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 .
Application Status
Applicant’s remarks and amendments to the claims filed November 17, 2025 are acknowledged. Claims 1, 10, 13, 20-21, and 25 were amended. Claims 1, 3-7, 10-26, and 30-35 are pending.
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Claims 30-35 remain withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention.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 1, 3-7, and 10-26 are under examination herein.
Priority
Applicant’s priority claims to foreign application CN201910322504.8 and PCT/CN2020/085366 are acknowledged. Claims 1, 3-7, and 10-26 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Withdrawn Rejections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 § 112(b) rejections raised in the prior action. These rejections are withdrawn, accordingly.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 § 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 1, and 3-5 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 maintained from the prior action with modification necessitated by Applicant’s amendments to the claims.
Claims 1 and 3 recite a tyrosine hydroxylase variant comprising “the amino acid sequence set forth in SEQ ID NO: 1,” wherein the tyrosine hydroxylase variant “comprises an N-terminal deletion… from SEQ ID NO: 1” of either “80 to 100” or “80 to 90” amino acid residues, respectively. The claims are interpreted as requiring a tyrosine hydroxylase variant lacking the N-terminal (i.e., first) I) 80, 81, 82,… 99, or 100 amino acids (claim 1) or II) 80, 81, 82,… 89, or 90 amino acids (claim 3) of SEQ ID NO: 1. For example, a tyrosine hydroxylase variant comprising amino acids 81-528 of SEQ ID NO: 1 is encompassed by both claims 1 and 3. On the other hand, a tyrosine hydroxylase variant comprising amino acids 79-528 of SEQ ID NO: 1 is excluded from both claims.
The claims are directed to a “pharmaceutical composition” comprising the tyrosine hydroxylase variant, an aromatic L-amino acid decarboxylase, and one, or a combination, of the recited pharmaceutically acceptable carriers, e.g., a “transfection facilitating agent,” which is interpreted as any agent which may be used for the purposes of transfection. The claim body fully sets forth all of the limitations of the claimed invention, such that the preamble term “pharmaceutical” merely states the purpose or intended use of the invention, i.e., that it be for pharmaceutical purposes. Accordingly, this term is considered a non-limiting intended use. See MPEP 2111.02(II).
Regarding claims 1 and 3, Nakashima teaches a composition (i.e., culture medium containing transfected AtT-20 cells) comprising a tyrosine hydroxylase variant ("del-52") and a pharmaceutically acceptable carrier, “FuGENE6 Reagent,” which is a “transfection facilitating agent” (“Cell Culture,” pg. 111-112; “Expression of hTH1 Enzymes in Cells”, pg. 112, left col.; “Accumulation of DA in AtT-20 Cells”, pg. 113, left col.; Fig. 1). Nakashima teaches the composition also comprises an aromatic L-amino acid decarboxylase (“The accumulation of DA and DOPAC within the AtT-20 cells reflects the ability of hTH1 to synthesize L-DOPA because aromatic L-amino acid decarboxylase (AADC) present in the cells converts L-DOPA to DA immediately”, pg. 113, left col.; Fig. 1).
It is noted that the reference sequence disclosed in Nakashima’s Fig. 5 is already missing ~30 amino acids from the N-terminus of SEQ ID NO: 1. Thus, Nakashima’s “del-52” variant is missing the first ~30 amino acids from the N-terminus of SEQ ID NO: 1, and an additional 52 amino acids (i.e., “del-52”), and thereby, reads on the instantly claimed tyrosine hydroxylase variant. Indeed, as evidenced by the alignment of record in Appendix II between instant SEQ ID NO: 1 and the sequence of the "del-52" variant in Fig. 5, Nakashima’s tyrosine hydroxylase variant comprises an N-terminal deletion of 83 amino acid residues from SEQ ID NO: 1 (Fig. 5, pg. 116; "Deletion mutations of the N-terminal amino acid residues of hTH1... The mutants missing the first 52 and 157 amino acid residues are referred to herein as del-52 and del-157, respectively", Material and Methods, pg. 111). As shown in the alignment of record, Nakashima’s tyrosine hydroxylase variant comprises 100% identity to amino acids 84-528 of SEQ ID NO: 1.
