DETAILED ACTION
Notice of Pre-AIA or AIA Status
1. The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
2. Applicant's arguments presented in the reply of 09 February 2026 have been fully considered but do not place the application in condition for allowance. All rejections not reiterated herein are hereby withdrawn. In particular, the previous rejection of the claims under 35 U.S.C. 112(a) has been obviated by the amendment to cancel claims 9-11, 13 and 20-22 and the amendment to claims 16, 18, and 19.
Claim Status
3. Claims 1-3, 5-8, 12, 14, 16, 18, 19 and 23-25 are pending.
Claims 7 and 8 are withdrawn from further consideration pursuant to 37 CFR 1.142(b), as being drawn to a nonelected species, there being no allowable generic or linking claim. Note that Applicant elected, without traverse, the species of the microbial macromolecule of DNA in the reply filed on 11 November 2024.
Claims 1-3, 5-6, 12, 14, 16, 18, 19, and 23-25 on the elected invention and have been examined herein.
Claim Objections
4. Claims 5 and 6 are objected to because of the following informalities:
Claims 5 and 6 are objected to because they depend from canceled claim 4, whereas claim 5 should depend from claim 1. Appropriate correction is required.
Modified Claim Rejections - 35 USC § 103
5. 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.
Claim(s) 1-3, 5-6, 12, 14, 18-19, and 23 is/are rejected under 35 U.S.C. 103 as being unpatentable over Kim (U.S. 20200354795), in view of in view of Skog et (20100196426; cited in the IDS of 06/25/2025) al and Schorey et al (U.S. 20170253916).
Claim(s) 1-3, 5-6, 12, 14, 18-19, and 23 is/are rejected under 35 U.S.C. 103 as being unpatentable over Kim (WO 2018124618, English translation included), in view of Skog et (20100196426; cited in the IDS of 06/25/2025) al and Schorey et al (U.S. 20170253916).
Note that U.S. 20200354795 is the National Stage application of PCT/KR2017/015174, which published as WO 2018124618. Citations below are with respect to U.S. 20200354795.
Kim teaches a method comprising: (a) obtaining a body fluid sample (e.g., blood) from a subject; (b) isolating an exosome / extracellular vesicle fraction from the body fluid sample; (c) “extracting DNA from extracellular vesicles isolated from a subject sample” (para [0012]); and (d) detecting a bacteria-derived (and archaea-derived) nucleic acids – i.e., 16S rRNA gene sequences - present in the exosome / extracellular vesicle fraction (e.g., para [0008-0009], [0011-0018]), [054-0055], Example 4 beginning at para [0057]).
Kim (para [0006]) states:
“Bacteria coexisting in human bodies secrete nanometer-sized vesicles to exchange information about genes, proteins, and the like with other cells. The mucous membranes form a physical barrier membrane that does not allow particles with the size of 200 nm or more to pass therethrough, and thus bacteria symbiotically living in the mucous membranes are unable to pass therethrough, but bacteria-derived extracellular vesicles have a size of approximately 100 nm or less and thus relatively freely pass through the mucous membranes and are absorbed into the human body.”
Kim (para [0029]) also states:
“Extracellular vesicles secreted from bacteria existing in the environment are absorbed into the human body, and thus may directly affect the occurrence of cancer, and it is difficult to diagnose pancreatic cancer early before the onset of symptoms so that efficient treatment thereof is difficult. Thus, according to the present invention, the causative factor and risk of pancreatic cancer can be diagnosed through metagenomic analysis of bacteria-derived extracellular vesicles using a human body-derived sample.”
