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 .
Status of the Application
Claims 1-13 and 15-21 are pending and under examination
Claims 14 and 22-28 are canceled
The following Office Action is in response to Applicant's communication dated 06/01/2026.
Applicant's arguments filed 06/01/2026 have been fully considered but they are not persuasive.
RE: Applicant argued that Liu and Picelli do not teach the alleged bidirectional FFPE-compatible fusion-detection invention. Applicant contends that Liu uses single directional workflow that can fail with fragmented RNA, while Picelli is directed to full length, oligo(dT) single cell transcript analysis, hence neither or in combination of references teach or suggest bidirectional fusion genes in degraded clinical samples.
In response: As presently drafted, claim 1 is much broader and not commensurate with the invention in Applicant’s argument. Applicant appears to rely on a specific two pools bidirectional primer architecture as the inventive distinction, in which template switching and random priming place different universal sequences at opposite ends of cDNA, followed by first PCR in two separated primer pools configure with different primer orientations to detect 5’ and 3’ fusion event. For illustration, Examiner citing the similar technological architecture from SMARTer™ RACE cDNA Amplification (Published June 2010, attached as NPL) below.
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However, claim 1 does not clearly recite those structural and bidirectional relationships. Instead, the claim broadly recites separate primer pools and a “first pair of primers” comprising a gene specific primer and a universal primer, without requiring that the respective pools contain different primer sets, different universal primers, different gene specific primer orientations, or primers that hybridize to opposite ends of the cDNA. Accordingly, under broadest reasonable interpretation, the limitation is broad enough to encompass separate or replicate pools using the same generic primer pair arrangement rather than specific reciprocal four primer arrangements shown in the disclosed workflow. While Examiner understand that claims are often drafted broadly, the claim should still reflect the overall inventive structure and workflow that Applicant is relying on to distinguish the prior art. Although the claims are interpreted in light of the specification, limitations from the specification are not read into the claims. See In re Van Geuns, 988 F.2d 1181, 26 USPQ2d 1057 (Fed. Cir. 1993). In this instant, if Applicant intends to rely on the disclosed bidirectional two-pool setup to distinguish the prior art, the claim need to expressly define the composition and orientation of each PCR pool and its relationship to 5’ and 3’ fusion detection.
New Claim Rejections - 35 USC § 112(b)-Necessitated by Amendment
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claim(s) 1-13 and 15-21 rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Claim 1 recites “the first pair of primers is used in separate primer pools configured for detection of a 5' fusion gene and a 3' fusion gene, respectively” wherein the “first pair of primers” comprises “a gene specific primer and a universal primer”. However, it is unclear what primers are required to be present in each of the separate primer pools. The claim does not clearly specify whether the same gene specific primer/universal primer pair is used in both primer pools, whether the pools are merely replicate pools, or whether each pool must contain a different primer set arranged for a different fusion detection orientation. Since one of ordinary skill in the art cannot determine with reasonable certainty the scope of protection, the metes and bounds of the claim are unascertainable.
For purpose of examination, claim 1 is interpreted as reciting replicate separate primer pools using the same first pair of primers because step (c) recites “a first pair of primers” and defines that pair as comprising “a gene specific primer” and “a universal primer”, thereby reasonably indicating only two primers rather than two distinct primer pairs (i.e. 4 primers) for respective pools. Accordingly, the prior art rejections are maintained because claim 1 is examined base on the limitations actually recited, rather than the more specific workflow Applicant appears to rely upon from the specification and drawings. See In re Van Geuns, 988 F.2d 1181, 26 USPQ2d 1057 (Fed. Cir. 1993).
Claim(s) 2-13 and 15-21 depend from claim 1 and is therefore similarly rejected.
Modified Claim Rejections - 35 USC § 103 - Necessitated by Amendments
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
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.
Liu et. al. and Picelli et al.
