, addition, etc.), wherein the variant comprises any two or more consecutive amino acid residues set forth in SEQ ID NO: 2 or SEQ ID NO: 3.
Laws of nature and natural phenomena, as identified by the courts, include naturally occurring principles/relations and nature-based products that are naturally occurring or that do not have markedly different characteristics compared to what occurs in nature. UniProt (“P07101, TY3H_HUMAN,” https://www.uniprot.org/uniprotkb/P07101/entry, accessed 2 June 2026) teaches a naturally-occurring tyrosine hydroxylase variant (“VAR_072862”) comprising an amino acid sequence set forth in each of SEQ ID NO: 2 and SEQ ID NO: 3 (“Disease & Variants,” “Features,” “Natural variant,” “VAR_072862,” “Position 19,” “Sequence: S [Wingdings font/0xE0] C,” pg. 6-7). See attached alignments in Appendix II. The tyrosine hydroxylase variants of claims 36-37 are, therefore, not markedly different from their naturally-occurring counterpart. Claims 36-37 recites a nature-based product.
The claims do not recite any additional elements. Thus, the judicial exception is not integrated into a practical application and does not include additional elements that are sufficient to amount to significantly more than the judicial exception.
Claim Rejections - 35 USC § 102 - Nakashima
The following is a quotation of the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
Claims 36-37 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Nakashima (Nakashima et al., 2005, Journal of Neuroscience Research, 81: 110-120; of record). The rejections that follow are new and necessitated by Applicant’s amendments to the claims.
As described above, the term “an amino acid sequence” is interpreted as any two or more consecutive amino acids. Claims 36-37, therefore, encompass tyrosine hydroxylase variants comprising any two or more consecutive amino acids set forth in SEQ ID NO: 2 or 3.
Nakashima teaches a tyrosine hydroxylase variant ("del-52"), which as shown in the alignment of record, comprises 100% identity to amino acids 84-528 of SEQ ID NO: 1 (see Appendix II dated February 3, 2025). Residues 84-528 of instant SEQ ID NO: 1 comprise two or more consecutive amino acids set forth in each of SEQ ID NOs: 2 and 3. Nakashima’s tyrosine hydroxylase variant meets the limitations of instant claims 36-37, accordingly,
Notice to Joint Inventors
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claim Rejections - 35 USC § 103 – During in view of Nakashima, Ota, and Palfi
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
Claims 19-22, 24, 26, and 36-39are rejected under 35 U.S.C. 103 as being unpatentable over During (During et al., 1998, Gene Therapy, (1998) 5, pg. 820-827), in view of Nakashima (Nakashima et al., 2005, Journal of Neuroscience Research, 81: 110-120; of record), Ota (Ota et al., 1995, Biochemical and Biophysical Research Communications, Vol. 213, No. 3, pg. 1099-1106; cited in IDS filed 13 April 2022; of record) and Palfi (Palfi et al., 2018, Human Gene Therapy Clinical Development, Vol. 29, No. 3, pg. 148-155; of record). The rejections that follow are new and necessitated by Applicant’s amendments to the claims.
Regarding claim 19, During teaches an AAV vector (“pAAVFLAGTH-IRES-hAADC”) comprising a polynucleotide construct, wherein the polynucleotide construct comprises a first polynucleotide encoding an N-terminally deleted human tyrosine hydroxylase variant (“truncated hTH”), and a second polynucleotide encoding a human aromatic L-amino acid decarboxylase (“hAADC”)(“(a) Diagram of the bicistronic AAVthIRESaadc plasmid… N-terminally deleted human TH (form II) cDNA together with the IRES… sequence… and a full-length human AADC cDNA,” Fig. 1 and description; “Preparation of viral vectors,” pg. 825). As shown in Fig. 1a, the AAV vector does not comprise any additional genes involved in dopamine synthesis.