Kim teaches that extracellular vesicles are isolated from blood by first generating a serum sample from a blood sample by centrifuging the blood sample at 3,500 x g at 4οC for 10 minute. Subsequently, the supernatant is filtered through a 0.22 um filter to remove “bacteria and impurities,” and resulting solution was subjected to ultra-high speed centrifugation at 150,000×g at 4ο C for 3 hours (e.g., para [0042-0043] and [0054])
In particular, Kim (para [0054]) states:
“To isolate extracellular vesicles and extract DNA, from blood, first, blood was added to a 10 ml tube and centrifuged at 3,500×g and 4 □ for 10 min to precipitate a suspension, and only a supernatant was collected, which was then placed in a new 10 ml tube. The collected supernatant was filtered using a 0.22 μm filter to remove bacteria and impurities, and then placed in centrifugal filters (50 kD) and centrifuged at 1500×g and 4 □ for 15 min to discard materials with a smaller size than 50 kD, and then concentrated to 10 ml. Once again, bacteria and impurities were removed therefrom using a 0.22 μm filter, and then the resulting concentrate was subjected to ultra-high speed centrifugation at 150,000×g and 4 □ for 3 hours by using a Type 90ti rotor to remove a supernatant, and the agglomerated pellet was dissolved with phosphate-buffered saline (PBS), thereby obtaining vesicles.”
In the absence of evidence to the contrary, the method of Kim for isolating extracellular vesicles per se from blood is considered to be a method that results in the isolation of a mixture of mammalian and bacterial exosomes. Note that Kim makes clear that the methods disclosed therein are for performing “metagenomic analysis of bacteria-derived vesicles isolated from a human-derived substance, such as blood” (para [0007]) and Kim teaches that the method is one for isolating extracellular vesicles per se (para [0038], and [0054]).
Kim does not teach treating the exosome fraction containing mammalian and bacterial exosomes with DNAse prior to detecting the microbial DNA.
However, Schorey et al teaches methods of detecting Mycobacterium nucleic acids in exosomes present in a bodily fluid sample of a mammalian subject (e.g., para [0010], [0013] and para [0055-0058]). Schorey teaches that the method comprises: obtaining a bodily fluid sample from a patient; (b) isolating an exosomes fraction of the bodily fluid sample; (c) treating the exosomes fraction of (b) with DNAse; and (d) detecting microbial RNA present in the exosomes fraction (e.g., para [0010-0011], [0013], [0016], [0055-0058], [0105] and [0123]). Schorey et al states “Exosomes were sequentially treated with RNase A and DNase I to remove any RNA or DNA not present within the exosome (i.e. attached to outside surface)” (para [0123]).
Further, Skog et al teaches methods of detecting DNA present in exosomes. Skog teaches that prior to extracting DNA from exosomes, the exosomes are treated with DNase (e.g., para [0178]). Skog states “The DNase treatment step was to remove DNA outside of the exosomes so that only DNA residing inside the exosomes was extracted” (para [0178]). At para [00223], Skog teaches that approximately 20% of the DNA isolated from exosomes not treated with DNase does not originate from inside the exosome.
In view of the teachings of Schorey et al and Skog et al, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of Kim so as to have treated the exosomes with DNase prior to lysing the exosomes so as to have removed any DNA present outside of the exosome, including DNA attached to the surface of the exosome. One would have been motivated to have done so for the advantages set forth by Schorey and Skog of avoiding contamination of the DNA isolated from within the exosome with DNA from outside of the exosome, thereby ensuring the accuracy of the method of detecting bacterial DNA from inside bacterial exosomes.
Regarding claim 12, Kim teaches methods wherein blood samples are obtained from a healthy subject (i.e., a normal control subject; see, e.g., para [0014], [0018-0022] [0044], [0033]; Example 4 beginning at para [0057] and Figure 4). For instance, Kim (para [0044] states:
Regarding claim 14, Kim teaches that the extracellular vesicles / exosomes are obtained from blood samples (e.g., para [0008], [0028], Example 2 beginning at para [0054], and Example 4 beginning at para [0057]).