Regarding claim 4, as shown in the alignment of record, Nakashima’s tyrosine hydroxylase variant comprises 100% identity to amino acids 84-528 of SEQ ID NO: 1 (see Appendix II). Residues 84-528 of instant SEQ ID NO: 1 comprise the amino acid sequence set forth in SEQ ID NO: 2, and thus, Nakashima’s tyrosine hydroxylase variant also comprises the amino acid sequence set forth in SEQ ID NO: 2.
Regarding claim 5, Nakashima teaches the tyrosine hydroxylase variant further comprises a tag protein attached to the N-terminus (“each of the mutants of hTH1 connected to the polyhistidine tag at their N-terminus”, pg. 113, left col.).
Response to Remarks - 35 USC § 102
Applicant’s remarks regarding the § 102 rejections over Nakashima have been reviewed. Applicant appears to argue that claim 1 “do[es] not encompass a culture medium containing transformed cells,” because it has been amended to require selection of one of, or a combination of, the recited pharmaceutically acceptable carriers. Applicant argues that Nakashima does not meet the scope of the amended claims, therefore.
Examiner respectfully disagrees with this characterization of Nakashima as it applies to amended claim 1. Claim 1 is directed to a pharmaceutical composition, wherein the term “pharmaceutical” is interpreted as a non-limiting intended use for the claimed composition. The composition must comprise a tyrosine hydroxylase with the recited structural elements, an aromatic L-amino acid decarboxylase, and one of, or a combination of, the pharmaceutically acceptable carriers recited in the claim. Nakashima teaches a composition comprising the claimed elements, wherein the composition is a culture medium containing transfected AtT-20 cells. The claim does not appear to limit the structure of the composition so as to exclude Nakashima’s culture medium. Nakashima’s composition comprises a tyrosine hydroxylase variant comprising an N-terminal deletion of between 80-100 amino acids from instant SEQ ID NO: 1, an aromatic L-amino acid decarboxylase, and a pharmaceutically acceptable carrier which is a “transfection facilitating agent” (i.e., FuGENE6 Reagent). There is no apparent deficiency in Nakashima with respect to the claims and the rejections are maintained and modified as necessitated by Applicant’s amendments.
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 – Azzouz in view of NM_199292.2, Palfi, Nakashima and Ota
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 1, 3-6, 14-15, 17, 19-22, and 24-26 are rejected under 35 U.S.C. 103 as being unpatentable over Azzouz (Azzouz et al., 2002, The Journal of Neuroscience, 22(23): 10302-10312; of record) as evidenced by ThermoFisher (ThermoFisher Scientific, Technical Resources, “NEUROBASAL™ Medium (1X) liquid,” available 19 November 2015 as evidenced by The WayBack Machine), in view of Palfi (Palfi et al., 2018, Human Gene Therapy Clinical Development, Vol. 29, No. 3, pg. 148-155; of record), 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) and NM_199292.2 (Homo sapiens tyrosine hydroxylase (TH), transcript variant 1, mRNA; NCBI Reference Sequence: NM_199292.2; available 21 October 2018; of record). The rejections that follow are maintained from the prior action with modification necessitated by Applicant’s amendments to the claims.
The interpretation of claims 1 and 3 is described in paragraph 7 above and applied hereinafter. The term “saline” is interpreted as encompassing “bicarbonate buffer,” “salts solution,” etc. (“Illustrative examples of saline include, without limitation, buffer saline, normal saline, phosphate buffer, citrate buffer, acetate buffer, bicarbonate buffer, sucrose solution, salts solution and polysorbate solution,” [00134]).
Regarding claims 1 and 3, Azzouz teaches a composition (i.e., culture medium containing transduced neurons) comprising a tyrosine hydroxylase variant comprising an N-terminal deletion of 160 amino acids (“Human tyrosine hydroxylase type 2… A truncated form of hTH-2 that lacks the first 160 amino acids (TH) was amplified,” pg. 10303, right col.), and an aromatic L-amino acid decarboxylase (“human aromatic L-amino acid dopa decarboxylase,” pg. 10303, left col.)(“Viral vector construction,” pg. 10303; In vitro transduction, pg. 10303-10304; pg. 10305-10306; Fig. 1-2). Azzouz teaches the truncation removes the regulatory domain, while leaving the catalytic domain unaffected, in an effort to “prevent[] any inhibition of the enzyme by the synthesized cytoplasmic dopamine” (pg. 10305, left col.). Azzouz teaches the culture medium is “Neurobasal medium,” which as evidenced by ThermoFisher comprises “saline,” as interpreted directly above (“Inorganic Salts,” “Sodium Bicarbonate,” “Sodium Chloride,” etc.). Thus, Azzouz’s composition comprises an N-terminally truncated tyrosine hydroxylase, an aromatic L-amino acid decarboxylase, and a pharmaceutically acceptable carrier.