During demonstrates that the vector is sufficient to produce dopamine in transfected cells in vitro (“the biscistronic construct released both L-Dopa… and dopamine… into the medium… numerous FLAG-IR cells were detected,” pg. 821). During also demonstrates that the vector is sufficient for use in gene transfer and to increase dopamine production in primate models (“One monkey in this group demonstrated a greater than two-fold elevation of tissue dopamine concentration in a punch that could be visualized as in the vicinity of the injection site… Immunocytochemistry revealed the presence of FLAG-IR cells surrounding the injection sites,” pg. 821; “there was clear TH antibody labeling of cells near the injection tracts… immunocytochemistry appeared to show the FLAG epitope in areas near the injections in other MPTP-treated monkeys… two monkeys that received AAVflag∆thIRESaadc and also showed the most complete behavioral recovery, the tissue dopamine concentrations were elevated in the sites adjacent to the injection… evidence of sprouting of dopaminergic terminals,” pg. 822-823). During demonstrates that the vector elicits behavioral improvement in three severely parkinsonian monkeys (“All three remaining severely parkinsonian monkeys treated with AAVflag∆thIRESaadc showed evidence of behavioral improvement,” pg. 822).
During concludes that their study “shows the successful transfection of primate neurons over a period up to 2.5 months with suggestive evidence of biochemical phenotypic effects and without significant toxicity” (Abstract). During teaches that their study “support[s] the idea of an in vivo gene therapy for Parkinson’s disease,” but that “more consistent and longer lasting biochemical and behavioral effects will be necessary” to further establish the therapeutic approach (Abstract).
As described above, During’s vector comprises an N-terminally deleted human tyrosine hydroxylase variant. The skilled artisan would have understood that During intended to increase the production of dopamine by using the N-terminal deleted variant (“Previous studies have shown that the cleavage of the regulatory N-terminal of the TH enzyme results in approximately 2.5-fold greater enzymatic activity. We therefore constructed a vector with an N-terminal deletion,” pg. 821). However, During does not teach the sequence of the N-terminally deleted human tyrosine hydroxylase variant.
Nakashima and Ota, provide means and motivation to the skilled artisan to arrive at a tyrosine hydroxylase variant consisting from N- to C-terminus of the amino acid methionine and amino acids 1-437 of SEQ ID NO: 2 (i.e., amino acids 92-528 of SEQ ID NO: 1). Nakashima demonstrates that two N-terminally deleted tyrosine hydroxylase variants, del-52 and del-157 produce significantly more dopamine (“DA”) in mammalian cells (Fig. 1), despite having lower catalytic activity than wildtype tyrosine hydroxylase when measured as substrate conversion using protein prepared from cell lysates (Table 1; “Assay of Enzyme Activity and Measurement of Catecholamine Contents,” pg. 112, left col.). Specifically, Nakashima’s del-52 variant produces approximately 400% more dopamine than full-length tyrosine hydroxylase (“wild-type” - ~100, “del-52” = ~400, Fig. 1A), whereas Nakashima’s del-157 variant produces approximately 250% more dopamine than full-length tyrosine hydroxylase (“wild-type” - ~100, “del-157” = ~250, Fig. 1A). Nakashima attributes the increased levels of dopamine production in mammalian cells to the “high stability” of the deletion variants (see at least pg. 119, left col.; “Degradation Rate of His-hTH1 Proteins Within AtT-20 Cells,” pg. 114).
Ota teaches additional N-terminally deleted tyrosine hydroxylase variants to the del-52 and del-157 variants of Nakashima (Fig. 1). Ota teaches that enzyme activity “abruptly decreased along with the advance of the deletion of N-terminal and resulted in hardly estimated levels at del-77” through “del-109” (pg. 1101; Fig. 2). Ota attributes the reduced/absent enzyme activity in lysate supernatants of the del-74 through del-109 variants to “the inability of the proteins to fold properly during expression result[ing] in their precipitation into aggregates termed inclusion bodies” (pg. 1103). Ota teaches that “enzyme activity turned round toward increase at del-117 (pg. 1101; Fig. 2).
Based on Nakashima and Ota, the skilled artisan would understand that the N-terminal domain of tyrosine hydroxylase influences its activity, and therefore, dopamine production. The skilled artisan would also understand that particular N-terminal truncations would not be suitable for use in the method of During, i.e., del-74 through del-109, due to improper folding. The skilled artisan would have also recognized based on Ota, that deletions between del-37 and del-73, or del-131 and del-157 could be particularly efficacious for the method of During based on the results of Ota (Fig. 2), and Nakashima (Fig. 1).
