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 .
1. The Amendment filed July 17, 2026 in response to the Office Action of February 26, 2026, is acknowledged and has been entered. Claims 97-109, 111-116 are pending. Claims 1-96, 110 are canceled. Claims 97 and 109 are amended. Claims 101, 102, 104, 111-115 remain withdrawn. Claims 97-100, 103, 105-109, 116 are currently being examined as drawn to the species of:
A. species of cell penetrating polypeptide: (i) conjugated to a therapeutic agent (claim 99, withdraw claims 104 and 115);
B. species of cell penetrating polypeptide: (ii) NOT joined to sequence AGIH (withdraw claim 101);
C. species of cell penetrating polypeptide: (ii) NOT joined to or comprises a peptide linker comprising a portion of CH1 and a swivel sequence (withdraw claim 102);
D. species of cell penetrating peptide: (i) comprises an amino substitution to asparagine at residue 31 (D31N) (claim 103); and
E. site of interest: (ii) is NOT an injury site (withdraw claims 111-114).
The claims are amended to recite the method comprises administering a complex comprising 3E10 antibody bound to purified sheared extracellular DNA.
Maintained Rejections
(amendments addressed with the addition of Yoo reference)
Claim Rejections - 35 USC § 103
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.
2. Claim(s) 97-100, 103, 105-109, and 116 remain rejected under 35 U.S.C. 103 as being unpatentable over WO 1997032602, Weisbart, in view of QIAGEN® Plasmid Purification Handbook (“Qiagen”), second edition, August 2003; and Yoo et al (Journal of Biophysical Chemistry, 2011, 2:102-110); and as evidenced by US Patent Application Publication 2008/0292618, Weisbart.
Weisbart teaches administering anti-dsDNA 3E10 antibody to live targeted cells (which is a site of interest), wherein the 3E10 antibody is complexed/conjugated with a therapeutic biologically active compound or biological compound of interest for transport into the cells (p. 3-5; p. 12, line 34 to p. 13, line 1; claims 1-20); wherein the biologically active compound is a carrier with, or conjugated to, DNA, DNA plasmid containing genes of interest, a gene, or genetic material, which would encompass extracellular DNA (p. 5, lines 7-18; p. 7, lines 18-28; p. 8, lines 5-15; claims 2, 7, 9, and 16); wherein the 3E10 antibody can be a functional fragment thereof such as an scFv (p. 6, lines 9-13); wherein the 3E10 antibody is humanized (p. 6, lines 14-34); wherein the 3E10 antibody has a mutation at position 31 of aspartic acid to asparagine (D31N) (p. 7, lines 3-17; p. 20, lines 9-20); wherein the 3E10 antibody complex/conjugate is administered to a patient or animal by injection (p. 9, lines 16-32).
Weisbart teaches: “Recent work indicates that cellular penetration requires complexes of antibody and DNA. MAb 3E10 can penetrate many different types of cell lines in tissue culture. In contrast, mAb 3E10 may be restricted in the cells it can bind and penetrate in vivo, where there is an absence of free DNA to facilitate its penetration” (p. 8-9). Weisbart teaches: “The results showed that residues required for binding DNA but not HP8/HEVIN were necessary for antibody penetration, indicating that cellular penetration required the presence of DNA or binding of antibody to a membrane determinant precisely resembling DNA” (abstract; p. 10, lines 30-32). Therefore, Weisbart determined that cellular penetration of 3E10 antibody requires the presence of, and binding to, DNA.
As evidenced by US 2008/0292618, Weisbart, the 3E10 antibody is transported into cells via the nucleoside salvage pathway through ENT2-mediated nucleoside salvage pathway (Example 1). Therefore, the method of WO 1997032602, Weisbart, delivering 3E10 complexes into cells necessarily depends on ENT2-mediated nucleoside salvage pathway.
Weisbart does not teach the genetic material, extracellular DNA, or DNA plasmid containing genes of interest was purified away from non-DNA nucleic acid, nucleotide, and nucleosides and contains sheared extracellular DNA.
Qiagen sells commercially available kits for rapidly isolating and purifying plasmid DNA, particularly for cell transfection purposes. Qiagen teaches the process for purification of plasmids in the figure on p. 12, which involves binding, wash, and elution steps utilizing QIAGEN Anion-Exchange Resin. Qiagen teaches the Anion-Exchange Resin purifies the plasmid DNA away from RNA, proteins, dyes, and low-molecular-weight impurities (i.e. nucleotides and nucleosides) (p. 13):
Principle and procedure
QIAGEN plasmid purification protocols are based on a modified alkaline lysis procedure, followed by binding of plasmid DNA to QIAGEN Anion-Exchange Resin under appropriate low-salt and pH conditions. RNA, proteins, dyes, and low-molecular-weight impurities are removed by a medium-salt wash. Plasmid DNA is eluted in a high-salt buffer and then concentrated and desalted by isopropanol precipitation.
Page 70:
DNA binding and washing on the QIAGEN-tip
The cleared lysate is loaded onto a pre-equilibrated QIAGEN-tip by gravity flow. The salt and pH conditions of the lysate and the superior selectivity of the QIAGEN Resin
ensure that only plasmid DNA binds, while degraded RNA, cellular proteins, and
metabolites are not retained and appear in the flow-through fraction. The QIAGEN-tip is then washed with medium-salt buffer (Buffer QC) which completely removes any remaining contaminants, such as traces of RNA and protein (e.g., RNase A), without affecting the binding of the plasmid DNA (see Figure 4 on page 61). Buffer QC also disrupts nonspecific interactions, and allows removal of nucleic acid-binding proteins without the use of phenol. The low concentration of alcohol in the wash buffer eliminates nonspecific hydrophobic interactions, further enhancing the purity of the bound DNA. The plasmid DNA is then efficiently eluted from the QIAGEN-tip with high salt buffer (Buffer QF or QN). For further information about QIAGEN Anion-Exchange Resin, see Appendix E: QIAGEN Anion-Exchange Resin, pages 76–78.