Regarding claims 14 and 19, Azzouz teaches a lentiviral vector comprising a polynucleotide construct comprising a first polynucleotide encoding the tyrosine hydroxylase variant, and a second polynucleotide encoding the aromatic L-amino acid decarboxylase (“Viral vector construction,” pg. 10303). Azzouz teaches the vector is designed as a long-term gene therapy to achieve dopamine replacement in Parkinson’s disease (PD) patients and overcome some of the disadvantages of systemic L-dopa administration, e.g., that it becomes less effective in later stages of disease, and is accompanied by “marked deleterious side effects” (Introduction, pg. 10302-10303; Discussion, pg. 10311).
The vector of Azzouz encodes an N-terminal deleted variant of “GenBank accession number X05290” (pg. 10303, right col.). Azzouz does not teach that the tyrosine hydroxylase variant comprises the sequence of SEQ ID NO: 1, comprising an N-terminal deletion between 80-100 or 80-90 amino acid residues.
However, NM_199292.2 teaches a tyrosine hydroxylase consisting of the sequence of instant SEQ ID NO: 1. See alignment of record showing 100% identity between the two sequences (pg. 1-2 of Appendix I). Palfi and Ota, as described below, provide means and motivation to the skilled artisan to prepare an N-terminal deletion of NM_199292.2 between 80-100 or 80-90 amino acid residues.
Palfi describes the results of a Phase I/II human study using a substantially identical lentiviral vector to Azzouz (“ProSavin vector”). Palfi teaches that the vector was “safe and well tolerated in patients with Parkinson’s disease” (Abstract, pg. 148; Discussion, pg. 154). Palfi teaches that “[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.). Palfi teaches that “[f]urther dose escalation using ProSavin would be challenging due to limitations on vector titers… and the volume of vector that can be safely administered into the human striatum” (pg. 154, right col.).
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 teaches that “enzyme activity turned round toward increase at del-117 (pg. 1101; Fig. 2). Ota’s enzyme activity is determined by comparing L-Dopa production directly from substrate (“L-tyrosine”) using protein prepared from E. coli lysate supernatants (“Measurement of Enzyme Activity,” pg. 1100). As shown in Fig. 2 of Ota, and consistent with the observations of Nakashima in Table 1, the del-52 and del-157 variants have less enzyme activity than wildtype tyrosine hydroxylase (Fig. 2). Other variants, e.g., del-37, del-67, and del-78 have roughly equivalent activity to wildtype tyrosine hydroxylase (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 references Daubner et al. as supporting these findings regarding this portion of the N-terminus (“These observations were very similar to the recent report concerning the deletion mutatgenesis of rat TH (20) in which mutants with the deletions in the residues 85-128 of rat TH gave the similar difficulty in obtaining the soluble proteins,” pg. 1104-1105).
Critical to the rationale herein, the residue numbering of Nakashima and Ota differs from that of instant SEQ ID NO: 1/NM_199292.2, or X05290 of Azzouz. As shown in Fig. A below, the N-terminal deletion variants encompassed by the instantly claimed ranges, i.e., 80-100 residues or 80-90 residues relative to SEQ ID NO: 1, lie N-terminal of the region identified by Ota at which enzyme activity “abruptly decrease[s]” (compare each N-terminal deletion variant of Ota with positions in SEQ ID NO: 1, where “*” marks E81, E91, and A101 relative to SEQ ID NO: 1). Indeed, as shown below, the instantly claimed range effectively encompasses the following N-terminal deletion variants described by Ota: del-52, del-67, and del-68. Azzouz’s N-terminal deletion variant lies C-terminal of Ota’s region of inactivity, and begins only 2 residues C-terminal to the del-157 variant, which Nakashima teaches produces approximately 250% more dopamine than full-length tyrosine hydroxylase in mammalian cells.