Nakashima teaches the sequence of the tyrosine hydroxylase from which the N-terminal deletion variants were prepared (Fig. 5). Nakashima’s tyrosine hydroxylase sequence comprises 100% identity to residues 62-528 of SEQ ID NO: 1. It is noted that the numbering of Nakashima’s and Ota’s tyrosine hydroxylase sequences differ from that of instant SEQ ID NO: 1. Nakashima’s del-57 and del-157 correspond to tyrosine hydroxylase variants comprising residues 84-528 and 189-528 of SEQ ID NO: 1 (i.e., an N-terminal deletion of 83 and 188 amino acids from SEQ ID NO: 1, respectively), and Ota’s del-37, del-67, del-68, del-70, del-73, and del-131 correspond to tyrosine hydroxylase variants comprising residues 70-528, 100-528, 101-528, 103-528, 106-528, and 164-528 of SEQ ID NO: 1, respectively (i.e., N-terminal deletions of 69, 99, 102, 105, and 163 amino acids from SEQ ID NO: 1, respectively). See attached alignment in Appendix III.
Finally, Palfi establishes that as of the effective filing date of the claimed invention, gene therapy approaches similar to During’s (i.e., a viral vector approach delivering at least tyrosine hydroxylase and AADC), still struggled to achieve consistent and long lasting biochemical and behavioral effects in the treatment of Parkinson’s disease, owing to suboptimal levels of dopamine replacement (“[a]lthough the results are encouraging, the data suggest that the optimal level of dopamine replacement may not have been achieved, since patients continued to require L-Dopa therapy to obtain maximal benefit, and some of the more severely affected patients required DBS 2-6 years following ProSavin administration,” pg. 154, right col.).
It would have been obvious to one of ordinary skill before the effective filing date of the claimed invention to have prepared the vector of During to encode a tyrosine hydroxylase variant consisting from N- to C-terminus of the amino acid methionine and amino acids 1-437 of SEQ ID NO: 2 (i.e., amino acids 92-528 of SEQ ID NO: 1) in view of Nakashima and Ota. It would have amounted to applying a known technique (N-terminal deletion) to a known tyrosine hydroxylase sequence, by known means to yield predictable results, and including a resulting variant in a known vector. Based on the prior art, gene therapy approaches delivering tyrosine hydroxylase and AADC, while promising for treatment of Parkinson’s disease, struggled to achieve levels of dopamine replacement necessary to produce consistent and long lasting effects. Based on the teachings of Nakashima and Ota, the skilled artisan would have recognized that the levels of dopamine produced using During’s vector could be increased by preparing the vector with one of range of N-terminally deleted tyrosine hydroxylase variants.
Based on the prior art’s characterization of the N-terminal region of tyrosine hydroxylase, there were 63 predictable, potential solutions (i.e., del-37, del-39, del-39… del-73, and del-131, del-132, del-133… del-157). Among these solutions is a tyrosine hydroxylase variant consisting from N- to C-terminus of the amino acid methionine and amino acids 1-437 of SEQ ID NO: 2 (i.e., amino acids 92-528 of SEQ ID NO: 1). The skilled artisan would reasonably conclude that a tyrosine hydroxylase variant with an N-terminal deletion within the range of del-37 and del-73, or del-131 and del-157, would produce significantly more dopamine in mammalian cells than wildtype tyrosine hydroxylase, while retaining enzyme activity. Indeed, each of the variants characterized by Ota within the ranges are active, and Nakashima demonstrates that the del-52 and del-157 variants produce approximately 250% and 400% more dopamine in mammalian cells, respectively, than wildtype tyrosine hydroxylase.
As evidenced by the prior art cited herein, it was well within the purview of the skilled artisan to prepare N-terminal deletion variants of a known sequence, encode such variants in vectors, and examine their effects in vitro and in vivo. The skilled artisan would have been motivated to prepare the vector of During with a N-terminally deleted tyrosine hydroxylase variant within the ranges characterized by Nakashima and Ota because During is silent as to the sequence of the N-terminally deleted variant in the vector, and because Nakashima and Ota provide evidence that the skilled artisan could improve outcomes in therapeutic methods of using the vector if the optimal N-terminally deleted tyrosine hydroxylase variant were delivered.
