Methods and compositions for radioresistant cancer

Abstract

In general, the disclosure relates to the discovery of certain radiotherapy-resistance markers can prognose radiotherapy resistance in a cancer patient.

Description

METHODS AND COMPOSITIONS FOR RADIORESISTANT CANCER

[0001] This application claims priority of U.S. Provisional Application No. 63 / 682,302, filed August 12, 2024, which is hereby incorporated by reference in its entirety. BACKGROUND OF THE INVENTION

[0002] This invention was made with government support under CA271894, CA248265, and CA214194 awarded by the National Institutes of Health. The government has certain rights in the invention. I. Field of the Invention

[0003] This invention relates to the field of oncology, proteomics, and medicine. II. Background

[0004] Prostate cancer is the second most common cause of cancer death in men1. It is usually curable when localized, so standard of care is curative intent treatment with either surgery (radical prostatectomy, RP) or image-guided radiotherapy (RT). Both are equally effective in this setting2, and decisions are often made based on side effects and comorbidities. For example, in patients of advanced age, radiotherapy is generally preferred3.

[0005] Radiotherapy involves the delivery of targeted ionizing radiation with the intent of damaging intracellular molecules4. The common assumption is that DNA is the main molecular target of radiotherapy, although proteins and lipids are also ionized5. Radiotherapy may also trigger the immune system and modulate the tumor microenvironment. Indeed tumor metastases can be attacked by the immune system after RT as part of a rare phenomenon called the abscopal effect6. Thus, radiotherapy exerts its therapeutic effects through a complex set of molecular and cellular responses.

[0006] Classical radiotherapy used a limited number of larger treatment fields, which would expose normal cells to significant amounts of radiation, causing dose-limiting toxicities4. Two main strategies have been taken to reduce radiotoxicities. First, advanced image- guidance is used to precisely target tumours and reduce the dose of radiation that normal cells experience7. Second, conventional fractionation (CF) schedules deliver the total prescribed299183876.1 - 1 -radiation dose in multiple smaller daily doses (~2 Gray, Gy) known as fractions, over several weeks to allow for normal tissue recovery between treatments8. SUMMARY OF THE INVENTION

[0007] The disclosure relates, in part, to the discovery that radiotherapy can lead to genomic instability and produce proteogenomic signatures. The disclosure also relates, in part, to the discovery that polymerase theta (POLQ) can drive radiotherapy resistance, and its inhibition can sensitize cancer cells to radiotherapy.

[0008] Disclosed herein are methods of treating cancer in a patient, methods of treating prostate cancer in a patient, methods of treating a cancer patient, methods of treating a prostate cancer patient, methods of treating a patient determined to be resistant to radiotherapy, methods of treating a patient determined to be sensitive to radiotherapy. In certain aspects, the method comprises one or more steps including administering a non-radiotherapy treatment to the patient. In certain aspects, the patient has been determined to have a change in at least one radiotherapy- resistance marker. In some aspects, the change is a biomolecular change. A biomolecular change includes a change in the amount or structure of a biomolecule, such as an amplification, deletion, or mutation in a gene and / or an increase, decrease, or modification to a gene product. In some aspects, the change is a proteomic change. In some aspects, the change is a transcriptomic change. In some aspects, the change is a genomic change.

[0009] In some aspects, the radiotherapy-resistance marker is a gene or gene product associated with a cellular response to radiotherapy, including any gene or gene product disclosed herein. In some aspects, the radiotherapy-resistance marker comprises NCOA2, MAP3K1, PCMTD1, PIK3CB, LPP, IL7R, POLQ, SALL4, RBM38, FANCC, TNC, GNAQ, NF2, PTCH1, NUP214, EWSR1, GNAS, PIM1, MYH9, RBFOX2, EP300, or a combination thereof. In some aspects, the radiotherapy-resistance marker is one or more of NCOA2, MAP3K1, PCMTD1, PIK3CB, LPP, IL7R, POLQ, SALL4, RBM38, FANCC, TNC, GNAQ, NF2, PTCH1, NUP214, EWSR1, GNAS, PIM1, MYH9, RBFOX2, or EP300. In some aspects, the radiotherapy-resistance marker includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 (or any range derivable therein) of NCOA2, MAP3K1, PCMTD1, PIK3CB, LPP, IL7R, POLQ, SALL4,299183876.1 - 2 -RBM38, FANCC, TNC, GNAQ, NF2, PTCH1, NUP214, EWSR1, GNAS, PIM1, MYH9, RBFOX2, or EP300. In certain aspects, the radiotherapy-resistance marker comprises MYC, RAD21, or a combination thereof. In some aspects, the radiotherapy-resistance marker is POLQ. In certain aspects, one or more of the radiotherapy-resistance markers is specifically excluded from the method.