Qiagen teaches protocols for processing extracellular DNA for purification that requires pipetting, mixing, shaking, centrifuging, and additional manipulations (p. 16-51).
Yoo teaches demonstrates that sheared (fragmented) DNA is commonly present in DNA samples resulting from basic laboratory handling and processing, including pipetting, vortexing, rocking, freeze-thawing, ultrasonication and ultrafiltration (abstract; sections 2.3 and 3.2; Figures 2-7).
It would have been prima facie obvious to one of ordinary skill in the art at the time the invention was filed to purify the extracellular DNA or DNA plasmid containing genes of interest away from non-DNA nucleic acid, nucleotide, and nucleosides, and for the plasmid DNA to comprise sheared DNA in the method and 3E10 antibody complex of Weisbart. One would have been motivated to, and have a reasonable expectation of success to because: (1) Weisbart teaches 3E10 antibody requires the presence of, and binding to, DNA for transport into a cell, and teaches complexing 3E10 with DNA or DNA plasmid containing genes of interest or genetic material for delivery into a cell, therefore any DNA present and contacted with 3E10 antibody is expected to bind to 3E10 antibody including sheared DNA; (2) Qiagen teaches and demonstrates isolation and purification of extracellular DNA or DNA plasmid containing genes of interest, away from non-DNA nucleic acid, nucleotide, and nucleosides, for use in cell delivery is routine and commercially available and discloses routine protocols for doing so including manipulations by pipetting, vortexing, centrifuging, rocking, and such; and (3) Yoo demonstrates that sheared DNA is a common by-product present in DNA samples due to routine laboratory manipulations including pipetting, vortexing, rocking, freeze-thawing, and physical agitations.
3. Claim(s) 97-100, 103, 105-109, and 116 remain rejected under 35 U.S.C. 103 as being unpatentable over US Patent 7,189,396, Weisbart, in view of QIAGEN® Plasmid Purification Handbook, second edition, August 2003; and Yoo et al (Journal of Biophysical Chemistry, 2011, 2:102-110); and as evidenced by US Patent Application Publication 2008/0292618, Weisbart.
US Patent 7,189,396, Weisbart claims priority to WO 1997032602, Weisbart, cited above, and has the same disclosure that anticipates the instant claims for the same reasons set forth above. US Patent 7,189,396 further patented a method for transporting a biologically active molecule into the nucleus of a target cell in vitro, comprising: a) combining said biologically active molecule in vitro with monoclonal antibody (mAb) 3E10 as produced by a hybridoma having ATCC accession number PTA 2439 or an scFv fragment thereof, to form an antibody-biologically active molecule conjugate or fusion protein, and b) contacting the target cell in vitro under suitable conditions with the antibody-biologically active molecule conjugate of a),
thereby transporting the biologically active molecule into the nucleus of the target cell; wherein said biologically active molecule is genetic material.
Wesibart further demonstrates and teaches cellular penetration of 3E10 antibody requires the presence of, and binding to, DNA, as stated above.
As evidenced by US 2008/0292618, Weisbart, the 3E10 antibody is transported into cells via the nucleoside salvage pathway through ENT2-mediated nucleoside salvage pathway (Example 1). Therefore, the method disclosed and claimed by the US Patent for delivering 3E10 complexes into cells necessarily depends on ENT2-mediated nucleoside salvage pathway.
Weisbart does not teach the extracellular DNA or DNA plasmid containing genes of interest was purified away from non-DNA nucleic acid, nucleotide, and nucleosides, and comprises sheared DNA.
Qiagen sells commercially available kits for rapidly isolating and purifying plasmid DNA, particularly for cell transfection purposes. Qiagen teaches the process for purification of plasmids in the figure on p. 12, which involves binding, wash, and elution steps utilizing QIAGEN Anion-Exchange Resin. Qiagen teaches the Anion-Exchange Resin purifies the plasmid DNA away from RNA, proteins, dyes, and low-molecular-weight impurities (i.e. nucleotides and nucleosides) (p. 13):
Principle and procedure
QIAGEN plasmid purification protocols are based on a modified alkaline lysis procedure, followed by binding of plasmid DNA to QIAGEN Anion-Exchange Resin under appropriate low-salt and pH conditions. RNA, proteins, dyes, and low-molecular-weight impurities are removed by a medium-salt wash. Plasmid DNA is eluted in a high-salt buffer and then concentrated and desalted by isopropanol precipitation.
Page 70:
DNA binding and washing on the QIAGEN-tip
The cleared lysate is loaded onto a pre-equilibrated QIAGEN-tip by gravity flow. The salt and pH conditions of the lysate and the superior selectivity of the QIAGEN Resin
ensure that only plasmid DNA binds, while degraded RNA, cellular proteins, and
metabolites are not retained and appear in the flow-through fraction. The QIAGEN-tip is then washed with medium-salt buffer (Buffer QC) which completely removes any remaining contaminants, such as traces of RNA and protein (e.g., RNase A), without affecting the binding of the plasmid DNA (see Figure 4 on page 61). Buffer QC also disrupts nonspecific interactions, and allows removal of nucleic acid-binding proteins without the use of phenol. The low concentration of alcohol in the wash buffer eliminates nonspecific hydrophobic interactions, further enhancing the purity of the bound DNA. The plasmid DNA is then efficiently eluted from the QIAGEN-tip with high salt buffer (Buffer QF or QN). For further information about QIAGEN Anion-Exchange Resin, see Appendix E: QIAGEN Anion-Exchange Resin, pages 76–78.
Qiagen teaches protocols for processing extracellular DNA for purification that requires pipetting, mixing, shaking, centrifuging, and additional manipulations (p. 16-51).
Yoo teaches demonstrates that sheared DNA is commonly present in DNA samples resulting from basic laboratory handling and processing, including pipetting, vortexing, rocking, freeze-thawing, ultrasonication and ultrafiltration (abstract; sections 2.3 and 3.2; Figures 2-7).