Claim(s) 1, 2, 7, 9-12, 13, and 18-21 is/are rejected under 35 U.S.C. 103 as being unpatentable over Liu et al. (US20210180051Al, Filed Dec 31st , 2018, of record) in view of Picelli et al. (Nat Protoc 9, 171-181 (2014) , of record)
Regarding claims 1, Liu discloses a method for detecting a gene alteration in a biological sample, comprising steps of: (a) mixing a target RNA obtained from the biological sample, a template-switching oligo, a reverse transcriptase, and a reverse transcription (RT) primer in a mixture, wherein the template
switch oligo comprises a first universal primer sequence; (b) subjecting the mixture to a condition under
which reverse transcription occurs to provide a target DNA complementary to the target RNA, (c)
amplifying the target DNA with a first pair of primers to obtain a first PCR product, wherein the first pair of primers is used in separate primer pools configured for detection of a 5' fusion gene and a 3' fusion gene, respectively, wherein the first pair of primers comprise a gene specific primer and a universal primer, the gene specific primer comprises a 5'-end first or second adapter sequence and a 3'-end sequence that hybridizes to a target gene under stringent conditions, the universal primer comprises a 5'-end first or second adapter sequence and a 3'-end sequence comprising the first or second universal primer sequence, and the first and second adapter sequences are different from each other; (e.g. method generates cDNA from RNA using one universal primer at 5' end and one target specific primer at 3' end. A universal adapter is added on the 3' end of the newly synthesized cDNA by template switching while the RNA is converted into cDNA from random hexamers in reverse transcription. The resulting cDNA serves as template for target amplification in multiplex PCR with a universal primer and a panel of target specific primers. The resulting cDNA serves as template for target amplification in multiplex PCR with a universal primer and a panel of target specific primers. [paragraph 0026]. Molecular barcodes are random nucleotides and can be as long as 3 to 50 nucleotides, or even longer. They are usually synthesized as a part of the primer or adapter" [paragraph 0026]. Reference further discloses that PCR reactions ).
(d) amplifying the first PCR product with a second pair of primers to obtain a second PCR product, wherein each of the second pair of primers comprises a 5'-end third or fourth adapter sequence and a 3'-end sequence that hybridizes to the first or second adaptor sequence, respectively, and the third and fourth adapter sequences are different from each other; and (e) analyzing the second PCR product to detect the presence of the gene alteration (e.g. The amplification products can be further amplified while sample barcodes and sequencing adapters are added. The final library can then be used in downstream analysis, such as NGS sequencing.[paragraph 0014]. Further, fig. 3 shows cDNA in a multiplex PCR reactions.)
However, Liu does not disclose the RT primer comprises a second universal primer sequence;
Picelli discloses initiating reverse transcription using oligo(T) primer that includes a 5' universal
adapter sequence, thereby incorporating adapter into cDNA during first-strand synthesis. The
incorporated adapter sequence subsequently serves as universal priming site for PCR amplification of
cDNA [Fig l].
As of the application' s effective filing date, one of ordinary skill in the art would have had a
reasonable expectation of success and motivated to combine these teachings to incorporate the RT primer design of Picelli into Liu's method in order to introduce reverse transcription , thereby simplifying
downstream amplification and enabling efficient PCR amplification and sequencing of the resulting cDNA products. Additionally, a one of ordinary skill in the art would have recognize that the oligo(T) portion of the RT primer in Picelli's protocol could be replaced with target specific sequence taught by Liu, depending on the desired transcript capture strategy. Substitution of primer binding regions to achieve desirable cDNA sequence was well known in the art of reverse transcription and represent routine design choice. Accordingly, a POSIT A would have reasonably expected success in modifying Pi cell i's primer to use in Liu's method and simplifying downstream cDNA amplification. See KSR International Co. v. Teleflex Inc., 550 U.S. 398, 415-421, USPQ2d 1385, 1395 - 97 (2007) (see MPEP § 2143, A and B).
Regarding claim 7, Picelli further discloses second universal primer is less than 30 bp in length. (e.g. T7 primer as shown in Fig 1 is 25 nucleotides long)
Regarding claim 9, Liu further discloses at least one of the first adapter sequence or the second adapter sequence comprises a barcode sequence.[paragraph 0023]
Regarding claim 10-12, Liu further discloses target DNA is at least 100 bp/ 100 bp to 4000 bp/ 100 bp to 500 bp in length. (e.g. target on DNA fragments of 160 bp in length [paragraph 0015]).
Regarding claim 13, Liu further discloses the target DNA is amplified by multiplex PCR with at least two gene specific primers (e.g. "A plurality of target-specific primers (e.g., >6, >10, >100, >1000, >10,000, etc.) can be used."[paragraph 0014]).
