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 .
The amendment filed on 06/26/2026 has been entered.
Claim 2, 6, 7, 12, 14, 20, 23, 30, 31, 41, and 53 were amended in the claim set filed on 06/26/2026.
Applicant’s election without traverse of Group I, claims 2, 6, 7, 12, 14, 18, 20, 25-26, 28, 30, 31, 34, 35, 39-41, 53, and 54, drawn to a method of detecting an abnormal level of mature and/or active mitochondrial transcription factor A (TFAM) protein in a sample and species steps of claim 14 in the reply filed on 02/26/2026 is acknowledged.
Claims 12 and 23 are withdrawn drawn to a nonelected Group II and non-elected species.
Claims 55-56 were added in the claim set filed on 06/26/2026. No new matter was added.
Claims 18, 25-26, and 28, were canceled in the claim set filed on 06/26/2026.
Claims 2, 6-7, 14, 20, 30-31, 34-35, 39-41 and 53-56 in the claim set filed on 06/26/2026 are currently under examination.
Response to the Arguments
Objections to the Specification and Claims in the previously mailed non-final have been withdrawn in light of applicants Specification and Claim amendments.
Applicant’s arguments regarding previous rejection(s) of claim(s) 6-7 under 35 U.S.C. 112 (b) have been fully considered and are persuasive. The 35 U.S.C. 112 (b) rejections documented in the previously mailed non-final have been withdrawn in light of applicants claim amendments and arguments on Pg. 9.
Applicant’s arguments regarding previous rejection(s) of claim(s) 7 under 35 U.S.C. 112 (d) have been fully considered and are persuasive. The 35 U.S.C. 112 (d) rejections documented in the previously mailed non-final have been withdrawn in light of applicants claim amendments and arguments on Pg. 10.
As necessitated by amendment, new grounds of rejection are made under 35 U.S.C. 112 as documented below in the 35 U.S.C. 112 rejection in this office action on Pg. 4.
Applicants argument against rejection of claims 18, 25-26 and 28 under 35 U.S.C. 103 are moot as they have been canceled.
As necessitated by amendment, the 35 U.S.C. 103 rejections of claim(s) 2, 6, 14, 18, 20, 25-26, 28, 30-31, 34-35, 39-41, 53, and 54 documented in the previously mailed non-final have been withdrawn in light of applicants claim amendments. However, upon further consideration and search, new grounds of rejection are made as documented below in the 35 U.S.C. 103 rejection in this office action on Pg. 5-23.
The new grounds of rejection for claims 2, 6-7, 14, 20, 30-31, 34-35, 39-41 and 53-56 are documented below in this Final Office Action are necessitated by claim amendments filed on 06/26/2026.
Priority
This application is the National Phase Application of International Application No. PCT/EP2021/082385, filed November 19, 2021, which claims priority to U.S. Provisional Application No. 63/126,703, filed December 17, 2020, and European Patent Application No. 20208932.2, filed November 20, 2020. Accordingly, the priority date of instant set of claims is determined to be November 20, 2020.
Claim Rejections - 35 USC § 112(b)
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.
Claims 2, 6-7, 14, 20, 30-31, 34-35, 39-41 and 53-56 are 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 14 is indefinite over the limitations “wherein the level of mature and/or active TFAM protein corresponds to (i) the amount of mature TFAM protein, (ii) the number of interactions of TFAM protein with TFB2M protein, and/or (iii) the ratio of the amount of mature TFAM protein over the amount of immature TFAM protein” (ln 4-9). It is unclear as to what the metes and bounds of the level of mature and/or active TFAM protein are. Is a “mature TFAM protein” structurally different from “an active TFAM protein”? How is “the number of interaction of TFAM protein with TFB2M protein” quantified? Furthermore, it is unclear if the level of mature and/or active TFAM protein is determined using a single option out of step i, ii, or iii recited in step (a), and if the other two steps not selected remain optional. Claims 2, 6-7, 20, 30-31, 34-35, 39-41 and 53-56 depend on claim 14.
Response to Arguments
Applicant's arguments filed 06/26/2026 (Pg. 10-13) with respect to claims 1-19 have been fully considered but do not apply to the new grounds of rejection.
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.
Claim Interpretations: Regarding claims 2,6,14, 20 ,30-31,34-35,39-40-41 and 53-54, “and/or” reads on any the limitation clause as an individual limitation or in combination with the any other limitation clause listed in the claim.