[0010] In some aspects, the patient is a human patient. In some aspects the patient has, is suspected of having, has been diagnosed with having, or has symptoms of prostate cancer. In some aspects, the patient has or has not received conventional fractionation radiotherapy. In some aspects, the patient has or has not received hypofractionation radiotherapy.

[0011] In some aspects, the change comprises an amplification of the radiotherapy-resistance marker, such as an increase in copy number of the radiotherapy-resistance marker. In some aspects, the change comprises a change in a gene product amount or modification relative to a control. In some aspects, the control is from a patient that is sensitive to radiotherapy. In some aspects, the control is from a patient that has not received a radiotherapy. In some aspects, the control is from a healthy individual. In some aspects, the change comprises a gene amplification or deletion in the marker. In some aspects, the change comprises an increase in RNA of the radiotherapy-resistance marker. In some aspects, the change comprises an increase in RNA of the radiotherapy-resistance marker relative to a control. In some aspects, the change comprises an increase in protein of the radiotherapy-resistance marker. In some aspects, the change comprises an increase in protein of the radiotherapy-resistance marker relative to a control. In some aspects, the change comprises an amplification of POLQ. In some aspects, the change comprises an increase in POLQ protein. In some aspects, the change comprises an increase in POLQ protein relative to a control. In some aspects, the change comprises a change in RNA abundance of the radiotherapy-resistance marker. In some aspects, the change comprises a change in the RNA abundance of POLQ. In some aspects, the change comprises an increase in POLQ RNA.

[0012] Also disclosed are methods of treating cancer in a patient and / or methods of sensitizing a cancer to a radiotherapy. In some aspects, the method comprises administering a POLQ inhibitor to the patient. In certain aspects, the patient has received a radiotherapy treatment. In certain aspects, the patient receives a radiotherapy treatment after the administering of the POLQ inhibitor. In certain aspects, the method comprises administering a radiotherapy treatment to the patient after administering the POLQ inhibitor to the patient. In some aspects, the radiotherapy treatment299183876.1 - 3 -comprises conventional fractionated radiotherapy. In some aspects, the radiotherapy treatment comprises hypofractionated or stereotactic radiotherapy. In some aspects, the POLQ inhibitor comprises a molecule known to inhibit POLQ. In some aspects, the POLQ inhibitor comprises an RNAi inhibitor, including an siRNA that binds to and / or inhibits POLQ translation. In some aspects, the POLQ inhibitor comprises novobiocin and / or ART4215. In some aspects, the administration of the POLQ inhibitor results in a specific proteomic signature. In certain aspects, POLQ inhibition results in a change in one or more gene products, including AGPS, CLPX, DVL2, EZR, GNG5, HNRNPDL, LTA4H, NHP2, PDS5B, PSAT1, QKI, TXNDC5, or a combination thereof.

[0013] Also disclosed are methods of measuring radiotherapy-resistance markers. In some aspects, the method comprises measuring at least one gene product of a radiotherapy-resistance marker in a biological sample obtained from a patient, including any radiotherapy-resistance marker disclosed herein. In some aspects, the radiotherapy-resistance marker is NCOA2, MAP3K1, PCMTD1, PIK3CB, LPP, IL7R, POLQ, SALL4, RBM38, FANCC, TNC, GNAQ, NF2, PTCH1, NUP214, EWSR1, GNAS, PIM1, MYH9, RBFOX2, EP300, or a combination thereof. In some aspects, the measuring comprises measuring by proteomics. In some aspects, the measuring comprises DNA or RNA sequencing. In some aspects, the measuring is done by mass spectrometry. The measuring may be done by any method known in the art.

[0014] Also disclosed is a method of treating a prostate cancer patient, the method comprising administering a non-radiotherapy to the patient, wherein the patient is determined to have a POLQ amplification in cancer cells from the patient. In some aspects, the POLQ amplification is an amplification in POLQ DNA. In some aspects, the POLQ amplification results in an increase in a POLQ gene product. In some aspects, the patient is determined to have an increase in a POLQ gene product. In some aspects, the patient is determined to have an increase in a POLQ mRNA. In some aspects, the patient is determined to have an increase in a POLQ protein.

