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 Status
Claims 1-6, 7-14, 23, and 32-36 are pending and under examination. Claims 1 and 35 are independent claims. Claim 31 has been canceled.
Response to Arguments
Rejections Withdrawn
The rejection of Claims 1-5, 7-11, 32, 35, and 36 under 35 U.S.C. 102(a)(2) as being clearly anticipated by Peterson et al. has been withdrawn following the applicants’ amendments.
The rejection of claims 1-5, 7-14, 23, and 31-35 under 35 U.S.C. 103 as being unpatentable over Peterson, as applied to claims 1-5, 7-11, 32, 35, and 36 above, in view of Gunderson et al. is withdrawn following the applicants’ amendments.
New Rejections
Claim Rejections - 35 USC § 103
The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
Claims 1-6, 7-14, 23, and 32-36 are rejected under 35 U.S.C. 103 as being unpatentable over Peterson et al. (US 2018/0208975 Al, published July 26, 2018) in view of Stoeckius et al. (US 2018/0251825 Al, published Sep. 6, 2018) and further in view of Gunderson (US 2018/0273933 Al, published September 27, 2018).
Peterson teaches methods for simultaneous detection of multiple analytes from the same biological sample, including protein analytes using antibody-oligonucleotide conjugates and RNA analytes using sequencing-based detection. Peterson discloses that the antibody-conjugated oligonucleotides include identifying sequences that permit differentiation and quantification of bound protein targets following amplification and sequencing, and further teaches sequencing-based analysis of nucleic acids derived from RNA obtained from the same sample.
In regards to claims 1 and 2, Peterson teaches biochemical assays for simultaneous genomic and proteomic analysis. Peterson teaches a method for detecting an analyte in which a two proximity probes attach to a target analyte in a sample, wherein each proximity probe comprises an analyte binding domain and an oligonucleotide domain. Wherein each proximity probe comprises a universal primer region (for amplification), a probe specific barcode, and a complementary sequence for interacting with corresponding paired probe (inter-molecular reacting region; see Figs. 1-4, para. [0006], However, Peterson does not expressly disclose that each oligonucleotide includes a unique molecular identifier (UMI). However, Stoeckius, who is also in the field of analyte detection oligo-conjugated antibodies, teaches antibody-oligonucleotide conjugates and related oligonucleotide constructs in which each oligonucleotide includes both an identifier sequence and a unique molecular identifier to enable digital counting of individual molecular events and to reduce amplification bias inherent in sequence-based assays (see Figs. 1, 2, and 6, para.[0005],[0022],[0064],[0066]-[0067]).
It would have been prima facie obvious to one of ordinary skill in the art at the time of filing to utilize a unique molecular identifier in each oligonucleotide as taught by Stoeckius with the proximity ligation-based assay taught by Peterson. A person of ordinary sill in the art would have been motivated to modify the oligonucleotide-antibody constructs of Peterson to include UMIs as taught by Stoeckius in order to improve quantitative accuracy, distinguish original binding events from PCR duplicates, and enhance the reliability of sequencing-based protein detection. Such a modification would have been straightforward and predictable, as Peterson already employs sequencing and amplification workflows compatible with UMI-containing oligonucleotides, and Stoeckius teaches the use of UMIs for precisely this purpose. The combination merely applies a known techniques for improving molecular counting to a similar system performing the same type of sequencing-based analysis, with no teaching away in either reference. Accordingly the claimed invention would have been obvious to a person of ordinary skill in the art at the time of the invention.
Subsequently, this also reads on claims 35, and 36 which outline a composition of these components.
In regards to claims 3 and 5, Peterson teaches utilizing antibodies, aptamers, or ligands as the analyte binding domain for binding small molecules of interest (Fig. 1-4, [0027]).
In regards to claim 4, Peterson teaches that the oligonucleotide domains may be conjugated to the analyte binding domains of the probes via a multiple different means including chemical bonds, photocleavable linkers, click chemistry among others (see [0022]- [0027]).