Claims 2, 6-7, 14, 20 30, 40-41, 53 and 56 is rejected under 35 U.S.C. 103 as being unpatentable over Kraft et al. (“Kraft”; (2019). Peripheral Blood Mononuclear Cells Demonstrate Mitochondrial Damage Clearance During Sepsis. Critical care medicine, 47(5), 651–658.) in view of T. I. Alam et al. (“Alam”; (2003). Human mitochondrial DNA is packaged with TFAM. Nucleic acids research, 31(6), 1640–1645.), Pooreydy et al. (“Pooreydy”; (2019). (2013). Organelle isolation for proteomics: mitochondria from peripheral blood mononuclear cells. Archives of Advances in Biosciences, 4.) and Gunst et al. (“Gunst”; (2013). Insufficient autophagy contributes to mitochondrial dysfunction, organ failure, and adverse outcome in an animal model of critical illness. Critical care medicine, 41(1), 182-194.).
Kraft discloses “Objective: Metabolic derangements in sepsis stem from mitochondrial injury and contribute significantly to organ failure and mortality; however, little is known about mitochondrial recovery in human sepsis. We sought to test markers of mitochondrial injury and recovery (mitochondrial biogenesis) non-invasively in peripheral blood mononuclear cells (PBMCs) from patients with sepsis and correlate serial measurements with clinical outcomes…Patients: Uninfected control patients (n=20) and septic ICU patients (n=37). Interventions: Blood samples were collected once from control patients and serially with clinical data on days 1, 3, and 5 from septic patients. Gene products for HMOX1, NRF1, PPARGC1A, and TFAM, and mitochondrial DNA (mtDNA) ND1 and D-loop were measured by qRT-PCR. Pro-inflammatory cytokines were measured in plasma and neutrophil lysates…Conclusions: Our findings support data that sepsis-induced mitochondrial damage is reversed by activation of mitochondrial biogenesis and that gene transcripts measured non-invasively in PMBCs can serve as novel biomarkers of sepsis recovery.” (Abstract).
Regarding claims 14, 30 and 40-41, Kraft teaches a method wherein “Whole blood was collected once from control subjects and serially from septic subjects on study days 1, 3, and 5. … PBMCs… were isolated by gradient centrifugation. qRT-PCR was performed for key nuclear-encoded genes that regulate transcriptional activation of mitochondrial biogenesis (HMOX1, NRF1, PPARGC1A, and TFAM), and for mitochondrial DNA content” (Pg. 652, Data Collection and Analysis, Para. 1). Kraft teaches a method wherein “We found a statistically significant decrease in levels of all four transcripts at day 1 in the sepsis group compared with controls (P<0.05), further supporting an early sepsis-mediated disruption of mitochondrial quality control” (Pg. 6521, Results, Para.3). Kraft teaches a method wherein “Future work is needed to validate these findings and to develop further understanding of mitochondrial quality control in the restoration of mitochondrial homeostasis in PBMCs and other readily retrievable cell types during sepsis.” (Pg. 657, Conclusions, Para. 1). Thus, Kraft teaches a method comprising (a) detecting an abnormal level of mitochondrial transcription factor A (TFAM) transcript in a blood sample from a patient and (b) determining whether the level in the sample is abnormal, wherein the level in the sample is determined to be abnormal if the level of TFAM transcript is at least about 20% lower than the amount determined for a reference sample, wherein an abnormal level of TFAM transcript in the sample is indicative of the presence of sepsis; and wherein said blood sample comprises at least one PBMC.
However, Kraft does not explicitly teach the limitation (a) measuring the level of mature and/or active TFAM protein in the sample, wherein the level of mature and/or active TFAM protein corresponds to (i) the amount of mature TFAM protein,(ii) the number of interactions of TFAM protein with TFB2M protein, and/or (iii) the ratio of the amount of mature TFAM protein over the amount of immature TFAM protein; and (b) determining whether the level in the sample is abnormal, wherein the level in the sample is determined to be abnormal if the level of mature and/or active TFAM protein is at least about 20%, at least about 50%, or at least about 70% lower than the amount determined for a reference sample, wherein an abnormal level of mature and/or active TFAM protein in the sample is indicative of the presence of sepsis.