It would have been prima facie obvious to one of ordinary skill in the art at the time the invention was filed to purify the extracellular DNA or DNA plasmid containing genes of interest away from non-DNA nucleic acid, nucleotide, and nucleosides, and for the plasmid DNA to comprise sheared DNA in the method and 3E10 antibody complex of Weisbart. One would have been motivated to, and have a reasonable expectation of success to because: (1) Weisbart teaches 3E10 antibody requires the presence of, and binding to, DNA for transport into a cell, and teaches complexing 3E10 with DNA or DNA plasmid containing genes of interest or genetic material for delivery into a cell, therefore any DNA present and contacted with 3E10 antibody is expected to bind to 3E10 antibody including sheared DNA; (2) Qiagen teaches and demonstrates isolation and purification of extracellular DNA or DNA plasmid containing genes of interest, away from non-DNA nucleic acid, nucleotide, and nucleosides, for use in cell delivery is routine and commercially available and discloses routine protocols for doing so including manipulations by pipetting, vortexing, centrifuging, rocking, and such; and (3) Yoo demonstrates that sheared DNA is a common by-product present in DNA samples due to routine laboratory manipulations including pipetting, vortexing, rocking, freeze-thawing, and physical agitations.
Response to Arguments
5. Applicants argue that Weisbart teaches plasmid DNA and not purified sheared DNA that is purified away from non-DNA nucleic acid, nucleotide, nucleoside purine, or pyrimidine base. Applicants argue that Weisbart fails to demonstrate purified extracellular sheared DNA complexed to the 3E10 antibody. Applicants argue there is no reasonable expectation of success to combine Qiagen with Weisbart to arrive at the claimed invention because Qiagen is directed to plasmid purification protocols for isolating only plasmid DNA and not sheared DNA.
6. The arguments have been considered but are not persuasive. Applicants are arguing limitations not recited in the claims. The claimed complex of 3E10 antibody with sheared/fragmented extracellular DNA purified away from non-DNA nucleic acid, nucleotide, nucleoside purine or pyrimidine bases does not exclude the presence of plasmid DNA. Yoo teaches that sheared DNA is commonly present in DNA samples through routine laboratory manipulation, providing an expectation that purified plasmid DNA through routine and commerical protocols and kits like Qiagen’s would comprise sheared DNA in the purified sample, as well as during any freeze/thaw or handling of the DNA sample during complexing with 3E10 antibody. Weisbart teaches the known binding function of 3E10 to DNA and intracellular transport, providing a reasonable expectation of success for 3E10 to complex with any DNA in the composition administered, including plasmid DNA and fragmented/sheared DNA. Further, Weisbaert explicitly suggests complexing genetic material to 3E10 for cell transport, which broadly encompasses any DNA including sheared.
Maintained Rejections
(Yoo reference and/or arguments added to address amendments)
Double Patenting
The nonstatutory double patenting rejection is based on a judicially created doctrine grounded in public policy (a policy reflected in the statute) so as to prevent the unjustified or improper timewise extension of the “right to exclude” granted by a patent and to prevent possible harassment by multiple assignees. A nonstatutory double patenting rejection is appropriate where the conflicting claims are not identical, but at least one examined application claim is not patentably distinct from the reference claim(s) because the examined application claim is either anticipated by, or would have been obvious over, the reference claim(s). See, e.g., In re Berg, 140 F.3d 1428, 46 USPQ2d 1226 (Fed. Cir. 1998); In re Goodman, 11 F.3d 1046, 29 USPQ2d 2010 (Fed. Cir. 1993); In re Longi, 759 F.2d 887, 225 USPQ 645 (Fed. Cir. 1985); In re Van Ornum, 686 F.2d 937, 214 USPQ 761 (CCPA 1982); In re Vogel, 422 F.2d 438, 164 USPQ 619 (CCPA 1970); In re Thorington, 418 F.2d 528, 163 USPQ 644 (CCPA 1969).
A timely filed terminal disclaimer in compliance with 37 CFR 1.321(c) or 1.321(d) may be used to overcome an actual or provisional rejection based on nonstatutory double patenting provided the reference application or patent either is shown to be commonly owned with the examined application, or claims an invention made as a result of activities undertaken within the scope of a joint research agreement. See MPEP § 717.02 for applications subject to examination under the first inventor to file provisions of the AIA as explained in MPEP § 2159. See MPEP § 2146 et seq. for applications not subject to examination under the first inventor to file provisions of the AIA . A terminal disclaimer must be signed in compliance with 37 CFR 1.321(b).
The filing of a terminal disclaimer by itself is not a complete reply to a nonstatutory double patenting (NSDP) rejection. A complete reply requires that the terminal disclaimer be accompanied by a reply requesting reconsideration of the prior Office action. Even where the NSDP rejection is provisional the reply must be complete. See MPEP § 804, subsection I.B.1. For a reply to a non-final Office action, see 37 CFR 1.111(a). For a reply to final Office action, see 37 CFR 1.113(c). A request for reconsideration while not provided for in 37 CFR 1.113(c) may be filed after final for consideration. See MPEP §§ 706.07(e) and 714.13.
The USPTO Internet website contains terminal disclaimer forms which may be used. Please visit www.uspto.gov/patent/patents-forms. The actual filing date of the application in which the form is filed determines what form (e.g., PTO/SB/25, PTO/SB/26, PTO/AIA /25, or PTO/AIA /26) should be used. A web-based eTerminal Disclaimer may be filled out completely online using web-screens. An eTerminal Disclaimer that meets all requirements is auto-processed and approved immediately upon submission. For more information about eTerminal Disclaimers, refer to www.uspto.gov/patents/apply/applying-online/eterminal-disclaimer.