Regarding claims 18 and 19, Kim discloses that the 16S rRNA gene sequences are analyzed to identify the bacteria from which the extracellular vesicles / exosomes are derived, determining if there is an increase or decrease in the quantity of particular bacteria-derived extracellular vesicles / exosomes, as compared to a normal control sample and as compared to control samples from subjects having colon cancer or having colon polyps, and diagnosing the subject as having pancreatic cancer based on the detection of the increase or decrease in the quantity of the particular bacteria-derived extracellular vesicles / exosomes (e.g., para [0015], [0038-0040] and Example 4 beginning at para [0057]). Kim identified bacteria belonging to particular phylum, orders, families and genera that were increased in extracellular vesicles isolated from serum samples from patients having pancreatic cancer as compared to serum samples healthy patients (see Example 4, beginning at para [0057])..
Thus, Kim teaches detecting a microbial signature that is indicative of the disease of pancreatic cancer.
Kim does not specifically recite that the patient diagnosed with an increased risk of pancreatic cancer is administered an anti-cancer therapy.
However, Kim (para [0041]) does teach:
“The method of the present invention may be used to delay the onset of pancreatic cancer through special and appropriate care for a specific patient, which is a patient having a high risk for pancreatic cancer or prevent the onset of pancreatic cancer. In addition, the method may be clinically used to determine treatment by selecting the most appropriate treatment method through early diagnosis of pancreatic cancer.”
It is further stated (para [0029]):
“according to the present invention, the causative factor and risk of pancreatic cancer can be diagnosed through metagenomic analysis of bacteria-derived extracellular vesicles using a human body-derived sample, and thus a risk group of pancreatic cancer can be diagnosed early, thereby delaying the onset of pancreatic cancer or preventing pancreatic cancer through appropriate management, and even after pancreatic cancer occurs, early diagnosis for pancreatic cancer can be implemented, thereby lowering the incidence of pancreatic cancer and increasing therapeutic effects..”
In view of these teachings of Kim, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of Kim so as to have included a step of treating the patient diagnosed with risk of pancreatic cancer by administering an anti-cancer agent in order to have provided an early intervention to prevent the further onset of pancreatic cancer or ameliorate one or more symptoms or outcomes of pancreatic cancer.
Regarding claim 23, Kim teaches that the subjects from which the blood samples containing the extracellular vesicles / exosomes are obtained are human subjects / individuals (e.g., para [0057] and [0065]).
6. Claim(s) 16, and 24-25 is/are rejected under 35 U.S.C. 103 as being unpatentable over Kim (U.S. 20200354795), in view of in view of Skog et (20100196426; cited in the IDS of 06/25/2025) al and Schorey et al (U.S. 20170253916), and further in view of Kalluri et al (U.S. 20170059572; cited in the IDS).
Claim(s) 16 and 24-25 is/are rejected under 35 U.S.C. 103 as being unpatentable over Kim (WO 2018124618, English translation included), in view of Skog et (20100196426; cited in the IDS of 06/25/2025) al and Schorey et al (U.S. 20170253916), and further in view of Kalluri et al (U.S. 20170059572; cited in the IDS).
Note that U.S. 20200354795 is the National Stage application of PCT/KR2017/015174, which published as WO 2018124618. Citations below are with respect to U.S. 20200354795.
The teachings of Kim (U.S. 20200354795 and WO 2018124618), Skog et al and Schorey et al are presented above.
As discussed above, Kim teaches a method of diagnosing pancreatic cancer in a subject based on the detection of the bacterial signature in the extracellular vesicles / exosomes isolated from a blood sample of the subject.
Regarding claim 16, Kim does not teach that the method is one that further comprises isolating glypican 1-containing exosomes from the exosome fraction.
However, Kalluri teaches “a method of isolating cancer cell-derived exosomes comprising (a) obtaining a body fluid sample from a cancer patient; (b) isolating an exosomes fraction of the body fluid sample; and (c) isolating exosomes comprising glypican 1 from the exosomes fraction, thereby isolating cancer cell-derived exosomes” (para [0026]). It is disclosed that glypican 1 (GPC1) is a surface marker present on exosomes derived from cancer cells but not normal cells (para [0207]). It is further disclosed in Example 6 (beginning at para [0210]) that GPC1-positive exosomes are a biomarker for the presence of cancer.