Figure A.
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It would have been obvious to one of ordinary skill before the effective filing date of the claimed invention to have prepared the vector and composition of Azzouz, using the tyrosine hydroxylase sequence taught by NM_199292.2 which is 100% identical to instant SEQ ID NO: 1, such that the protein comprises an N-terminal deletion between 80-100, or 80-90 residues in view of Nakashima and Ota. It would have amounted to applying a known technique (N-terminal deletion) to a known, full-length tyrosine hydroxylase sequence, ready for improvement to yield predictable results.
Based on the teachings of Nakashima and Ota, the skilled artisan would reasonably conclude that a tyrosine hydroxylase sequence 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, 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. The skilled artisan, based on the teachings of Nakashima and Ota, would understand that the increased dopamine production would occur despite the lower catalytic activity of some variants in the ranges, when measured as substrate conversion using protein prepared from cell lysates (see Ota, Fig. 2). Furthermore, 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.
Azzouz’s N-terminal deletion variant lies 2 residues C-terminal to the del-157 variant (see Fig. A above). Nakashima teaches the del-157 variant produces approximately 250% more dopamine than full-length tyrosine hydroxylase in mammalian cells, whereas the del-52 variant produces approximately 400% more dopamine. These teachings, coupled with Palfi’s teaching that a substantially identical vector to Azzouz’s, while “safe and well tolerated in patients with Parkinson’s disease,” fails to achieve enough dopamine production in vivo to render L-Dopa administration obsolete, would have motivated the skilled artisan to prepare the vector and composition with a tyrosine hydroxylase variant comprising an N-terminal deletion in the claimed range, i.e., which encompasses the del-52 variant, as well as variants in close proximity thereto.
Regarding claim 4, the obviousness of arriving at a tyrosine hydroxylase variant comprising the sequence set forth in SEQ ID NO: 2, i.e., amino acids 92-528 of instant SEQ ID NO: 1, is described above and applied hereinafter.
Regarding claims 5-6, the tyrosine hydroxylase variant of Azzozu comprises a tag protein attached to the N- or C-terminus, wherein the tag protein is HA, Myc, or Flag (Fig. 1).
Regarding claim 15, NM_199292.2 teaches the nucleotide sequence encoding the tyrosine hydroxylase, which as shown in the alignment of record is 100% identical to instant SEQ ID NO: 12 (see pg. 3-6 of Appendix I).
Regarding claim 17, Azzouz teaches the polynucleotide construct further comprises a promoter operably linked to the first and/or second polynucleotide (Fig. 1)
Regarding claim 20, Azzouz teaches a set of vectors, comprising a first vector comprising a first polynucleotide encoding a tyrosine hydroxylase variant (“A truncated form of hTH-2 that lacks the first 160 amino acids (TH) was… cloned directly into pcDNA3.1zeo… The new plasmid that expresses TH is called pNE4a” pg. 10303, right col.), and a second vector comprising a second polynucleotide encoding an aromatic L-amino acid decarboxylase (“The human aromatic L-amino acid dopa decarboxylase… was cloned from a human liver… and cloned into pcDNA3.1neo… to generate the plasmid pNE2,” pg. 10303, left col.). The obviousness of arriving at a tyrosine hydroxylase variant comprising the sequence set forth in SEQ ID NO: 1 comprising an N-terminal deletion of 80 to 100 amino acid residues is described above and applied hereinafter.
Regarding claims 21-22, as shown in Fig. 1 of Azzouz, 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 claim 24, Azzouz teaches an in vitro host cell comprising or transfected by the vector (see at least “Viral vector production,” pg. 10303, right col.; “In vitro transduction,” pg. 10303-10304).
Regarding claim 25, Azzouz teaches a virus comprising a virus genome wherein the virus genome comprises the polynucleotide construct (see “Viral vector construction,” and “Viral vector production,” pg. 10303).
Regarding claim 26, as evidenced by ThermoFisher, Azzouz teaches a composition comprising the virus and a pharmaceutically acceptable carrier (“Transduction was performed by diluting concentrated virus in supplemented Neurobasal medium,” pg. 10304, left col.).