7. Claims 97-100, 103, 105-109, and 116 remain rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1-20 of U.S. Patent No. 10,383,945 in view of QIAGEN® Plasmid Purification Handbook, second edition, August 2003 and Yoo et al (Journal of Biophysical Chemistry, 2011, 2:102-110). Although the claims at issue are not identical, they are not patentably distinct from each other because the US Patent is claiming methods of delivering complexes/conjugates of 3E10 antibody or an scFv fragment of 3E10 bound to isolated DNA to live cells at a site of interest, that overlap with the instant claims, rendering the instant claims obvious.
The US Patent claims:
1. A method for selectively targeting live cells at a site of interest with a cell-penetrating polypeptide, the method comprising:
(a) introducing an isolated DNA near or around the live cells at the site of interest;
(b) administering the cell-penetrating polypeptide comprising cell-penetrating determinants, before, after or concurrently with the DNA of step (a);
(c) contacting the DNA or its degradation product with the cell-penetrating polypeptide so that the cell-penetrating polypeptide binds the DNA or its degradation product near or around the live cells so as to form a complex;
(d) contacting one of the live cells with the complex in (c) so as to bind and penetrate the live cell; and
(e) permitting additional complexes to form as in (c) and contacting additional cells with said complexes so as to bind and penetrate additional live cells at the site of interest;
wherein introducing an isolated DNA near or around the live cells at the site of interest is or comprises administration of the isolated DNA at the site of interest by injection, microinjection, microprojectile or implantation or a combination thereof, or alternatively, implantation of an object or substance containing DNA or artificial DNA, wherein the implant releases DNA or artificial DNA in a single burst or over an extended period; thereby, selectively targeting live cells at the site of interest with a cell-penetrating polypeptide, wherein the cell-penetrating polypeptide is a 3E10 antibody or a fragment thereof, wherein the fragment is a single chain Fv (scFv) fragment of mAb 3E10 antibody.
2. A method for inhibiting a tumor associated with ischemia, cellular/tissue necrosis or cellular/tissue apoptosis by selective targeting of live cells at a site of interest by the method of claim 1.
3. The method of claim 1, wherein the anti-DNA antibody binds single-stranded DNA, double-stranded DNA or both.
4. The method of claim 1, wherein the 3E10 antibody has a binding specificity of an antibody as produced by a hybridoma having ATCC accession number PTA 2439 or a fragment thereof.
5. The method of claim 1, wherein cell penetration is dependent on a salvage pathway.
6. The method of claim 5, wherein the salvage pathway is a nucleoside salvage pathway mediated by equilibrative nucleoside transporters (ENTs) or SLC29 family of integral membrane proteins.
7. The method of claim 6, wherein the equilibrative nucleoside transporter (ENT) or a member of the SLC29 family of integral membrane proteins is an equilibrative nucleoside transporter ENT2.
8. The method of claim 1, wherein the cells are present in a mammal or derived from a mammal.
9. The method of claim 1, wherein the cell-penetrating polypeptide is or comprises a therapeutic agent.
10. The method of claim 1, wherein the 3E10 antibody is a humanized variant of an antibody produced by the hybridoma having ATCC accession number PTA 2439.
11. The method of claim 1, wherein the 3E10 antibody, the fragment of the mAb 3E10 antibody, or the single chain Fv (scFv) fragment of the mAb 3E10 antibody is joined to an amino acid sequence AGIH at its amino terminus.
12. The method of claim 1, wherein the 3E10 antibody, the fragment of the mAb 3E10 antibody or the single chain Fv (scFv) fragment of the mAb 3E10 antibody has a change in its amino acid sequence, wherein the change in the amino acid sequence enhances cell penetration and comprises an amino acid substitution in the 3E10 variable heavy chain at amino acid 31 from an aspartic acid to an asparagine (D31N).
13. The method of claim 1, wherein the isolated DNA is single-stranded, double-stranded, triple-stranded, or four-stranded or a combination thereof, comprising a phosphodiester bond, a phosphorothioate bond or a methylphosphonate bond or a combination thereof or an artificial DNA comprising a pseudopeptide backbone, comprising an ethylglycine, a propylglycine, an ethyl-β-alanine, a propionyl linker, a retro inverso linker, a (S,S)-cyclohexyl linker, a (R,R)-cyclohexyl linker, an L-ornithine, a 2-me-ethyl-glycine, an ethyl-lysine, a L-proline, a n-proline, a glycine backbone/ethyl linker, a L-4-trans-amino proline, a L-4-cis-amino proline, a D-4-trans-amino proline, a P-alanine/proline, a glycylglycine/ethyl linker, a glyeine/ethyl linker, a proline-glycine, a f3-amino-alanine, E-OPA, Z-OPA, APNA, a serinol-ethyl-methyl linker, a serinol-ethyl-ethyl linker, an α-methyl- serinol-ethyl-ethyl linker, an aminopentan, a hydroxyethyl phosphono glycine, an aminoethyl phosphono glycine, a lysine, an aminoethyl prolyl or a serinyl methylene or a combination thereof.
14. The method of claim 1, wherein the cell-penetrating polypeptide further comprises a cytotoxic agent selected from the group consisting of a chemotherapeutic agent, a drug, a growth inhibitory agent, a toxin and a radioactive isotope.
15. The method of claim 1, wherein the cell penetrating polypeptide further comprises a cytoprotective agent selected from the group consisting of a heat shock protein, stress protein and chaperone protein.
16. The method of claim 1, wherein the cell-penetrating polypeptide is further linked or bound to an imaging agent or detectable marker.
17. The method of claim 16, wherein the imaging agent or detectable marker is selected from the group consisting of a radioisotope, a fluorophore, a fluorescent quencher, an enzyme, a luminescent compound, a chemiluminescent compound, a bioluminescent compound, a photon emitter, a heavy metal, a ferromagnetic agent, a contrast agent, a metal chelator, and an epitope.
18. The method of claim 1, wherein the mAb 3E10 antibody, the fragment of the mAb 3E10 antibody, or the single chain 17v (scFv) fragment of the mAb 3E10 antibody is a humanized antibody or fragment from a hybridoma having ATCC accession number PTA 2439.