Regarding claim 20, During teaches a set of AAV vectors in solution (“Fifteen microliters of the AAV vector (titer 107/ml transducing units) were injected through a 22-gauge needle into each of the three sites in the monkey caudate…,” pg. 826). As shown in Fig. 1a, the set of vectors would comprise an AAV vector comprising a first polynucleotide comprising a gene encoding a tyrosine hydroxylase variant, and an AAV vector comprising a second polynucleotide comprising a gene encoding an aromatic L-amino acid decarboxylase. As shown in Fig. 1a, the set of AAV vectors does not comprise any additional genes involved in dopamine synthesis. The obviousness of preparing an AAV vector of During to encode a tyrosine hydroxylase variant consisting from N- to C-terminus of the amino acid methionine and amino acids 1-437 of SEQ ID NO: 2 (i.e., amino acids 92-528 of SEQ ID NO: 1) is described above and applied here.
Regarding claims 21-22, as shown in Fig. 1a of During, the vector also comprises a third polynucleotide inserted between the first and second polynucleotide, wherein the third polynucleotide encodes an internal ribosome entry site (IRES)(Fig. 1).
Regarding claims 24 and 38, During teaches an in vitro host cell comprising or transfected by the vector or set of vectors (see “Characterization of the vector in vitro,” pg. 821).
Regarding claims 26 and 39, During teaches a pharmaceutical composition comprising the vector or set of vectors and a pharmaceutically acceptable carrier (“Fifteen microliters of the AAV vector (titer 107/ml transducing units) were injected through a 22-gauge needle into each of the three sites in the monkey caudate…,” “Infusion of vector into striatum,” pg. 826).
Regarding claims 36-37, the claims encompass a tyrosine hydroxylase variant comprising any two or more consecutive amino acids set forth in SEQ ID NOs: 2 or 3. The rejection above renders obvious a tyrosine hydroxylase variant consisting from N- to C-terminus of the amino acid methionine and amino acids 1-437 of SEQ ID NO: 2 (i.e., amino acids 92-528 of SEQ ID NO: 1). SEQ ID NO: 3 comprises amino acids 1-437 of SEQ ID NO: 2. Therefore, the rejection above also renders obvious claims 36-37.
Claim Rejections - 35 USC § 103 – Fan in view of Nakashima, Ota, and Palfi
Claims 20, and 36-39 are rejected under 35 U.S.C. 103 as being unpatentable over Fan (Fan et al., 1998, Human Gene Therapy, 9:2527-2535), in view of Nakashima (Nakashima et al., 2005, Journal of Neuroscience Research, 81: 110-120; of record), Ota (Ota et al., 1995, Biochemical and Biophysical Research Communications, Vol. 213, No. 3, pg. 1099-1106; cited in IDS filed 13 April 2022; of record) and Palfi (Palfi et al., 2018, Human Gene Therapy Clinical Development, Vol. 29, No. 3, pg. 148-155; of record). The rejections that follow are new and necessitated by Applicant’s amendments to the claims.
The rejections that follow address an embodiment encompassed by claim 20, wherein a first AAV vector comprises a gene encoding a tyrosine hydroxylase variant and no other genes involved in dopamine synthesis, and a second vector comprises a gene encoding an aromatic L-amino acid decarboxylase and no other genes involved in dopamine synthesis.
Regarding claim 20, Fan teaches a set of AAV vectors, wherein a first vector comprises a first polynucleotide encoding a human tyrosine hydroxylase variant, and a second vector comprises a second polynucleotide encoding a human aromatic L-amino acid decarboxylase (“we coexpressed TH and AADC in striatal cells using two separate AAV vectors,” pg. 2528; “1.8-kb human TH (type 1) and 1.9-kb human AADC cDNAs,” pg. 2528). The set of vectors do not comprise any additional genes involved in dopamine synthesis (see “Plasmids and a 293 cell line,” pg. 2528).
Fan demonstrates that the vectors are sufficient to produce dopamine in transfected cells in vitro (“A significant amount of dopamine was produced subsequent to transduction with both vectors,” pg. 2530; “cultures cotransduced with the AAV-TH/AAV-AADC vector mixture (ratio 1:1) showed a significant increase in dopamine level,” pg. 2532; Table 1). Fan also demonstrates that the vector is sufficient for use in gene transfer and to increase dopamine production in rodent models (“the expression of TH and AADC in the striatum was examined… To confirm the behavioral recovery following AAV-TH/AAV-AADC cotransduction was caused by enhanced dopamine production in vivo…,” pg. 2532; Fig. 5). Fan demonstrates that the vector elicits behavioral improvement in rodent models of Parkinson’s disease (“significant decreases… in the rotational rate were observed in both groups of rats treated with AAV-TH/AAV-AADC mixture… the decrease was more remarkable in rats injected with the… mixture,” pg. 2532; Fig. 4).