Claim Rejections - 35 USC § 103 – Azzouz in view of NM_199292.2, Palfi, Nakashima and Ota as evidenced by M76180
Claims 10-12, and 16 are rejected under 35 U.S.C. 103 as being unpatentable over Azzouz (Azzouz et al., 2002, The Journal of Neuroscience, 22(23): 10302-10312; of record) as evidenced by ThermoFisher (ThermoFisher Scientific, Technical Resources, “NEUROBASAL™ Medium (1X) liquid,” available 19 November 2015 as evidenced by The WayBack Machine), in view of Palfi (Palfi et al., 2018, Human Gene Therapy Clinical Development, Vol. 29, No. 3, pg. 148-155; of record), 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) and NM_199292.2 (Homo sapiens tyrosine hydroxylase (TH), transcript variant 1, mRNA; NCBI Reference Sequence: NM_199292.2; available 21 October 2018; of record), as evidenced by M76180 (Human aromatic amino acid decarboxylase (ddc) mRNA, complete cds, NCBI Reference Sequence: M76180.1, available 31 December 1994; of record). The rejections that follow are maintained from the prior action with modification necessitated by Applicant’s amendments to the claims.
Azzouz teaches that the aromatic L-amino acid decarboxylase corresponds to “GenBank accession number M76180” (“human aromatic L-amino acid dopa decarboxylase (GenBank accession number M76180; hAADC480),” pg. 10303, left col). However, Azzouz does not teach the amino acid sequence of the AADC or the nucleotide sequence encoding the AADC corresponding to “GenBank accession number M76180.”
Regarding claim 10, M76180 teaches the amino acid sequence of Azzouz’s AADC, which as shown in the alignment of record is 100% identical to instant SEQ ID NO: 4 (see pg. 7-8 of Appendix I). Thus, as evidenced by M76180, Azzouz’s composition comprising an AADC which meets the limitations of instant claim 10, and therefore, the claim is also obvious for the reasons described above.
Regarding claims 11-12, the AADC of Azzozu comprises a tag protein attached to the N- or C-terminus, wherein the tag protein is HA, Myc, or Flag (Fig. 1).
Regarding claim 16, M76180 teaches the nucleotide sequence encoding the AADC, which as shown in the alignment of record is 100% identical to instant SEQ ID NO: 15 (see pg. 9-12 of Appendix I). Thus, as evidenced by M76180, Azzouz’s vector comprises a nucleotide sequence which meets the limitations of instant claim 16, and therefore, the claim is also obvious for the reasons described above.
Claim Rejections - 35 USC § 103 –
Azzouz, NM_199292.2, Palfi, Nakashima, and Ota, in further view of G-Biosciences
Claim 7 is rejected under 35 U.S.C. 103 as being unpatentable over Azzouz (Azzouz et al., 2002, The Journal of Neuroscience, 22(23): 10302-10312; of record) as evidenced by ThermoFisher (ThermoFisher Scientific, Technical Resources, “NEUROBASAL™ Medium (1X) liquid,” available 19 November 2015 as evidenced by The WayBack Machine), in view of Palfi (Palfi et al., 2018, Human Gene Therapy Clinical Development, Vol. 29, No. 3, pg. 148-155; of record), 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) and NM_199292.2 (Homo sapiens tyrosine hydroxylase (TH), transcript variant 1, mRNA; NCBI Reference Sequence: NM_199292.2; available 21 October 2018; of record) as applied to claims 1, 3-6, 14-15, 17, 19-22, and 25-26, and in further view of G-Biosciences (G-Biosciences, 10 June 2014, “8 Protein Tags Explained,” accessed 23 January 2025; of record). The rejection that follows is maintained from the prior action with modification necessitated by Applicant’s amendments to the claims.
Regarding claim 7, instant SEQ ID NO: 3 comprises SEQ ID NO: 2 (i.e., amino acids 92-528 of SEQ ID NO: 1) joined at its N-terminus to the sequence: “YPYDVPDYAYPYDVPDYA.”
The teachings of Azzouz as evidenced by ThermoFisher, NM_199292.2, Palfi, Nakashima, and Ota are described above and applied hereinafter. As stated therein, the tyrosine hydroxylase variant and AADC of Azzouz each comprise a tag protein at their N-terminus (Fig. 1). The N-terminal tag protein on the tyrosine hydroxylase variant is a Myc tag (Fig. 1). The N-terminal tag protein on the AADC is an HA tag (Fig. 1).