19. The method of claim 1, wherein the site of interest is an injury site.
20. The method of claim 19, wherein the injury site is an intracranial injury, brain injury, myocardial infarction, skin injury, liver injury, gastrointestinal injury, lung injury, eye injury, kidney injury, pancreas injury, peritoneal injury, bone injury, nasopharyngeal injury, uterine injury, cervical injury, breast injury, organ injury, tissue injury, burn or radiation injury.
The US Patent claims do not recite that the isolated DNA was purified away from non-DNA nucleic acid, nucleotide, and nucleoside purine and pyrimidine bases and comprises sheared extracellular DNA.
The purified sheared extracellular DNA is obviated for reasons under (A) and (B):
(A) The US Patent discloses that purified sheared extracellular DNA is encompassed by isolated DNA (col. 8, lines 5-19):
In accordance with the practice of the invention, “extracellular DNA” is DNA free of a cell or cell-free DNA. Extracellular DNA may be introduced or administered by methods known in the art such as, for example, microinjection of DNA into extracellular space or outside of a cell or cells, cell or cells in a tissue, or cell or cells in an organ, so long as the DNA is not introduced or administered into a cell. The delivered DNA in the extracellular space or outside of a cell may be in any physical state, including in a solution, as a solid, as a colloidal form, as a semi-crystalline state, as a nanoparticle or combination thereof. The delivered DNA may be isolated DNA from nature, either total, fractionated, intact or sheared, or synthesized by the hand of man such as through synthetic chemistry or in vitro enzymatic methods as known in the art.
The US Patent also discloses that isolated DNA encompasses DNA purified away from non-DNA nucleic acid, nucleotide, nucleoside or purine or pyrimidine base (col. 15, lines 66 to col. 16, line 13).
In NSDP rejection, “while the earlier patent’s specification cannot be used as prior art to the later patent application claims, it can be consulted to ascertain the meaning of the words in the claims to interpret the patent claim’s proper scope (Sun Pharmaceutical Industries Ltd v. Eli Lilly and Co., 611 F.3d 1381, 1387 (Fed. Cir. 2010)”.
“A patent’s disclosure may be used to determine whether an application claim is merely an obvious variation of an invention claimed in a patent”In Re Basell Poliolefine Italia S.P.A., 547 F.3d 1371, 1378 (Fed. Cir. 2008) and Sun Pharmaceutical Industries Ltd v. Eli Lilly and Co., 611 F.3d 1381, 1387 (Fed. Cir. 2010)”.
In the instant case, the specification of the US Patent discloses that purified sheared extracellular DNA is encompassed by isolated DNA. Therefore, purified sheared extracellular DNA is encompassed by and obviated by the claimed isolated DNA in the method of the US Patent claims. The instantly claimed purified sheared extracellular DNA is not patentably distinct from the US Patent’s isolated DNA.
(B) Qiagen teaches commercially available kits for rapidly isolating and purifying plasmid DNA, particularly for cell transfection purposes as set forth above.
Yoo teaches routine laboratory manipulations, including those employed to isolate DNA, commonly result in the presence of sheared/ fragmented DNA, as set forth above.
It would have been prima facie obvious to one of ordinary skill in the art at the time the invention was filed to purify the DNA or DNA plasmid containing genes of interest away from non-DNA nucleic acid, nucleotide, and nucleosides in the method of the US Patent and for the DNA to comprise sheared DNA. One would have been motivated to, and have a reasonable expectation of success to because: (1) the US Patent claims complexing isolated DNA with 3E10 antibody for cell delivery; (2) Qiagen teaches and demonstrates isolation and purification of DNA or DNA plasmid containing genes of interest, away from non-DNA nucleic acid, nucleotide, and nucleosides, for use in cell delivery is routine and commercially available, and discloses routine protocols for doing so including manipulations by pipetting, vortexing, centrifuging, rocking, and such; and (3) Yoo demonstrates that sheared DNA is a common by-product present in DNA samples due to routine laboratory manipulations including pipetting, vortexing, rocking, freeze-thawing, and physical agitations.
8. Claims 97-100, 103, 105-109, and 116 remain rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1-40 of U.S. Patent No. 11,453,725. Although the claims at issue are not identical, they are not patentably distinct from each other because the US Patent is claiming methods of delivering complexes/conjugates of 3E10 antibody or an scFv fragment of 3E10 bound to purified DNA to live cells at a site of interest.
The US Patent claims:
1. A method for selectively targeting live cells at a site of interest with a cell-penetrating polypeptide, the method comprising:
(a) introducing purified DNA near or around the live cells at the site of interest;
(b) administering the cell-penetrating polypeptide comprising cell-penetrating determinants, before, after or concurrently with the purified DNA of step (a);
(c) contacting the purified DNA or its degradation product with the cell-penetrating polypeptide so that the cell-penetrating polypeptide binds the purified DNA or its degradation product near or around the live cells so as to form a complex;
(d) contacting one of the live cells with the complex in (c) so as to bind and penetrate the live cell; and
(e) permitting additional complexes to form as in (c) and contacting additional cells with said complexes so as to bind and penetrate additional live cells at the site of interest;
wherein the cell penetrating polypeptide is a 3E10 antibody or a fragment thereof, wherein the fragment thereof is a single chain Fv (scFv) fragment of mAb 3E10 antibody,
thereby, selectively targeting live cells at the site of interest with a cell-penetrating polypeptide.
4. The method of claim 1 wherein the cell-penetrating polypeptide is conjugated to a therapeutic agent.
7. The method of claim 1, wherein the m Ab 3E10 antibody, the fragment of the mAb 3E10 antibody, or the scFv fragment of the mAb 3E10 antibody is a humanized antibody or fragment from a hybridoma having ATCC accession number PTA 2439.
8. The method of claim 1, wherein the 3E10 antibody is a humanized variant of an antibody produced by the hybridoma having ATCC accession number PTA 2439.