In regards to claim 7-11, Peterson teaches using a proximity extension or proximity ligation assays to generate a third a third oligonucleotide, which has an adapter sequence attached (Fig. 2 – 3, [0011] – [0012], [0029], and throughout).
In regards to claims 12 and 33, Peterson and Stoeckius teach the limitations of claim 1 for which claim 12 depends. Peterson teaches preparing libraries of cDNA from the sample, comprising PCR based tagging methods and performing reverse transcription of RNA molecules in the presence of a RNA tag. Wherein the RNA tag contains an identifier, a UMI, and a poly(T) sequence (see [0030]-[0035]). While Peterson teaches that their methods may be used with aptamers and antibodies towards other small molecules, they do not specifically teach detecting DNA with their methods (see [0027]). Stoeckius teaches generating cDNA and DNA libraries from the samples (see [0015], [0135], [0402]-[0404], [0429]-[0430], and throughout), though they utilizing different methods of placing tags. However, neither Stoeckius nor Peterson teach ligating the tags onto the DNA or RNA molecules or purifying the nucleic acids from the sample.
Like Peterson and Stoeckius, Gunderson also teaches methods and compositions for analyzing cellular components, including methods in which they utilize proximity ligation and proximity elongation assays to detect analytes (see [0049], [0119]-[0120]). One skilled in the art would recognize that this process utilizes probes that each consist of an analyte binding domain and an oligonucleotide domain and would further recognize that for each PEA or PLA the oligonucleotide region would contain a region that interacts with the other oligonucleotide on the corresponding probe (intermolecular reacting region). Gunderson incorporates a variety of reporter moieties into their oligonucleotide tags, including any arrangement of tags that allow them to determine the composition, identity, and/or source of the analyte being investigated and teaches the use of multiple reporter moieties simultaneously for detecting multiple individual analytes in a sample ([0045] – [0048]) Gunderson teaches methods of detecting any nucleic acid of interest, including but not limited to RNA and DNA in the same sample (Fig. 5, [0066]-[0068]), and that the nucleic acids may be obtained from any source, and prepared, purified and/or enriched before analysis ([0069]-[0071]). Gunderson further teaches ligating a barcodes or amplification tags to polynucleotides to prepare them for detection or amplification (Fig. 1, [0082]-[0083]).
It would have been prima facie obvious to one of ordinary skill in the art at the time of filing to utilize the ligation methods taught by Gunderson with a tag containing an identifier, UMI, and poly(T) as taught by Peterson and Stoeckius. Stoeckius teaches that the inclusion of a UMI in a tag permits distinguishing original molecules from amplification duplicates and thereby improves accuracy of downstream quantification. Gunderson teaches ligating oligonucleotides to target nucleic acids to generate tagged products suitable for subsequent amplification and sequencing/readout. A person of ordinary skill in the art would have been motivated to incorporate the UMI containing tag using Gunderson’s ligation workflow because ligation-based tagging followed by amplification is susceptible to PCR duplication and related quantification artifacts, and UMIs were a known solution to mitigate those artifacts. The combination would have been a simple substitution o one known tag design for another in known tagging context., yielding the predictable results off producing ligated products that retain UMI information for improved molecule counting/quantification.
In regards to claim 13, Peterson teaches binding RNA with a second probe comprising a RNA specific tag, and UMI and template switching oligonucleotide (see [0019]).
In regards to claim 14 and 23, Peterson teaches amplifying tagged cDNA (see [0032]) or RNA ([0033]) for enrichment utilizing gene specific primers.
In regards to claim 32, Peterson teaches producing a library of DNA corresponding to the unique protein or RNA tags used ([0031]-[0033], [0037]-[0038]).
Conclusion
No claim is allowed.
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/MATTHEW HAROLD RAYMONDA/Examiner, Art Unit 1684
/AARON A PRIEST/Primary Examiner, Art Unit 1681