None of the above references teach that the tyrosine hydroxylase variant comprises the sequence “YPYDVPDYAYPYDVPDYA” at its N-terminus.
G-Biosciences teaches that Myc and HA tags are readily recognized by antibodies, and due to their small size, have little or no effect on the structure of the protein to which they are attached. G-Biosciences teaches the sequence of the HA tag: “YPYDVPDYA.”
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have substituted the N-terminal Myc tag on the tyrosine hydroxylase variant rendered obvious above for the tag protein sequence “YPYDVPDYAYPYDVPDYA” in view of G-Biosciences, in order to arrive at the sequence of instant SEQ ID NO: 3. It would have amounted to substituting one N-terminal tag protein, for two repeats of another, known tag protein, by known means to yield predictable results. As demonstrated by Azzouz and G-Biosciences, it was well within the purview of the skilled artisan to tag a protein of interest using recombinant techniques, and thus, the skilled artisan would have had a reasonable expectation of success in applying two repeats of the HA tag sequence disclosed by G-Biosciences to the tyrosine hydroxylase variant rendered obvious above. Tag proteins, e.g., HA and Myc, are art-recognized equivalents for their purpose. Indeed, the specification states “the term[] “tag protein”… refer[s] to a polypeptide or protein that is fused with a target protein... to facilitate expression, detection, tracing, or purification of the target protein. Protein tags include, but are not limited to… HA… or Myc” ([0079]). Accordingly, “An express suggestion to substitute one equivalent component or process for another is not necessary to render such substitution obvious. In re Fout, 675 F.2d 297, 213 USPQ 532 (CCPA 1982).” See MPEP 2144.06(II).
Claim Rejections - 35 USC § 103 –
Azzouz, NM_199292.2, Palfi, Nakashima, Ota, and M76180.1, in further view of G-Biosciences
Claim 13 is rejected under 35 U.S.C. 103 as being unpatentable over Azzouz (Azzouz et al., 2002, The Journal of Neuroscience, 22(23): 10302-10312; of record) as evidenced by ThermoFisher (ThermoFisher Scientific, Technical Resources, “NEUROBASAL™ Medium (1X) liquid,” available 19 November 2015 as evidenced by The WayBack Machine), in view of Palfi (Palfi et al., 2018, Human Gene Therapy Clinical Development, Vol. 29, No. 3, pg. 148-155; of record), 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), NM_199292.2 (Homo sapiens tyrosine hydroxylase (TH), transcript variant 1, mRNA; NCBI Reference Sequence: NM_199292.2; available 21 October 2018; of record), and M76180 (Human aromatic amino acid decarboxylase (ddc) mRNA, complete cds, NCBI Reference Sequence: M76180.1, available 31 December 1994; of record), as applied to claims 1, 3-6, 10-12, 14-16, 17, 19-22, and 25-26, and in further view of G-Biosciences (G-Biosciences, 10 June 2014, “8 Protein Tags Explained,” accessed 23 January 2025; of record). The rejection that follows is maintained from the prior action with modification necessitated by Applicant’s amendments to the claims.
Regarding claim 13, instant SEQ ID NO: 10 consists of amino acids 2-480 of instant SEQ ID NO: 4, joined at its C-terminus to the sequence: “EQKLISEEDL.”
The teachings of Azzouz as evidenced by ThermoFisher, NM_199292.2, Palfi, Nakashima, Ota, and M76180.1 are described above and applied hereinafter. As stated therein, the tyrosine hydroxylase variant and AADC of Azzouz each comprise a tag protein at their N-terminus (Fig. 1). The N-terminal tag protein on the tyrosine hydroxylase variant is a Myc tag (Fig. 1). The N-terminal tag protein on the AADC is an HA tag (Fig. 1).
None of the above references teach that the AADC comprises the sequence “EQKLISEEDL” at its C-terminus.
G-Biosciences teaches that Myc and HA tags are readily recognized by antibodies, and due to their small size, have little or no effect on the structure of the protein to which they are attached. G-Biosciences teaches the sequence of the Myc tag: “EQKLISEEDL.”