11. The method of claim 1, wherein the fragment of the mAb 3E10 antibody, or the scFv fragment of the mAb 3E10 antibody has a change in its amino acid sequence, wherein the change in the amino acid sequence enhances cell penetration and comprises an amino acid substitution in the 3E10 variable heavy chain at amino acid 31 from an aspartic acid to an asparagine (D31N).
12. The method of claim 1, wherein cell penetration is dependent on a salvage pathway.
13. The method of claim 12, wherein the salvage pathway is a nucleoside salvage pathway.
14. The method of claim 13, wherein the nucleoside salvage pathway is a pathway mediated by equilibrative nucleoside transporters (ENTs) or SLC29 family of integral membrane proteins.
15. The method of claim 14, wherein the equilibrative nucleoside transporter (ENT) or a member of the SLC29 family of integral membrane proteins is a transporter for purine and pyrimidine nucleosides and nucleobases or a metabolite thereof.
16. The method of claim 15, wherein the transporter for purine and pyrimidine nucleosides and nucleobases or a metabolite thereof is an equilibrative nucleoside transporter ENT2.
17. The method of claim 1, wherein the purified DNA is single-stranded, double-stranded, triple-stranded, or four-stranded or a combination thereof.
26. The method of claim 1, wherein introducing the purified DNA near or around the live cells at the site of interest is or comprises administration of the purified DNA at the site of interest by injection, microinjection, microprojectile or implantation or a combination thereof, or alternatively, implantation of an object or substance containing the purified DNA or artificial DNA, wherein the implant releases the purified DNA or artificial DNA in a single burst or over an extended period.
27. A method for inhibiting a tumor associated with ischemia, cellular/tissue necrosis or cellular/tissue apoptosis by selective targeting of live cells at a site of interest by the method of claim 1.
28. The method of claim 27, wherein the cellular/tissue necrosis or cellular/tissue apoptosis is associated with an ischemia, a chemical burn, a fire, radiation, hypothermia, freezing, a physical trauma, hypoxia, a poison, a chemotherapeutic agent, a cell damaging agent, a tumor or a cancer.
29. A method for selective targeting of live cells at a site of interest, wherein the site of interest has undergone or is undergoing radiation or chemotherapy, with a cell-penetrating polypeptide, the method comprising:
(a) administering purified DNA to the site of interest in an extracellular space;
(b) contacting a live cell at the site of interest with a cell-penetrating polypeptide comprising cell-penetrating determinants so that the cell-penetrating polypeptide binds the purified DNA or its degradation product so as to form a complex;
(c) contacting one of the live cells at the site of interest with the complex in (b) so as to bind and penetrate the live cell;
(d) permitting additional complexes to form as in b and contacting additional cells with said complexes so as to bind and penetrate additional live cells at the site of interest;
wherein the cell penetrating polypeptide is a 3E10 antibody or a fragment thereof, wherein the fragment thereof is a single chain Fv (scFv) fragment of mAb 3E10 antibody, and
thereby, selectively targeting live cells at the site of interest, which has undergone or is undergoing radiation or chemotherapy, with a cell-penetrating polypeptide.
30. The method of claim 29, wherein the cell-penetrating polypeptide is conjugated to a therapeutic agent.
31. A method for inhibiting cellular injury in a subject, the method comprising:
(a) administering directly to the live cells at or near a site of cellular injury of the subject a cell-penetrating polypeptide comprising cell-penetrating determinants joined to a therapeutic agent;
(b) contacting purified DNA with the cell-penetrating polypeptide so that the cell-penetrating polypeptide binds the purified DNA or its degradation product near or around the live cells so as to form a complex;
(c) contacting one of the live cells with the complex in (b) so as to bind and penetrate the live cell; and
(d) permitting additional complexes to form as in (b) and contacting additional cells with said complexes so as to bind and penetrate additional live cells at the site of cellular injury;
wherein the cell penetrating polypeptide is a 3E10 antibody or a fragment thereof, wherein the fragment thereof is a single chain Fv (scFv) fragment of mAb 3E10 antibody,
thereby, inhibiting cellular injury in the subject.
32. The method of claim 31, wherein introducing the purified DNA near or around the live cells at the site of interest is or comprises administration of the purified DNA at the site of interest by injection, microinjection, microprojectile or implantation or a combination thereof, or alternatively, implantation of an object or substance containing the purified DNA or artificial DNA, wherein the implant releases the purified DNA or artificial DNA in a single burst or over an extended period.
33. A method for inducing cell death in a subject, the method comprising:
(a) administering directly to the live cells at or near a site of injury of the subject a cell-penetrating polypeptide comprising cell-penetrating determinants joined to a therapeutic agent;
(b) contacting purified DNA near or around the live cells with the cell-penetrating polypeptide so that the cell-penetrating polypeptide binds the purified DNA or its degradation product near or around the live cells so as to form a complex;
(c) contacting one of the live cells with the complex in (b) so as to bind and penetrate the live cell which induces cell death; and
(d) permitting additional complexes to form as in (b) and contacting additional cells with said complexes so as to bind and penetrate additional live cells inducing additional cell death at the site of injury;
wherein the cell penetrating polypeptide is a 3E10 antibody or a fragment thereof, wherein the fragment thereof is a single chain Fv (scFv) fragment of mAb 3E10 antibody,
thereby inducing cell death at or near a site of injury with the cell-penetrating polypeptide.
34. The method of claim 33, wherein introducing the purified DNA near or around the live cells at the site of interest is or comprises administration of the purified DNA at the site of interest by injection, microinjection, microprojectile or implantation or a combination thereof, or alternatively, implantation of an object or substance containing the purified DNA or artificial DNA, wherein the implant releases the purified DNA or artificial DNA in a single burst or over an extended period.