Alam discloses “Mitochondrial transcription factor A (TFAM), a member of the high mobility group proteins, is essential for maintenance of mitochondrial DNA (mtDNA). Most TFAM and mtDNA (both of which are normally soluble) was recovered from the particulate fraction of human placental mitochondria when extracted with the non-ionic detergent Nonidet P-40. mtDNA and TFAM were co-immunoprecipitated by anti-TFAM antibodies. TFAM was released into the supernatant by DNase I digestion of mtDNA in the particulate fraction. Thus, TFAM and mtDNA are tightly associated with each other, and it is likely that few TFAM or mtDNA molecules exist in an unbound form in mitochondria. Based on the fact that TFAM is abundant enough to wrap mtDNA entirely, these results suggest that human mtDNA is packaged with TFAM.” (Abstract)
Regarding claims 14, 30 and 40-41, Alam teaches a method wherein “mitochondria were prepared by a combination of differential and…gradient centrifugation. Mitochondria were solubilized” (Pg. 1641). Alam teaches a method wherein “TFAM was detected by western blotting” (Pg. 1643, Fig. 3). Alam teaches a method wherein “mtDNA fragments eluted almost in parallel with TFAM” (Pg. 1644). “ mtDNA fragments eluted almost in parallel with TFAM” reads on active TFAM Thus, Alam teaches a method comprising (a) measuring the level of mature and/or active TFAM protein in the sample, wherein the level of mature and/or active TFAM protein corresponds to (i) the amount of mature TFAM protein; and suggests a method wherein the level of mature and/or active TFAM protein corresponds to the amount of mature TFAM protein in the mitochondria.
Further in support, Pooreydy discloses “This protocol focuses on step-by-step procedure of mitochondrial proteome extraction from peripheral blood mononuclear cell (PBMC) mitochondria” (Abstract).
Regarding claims 14, 30 and 40-41, Pooreydy teaches a method comprising “Blood for isolation peripheral blood mononuclear cell (PBMC)” (Pg. 79) and “Mitochondria isolation” (Pg. 81). Thus, Pooreydy teaches a method comprising mitochondrial isolation from PBMCs. The ordinary artisan would be motivated to use the method of mitochondrial isolation to assess the levels of active TFAM in the mitochondria. In addition, Pooreydy mentions the high mitochondrion concentration cells such as muscle, liver and cultured cells. Thus, the ordinary artisan would expect levels of protein in mitochondria-rich liver to be similar to those in the mitochondria-rich PBMCs, likewise, comparable to levels in the mitochondria of PBMCs.
Lastly, Gunst discloses “Our present data… support a role for mitochondrial biogenesis in mediating recovery. Mitochondrial transcription factor A levels were lowest in the liver of nonsurviving animals.” (Pg. 192, last two lines -193, first two lines)
Regarding claims 14, 30 and 40-41, Gunst teaches a method comprising “Antibodies were purchased from Abcam, Cambridge, UK…. mitochondrial transcription factor A… DakoCytomation, Glostrup, Denmark (horseradish peroxidase–conjugated goat–antirabbit or goat–antimouse antibodies).” (Pg. 185, Immunoblotting, Para. 1). Gunst teaches a method comprising “Relative expression levels of key proteins… TFAM = mitochondrial transcription factor A” (Pg. 187, Figure 4 legend). Furthermore, as depicted in Figure 4, the levels of TFAM protein expression were decreased about 20%-60% in Non-survivor patient samples compared to the Healthy patient sample. Gunst teaches a method comprising “Expression of mitochondrial transcription factor A, a key transcription factor in mitochondrial biogenesis, was lowered in the liver of nonsurviving critically ill rabbits, whereas in survivors it remained low-normal (Fig. 4)” (Pg. 188, Mitochondrial Biogenesis, Para. 1; Fig. 4). Gunst also mentions that “A study on muscle biopsies found an early biogenesis response in surviving but not in nonsurviving patients with severe sepsis” (Pg. 192). Thus, Gunst teaches a method comprising measuring the level of TFAM protein in the sample; and (b) determining whether the level in the sample is abnormal, wherein the level in the sample is determined to be abnormal if the level of TFAM protein is at least about 20% or at least about 50% lower than the amount determined for a reference sample.
Thus, Kraft, Alam, Pooreydy and Gunst suggest a method of detecting an abnormal level of mature and/or active mitochondrial transcription factor A (TFAM) protein in a blood sample from a patient, wherein said method comprises (a) measuring the level of mature and/or active TFAM protein in the sample, wherein the level of mature and/or active TFAM protein corresponds to (i) the amount of mature TFAM protein,(ii) the number of interactions of TFAM protein with TFB2M protein, and/or (iii) the ratio of the amount of mature TFAM protein over the amount of immature TFAM protein; and (b) determining whether the level in the sample is abnormal, wherein the level in the sample is determined to be abnormal if the level of mature and/or active TFAM protein is at least about 20%, at least about 50%, or at least about 70% lower than the amount determined for a reference sample, wherein an abnormal level of mature and/or active TFAM protein in the sample is indicative of the presence of sepsis; (b) determining whether the level in the sample is abnormal, wherein the level in the sample is determined to be abnormal if the level of mature and/or active TFAM protein is at least about 20%, at least about 50%, or at least about 70% lower than the amount determined for a reference sample, wherein an abnormal level of mature and/or active TFAM protein in the sample is indicative of the presence of sepsis.
