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 .
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.
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(s) 1, 2, 5, 6, 9, 11-12, 15-17, 21-23, 25, 28-29, 44, 66, and 106 is/are rejected under 35 U.S.C. 103 as being unpatentable over Abdelaal et al. (Int. J. Mol. Sci. 2019, 20, 5165) in view of Dahotre et al. (Anal. Chem. 2019, 91, 2695−2700 and Supplemental Methods) and Black et al (Nature Protocols; Vol. 16, August 2021; pages 3802-3825).
Abdelaal et al. teaches detection of antigen specific T cells using in situ MHC tetramer staining.
The method comprises steps of contacting a tissue sample with an antigen and a detection complex wherein the detection complex comprises an antigen presenting molecule (APM) multimer that binds to the antigen to form an antigen loaded detection complex. See Figure 1, “MHC Class I tetramer.”
The reference further teaches that the antigen-loaded detection complex binds to a receptor on or in a cell at a location in the biological tissue sample. See Figure 1, “CD8+T Cell.”
The reference teaches detecting a signal from the bound antigen-loaded complex. See Figure 1.
With regard to claim 2, Abdelaal et al. teaches the antigen bound to the MHC Class I tetramer prior to contacting the tetramer with the tissue. See Figure 1.
With regard to claim 5, the antigen-loaded detection complex taught by Abdelaal is a tetramer of the antigen-APM monomer. See Figure 1.
With regard to claim 6, Abdelaal teaches a peptide antigen. See Figure 1, “viral peptide,” for example.
With regard to claim 9, Abdelaal teaches a Class I MHC molecule which inherently comprises an alpha subunit and a Beta2 subunit.
With regard to claims 11-12 and 15-16, the reference teaches the antigen loaded detection complex comprises a streptavidin scaffold that has four MHC molecules each linked to biotin that binds the streptavidin. With regard to claim 16, each of the four MHC molecules is bound to one molecule of the antigen. See Figure 1 and Figure 1 description.
Regarding claim 21, the cell is a T cell, and regarding claim 22 the receptor is a T cell receptor. See title.
Regarding claim 29, Abdelaal teaches labeling fresh tissue. See Figure 1.
Abdelaal et al. does not teach a method wherein the antigen-loaded detection complex is conjugated to a reporter oligonucleotide. Nor does Abdelaal et al. teach contacting the tissue sample with a probe or probe set.
With regard to claim 17, Abdelaal teaches that tetramers consist of four MHC monomers loaded with a specific peptide to interact with T cells specific to that peptide. Abdelaal does not teach the size of the peptides.
With regard to claim 38, Abdelaal does not teach fixing the cells after adding the MHC tetramer.
With regard to claim 44, Abdelaal does not teach a tetramer that is not fluorescently labeled.
Regarding claim 106, Abdelaal teaches that using a dextran scaffold allows more MHC moleucles to bind targets which helps increase the detection of antigen-specific T cells. See page 7 of 11.
Dahotre et al. teach MHC multimers for the detection of antigen specific T cells. In the method taught by Dahotre et al. the MHC multimer is labeled with a DNA barcode, See Figure 1.
With regard to claim 2, Dahotre et al. also teach loading the MHC prior to contacting with the sample (see Supplemental methods, p. 3).
With regard to claim 17, Dahotre et al. exemplifies antigens that are 9-11 amino acids in length.
Regarding claim 23, the reporter oligonucleotide is conjugated to the antigen via the scaffold of the tetramer.
Regarding claim 25, the reporter oligonucleotide is covalently conjugated to a scaffold of the detection complex.
Regarding claim 28, the reporter oligonucleotide comprise a barcode region for identifying the antigen and the barcode region comprises one or more barcode sequence, see Supplemental Table S1.
Further, Black et al. teach a method for multiplexed tissue imaging with DNA conjugated antibodies. Like Abdelaal, the Black et al. teach tissue imaging, that is detecting targets in the context of the tissue sample itself. Like Dahotre, the method employs a barcoded target binding molecule. Black et al. teach contacting the tissue sample with a detection complex, wherein the detection complex is conjugated to a reporter oligonucleotide followed by contacting the tissue sample with a probe or probe set that hybridizes to the reporter oligonucleotide and detecting a signal associated with the probe or probe set or a product thereof at the location in the tissue sample.
Regarding claim 38, Black teaches fixing the tissue sample after contacting the detection reagent and before contacting the probes. See Figure 5, “Post-staining antibody fixation.”
Regarding claim 44, Black teaches that the detection reagent is not fluorescently labeled.
Regarding claim 66, the detecting in c) is by sequential hybridization. See Figure 2. Further, the detecting in c) taught by Black does not comprise contacting the tissue smale with an antibody that binds to the detection complex.
It would have been obvious to have modified the in situ method taught by Abdelaal by using the barcoded MHC tetramer taught by Dahtore et al. in an in situ method for detecting antigen specific T cells. One would have been motivated to use the barcoded detection reagent for in situ detection by Black et al. who demonstrates an analogous method for using a barcoded detection agent for in situ detection. Black teaches that immunohistochemistry and fluorescence detection, like that used by Abdelaal, have limitations with regard to the ability to multiplex, where the method that uses barcoded binding molecules could be scaled for the quantification of up to 60 markers in a single tissue sample (p. 3802). Therefore, one would have been motivated to use the barcoded MHC tetramer taught by Dahotre and the detection technique taught by Black instead of the staining combined with immunohistochemistry taught by Abdelaal in order to increase the multiplexing ability of the method taught by Abdelaal et al.
Further, with regard to claim 17, it would have been obvious to have used appropriately sized antigens, such as those demonstrated by Dahotre et al. that are in the size range claimed. One would have been motivated to do so by the exemplification of Dahotre et al. that antigens of this size are effective for detecting target T cells via binding to T cell receptors.
Regarding claim 38, it would have been obvious to have included the fixing step taught by Black et al. into the method for detecting T cells using barcoded MHC tetramers. This would have been obvious because Black teaches a fixation step prior to rounds of hybridizing and stripping probes. The fixing of the detection reagent into place would prevent the reagent from being stripped during the sequential hybridization process.
Regarding claim 44, when modifying the teachings of Abdelaal to practice an in situ method with a barcoded MHC tetramer, analogous to that taught by Black where the detection reagent is an antibody, it would have been obvious to provide an MHC multimer that was not fluorescently labeled since the method taught by Black relies on fluorescently labeling the oligonucleotide barcodes for identification of the antigen and the fluorescent labeling of the detection reagent directly would have provided background signal.
Regarding claim 11, it would have been obvious to have employed a dextran scaffold in the method T cell staining because using dextran allows more MHC molecules to bind a target, as taught by Abdelaal which helps increase the signal of antigen specific T cells. See Abdelaal page 7 of 11.
Thus, the claimed invention is prima facie obvious.
Response to Remarks
The remarks are persuasive; the prior art references do not teach detecting a signal at a location in a tissue sample as recited in instant claim 1.
Applicant further points out that claims 2, 15-17, 22, 28, 38, 44 and 46 were not addressed in the prior Office action. These claims are addressed herein.
Conclusion
The prior art made of record and not relied upon is considered pertinent to applicant's disclosure. Liu et al. teaches high spatial resolution multi-omics by barcoding in tissue. The reference deploying oligonucleotide conjugated detection reagents followed by hybridization and imaging to detect the successful labeling with the barcodes. See Figure 1.
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Juliet Switzer
Primary Examiner
Art Unit 1682
/JULIET C SWITZER/Primary Examiner, Art Unit 1682