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have substituted the N-terminal HA tag on the AADC of Azzouz, for a C-terminal Myc tag with the sequence “EQKLISEEDL” in view of G-Biosciences, in order to arrive at the sequence of instant SEQ ID NO: 10. It would have amounted to substituting an N-terminal tag protein, for another known tag protein at the C-terminus, by known means to yield predictable results. As demonstrated by Azzouz and G-Biosciences, it was well within the purview of the skilled artisan to tag a protein of interest using recombinant techniques. G-Bioscience also teaches that tag proteins have “extremely little or no effect on the structure of the resulting fusion protein.” Thus, the skilled artisan would have had a reasonable expectation of success in applying the Myc tag disclosed by G-Biosciences to the C-terminus of the AADC of Azzouz. Tag proteins, e.g., HA and Myc, are art-recognized equivalents for their purpose. Indeed, the specification states “the term[] “tag protein”… refer[s] to a polypeptide or protein that is fused with a target protein... to facilitate expression, detection, tracing, or purification of the target protein. Protein tags include, but are not limited to… HA… or Myc” ([0079]). Accordingly, “An express suggestion to substitute one equivalent component or process for another is not necessary to render such substitution obvious. In re Fout, 675 F.2d 297, 213 USPQ 532 (CCPA 1982).” See MPEP 2144.06(II).
Claim Rejections - 35 USC § 103 –
Azzouz, NM_199292.2, Palfi, Nakashima, and Ota, in further view of Li
Claim 18 is rejected under 35 U.S.C. 103 as being unpatentable over Azzouz (Azzouz et al., 2002, The Journal of Neuroscience, 22(23): 10302-10312; of record) as evidenced by ThermoFisher (ThermoFisher Scientific, Technical Resources, “NEUROBASAL™ Medium (1X) liquid,” available 19 November 2015 as evidenced by The WayBack Machine), in view of Palfi (Palfi et al., 2018, Human Gene Therapy Clinical Development, Vol. 29, No. 3, pg. 148-155; of record), 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) and NM_199292.2 (Homo sapiens tyrosine hydroxylase (TH), transcript variant 1, mRNA; NCBI Reference Sequence: NM_199292.2; available 21 October 2018; of record) as applied to claims 1, 3-6, 14-15, 17, 19-22, and 25-26, and in further view of Li (Li et al., 2010, Journal of Neuroscience Methods, 189 (2010), pg. 56-64; of record). The rejection that follows is maintained from the prior action with modification necessitated by Applicant’s amendments to the claims.
The teachings of Azzouz as evidenced by ThermoFisher, NM_199292.2, Palfi, Nakashima, and Ota are described above and applied hereinafter. Azzouz teaches that the vector uses “the CMV promoter rather than the phosphoglycerate kinase promoter” (PGK) (pg. 10311, right col.). Azzouz teaches that use of “alternative promoters might further increase production of dopamine… probably leading to increased behavioral improvement” (pg. 10311, right col.).
None of the references teach that the promoter comprises a “neuron-specific promoter.”
However, Li teaches a variety of promoters suitable for use in lentiviral vectors, including CMV, PGK, and neuron-specific promoters, e.g., GFAP (Fig. 1; Discussion). Li teaches that different promoters have different expression profiles in target cells, and therefore, identification of a suitable promoter for the application is desirable (“promoters exhibit different relative levels of expression in different types of neurons and in astrocytes. The success of future studies will depend on identification and use of the appropriate promoter for expressing genes of interest delivered by lentiviral vectors in the target cell type,” section 1, Introduction).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have substituted the promoter in the vector of Azzouz for a neuron-specific promoter, e.g., GFAP, in view of Li. It would have amounted to a simple substitution of two known promoters suitable for use in lentiviral vectors, by known means to yield predictable results. The skilled artisan would have had a reasonable expectation of success because as evidenced by Azzouz and Li it was well within the purview of the skilled artisan to prepare vectors with different promoter sequences. Azzouz teaches that use of “alternative promoters might further increase production of dopamine… probably leading to increased behavioral improvement.” Li teaches that, due to their different expression profiles in target cells, identification of an appropriate promoter is desirable. The skilled artisan, therefore, would have been motivated to substitute the promoter in the vector of Azzouz for a neuron-specific promoter in an effort to identify the appropriate promoter for expression in target cells, and further increase production of dopamine and improve behavior.