Kraft and Gunst are both considered to be analogous to the claimed invention because they are in the same field of mitochondrial injury and recovery (mitochondrial biogenesis), organ failure and TFAM levels. Kraft is also analogous to the claimed invention because the prior art is in the same field of non-invasive detection of levels of TFAM from PBMCs of human patients. Alam is considered to be analogous to the claimed invention because the prior art is in the same field of active TFAM levels in the mitochondria and Pooreydy is related to the claimed invention in support of detecting active TFAM protein based on the field of PBMC mitochondria isolation.
Therefore, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method comprising (a) detecting an abnormal level of mitochondrial transcription factor A (TFAM) transcript in a blood sample from a patient and (b) determining whether the level in the sample is abnormal, wherein the level in the sample is determined to be abnormal if the level of TFAM transcript is at least about 20% lower than the amount determined for a reference sample, wherein an abnormal level of TFAM transcript in the sample is indicative of the presence of sepsis as suggest by Kraft to incorporate the method of detecting levels of active/mature protein levels TFAM as suggested by Alam wherein the detection levels of active TFAM (e.g. protein levels in mitochondria of PBMCs) is supported by Pooreydy and wherein the level of TFAM protein levels is abnormal in critically ill subjects compared to controls is supported by Gunst; and provide a method according to the claim limitations of claims 14, 30 and 40-41. These claim elements were known in the art and one of skill in the art could have combined these elements by known methods with no change in their respective functions, and the combination would have yielded the predictable outcome according to the limitations of claims 14, 30 and 40-41. Doing so would allow improved method of detection of TFAM in mitochondrial biogenesis.
The teachings of Kraft, Alam, Pooreydy and Gunst are documented above in the rejection of claims 14, 30 and 40-41 under 35 U.S.C. 103. Claims 1, 20, 30, 40, 41 and 53 depend on claim 14. Claim 7 depends on claim 6. Claims 2 and 6 depend on claim 53, which depend on claim 14.
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Regarding claim 20, Kraft teaches a method wherein “in subjects that were not ICU-free by 1 week, day 1 PBMC levels of HMOX1, NRF1, PPARGC1A, and TFAM were significantly reduced (P<0.05) (Figure 3, Day 1, shown below)” (Pg. 653, Col. 2 Para. 2). TFAM levels are decreased by about 40% in not ICR-free at 1 week subject. “not ICU free at 1-week” reads on the increased severity of patient complications (i.e., sepsis).
Thus, Kraft, Alam, Pooreydy and Gunst suggest a method wherein a level of mature and/or active TFAM protein that is at least about 40% or at least about 80% lower in the sample from said patient than the amount determined for said reference sample is indicative of the presence of very severe sepsis
Regarding claim 53, Kraft teaches Fig. 1 (TFAM, lower right) which depicts the median gene expression level of TFAM in PBMCs of control subjects compared to a control mRNA, which is not considered significantly different from the level of the control. (Pg. 653. Figure 1). Thus, Kraft, Alam, Pooreydy and Gunst suggest a method further comprising (c) determining that sepsis is present in the patient when the level of mature and/or active TFAM protein is abnormal, and/or determining that no sepsis is present in the patient when the level of mature and/or active TFAM protein is not abnormal.
Regarding claims 2 and 6, Kraft teaches a method wherein “in subjects that were not ICU-free by 1 week, day 1 PBMC levels of HMOX1, NRF1, PPARGC1A, and TFAM were significantly reduced (P<0.05) (Figure 3, Day 1, shown above)” (Pg. 653, Col. 2 Para. 2). TFAM levels are decreased by about 40% in not ICR-free at 1 week subject. “not ICU free at 1-week” reads on the increased severity of patient complications (i.e., sepsis). Furthermore, Gunst teaches a method comprising “Expression of mitochondrial transcription factor A, a key transcription factor in mitochondrial biogenesis, was lowered in the liver of nonsurviving critically ill rabbits, whereas in survivors it remained low-normal (Fig. 4)” (Pg. 188, Mitochondrial Biogenesis, Para. 1; Fig. 4). Furthermore, as depicted in Figure 4, the levels of TFAM protein expression decreased by about 20%-60% in Non-survivor patient samples compared to the Healthy patient sample. “nonsurvivor” reads on negative outcome. Thus, Kraft, Alam, Pooreydy and Gunst suggest a method further comprising d) determining a degree of severity of sepsis in the patient, wherein (i) the degree of severity is low when the level of mature and/or active TFAM protein is abnormal but less than 40% lower in the sample from said patient than the amount determined for said reference sample, and/or (ii) the degree of severity is high when the level of mature and/or active TFAM protein is at least about 40% or at least about 80% lower in the sample from said patient than the amount determined for said reference sample; and further comprising d) prognosing the outcome of said sepsis in the patient, wherein (i) when the level of mature and/or active TFAM protein is abnormal but less than 40% lower in the sample from said patient than the amount determined for said reference sample, then a positive outcome is prognosed, and/or (ii) when the level of mature and/or active TFAM protein is at least about 40% or at least about 80% lower in the sample from said patient than the amount determined for said reference sample, then a negative outcome is prognosed.