35. A method for selective targeting of live cells at or near a site of cellular injury with a cell-penetrating polypeptide which comprises cell-penetrating determinants joined to a therapeutic agent, the method comprising:
(a) contacting the live cells with a composition comprising
(i) a cell-penetrating polypeptide which comprises cell-penetrating determinants joined to a therapeutic agent and
(ii) purified DNA so that the cell-penetrating polypeptide binds the purified DNA or its degradation product near or around the live cells so as to form a complex such that the complex so formed binds one of the live cells and penetrates the live cell; and
(b) permitting additional complexes to form as in (a) and contacting additional cells with said complexes so as to bind and penetrate additional live cells at the site of cellular injury,
wherein the cell penetrating polypeptide is a 3E10 antibody or a fragment thereof, wherein the fragment thereof is a single chain Fv (scFv) fragment of mAb 3E10 antibody,
thereby selectively targeting live cells at a site of cellular injury with a cell-penetrating polypeptide.
36. The method of claim 35, wherein introducing the purified DNA near or around the live cells at the site of interest is or comprises administration of the purified DNA at the site of interest by injection, microinjection, microprojectile or implantation or a combination thereof, or alternatively, implantation of an object or substance containing the purified DNA or artificial DNA, wherein the implant releases the purified DNA or artificial DNA in a single burst or over an extended period.
37. A method for enhancing the effects of chemotherapy or radiation therapy by selectively targeting live cells by the method of claim 1.
38. The method of claim 37, wherein introducing the purified DNA near or around the live cells at the site of interest is or comprises administration of the purified DNA at the site of interest by injection, microinjection, microprojectile or implantation or a combination thereof, or alternatively, implantation of an object or substance containing the purified DNA or artificial DNA, wherein the implant releases the purified DNA or artificial DNA in a single burst or over an extended period.
39. A method for increasing uptake and enhancing efficacy of a cell-penetrating polypeptide comprising an anti-DNA antibody or a fragment in targeting tumor or cancer cells, the method comprising:
(a) introducing purified DNA or artificial DNA at or near the tumor or cancer cells;
(b) administering the cell-penetrating polypeptide;
(c) allowing the cell-penetrating polypeptide to form additional complexes with the additional purified DNA or artificial DNA; and
(d) permitting the additional complexes in (c) to contact the tumor or cancer cells,
wherein the cell penetrating polypeptide is a 3E10 antibody or a fragment thereof, wherein the fragment thereof is a single chain Fv (scFv) fragment of mAb 3E10 antibody,
thereby increasing uptake and enhancing efficacy of a cell-penetrating polypeptide comprising an anti-DNA antibody or a fragment or a variant thereof in targeting tumor or cancer cells.
40. The method of claim 39, wherein introducing the purified DNA or artificial DNA near or around the live cells at the site of interest is or comprises administration of the purified DNA or artificial DNA at the site of interest by injection, microinjection, microprojectile or implantation or a combination thereof, or alternatively, implantation of an object or substance containing the purified DNA or artificial DNA, wherein the implant releases the purified DNA or artificial DNA in a single burst or over an extended period.
The US Patent does not recite that the purified DNA is purified sheared extracellular DNA.
However, the US Patent discloses that purified sheared extracellular DNA is encompassed by purified DNA (col. 8, lines 8-22):
In accordance with the practice of the invention, “extracellular DNA” is DNA free of a cell or cell-free DNA. Extracellular DNA may be introduced or administered by methods known in the art such as, for example, microinjection of DNA into extracellular space or outside of a cell or cells, cell or cells in a tissue, or cell or cells in an organ, so long as the DNA is not introduced or administered into a cell. The delivered DNA in the extracellular space or outside of a cell may be in any physical state, including in a solution, as a solid, as a colloidal form, as a semi-crystalline state, as a nanoparticle or combination thereof. The delivered DNA may be isolated DNA from nature, either total, fractionated, intact or sheared, or synthesized by the hand of man such as through synthetic chemistry or in vitro enzymatic methods as known in the art.
The US Patent also discloses that purified DNA encompasses DNA purified away from non-DNA nucleic acid, nucleotide, nucleoside or purine or pyrimidine base (col. 16, lines 15-17).
In NSDP rejection, “while the earlier patent’s specification cannot be used as prior art to the later patent application claims, it can be consulted to ascertain the meaning of the words in the claims to interpret the patent claim’s proper scope (Sun Pharmaceutical Industries Ltd v. Eli Lilly and Co., 611 F.3d 1381, 1387 (Fed. Cir. 2010)”.
“A patent’s disclosure may be used to determine whether an application claim is merely an obvious variation of an invention claimed in a patent”In Re Basell Poliolefine Italia S.P.A., 547 F.3d 1371, 1378 (Fed. Cir. 2008) and Sun Pharmaceutical Industries Ltd v. Eli Lilly and Co., 611 F.3d 1381, 1387 (Fed. Cir. 2010)”.
In the instant case, the specification of the US Patent discloses that purified sheared extracellular DNA is encompassed by purified DNA. Therefore, purified sheared extracellular DNA is encompassed by and obviated by the claimed purified DNA in the method of the US Patent claims. The instantly claimed purified sheared extracellular DNA is not patentably distinct from the US Patent’s purified DNA.
Response to Arguments
9. Applicants state they do not agree with the double patenting rejections over U.S. Patent No. 10,383,945 and U.S. Patent No. 11,453,725.
10. The arguments have been considered but are not persuasive because no specific reasons for traversal have been presented. The claim amendments are obviated by the US Patent claims for the reasons stated above.
11. Claims 97-100, 103, 105-109, and 116 remain rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1, 2, 4-8 of U.S. Patent No. 7,189,396, in view of WO 1997032602, Weisbart; QIAGEN® Plasmid Purification Handbook, second edition, August 2003; and Yoo et al (Journal of Biophysical Chemistry, 2011, 2:102-110); as evidenced by US Patent Application Publication 2008/0292618, Weisbart. Although the claims at issue are not identical, they are not patentably distinct from each other because the US Patent is claiming methods of delivering complexes/conjugates of 3E10 antibody or an scFv fragment of 3E10 bound to genetic material to target live cells that overlap with the instant claims, rendering the instant claims obvious.