The skilled artisan would understand that Fan is concerned with using a gene therapy approach to increase the production of dopamine in vivo, to remedy dopamine loss causal to Parkinson’s disease (Abstract). Fan does not teach that the first AAV vector encodes an N-terminally deleted human tyrosine hydroxylase variant consisting from N- to C-terminus of the amino acid methionine and amino acids 1-437 of SEQ ID NO: 2 (i.e., amino acids 92-528 of SEQ ID NO: 1).
Nakashima and Ota, provide means and motivation to the skilled artisan to arrive at a tyrosine hydroxylase variant consisting from N- to C-terminus of the amino acid methionine and amino acids 1-437 of SEQ ID NO: 2 (i.e., amino acids 92-528 of SEQ ID NO: 1). Nakashima demonstrates that two N-terminally deleted tyrosine hydroxylase variants, del-52 and del-157 produce significantly more dopamine (“DA”) in mammalian cells (Fig. 1), despite having lower catalytic activity than wildtype tyrosine hydroxylase when measured as substrate conversion using protein prepared from cell lysates (Table 1; “Assay of Enzyme Activity and Measurement of Catecholamine Contents,” pg. 112, left col.). Specifically, Nakashima’s del-52 variant produces approximately 400% more dopamine than full-length tyrosine hydroxylase (“wild-type” - ~100, “del-52” = ~400, Fig. 1A), whereas Nakashima’s del-157 variant produces approximately 250% more dopamine than full-length tyrosine hydroxylase (“wild-type” - ~100, “del-157” = ~250, Fig. 1A). Nakashima attributes the increased levels of dopamine production in mammalian cells to the “high stability” of the deletion variants (see at least pg. 119, left col.; “Degradation Rate of His-hTH1 Proteins Within AtT-20 Cells,” pg. 114).
Ota teaches additional N-terminally deleted tyrosine hydroxylase variants to the del-52 and del-157 variants of Nakashima (Fig. 1). Ota teaches that enzyme activity “abruptly decreased along with the advance of the deletion of N-terminal and resulted in hardly estimated levels at del-77” through “del-109” (pg. 1101; Fig. 2). Ota attributes the reduced/absent enzyme activity in lysate supernatants of the del-74 through del-109 variants to “the inability of the proteins to fold properly during expression result[ing] in their precipitation into aggregates termed inclusion bodies” (pg. 1103). Ota teaches that “enzyme activity turned round toward increase at del-117 (pg. 1101; Fig. 2).
Based on Nakashima and Ota, the skilled artisan would understand that the N-terminal domain of tyrosine hydroxylase influences its activity, and therefore, dopamine production. The skilled artisan would also understand that particular N-terminal truncations would not be suitable for use in the method of Fan, i.e., del-74 through del-109, due to improper folding. The skilled artisan would have also recognized based on Ota, that deletions between del-37 and del-73, or del-131 and del-157 could be particularly efficacious for the method of Fan based on the results of Ota (Fig. 2), and Nakashima (Fig. 1).
Nakashima teaches the sequence of the tyrosine hydroxylase from which the N-terminal deletion variants were prepared (Fig. 5). Nakashima’s tyrosine hydroxylase sequence comprises 100% identity to residues 62-528 of SEQ ID NO: 1. It is noted that the numbering of Nakashima’s and Ota’s tyrosine hydroxylase sequences differ from that of instant SEQ ID NO: 1. Nakashima’s del-57 and del-157 correspond to tyrosine hydroxylase variants comprising residues 84-528 and 189-528 of SEQ ID NO: 1 (i.e., an N-terminal deletion of 83 and 188 amino acids from SEQ ID NO: 1, respectively), and Ota’s del-37, del-67, del-68, del-70, del-73, and del-131 correspond to tyrosine hydroxylase variants comprising residues 70-528, 100-528, 101-528, 103-528, 106-528, and 164-528 of SEQ ID NO: 1, respectively (i.e., N-terminal deletions of 69, 99, 102, 105, and 163 amino acids from SEQ ID NO: 1, respectively). See attached alignment in Appendix III.