Regarding claim 7, Kraft teaches a method wherein as described in the legend of Fig.3 “Open black circles depict individual subjects with sepsis that had prolonged ICU stays (≥ 1 wk) or early death (within 1 wk of presentation)” Thus, Kraft, Alam, Pooreydy and Gunst suggest a method wherein the positive outcome comprises survival for at least one month, and/or the negative outcome comprises death within one month.
Regarding claim 56, Kraft teaches a method wherein “This is the first human study to measure transcriptional activation of the mitochondrial biogenesis program in sepsis 1) non-invasively in peripheral blood” (Pg. 654, Discussion, Para. 1). Thus, Kraft, Alam, Pooreydy and Gunst suggest a method wherein the patient is a human.
Response to Arguments
Applicant's arguments filed 06/26/2026 (Pg. 10-13) with respect to claims 2, 6-7, 14, 20 30, 40-41, 53 and 56, have been fully considered but do not apply to the new grounds of rejection. To clarify some instances argued in the response filed 06/26/2026 see responses to each argument made by Applicant below:
Applicants’ argument: “The present application found, for the first time, an approximately 80% decrease in mature intramitochondrial TFAM protein, a marked decrease in TFAM-TFB2M interactions, an inverse correlation between TFAM-TFB2M interactions and SOFA score, and a survival association based on TFAM-TFB2M interaction thresholds in blood-derived PBMCs from septic patients.” (Pg. 12)
Response: In response to applicant's argument that stated above, the fact that the inventor has recognized another advantage which would flow naturally from following the suggestion of the prior art cannot be the basis for patentability when the differences would otherwise be obvious. See Ex parte Obiaya, 227 USPQ 58, 60 (Bd. Pat. App. & Inter. 1985).
Claim 54 is rejected under 35 U.S.C. 103 as being unpatentable over Kraft et al. (“Kraft”; (2019). Peripheral Blood Mononuclear Cells Demonstrate Mitochondrial Damage Clearance During Sepsis. Critical care medicine, 47(5), 651–658.) in view of T. I. Alam et al. (“Alam”; (2003). Human mitochondrial DNA is packaged with TFAM. Nucleic acids research, 31(6), 1640–1645.), Pooreydy et al. (“Pooreydy”; (2019). (2013). Organelle isolation for proteomics: mitochondria from peripheral blood mononuclear cells. Archives of Advances in Biosciences, 4.) and Gunst et al. (“Gunst”; (2013). Insufficient autophagy contributes to mitochondrial dysfunction, organ failure, and adverse outcome in an animal model of critical illness. Critical care medicine, 41(1), 182-194.) as applied to claim 14 above, and further in view of Hood et al. (“Hood”; Patent Pub. US 8586006 B2, Nov. 19, 2013).
The teachings of Kraft, Alam, Pooreydy and Gunst are documented above in the rejection of claims 2, 6-7, 14, 20 30, 40-41, 53 and 56 under 35 U.S.C. 103. Claim 54 depends on claim 14. Kraft, Alam, Pooreydy and Gunst do not explicitly teach the limitations of claim 54.
Hood discloses methods for identifying and using organ-specific proteins and transcripts. The present invention further provides compositions comprising organ-specific proteins and transcripts encoding the same, detection reagents for detecting such proteins and transcripts, and diagnostic panels, kits and arrays for measuring organ-specific proteins/transcripts in blood, biological tissue or other biological fluid. (Abstract)
Regarding claim 54, Hood teaches SEQ ID No: 4625 which has 100% sequence identity to SEQ ID 4 as shown below in the search result.
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“and/or” reads on the limitation “ and/or wherein the mature and/or active TFAM protein does not contain at the N-terminus the sequence set forth in SEQ ID NO:40 or a sequence that has at least 90% sequence identity to the sequence set forth in SEQ ID NO:40 as an optional limitation. Thus, Kraft, Alam, Pooreydy, Gunst and Hood suggest a method wherein the mature and/or active TFAM protein is a human mature TFAM protein with the sequence set forth in SEQ ID NO:4 or SEQ ID NO:8, or a protein that has at least 90% sequence identity to the sequence set forth in SEQ ID NO:4 or SEQ ID NO:8; and/or wherein the mature and/or active TFAM protein does not contain at the N-terminus the sequence set forth in SEQ ID NO:40 or a sequence that has at least 90% sequence identity to the sequence set forth in SEQ ID NO:40.
