Ex Parte Woods et alDownload PDFPatent Trial and Appeal BoardSep 28, 201713148289 (P.T.A.B. Sep. 28, 2017) Copy Citation United States Patent and Trademark Office UNITED STATES DEPARTMENT OF COMMERCE United States Patent and Trademark Office Address: COMMISSIONER FOR PATENTS P.O.Box 1450 Alexandria, Virginia 22313-1450 www.uspto.gov APPLICATION NO. FILING DATE FIRST NAMED INVENTOR ATTORNEY DOCKET NO. CONFIRMATION NO. 13/148,289 10/28/2011 Robert J. Woods 235.01890101 1082 26813 7590 10/02/2017 MUETING, RAASCH & GEBHARDT, P.A. P.O. BOX 581336 MINNEAPOLIS, MN 55458-1336 EXAMINER PROUTY, REBECCA E ART UNIT PAPER NUMBER 1652 NOTIFICATION DATE DELIVERY MODE 10/02/2017 ELECTRONIC Please find below and/or attached an Office communication concerning this application or proceeding. The time period for reply, if any, is set in the attached communication. Notice of the Office communication was sent electronically on above-indicated "Notification Date" to the following e-mail address(es): ptodocketing @ mrgs .com PTOL-90A (Rev. 04/07) UNITED STATES PATENT AND TRADEMARK OFFICE BEFORE THE PATENT TRIAL AND APPEAL BOARD Ex parte ROBERT J. WOODS, LORETTA YANG, and KAUSARN. SAMLI1 Appeal 2017-001673 Application 13/148,289 Technology Center 1600 Before JOHN G. NEW, TAWEN CHANG, and DEVON ZASTROW NEWMAN, Administrative Patent Judges. NEW, Administrative Patent Judge. 1 Appellants state that the real party-in-interest is the University of Georgia Research Foundation, Inc. App. Br. 3. Appeal 2017-001673 Application 13/148,289 DECISION ON APPEAL Appellants file this appeal under 35 U.S.C. § 134(a) from the Examiner’s Final Rejection of Appellants’ claims 51—53, 55—62, 65—69, 72, and 73 as unpatentable under 35 U.S.C. § 103(a) as being obvious over the combination of A. Jokilammi et al., Construction of Antibody Mimics from a Noncatalytic Enzyme-detection of Polysialic Acid, 295 J. Immunol. Meth. 149-60 (2004) (“Jokilammi”)), K. Karaveg and K.W. Moremen, Energetics of Substrate Binding and Catalysis by Class 1 (Glycosylhydrolase Family 47) a-Mannosidases Involved in N-Glycan Processing and Endoplasmic Reticulum Quality Control, 280(33) J. Biol. Chem. 29837-48 (2005) (“Karaveg”), W.M. Patrick and A.E. Firth, Strategies and Computational Tools for Improving Randomized Protein Libraries, 22 Biolmol. Eng. 105— 12 (2005) (“Patrick”), N.H. Barakat and J.J. Love, Molecular Diversity in Engineered Protein Libraries, 11 Curr. Opin. Chem. Biol. 335—41 (2007) (“Barakat”), and M.L. DeMarco et al. R.J. Woods, Structural Glycobiology: A Game of Snakes and Ladders, 18(6) Glycobiol. 426-40 (2008) (“DeMarco”). Claim 69 also stands rejected as unpatentable under 35 U.S.C. § 103(a) as being obvious over Jokilammi, Karaveg, Patrick, Barakat, DeMarco, and P. Kuhn et al. Active Site and Oligosaccharide Recognition Residues of Peptide-N4-(N-acetyl-f-D-glucosaminyl)asparagine Amidase F, 270(49) J. Biol. Chem. 29493-97 (1995) (“Kuhn”). We have jurisdiction under 35 U.S.C. § 6(b). We REVERSE. 2 Appeal 2017-001673 Application 13/148,289 NATURE OF THE CLAIMED INVENTION Appellants’ invention is directed to “lectenz” molecules, which are mutated carbohydrate processing enzymes that are catalytically inactive and that have had their substrate affinity increased by at least 1.2 fold. Abstract. REPRESENTATIVE CLAIM Claim 51 is representative of the claims on appeal and recites: 51. A method for generating a lectenz comprising an inactivated mutated carbohydrate-processing enzyme having enhanced affinity for its substrate compared to a corresponding wild-type carbohydrate-processing enzyme, the method comprising: (a) providing a 3D structure of an enzyme-substrate complex comprising a substrate bound to a catalytically inactive mutant carbohydrate-processing enzyme or a corresponding wild-type carbohydrate-processing enzyme, wherein the catalytically inactive carbohydrate-processing enzyme comprises an amino acid sequence comprising at least one inactivating mutation that eliminates catalytic activity of the enzyme; (b) performing a computational simulation on the enzyme- substrate complex to predict the per-residue contributions to total interaction energy (AEMm) and/or total binding free energy (AGBinding) for amino acid residues of the enzyme; (c) analyzing the per-residue energetic contributions to identify at least one amino acid residue as a potential mutation site for enhancing binding affinity of the inactivated enzyme for its substrate as compared to a