Regarding claim 18, Liu further discloses a gene fusion comprising a sequence of a known gene selected from the group consisting of ABLl, AKT3, ALK, ARV7, BCR, BRAF, CD74, EGFR, ERBB2, ERBB4, ERG, ESRl, ETVl, ETV4, ETV5, ETV6, EZR, FGFRl, FGFR2, FGFR3, KIT, KMT2A, MET, NRGl, NRG2, NTRKl, NTRK2, NTRK3, NUTMl, PDGFRA, PDGFRB, PIK3CA,RAF1, RARA, RET, ROSl, RSPO2, SDC4, SLC34A2 and TMPRSS2. [table 2]
Regarding claim 19, Liu further discloses the biological sample is from a solid tumor. [paragraph 0005]
Regarding claim 20, Liu further discloses the biological sample is a Formalin-Fixed Paraffin-Embedded (FFPE) tissue sample [paragraph 0017].
Regarding claim 21, Liu further discloses the second PCR product is analyzed by a next generation sequencing [paragraph 0024].
Liu et al., Picelli et al., and Iafrate et al.
Claim(s) 3-6, and 8 is/are rejected under 35 U.S.C. 103 as being unpatentable over Liu
et al. (US20210180051Al, Filed Dec 31st 2018) in view of Picelli et al. (Nat Protoc 9, 171-181 (2014)), and
Iafrate et al. (US10450597B2, Flied Jan 26th 2015)
Regarding claim 3, Liu does not disclose the RT primer comprises a stem-loop structure and an overhang structure having at least 5 random nucleotides.
Iafrate discloses the RT primer comprises a stem-loop structure [Fig. 3 and column 12, lines 65-67] and an overhang structure having at least 5 random nucleotides (e.g. molecular barcodes can be built into primers (e.g., RT primers, target-specific primers, extension sequence primers) such that each individual molecule produced by a primer obtains a unique barcode tag. [column 12, lines 42-49]. The reference also disclosed a variety of random nucleotides lengths [column 17, lines 3-27]).
As of the application' s effective filing date, one of ordinary skill in the art would have had a
reasonable expectation of success and motivated to modify the method of Liu by incorporating the stem loop RT primer with barcode taught by Iafrate as both references are directed to sequencing-base detection of genetic alterations using reverse transcription, barcoding, PCR amplification, and address the common goal of improving molecular identification accuracy and downstream sequencing performance. Iafrate teaches that RT primers comprise stem-loop structures and random nucleotide barcodes, Liu similarly teaches the use of random nucleotide barcode sequence in RT primers. Stem-loop RT primers were well-known in the art at the time of the invention as a design choice that provides improved priming specificity, structural stability, and efficient incorporation of barcode and adapter sequences. One of ordinary skill in the art would have recognized that substituting the RT primer of Liu with a stem-loop RT primer represents the use of a known alternative primer structure to perform the same function in the same manner. This reasoning is consistent with KSR International Co. v. Teleflex Inc., 550 U.S. 398, 415-421, USPQ2d 1385, 1395 - 97 (2007) (see MPEP § 2143, B).
Regarding claim 4, Iafrate further discloses the stem-loop structure is a hairpin stem-loop structure or a Y shape stem-loop structure. [Fig. 3].
Regarding claim 5, Iafrate further discloses the stem-loop structure comprises a barcode sequence. (e.g. primer complementary common regions base pair to form the stem of a hairpin having a loop portion that comprises the molecular barcode.[column 17, lines 33-40]).
Regarding claim 6, Iafrate further discloses the stem-loop structure is at least the length of the second universal primer sequence. (e.g. primer complementary common regions base pair to form the stem of a hairpin having a loop portion that comprises the molecular barcode. The loop and stem collectively having selectable lengths spanning at least 6-25 nucleotides or more. Because the applicant's universal primers are 7-23 nucleotides, the lengths disclosed in Iafrate meets the claimed length requirements [column 17, lines 3-54]).
Regarding claim 8, Iafrate further discloses the stem of the stem-loop structure is 8 bp in length. (e.g. primers comprise stem-loop (hairpin) structure formed by two complementary common sequences, wherein the stem length is selectable within the disclosed ranges including 6-12 nucleotides. As set forth in MPEP 2144.05; In re Peterson, 315 F.3d 1325 (Fed. Cir. 2003); "[A] prior art reference that discloses a range encompassing a somewhat narrower claimed range is sufficient to establish a prima facie case of obviousness.").
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.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to Khai Quynh Tien Pham whose telephone number is (571)272-6998. The examiner can normally be reached M-T, 9-4 ET.
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If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Heather Calamita can be reached at (571) 272-2876. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
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/KHAI QUYNH TIEN PHAM/Examiner, Art Unit 1684
/JEREMY C FLINDERS/Primary Examiner, Art Unit 1684
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