Hood is considered to be analogous to the claimed invention because they are in the same field of detecting proteins in a biological sample. Therefore, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of detecting an abnormal level of mature and/or active mitochondrial transcription factor A (TFAM) protein in a sample from a patient as taught by Kraft, Alam, Pooreydy and Gunst to incorporate the method wherein TFAM sequence comprises SEQ ID NO:4 as taught by Hood and provide a method of detecting an abnormal level human mature TFAM protein with the sequence set forth in SEQ ID NO:4. These claim elements were known in the art and one of skill in the art could have combined these elements by known methods with no change in their respective functions, and the combination would have yielded the predictable outcome according to the limitations of claim 54. Doing so would allow for detection of active TFAM protein level.
Response to Arguments
Applicant's arguments filed 06/26/2026 (Pg. 10-13) have been fully considered but they do not apply to the new grounds of rejection.
Claim 55 is rejected under 35 U.S.C. 103 as being unpatentable over Kraft et al. (“Kraft”; (2019). Peripheral Blood Mononuclear Cells Demonstrate Mitochondrial Damage Clearance During Sepsis. Critical care medicine, 47(5), 651–658.) in view of T. I. Alam et al. (“Alam”; (2003). Human mitochondrial DNA is packaged with TFAM. Nucleic acids research, 31(6), 1640–1645.), Pooreydy et al. (“Pooreydy”; (2019). (2013). Organelle isolation for proteomics: mitochondria from peripheral blood mononuclear cells. Archives of Advances in Biosciences, 4.) and Gunst et al. (“Gunst”; (2013). Insufficient autophagy contributes to mitochondrial dysfunction, organ failure, and adverse outcome in an animal model of critical illness. Critical care medicine, 41(1), 182-194.) as applied to claim 14 above, and further in view of Yoshida et al. (“Yoshida”; (2003). P53 physically interacts with mitochondrial transcription factor A and differentially regulates binding to damaged DNA. Cancer research, 63(13), 3729–3734.).
The teachings of Kraft, Alam, Pooreydy and Gunst are documented above in the rejection of claims 2, 6-7, 14, 20 30, 40-41, 53 and 56 under 35 U.S.C. 103. Claim 55 depends on claim 14. Kraft, Alam, Pooreydy and Gunst do not explicitly teach the limitations of claim 55.
Yoshida teaches a method wherein “Cell Fractionation for Western Blotting… samples were… immunoblotted with anti-mtTFA” (Pg. 3729) Regarding claim 55, Yoshida teaches a method wherein “Immunoblots of nuclear extract, cytosol, and the mitochondrial fraction from … cells… are shown in Fig. 1” (Pg. 3730; Fig. 1, Top panel , mtFA). Yoshida depicts detection of TFAM protein levels in three different cellular fractions in Fig. 1.((Fig. 1, Top panel , mtFA(-), N.E(-), Cyto(-), Mito(-)) Hence, one of ordinary skill would be interested in determining active TFAM levels and further evaluating whether levels of TFAM precursor levels are also effected. Thus, Kraft, Alam, Pooreydy, Gunst and Yoshida suggest a method wherein the level of mature and/or active TFAM protein corresponds to the ratio of the amount of mature TFAM protein over the amount of immature TFAM protein.
Yoshida is considered to be analogous to the claimed invention because they are in the same field of detecting TFAM proteins in a biological sample. Therefore, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of detecting an abnormal level of mature and/or active mitochondrial transcription factor A (TFAM) protein in a sample from a patient as taught by Kraft, Alam, Pooreydy and Gunst to incorporate the method wherein the level of mature and immature TFAM protein is detected as suggested by Yoshida and provide a method wherein the level of mature and/or active TFAM protein corresponds to the ratio of the amount of mature TFAM protein over the amount of immature TFAM protein. These claim elements were known in the art and one of skill in the art could have combined these elements by known methods with no change in their respective functions, and the combination would have yielded the predictable outcome according to the limitations of claim 55. Doing so would allow for detection of active TFAM protein level.
Response to Arguments
Applicant's arguments filed 06/26/2026 (Pg. 10-13) have been fully considered but they do not apply to the new grounds of rejection.