wild-type enzyme, wherein an amino acid residue is identified as a potential mutation site for enhancing binding activity when o (i) for an amino acid residue located within 5 A of the substrate in the enzyme-substrate complex, the per- 3 Appeal 2017-001673 Application 13/148,289 residue contribution of the amino acid residue to at least one of AEmmot AGBinding is > -0.7 kcal/mol; and o (ii) for an amino acid residue located more than 5 A from the substrate in the enzyme-substrate complex, the per-residue contribution of the amino acid residue to at least one of is AEMm or AGBinding > 0.0 kcal/mol; (d) expressing a library of inactivated mutated carbohydrate-processing enzymes, each inactivated mutated enzyme comprising a plurality of amino acid mutations, wherein each inactivated mutated enzyme comprises at least one inactivating mutation that eliminates catalytic activity of the enzyme; and each inactivated mutated enzyme further independently comprises at least one potential affinity- enhancing mutation at a site identified in step (c); (e) assaying the inactivated mutated enzymes of step (d) for their ability to form the enzyme-substrate complex; and (f) identifying inactivated mutated enzymes from step (e) that exhibit binding affinities to the substrate that are at least 1.2- fold greater than those of the wild-type carbohydrate-processing enzyme. Claims App’x Br. 1—2.2 Issue Appellants argue that the Examiner erred by failing to identify any teaching or suggestion in the cited prior art of an inactivated, affmity- 2 The pages of Appellants’ Claims Appendix are not numbered. We therefore provisionally number them seriatim, beginning with “1” as the first page of the Appendix. 4 Appeal 2017-001673 Application 13/148,289 enhancing enzyme that has separate affinity-enhancing and inactivating mutations. App. Br. 14. Analysis The Examiner finds Jokilammi teaches, in relevant part, a catalytically-inactive enzyme that retains the substrate binding affinity of the wild type enzyme can be used as an antibody mimic. Non-Final Act. 5 (February 19, 2015). The Examiner finds Jokilammi also teaches that the advantages of such antibody mimics include ease of preparation, the “lack of animal use,” and the lack of cross-reactions with antibody binding proteins, as well as the ability to provide probes to any molecule that is poorly immunogenic. Id. at 5—6. Furthermore, the Examiner finds that, because the use of mutagenesis for the engineering of antibodies to increase their binding affinities to the antigen is well known in the art, it would therefore have been obvious to one of ordinary skill to use mutagenesis for the engineering of antibody mimics to increase their binding affinities to the substrate. Id. at 6. The Examiner finds Karaveg teaches methods of generating a mutant mannosidase that is catalytically inactive and has higher binding affinity for its substrate. Non-Final Act. 6. The Examiner finds Karaveg teaches: (1) using the 3-D X-ray structure of a co-complex of the mannosidase and an inhibitor to identify one or more amino acid residues that are hypothesized to be involved in catalysis and/or substrate binding; (2) mutating these residues; (3) screening the mutants produced for the catalytic activity and the binding affinity to the substrate; and (4) selecting a mutant that lacks substantial catalytic activity but has increased binding 5 Appeal 2017-001673 Application 13/148,289 affinity. Id. The Examiner acknowledges that the methods of Karavag differ from those claimed in that they do not specifically start with a catalytically-inactive enzyme and screen for mutations that further increase binding affinity and do not include a step of performing an in silico computational screening of the residues hypothesized to be involved in binding and/or catalysis prior to screening for a mutant with increased binding affinity. Id. Appellants argue that the Examiner failed to identify any teaching or suggestion of an enzyme that includes an inactivating mutation and a separate affinity-enhancing mutation. App. Br. 14. Appellants take issue with the Examiner’s assertions that “there is no requirement that the motivation or reason to modify the prior art be explicitly taught in the art itself’ and that, because Jokilammi analogizes catalytically-inactive enzymes to antibodies, “a skilled artisan would have found it obvious to use known antibody engineering techniques on the catalytically inactive enzyme which