Finally, Palfi establishes that as of the effective filing date of the claimed invention, gene therapy approaches similar to Fan’s (i.e., a viral vector approach delivering at least tyrosine hydroxylase and AADC), still struggled to achieve consistent and long lasting biochemical and behavioral effects in the treatment of Parkinson’s disease, owing to suboptimal levels of dopamine replacement (“[a]lthough the results are encouraging, the data suggest that the optimal level of dopamine replacement may not have been achieved, since patients continued to require L-Dopa therapy to obtain maximal benefit, and some of the more severely affected patients required DBS 2-6 years following ProSavin administration,” pg. 154, right col.).
It would have been obvious to one of ordinary skill before the effective filing date of the claimed invention to have prepared the first AAV vector of Fan to encode a tyrosine hydroxylase variant consisting from N- to C-terminus of the amino acid methionine and amino acids 1-437 of SEQ ID NO: 2 (i.e., amino acids 92-528 of SEQ ID NO: 1) in view of Nakashima and Ota. It would have amounted to applying a known technique (N-terminal deletion) to a known tyrosine hydroxylase sequence, by known means to yield predictable results, and including a resulting variant in a known vector. Based on the prior art, gene therapy approaches delivering tyrosine hydroxylase and AADC, while promising for treatment of Parkinson’s disease, struggled to achieve levels of dopamine replacement necessary to produce consistent and long lasting effects. Based on the teachings of Nakashima and Ota, the skilled artisan would have recognized that the levels of dopamine produced using Fan’s vector could be increased by preparing the vector with one of range of N-terminally deleted tyrosine hydroxylase variants.
Based on the prior art’s characterization of the N-terminal region of tyrosine hydroxylase, there were 63 predictable, potential solutions (i.e., del-37, del-39, del-39… del-73, and del-131, del-132, del-133… del-157). Among these solutions is a tyrosine hydroxylase variant consisting from N- to C-terminus of the amino acid methionine and amino acids 1-437 of SEQ ID NO: 2 (i.e., amino acids 92-528 of SEQ ID NO: 1). The skilled artisan would reasonably conclude that a tyrosine hydroxylase variant with an N-terminal deletion within the range of del-37 and del-73, or del-131 and del-157, would produce significantly more dopamine in mammalian cells than wildtype tyrosine hydroxylase, while retaining enzyme activity. Indeed, each of the variants characterized by Ota within the ranges are active, and Nakashima demonstrates that the del-52 and del-157 variants produce approximately 250% and 400% more dopamine in mammalian cells, respectively, than wildtype tyrosine hydroxylase.
As evidenced by the prior art cited herein, it was well within the purview of the skilled artisan to prepare N-terminal deletion variants of a known sequence, encode such variants in vectors, and examine their effects in vitro and in vivo. The skilled artisan would have been motivated to prepare the vector of Fan with a N-terminally deleted tyrosine hydroxylase variant within the ranges characterized by Nakashima and Ota, because because Nakashima and Ota provide evidence that the skilled artisan could improve outcomes in therapeutic methods of using the vector if an N-terminally deleted tyrosine hydroxylase variant were delivered.
Regarding claims 36-37, the claims encompass a tyrosine hydroxylase variant comprising any two or more consecutive amino acids set forth in SEQ ID NOs: 2 or 3. The rejection above renders obvious a tyrosine hydroxylase variant consisting from N- to C-terminus of the amino acid methionine and amino acids 1-437 of SEQ ID NO: 2 (i.e., amino acids 92-528 of SEQ ID NO: 1). SEQ ID NO: 3 comprises amino acids 1-437 of SEQ ID NO: 2. Therefore, the rejection above also renders obvious claims 36-37.
Regarding claim 38, Fan teaches an in vitro host cell comprising or transfected by the set of vectors (see “AAV vector-mediated coexpression of TH and AADC in vitro,” pg. 2530-2532).
Regarding claim 39, Fan teaches a pharmaceutical composition comprising the set of vectors and a pharmaceutically acceptable carrier (“AAV vector solution was injected slowly… Each injection was 2 ul (5 X 108 vector particles per microliter…),” pg. 2529).
Response to Remarks - 35 USC § 103
Applicant’s remarks regarding the § 103 rejections raised in the prior action have been reviewed. As described above, the § 103 rejections raised in the prior action have been withdrawn in view of Applicant’s amendments to the claims. The rejections above are new, and necessitated by Applicant’s amendments to the claims. The remarks relevant to the new rejections above are addressed hereinafter.