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of Kim so as to have further included a step of isolating glypican 1-containing exosomes from the exosome fraction, as taught by Kalluri. One would have been motivated to have done so because Kalluri teaches that GPC1-positive exosomes are a biomarker for the presence of cancer. Thereby, the ordinary artisan would have recognized that modification of the method of Kim as set forth above would have provided the benefit of confirming the presence of colon cancer in the subjects in which GPC1-positive exosomes were detected.
Regarding claims 24 and 25, Kim does not teach repeating the steps of obtaining the body fluid sample, isolating exosomes / extracellular vesicles from the body fluid sample, and detecting the 16S rDNA in the isolated exosomes / extracellular vesicles (claim 24), particularly at least one day, week or month after the initial obtaining, isolating and detecting steps (claim 25).
However, Kim (para [0006]) does state:
“A microbiota or microbiome is a microbial community that includes bacteria, archaea, and eukaryotes present in a given habitat. The intestinal microbiota is known to play a vital role in human's physiological phenomena and significantly affect human health and diseases through interactions with human cells. Bacteria coexisting in human bodies secrete nanometer-sized vesicles to exchange information about genes, proteins, and the like with other cells..”
Kim also teaches monitoring “the distribution pattern of bacteria and extracellular vesicles over time after intestinal bacteria and bacteria-derived extracellular vesicles (EVs) were orally administered to mice” (see para [0030] and Figure 1).
Thus, Kim teaches that there may be changes in the microorganisms present in patients over time and these changes may be associated with the development of pancreatic cancer. Also, as discussed above, the teachings of Kim suggest treating patients diagnosed as at risk of colon cancer.
Further, Kalluri teaches monitoring therapy by assaying for changes in exosomes over time in order to assess the progression of disease and therapeutic efficacy (e.g., para [0030], [0073] and [0087]). Kalluri (para [0076]) states:
“analysis of a subject's cancer cell-derived exosomes number and characterization may be made over a particular time course in various intervals to assess a subject's progression and pathology. For example, analysis may be performed at regular intervals such as one day, two days, three days, one week, two weeks, one month, two months, three months, six months, or one year, in order to track the level and characterization of cancer cell-derived exosomes as a function of time. In the case of existing cancer patients, this provides a useful indication of the progression of the disease.”
In view of the teachings of Kalluri of monitoring changes in exosomes over time, including at weekly or monthly intervals, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have obtained a second blood sample from the subject at a later time point, including conventional time points of at least a week or month after the first time point, and to have isolated exosomes / extracellular vesicles from the second body fluid and assayed have isolated exosomes / extracellular vesicles from the second body fluid to identify the bacteria 16S rDNA sequences present therein in order to have determined if there was a change in the patient’s risk of pancreatic cancer or prognosis of pancreatic cancer or to aid in monitoring the efficacy of treatment of pancreatic cancer.
Response to remarks regarding the rejections under 35 U.S.C. 103:
The response argues that the rejection is based on inherency and the Office has not met the high standard for inherency. The response states:
“Referencing Applicant's FIG. 1B (reproduced below), Applicant identified microbial DNA in healthy serum-derived exosomes. Serum-derived exosome samples were treated with DNAse prior to DNA extraction to remove any freely circulating nucleic acids. Isolated DNA was PCR amplified with universal primers for the bacterial 16S ribosomal RNA gene. DNA from E. coli was used as a positive control, and DNA from human cell lines Panc-1 and Fibroblasts BJ, as well as blank (no template DNA), were used as negative controls. As illustrated by the FIG. 1B gel analysis of the amplified DNA, not every exosomal DNA sample included both mammalian and bacterial DNA-the second lane, from left to right, of the "Exosomal DNA (healthy serum)" group lacks a PCR amplicon corresponding to detection of bacterial DNA.”