The US Patent claims:
1. A method for transporting a biologically active molecule into the nucleus of a target cell in vitro, comprising:
a) combining said biologically active molecule in vitro with monoclonal antibody (mAb) 3E10 as produced by a hybridoma having ATCC accession number PTA 2439 or an scFv fragment thereof, to form an antibody-biologically active molecule conjugate or fusion protein, and
b) contacting the target cell in vitro under suitable conditions with the antibody-biologically active molecule conjugate of a),
thereby transporting the biologically active molecule into the nucleus of the target cell.
2. The method of claim 1 wherein said biologically active molecule is a nuclear transcription factor, an enzyme, an enzyme inhibitor, genetic material, an inorganic or organic molecule, a pharmaceutical agent, a drug, or a polypeptide.
4. The method of claim 1, wherein the antibody is the scFv fragment.
6. The method of claim 1, wherein the contacting comprises incubating the cells in the presence of the conjugate or a fusion protein.
7. The method of claim 1, wherein the scFv fragment comprises the variable region of the heavy chain (VH) and variable region of the kappa light chain (Vκ) of mAb 3E10.
8. The method of claim 7, wherein the scFv fragment further comprises the signal peptide of the Vκ.
As evidenced by US 2008/0292618, Weisbart, the 3E10 antibody is transported into cells via the nucleoside salvage pathway through ENT2-mediated nucleoside salvage pathway (Example 1). Therefore, the method claimed by the US Patent delivering 3E10 complexes into cells necessarily depends on ENT2-mediated nucleoside salvage pathway.
The US Patent does not claim the genetic material is DNA purified away from non-DNA nucleic acid, nucleotide, and nucleoside purine and pyrimidine bases and comprises sheared DNA.
WO 1997032602, Weisbart, teaches complexing 3E10 scFv antibody to biologically active material that is genetic material including DNA, or DNA plasmids containing a gene of interest, for delivery into live cells, as set forth above.
Qiagen sells commercially available kits for rapidly isolating and purifying plasmid DNA genetic material, particularly for cell transfection purposes. Qiagen teaches the process for purification of plasmids in the figure on p. 12, which involves binding, wash, and elution steps utilizing QIAGEN Anion-Exchange Resin. Qiagen teaches the Anion-Exchange Resin purifies the plasmid DNA away from RNA, proteins, dyes, and low-molecular-weight impurities (i.e. nucleotides and nucleosides) (p. 13):
Principle and procedure
QIAGEN plasmid purification protocols are based on a modified alkaline lysis procedure, followed by binding of plasmid DNA to QIAGEN Anion-Exchange Resin under appropriate low-salt and pH conditions. RNA, proteins, dyes, and low-molecular-weight impurities are removed by a medium-salt wash. Plasmid DNA is eluted in a high-salt buffer and then concentrated and desalted by isopropanol precipitation.
Page 70:
DNA binding and washing on the QIAGEN-tip
The cleared lysate is loaded onto a pre-equilibrated QIAGEN-tip by gravity flow. The salt and pH conditions of the lysate and the superior selectivity of the QIAGEN Resin
ensure that only plasmid DNA binds, while degraded RNA, cellular proteins, and
metabolites are not retained and appear in the flow-through fraction. The QIAGEN-tip is then washed with medium-salt buffer (Buffer QC) which completely removes any remaining contaminants, such as traces of RNA and protein (e.g., RNase A), without affecting the binding of the plasmid DNA (see Figure 4 on page 61). Buffer QC also disrupts nonspecific interactions, and allows removal of nucleic acid-binding proteins without the use of phenol. The low concentration of alcohol in the wash buffer eliminates nonspecific hydrophobic interactions, further enhancing the purity of the bound DNA. The plasmid DNA is then efficiently eluted from the QIAGEN-tip with high salt buffer (Buffer QF or QN). For further information about QIAGEN Anion-Exchange Resin, see Appendix E: QIAGEN Anion-Exchange Resin, pages 76–78.
Qiagen teaches protocols for processing extracellular DNA for purification that requires pipetting, mixing, shaking, centrifuging, and additional manipulations (p. 16-51).
Yoo teaches demonstrates that sheared DNA is commonly present in DNA samples resulting from basic laboratory handling and processing, including pipetting, vortexing, rocking, freeze-thawing, ultrasonication and ultrafiltration (abstract; sections 2.3 and 3.2; Figures 2-7).
It would have been prima facie obvious to one of ordinary skill in the art at the time the invention was filed to utilize purified extracellular DNA or DNA plasmid containing genes of interest away from non-DNA nucleic acid, nucleotide, and nucleoside purine and pyrimidine bases and ocmprise sheared DNA in the method of the US Patent. One would have been motivated to, and have a reasonable expectation of success to because: (1) the US Patent claims complexing genetic material with 3E10 antibody for cell delivery; (2) Weisbart teaches the genetic material can be DNA or DNA plasmids containing a gene of interest; (3) Qiagen teaches and demonstrates isolation and purification of DNA or DNA plasmid containing genes of interest away from non-DNA nucleic acid, nucleotide, and nucleoside purine and pyrimidine bases for use in cell delivery is routine and commercially available and discloses routine protocols for doing so including manipulations by pipetting, vortexing, centrifuging, rocking, and such; and (4) Yoo demonstrates that sheared DNA is a common by-product present in DNA samples due to routine laboratory manipulations including pipetting, vortexing, rocking, freeze-thawing, and physical agitations.
Response to Arguments
12. Applicants state they do not agree with the double patenting rejection over US Patent 7,189,396.
13. The arguments have been considered but are not persuasive because no specific reasons for traversal have been presented. The claim amendments are obviated by the US Patent claims and secondary references for the reasons stated above.
14. Conclusion: No claim is allowed.
Conclusion
15. 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.
16. Any inquiry concerning this communication or earlier communications from the examiner should be directed to LAURA B GODDARD whose telephone number is (571)272-8788. The examiner can normally be reached Mon-Fri, 7am-3:30pm.
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/Laura B Goddard/Primary Examiner, Art Unit 1642