Applicant submits that the there was “no reasonable expectation that the TH/AADC AAV vector… would be successful for delivering the TH/AADC and treating PD.” Applicant alleges that the AAV vectors were “not considered an option,” and lentiviral vectors were preferred, owing at least to their packaging capacity. Applicant also alleges that there was no reasonable expectation that omitting CH1, would result in an AAV vector effective at treating PD.
These remarks have been considered, but are not found persuasive. First, the instant claims are directed to vectors, not methods of treatment; the instantly claimed vectors must be suitable to function as a vector (i.e., as a substance which carries genetic material into a cell/organism), but do not require any therapeutic effects. There is no evidence that the obvious AAV vector(s) would be unsuitable for carrying genetic material into a cell/organism. Furthermore, the prior art cited above describes AAV vector(s) encoding TH and AADC, and provides evidence that the vector(s) are functional for delivering TH and AADC to cells in vitro and in vivo, and for their intended use of treating PD. The prior art cited above with respect to N-terminally deleted TH variants also provides substantial evidence that a vector comprising the instantly claimed N-terminally deleted TH variant would function for its intended use. Given the prior art cited above, the Examiner does not need to render obvious the substitution of one vector type for another, or the omission of any other dopamine synthesis enzymes. Beyond argument, Applicant has provided no evidence that there was no reasonable expectation of success.
Applicant also submits that “it would not be obvious to modify the known, full-length TH sequence to obtain a sequence with a 90-aa N-terminal deletion as instantly claimed.” Applicant states that it would not be reasonably expected that “Nakashima’s and Ota’s constructs would produce the same results in vivo.” Applicant submits that the “range of N-terminal deletions relative to the full-length sequence disclosed in the cited references in very broad” and that it would not have been obvious to arrive at the specific 90-aa N-terminal deletion.
These remarks have been considered, but are not found persuasive. As stated above, the instant claims are directed to vectors, not methods of treatment or expressing genes in a particular context; the instantly claimed vectors must be suitable to function as a vector (i.e., as a substance which carries genetic material into a cell/organism), but do not require any therapeutic effects. There is no evidence that the obvious AAV vector(s) would be unsuitable for carrying genetic material into a cell/organism. The obviousness rejection above meets each requirement set forth in MPEP 2143(I)(E). The number of possible solutions is “small or easily traversed” based on the context of the art in question which shows that preparing and characterizing many N-terminally deleted tyrosine hydroxylase variants was routine and within the purview of the skilled artisan (see for example, Ota). A 90-aa N-terminal deletion is within the small or easily traversed number of possible solutions. Given the context of the art (i.e., the data provided by the prior art with respect to the activity of N-terminally deleted TH variants), the skilled artisan would have had a reasonable expectation of success. There is no requirement that a specific solution amongst the possible solutions be rendered obvious under the applied rationale.
Finally, Applicant alleges that the claimed vectors produce “unexpected results.” Applicant states that the mice receiving “the instantly claimed TH90del/AADC AAV vector showed significantly decreased net turns per minute of contralateral rotation in apomorphine-induced motor asymmetry tests.” Applicant alleges that in contrast to Azzouz et al., the instantly claimed vectors completely eliminate rotational behavior after 4 weeks of treatment. Examiner has considered Applicant’s evidence of alleged unexpected results with respect to the instant claims. Applicant’s evidence appears to correspond to a specific AAV vector comprising a specific N-terminally deleted TH sequence and a specific AADC sequence (see Examples 1 and 3, pgs. 33-34, and 37-38). Applicant’s evidence does not appear to correspond to any “set of AAV vectors” as recited in claim 20. Applicant’s evidence also does not appear to correspond to the generic AAV vector recited in claim 19. There is also insufficient evidence that the skilled artisan “could ascertain a trend in the exemplified data… to reasonably extend the probative value thereof,” i.e., to any AAV vector comprising any regulatory sequences, promoters, etc., comprising any AADC, and a TH variant with any number of modifications provided it comprises an N-terminal deletion of 90 amino acids from SEQ ID NO: 1. Applicant’s evidence of alleged unexpected results is at least not commensurate with the scope of the claims.
Conclusion
No claims are allowed.
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/JENNA L PERSONS/Examiner, Art Unit 1637
/Jennifer Dunston/Supervisory Patent Examiner, Art Unit 1637
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