Claim 31 is rejected under 35 U.S.C. 103 as being unpatentable over Kraft et al. (“Kraft”; (2019). Peripheral Blood Mononuclear Cells Demonstrate Mitochondrial Damage Clearance During Sepsis. Critical care medicine, 47(5), 651–658.) in view of T. I. Alam et al. (“Alam”; (2003). Human mitochondrial DNA is packaged with TFAM. Nucleic acids research, 31(6), 1640–1645.), Pooreydy et al. (“Pooreydy”; (2019). (2013). Organelle isolation for proteomics: mitochondria from peripheral blood mononuclear cells. Archives of Advances in Biosciences, 4.) and Gunst et al. (“Gunst”; (2013). Insufficient autophagy contributes to mitochondrial dysfunction, organ failure, and adverse outcome in an animal model of critical illness. Critical care medicine, 41(1), 182-194.) as applied to claim 14 above, and further in view of Sarkar et al. (“Sarkar”; Patent App. Pub. US 20110033480 A1, Feb. 10, 2011).
The teachings of Kraft, Alam, Pooreydy and Gunst are documented above in the rejection of claims 2, 6-7, 14, 20 30, 40-41, 53 and 56 under 35 U.S.C. 103. Claim 31 depends on claim 14. Kraft, Alam, Pooreydy and Gunst do not explicitly teach the limitations of claim 31.
Sarkar discloses “mitochondrial transcription stimulatory factors Tfam, TBF1M and TFB2M facilitate mtRNA-polymerase activity to initiate transcription of the mtDNA-encoded genes such as several subunits of the OxPhos subunits, tRNA and rRNA genes, essential for mitochondrial integrity, function and mitochondrial DNA replication.”(Para. 305).
Regarding claim 31, Sarkar teaches a method wherein “Tfam, and TFB2M were evaluated… by Western blotting” (Para. 306). “Tfam” reads on mature and/or active TFAM protein. “Western blotting” reads on measuring the levels of protein. “Tfam … and TFB2M facilitate mtRNA-polymerase activity” reads on TFAM interaction with TFB2M. Thus, Kraft, Alam, Pooreydy and Gunst and Sarkar suggests a method wherein the level of mature and/or active TFAM protein corresponds to the number of interactions of TFAM protein with TFB2M protein, and/or wherein measuring the level of mature and/or active TFAM protein comprises quantifying the interaction of TFAM protein with TFB2M protein.
Sarkar is considered to be analogous to the claimed invention because the prior art is in the same field of detecting proteins in a biological sample. Therefore, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of detecting an abnormal level of mature and/or active mitochondrial transcription factor A (TFAM) protein in a sample from a patient as taught by Kraft, Alam, Pooreydy and Gunst to incorporate the method of measuring the level TFAM protein with TFB2M protein as taught by Sarkar and provide a method for detecting an abnormal level TFAM protein in a sample from a patient, wherein measuring the level of mature and/or active TFAM protein comprises quantifying the interaction of TFAM protein with TFB2M protein. These claim elements were known in the art of protein quantification and one of skill in the art could have combined these elements by known methods with no change in their respective functions, and the combination would have yielded the predictable outcome according to the limitations of claim 31. Doing so would provide insight to the levels of TFAM protein with TFB2M that is able to facilitate mitochondrial function.
Response to Arguments
Applicant's arguments filed 06/26/2026 (Pg. 10-13) have been fully considered but they do not apply to the new grounds of rejection.
Claim 34-35 and 39 are rejected under 35 U.S.C. 103 as being unpatentable over Kraft et al. (“Kraft”; (2019). Peripheral Blood Mononuclear Cells Demonstrate Mitochondrial Damage Clearance During Sepsis. Critical care medicine, 47(5), 651–658.) in view of T. I. Alam et al. (“Alam”; (2003). Human mitochondrial DNA is packaged with TFAM. Nucleic acids research, 31(6), 1640–1645.), Pooreydy et al. (“Pooreydy”; (2019). (2013). Organelle isolation for proteomics: mitochondria from peripheral blood mononuclear cells. Archives of Advances in Biosciences, 4.), Gunst et al. (“Gunst”; (2013). Insufficient autophagy contributes to mitochondrial dysfunction, organ failure, and adverse outcome in an animal model of critical illness. Critical care medicine, 41(1), 182-194.), and Sarkar et al. (“Sarkar”; Patent App. Pub. US 20110033480 A1, Feb. 10, 2011), as applied to claim 31 above, and further in view of Alam, Muhammad (“Alam 2018”; "Proximity ligation assay (PLA)." Current protocols in immunology 123.1 (2018): e58.).
The teachings of Kraft, Alam, Pooreydy, Gunst and Sarkar are documented above in the rejection of claim 31 under 35 U.S.C. 103. Claim 39 depends on claim 35, which depends on claim 34, which depends on claim 31. Kraft, Alam, Pooreydy, Gunst and Sakar do not explicitly teach the limitations of claim 34-35 and 39.