These arguments have been fully considered but are not persuasive. First it is noted that Figure 1B of the present application is described as “another repeat” (see para [0018] of the specification). It appears that Figure 1B may be a repeat experiment using the same samples used in Figure 1A. As shown below, in Figure 1A, all samples of healthy serum contained detectable E. coli DNA. Thus, Applicant’s arguments establish only a lack of reproducibility or unpredictability in detecting bacterial exosome-derived E. coli DNA using the particular healthy serum samples.
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The results presented in Figure 1B of the present application do not establish a lack of inherency or an unpredictability in the inherency of the method of Kim et al which obtains an isolated exosome fraction containing a mixture of mammalian and bacterial exosomes. Kim makes clear that the methods disclosed therein are for performing “metagenomic analysis of bacteria-derived vesicles isolated from a human-derived substance, such as blood” (para [0007]) and Kim teaches that the method is one for isolating extracellular vesicles per se from a blood sample (e.g., para [0038], and [0054]). The method of Kim for isolating the extracellular vesicles from blood is nearly identical to the method disclosed in the present specification for isolating an extracellular vesicle fraction that is characterized as comprising a mixture of mammalian and bacterial exosomes. It is maintained that, in the absence of evidence to the contrary, the method of Kim is one that necessarily has the property of “isolating an exosomes fraction” from the body fluid sample (i.e., blood sample), “wherein the isolated exosomes fraction comprises a mixture of mammalian and bacterial exosomes.”
Note again that the method disclosed in the present specification for isolating the mixture of mammalian and bacterial exosomes is nearly identical to that of Kim. Applicant does not point to any differences in the method disclosed in the specification which distinguish the claimed method for isolating a mixture of mammalian and bacterial exosomes over that of Kim. Any such differences are not recited in the claims since steps (a) and (b) of claim 1 are recited at a very high level of generality: “(a) obtaining a body fluid sample from a patient;(b) isolating an exosomes fraction of the body fluid sample, wherein the isolated exosomes fraction comprises a mixture of mammalian and bacterial exosomes.” If Applicant is asserting that the treatment of the exosome fraction with DNase would result in the loss of detectable bacterial exosomes / DNA derived from bacterial exosomes, no evidence or cogent arguments have been provided to support such a contention. Note also that the rejection relies on the teachings of Schorey et al which establish that treatment of exosomes derived from a bodily fluid sample of a mammalian subject with DNase (and RNase) removes nucleic acids from the outside surface of the exosome but does not interfere with the detection of nucleic acids within the bacterial / microbial exosome.
Importantly, Kim provides examples in which bacterial exosomes were isolated from the blood / serum samples of patients. For instance, Kim (para [0057]) teaches “EVs were isolated from blood samples of 176 pancreatic cancer patients and 271 normal individuals, the two groups matched in age and gender, and then metagenomic sequencing was performed thereon using the method of Example 3.” Kim identified orders, family and genus of bacteria based on the detection of bacterial exosomal DNA that was differentially present in the blood samples of subjects having pancreatic cancer as compared to blood samples from normal control subjects (Figures 4-6). Thus, although Figure 1B of the present application shows that one of the healthy serum samples did not have detectable levels of E. coli exosome-derived DNA, Kim does establish that the methods disclosed therein detect bacterial exosome-derived DNA in blood samples from patients with cancer (as encompassed by claims 1-3, 5, 6, 14, 16, 18, 19, and 23-25) and in blood samples from healthy control subjects (as encompassed by claim 12).
The rejection is maintained for the reasons of record.
Conclusion
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). 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 CARLA J MYERS whose telephone number is (571)272-0747. The examiner can normally be reached M-Th 6:30-5:00 EST.
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/CARLA J MYERS/Primary Examiner, Art Unit 1682
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