retains [the] substrate binding affinity of Jokilammi.” App. Br. 14 (quoting Final Act. 5—6). In response, Appellants argue, first, that the Examiner failed to specifically identify the “known antibody engineering techniques” that a skilled artisan would have found it obvious to use. App. Br. 14. Second, Appellants argue that, because the Examiner failed to clearly articulate a specific technique or types of techniques, or the effects of those techniques, the Examiner failed to provide the “explicit” analysis and “articulated reasoning” required by KSR Int’l Co. v. Teleflex Inc., 550 U.S. 398, 418 (2007) to show that the claimed combination was obvious. Id. at 14—15. 6 Appeal 2017-001673 Application 13/148,289 Appellants argue further that, even if an artisan was motivated to use a known antibody engineering technique on an enzyme, the Examiner does not articulate why it would have been obvious to select an enzyme having an inactivating mutation and a separate affinity enhancing mutation, as required by the claims. App. Br. 15. Additionally, Appellants contend, even if a skilled artisan would have found it obvious to use known antibody engineering techniques on the catalytically inactive enzyme, the results of such techniques would not have been predictable. Id. The Examiner acknowledges that the rejection does not provide an express teaching by any of the references to modify an enzyme having an inactivating mutation to also and independently have a separate affinity enhancing mutation. Ans. 10. However, the Examiner finds that the teachings of Jokilammi, which explicitly analogizes a catalytically-inactive enzyme that retains substrate binding affinity to an antibody, combined with the knowledge of an ordinary artisan that engineering antibodies by mutagenesis to increase their binding affinities to the antigen is common in the art, provide a suggestion to modify an enzyme having an inactivating mutation to also have a separate affinity enhancing mutation. Ans. 10. The Examiner finds that a skilled artisan would therefore have found it obvious to use known antibody engineering techniques on the catalytically-inactive enzyme that retains substrate binding affinity, as taught by Jokilammi on similar catalytically-inactive enzymes, which retain substrate binding affinity, as taught by Karaveg. Id. With respect to Appellants’ argument that the Examiner failed to specifically identify the “known antibody engineering techniques” that a skilled artisan would have found it obvious to use, the Examiner concludes 7 Appeal 2017-001673 Application 13/148,289 that it would have been obvious to one of ordinary skill to use mutagenesis for the engineering of antibody mimics to increase their binding affinities to a substrate. App. Br. 14—15. The Examiner further concludes that it would have been obvious to one of ordinary skill in the art to use a directed evolution strategy (on the catalytically-inactive mutant of Karaveg) to produce mutants having further increased binding affinity. Id. at 11. Appellants reply that Jokilammi teaches that a catalytically inactive enzyme is functionally similar to an antibody, but not structurally similar, and the Examiner did not articulate how known antibody engineering techniques — which depend on the structure of the antibody — could be successfully applied to a catalytically inactive enzyme. Reply Br. 3. Appellants argue that, because of the structural differences between antibodies and enzymes, a skilled artisan would not have been motivated to use the antibody engineering techniques identified by the Examiner. Id. Appellants contend further that Karaveg teaches making several single-site mannosidase mutants, and characterizing their roles in catalysis and substrate binding. Reply Br. 6 (citing Karaveg Table I). Appellants contend that Karaveg teaches that per-residue contributions to total binding energy were evaluated for sugar residues within the oligosaccharide substrate bound to mutant enzyme E330Q, but there are no data shown for the substrate binding contributions of individual amino acid residues. Id. (citing Karaveg 29844; Fig. 3; 29845; Tables IV and V). We find that Appellants have the better position. Claim 51 expressly requires, inter alia, that: “[E]ach inactivated mutated enzyme comprises at least one inactivating mutation that eliminates catalytic activity of the enzyme; and each inactivated-mutated enzyme further independently 8 Appeal 2017-001673 Application 