Alam 2018 discloses “Proximity ligation assay (PLA), also referred to as Duolink® PLA technology, permits detection of protein-protein interactions in situ (at distances <40 nm) at endogenous protein levels. It exploits specific antibodies identifying (either directly or indirectly) the two proteins of interest and utilizes specific DNA primers covalently linked to the antibodies. A hybridization step followed by DNA amplification with fluorescent probes permit visualization of spots of proximity by fluorescence microscopy. Since the development of PLA in 2002, it has been increasingly used to detect the interaction between two proteins with high sensitivity and specificity. It is a simple and sensitive technique to study protein-protein interaction in cells”. (Abstract)
Regarding claim 34-35 and 39, Alam 2018 teaches a method wherein “Duolink R PLA technology, allows one to detect protein-protein interactions at endogenous protein levels with high sensitivity and specificity (Fredriksson et al., 2002; Gullberg et al., 2004; Soderberg et al., 2006). Cells must be stained with two immunohistochemistry or immunofluorescence compatible primary antibodies to the target proteins. The two primary antibodies must be generated from different species (i.e., mouse/rabbit, rabbit/goat, or mouse/goat). Cells are then stained with secondary antibodies (2°-Ab) known as PLA probes (one PLUS and one MINUS). The PLA probes that bind to the constant regions of the primary antibodies contain a unique DNA strand. If the proteins of interest interact with each other, the DNA probes hybridize to make circular DNA. This DNA can be amplified and visualized by fluorescently labeled complementary oligonucleotide probes. The schematic diagram of Duolink® PLA is shown in Figure 1. Interactions are visualized as dots, and the number and intensity of the dots can be quantified by fluorescence microscopy.” (Pg. 1, Para. 2). The teaching “primary antibodies to the target proteins” reads on a pair of binding molecules wherein one specifically bind TFAM protein and the other specifically binds TFB2M protein. Thus, Kraft, Alam, Pooreydy, Gunst, Sarkar and Alam 2018 suggest a method wherein quantifying the interaction of TFAM protein with TFB2M protein comprises the steps of (a) contacting said sample with a pair of binding molecules, wherein one of said binding molecules specifically binds TFAM protein, and wherein the other of said binding molecules specifically binds TFB2M protein, and(b) generating a detectable signal when said two binding molecules are in close proximity to each other; wherein one of the binding molecules is an antibody specifically binding TFAM protein and the other binding molecule is an antibody specifically binding TFB2M protein, wherein at least one of said binding molecules is an antibody conjugated to an oligonucleotide, and/or wherein at least one of said binding molecules is specifically bound by an antibody conjugated to an oligonucleotide; and/or wherein said step (b) of generating a detectable signal comprises the steps of (i) generating an oligonucleotide template when said two binding molecules are in close proximity to each other, and(ii) amplifying and/or extending said oligonucleotide template; and wherein said interaction is quantified by a proximity ligation assay (PLA) and/or a proximity- dependent initiation of hybridization chain reaction (proxHCR).
Alam 2018 is analogous to the claimed invention because the prior art is in the same field of detecting proteins in a biological sample. Therefore, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of detecting an abnormal level of mature and/or active mitochondrial transcription factor A (TFAM) protein in a sample from a patient as suggested by Kraft, Alam, Pooreydy Gunst and Sakar to incorporate the method of quantifying the interaction of TFAM protein with TFB2M protein as taught by Alam 2018 and provide a method for detecting an abnormal level TFAM protein in a sample from a patient, wherein measuring the level of mature and/or active TFAM protein comprises quantifying the interaction of TFAM protein with TFB2M protein and wherein quantifying the interaction of TFAM protein with TFB2M protein comprises the steps of (a) contacting said sample with a pair of binding molecules, wherein one of said binding molecules specifically binds TFAM protein, and wherein the other of said binding molecules specifically binds TFB2M protein, and (b) generating a detectable signal when said two binding molecules are in close proximity to each other. These claim elements were known in the art and one of skill in the art could have combined these elements by known methods with no change in their respective functions, and the combination would have yielded the predictable outcome according to the limitations of claim 34-35 and 39. Doing so would provide insight to the protein-protein interactions of TFAM protein with TFB2M that is able to facilitate mitochondrial function in situ.
Response to Arguments
Applicant's arguments filed 06/26/2026 (Pg. 10-13) have been fully considered but they do not apply to the new grounds of rejection.
Conclusion of Response to Arguments
In view of the amendments, new grounds of rejections and above responses to arguments, no claims are in condition for allowance.
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.
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/KENDRA R VANN-OJUEKAIYE/Examiner, Art Unit 1682
/WU CHENG W SHEN/Supervisory Patent Examiner, Art Unit 1682