Claim Rejections - 35 USC § 103 –
Azzouz, NM_199292.2, Palfi, Nakashima, and Ota, in further view of Gray
Claim 23 is rejected under 35 U.S.C. 103 as being unpatentable over Azzouz (Azzouz et al., 2002, The Journal of Neuroscience, 22(23): 10302-10312; of record) as evidenced by ThermoFisher (ThermoFisher Scientific, Technical Resources, “NEUROBASAL™ Medium (1X) liquid,” available 19 November 2015 as evidenced by The WayBack Machine), in view of Palfi (Palfi et al., 2018, Human Gene Therapy Clinical Development, Vol. 29, No. 3, pg. 148-155; of record), 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) and NM_199292.2 (Homo sapiens tyrosine hydroxylase (TH), transcript variant 1, mRNA; NCBI Reference Sequence: NM_199292.2; available 21 October 2018; of record) as applied to claims 1, 3-6, 14-15, 17, 19-22, and 25-26, and in further view of Gray (Gray et al., 2010, Ther Deliv. 1(4):517-534, available 21 July 2015 on PMC; of record). The rejection that follows is maintained from the prior action with modification necessitated by Applicant’s amendments to the claims.
The teachings of Azzouz as evidenced by ThermoFisher, NM_199292.2, Palfi, Nakashima, and Ota are described above and applied hereinafter.
None of the references teach that the vector is a herpes simplex vector, adenovirus vector, or adeno-associated virus vector.
However, Gray teaches various viral vectors which may be suitable for use in the central nervous system, including herpes simplex viral vectors and lentiviral vectors (“Introduction & overview of viral vectors,” pg. 2). Gray teaches that one of the greatest advantages of herpes simplex viral vectors is “the high packaging capacity” and “natural neurotropism” (pg. 2-3).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have substituted the lentiviral vector of Azzouz for a different viral vector, i.e., a herpes simplex virus vector, in view of Gray. It would have amounted to a simple substitution of two viral vectors, of sufficient capacity, by known means to yield predictable results. The skilled artisan would have had a reasonable expectation of success in substituting the vectors because as evidenced by Gray, herpes simplex viral vectors have a packaging capacity greater than lentiviral vectors, and therefore, would be suitable to carry the cargo of Azzouz’s vector. The skilled artisan would have been motivated to substitute the type of viral vector because Gray teaches that herpes simplex viral vectors have “high packaging capacity” and “natural neurotropism.”
Response to Remarks - 35 USC § 103
Applicant’s remarks regarding the § 103 rejections raised in the prior action have been reviewed. Applicant refers to the § 102 remarks, which as stated above, appear to argue that claim 1 “does not encompass culture medium containing transformed cells” because it has been amended to require selection of one of, or a combination of, the recited pharmaceutically acceptable carriers. Applicant argues that Azzouz’s composition does not meet the scope of the amended claims, and none of the secondary references resolve the alleged deficiencies of Azzouz.
Examiner respectfully disagrees with this characterization of Azzouz as it applies to amended claim 1. Claim 1 is directed to a pharmaceutical composition, wherein the term “pharmaceutical” is interpreted as a non-limiting intended use for the claimed composition. The composition must comprise a tyrosine hydroxylase with the recited structural elements, an aromatic L-amino acid decarboxylase, and one of, or a combination of, the pharmaceutically acceptable carriers recited in the claims, e.g., “saline.” The term “saline” is interpreted as, for example, “salt solutions” based on the specification ([00134]). As evidenced by ThermoFisher, Azzouz’s composition (i.e., culture medium containing transduced neurons) comprises “saline” (“Inorganic Salts,” “Sodium Bicarbonate,” “Sodium Chloride,” etc.). The claim does not appear to limit the structure of the composition so as to exclude Azzouz’s culture medium. There is no apparent deficiency in Azzouz with respect to a composition comprising a pharmaceutically acceptable carrier recited in the amended claims, and the rejections are maintained and modified as necessitated by Applicant’s amendments.
Conclusion
No claims are allowed.
THIS ACTION IS MADE FINAL. 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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/JENNA L PERSONS/Examiner, Art Unit 1637
/Soren Harward/Primary Examiner, TC 1600