13/148,289 comprises at least one potential affinity-enhancing mutation at a site identified in step (c)” (emphasis added). Jokilammi teaches “[a]n engineered noncatalytic bacteriophage-encoded endosialidase [enzyme that] binds specifically to polySia but does not degrade it and can thus be used as the equivalent of an immunohistological stain,” and which is fused to green fluorescent protein (“GFO”). Jokilammi, Abstr., 150. Jokilammi thus teaches a polysialic-binding enzyme that is inactivated, in that it binds to its substrate but does not catalyze the substrate, but is not binding affinity- enhanced. Karaveg teaches mutants of glycosylhadrase and, in particular, mutants containing a single-site mutation at the -1 subsite (E330Q), that “ha[ve] minimal catalysis under the conditions . . . routinely use[d] for SPR binding studies (10°C, pH 7.0), yet. . . retain[] high affinity glycan binding with reduced on- and off-rates.” Karaveg 29846. Furthermore, Karaveg teaches that “[mjutations in the -1 subsite or conditions that significantly reduce dMNJ binding affinity or catalysis either have a minimal effect on Man9GlcNAc2 glycan binding (F659A mutant or Ca2 + depletion) or actually increase glycan binding affinity (T688A and E330Q mutants).” Id. (emphases added). Karaveg thus teaches a single-site mutation that decreases catalysis and increases the binding affinity as the result of a single site (-1) subsite mutation. But neither Jokilammi nor Karaveg, upon which the Examiner relies, teaches an enzyme with a mutation that eliminates catalytic activity of the enzyme and further independently comprises at least one potential affinity- enhancing mutation, as required by claim 51. In other words, claim 51 requires that the enzyme contain a mutation that eliminates the catalytic 9 Appeal 2017-001673 Application 13/148,289 function of the enzyme and another, independent, mutation that enhances the affinity of the enzyme for the substrate. Jokilammi does not teach an affinity-enhancing mutation. Karaveg teaches a single-site mutation that both enhances affinity and decreases catalysis. Neither references teaches mutations that independently perform each required function. Nor are we persuaded by the Examiner’s position that, because Jokilammi functionally analogizes its fusion protein to an antibody, that a person of ordinary skill in the art would understand that the mutagenesis techniques employed in enhancing antibody binding affinity can be equally applied to enzymatic binding affinity. As Appellants explain in detail in their Reply Brief, the complex structural and functional differences between antibodies and enzymes is such that the mutagenic mechanisms for altering binding affinity in the former cannot be obviously employed to alter binding affinity in the latter. See Reply Br. 3—5. In the Answer, the Examiner adduces two additional pieces of prior art, A. Rajpul et al., A General Method for Greatly Improving the Affinity of Antibodies by Using Combinatorial Libraries, 102(24) Proc. Nat. Acad. Sci. 8466—71 (2005) (“Rajpul”) and P.S. Chowdhury and I. Pastan, Improving Antibody Affinity by Mimicking Somatic Hypermutation in vitro, 17 Nature Biotech. 568—72 (1999) (“Chowdury”). Both references are directed to means of enhancing binding affinity in antibodies via mutagenesis, and thus speak to the knowledge of a person of ordinary skill in that particular field. However, neither reference speaks to the state of the art of altering enzyme-substrate binding affinity, or to whether techniques directed to altering antibody binding affinity can be successfully directed to altering or increasing the binding affinity of enzymes. 10 Appeal 2017-001673 Application 13/148,289 We therefore conclude that the Examiner has failed to establish a prima facie case of obviousness because: (1) the references fail to teach or suggest all of the limitations of the claims; and (2) the Examiner has not adequately demonstrated that the knowledge of a person of ordinary skill in the art of enzyme chemistry would be sufficient to cure the deficiencies of the cited references. We consequently reverse the Examiner’s rejection of the claims. Because we find this issue to be dispositive of the rejections, we do not reach Appellants’ additional arguments. DECISION The Examiner’s rejection of claims 51—53, 55—62, 65—69, 72, and 73 as unpatentable under 35 U.S.C. § 103(a) is reversed. REVERSED 11 Copy with citationCopy as parenthetical citation