[0015] Also disclosed are methods of prognosing radiotherapy-resistance in a cancer patient. In some aspects, the method comprises measuring at least one gene product of a radiotherapy- resistance marker in a biological sample obtained from the patient. In some aspects, the radiotherapy-resistance marker is NCOA2, MAP3K1, PCMTD1, PIK3CB, LPP, IL7R, POLQ, SALL4, RBM38, FANCC, TNC, GNAQ, NF2, PTCH1, NUP214, EWSR1, GNAS, PIM1, MYH9, RBFOX2, EP300, or a combination thereof.299183876.1 - 4 -

[0016] Also disclosed are kits comprising one or more detection agents for determining levels of one or more radiotherapy-resistance markers, wherein the radiotherapy-resistance marker is NCOA2, MAP3K1, PCMTD1, PIK3CB, LPP, IL7R, POLQ, SALL4, RBM38, FANCC, TNC, GNAQ, NF2, PTCH1, NUP214, EWSR1, GNAS, PIM1, MYH9, RBFOX2, EP300, or a combination thereof. In some aspects, the kit comprises one or more negative or positive control samples and / or control detection agents. In some aspects, the kit comprises instructions for use. In some aspects, the kit excludes reagents for detection of other biomarkers.

[0017] Also disclosed are one or more of the following Aspects:

[0018] Aspect 1 includes a method of treating cancer in a patient, the method comprising administering a non-radiotherapy treatment to the patient, wherein the patient has been determined to have a change in at least one radiotherapy-resistance marker, wherein the radiotherapy- resistance marker comprises NCOA2, MAP3K1, PCMTD1, PIK3CB, LPP, IL7R, POLQ, SALL4, RBM38, FANCC, TNC, GNAQ, NF2, PTCH1, NUP214, EWSR1, GNAS, PIM1, MYH9, RBFOX2, EP300, or a combination thereof.

[0019] Aspect 2 depends upon Aspect 1, wherein the radiotherapy-resistance marker further comprises MYC, RAD21, or a combination thereof.

[0020] Aspect 3 depends upon any one of Aspects 1 or 2, wherein the patient is a human patient.

[0021] Aspect 4 depends upon any one of Aspects 1 to 3, wherein the patient has, is suspected of having, has been diagnosed with having, or has symptoms of prostate cancer.

[0022] Aspect 5 depends upon any one of Aspects 1 to 4, wherein the patient has received conventional fractionation radiotherapy.

[0023] Aspect 6 depends upon any one of Aspects 1 to 4, wherein the patient has received hypofractionation radiotherapy.

[0024] Aspect 7 depends upon any one of Aspects 1 to 6, wherein the radiotherapy-resistance marker is POLQ.

[0025] Aspect 8 depends upon any one of Aspects 1 to 7, wherein the change comprises a genetic amplification of the radiotherapy-resistance marker.

[0026] Aspect 9 depends upon any one of Aspects 1 to 8, wherein the change comprises an increase in RNA of the radiotherapy-resistance marker.299183876.1 - 5 -

[0027] Aspect 10 depends upon any one of Aspects 1 to 9, wherein the change comprises an increase in protein of the radiotherapy-resistance marker.

[0028] Aspect 11 depends upon any one of Aspects 1 to 10, wherein the change comprises an amplification of POLQ.

[0029] Aspect 12 depends upon any one of Aspects 1 to 11, wherein the change comprises an increase in POLQ protein.

[0030] Aspect 13 includes a method of treating cancer in a patient, the method comprising administering a POLQ inhibitor to the patient.

[0031] Aspect 14 depends upon Aspect 13, wherein the patient has received a radiotherapy treatment.

[0032] Aspect 15 depends upon Aspect 13 or 14, wherein the patient receives a radiotherapy treatment after the administering of the POLQ inhibitor.

[0033] Aspect 16 depends upon any one of Aspects 13 to 15, further comprising administering a radiotherapy treatment to the patient after administering the POLQ inhibitor to the patient.

[0034] Aspect 17 depends upon any one of Aspects 13 to 16, wherein the radiotherapy treatment comprises conventional fractionation radiotherapy.

[0035] Aspect 18 depends upon any one of Aspects 13 to 16, wherein the radiotherapy treatment comprises hypofractionation radiotherapy.

[0036] Aspect 19 depends upon any one of Aspects 13 to 18, wherein the patient is a human patient.

[0037] Aspect 20 depends upon any one of Aspects 13 to 19, wherein the patient has, is suspected of having, has been diagnosed with having, or has symptoms of prostate cancer.

[0038] Aspect 21 depends upon any one of Aspects 13 to 20, wherein the POLQ inhibitor comprises an RNAi inhibitor.

[0039] Aspect 22 depends upon any one of Aspects 13 to 21, wherein the POLQ inhibitor comprises novobiocin and / or ART4215.

[0040] Aspect 23 includes a method of treating a radiotherapy-resistant cancer in a patient, the method comprising administering a POLQ inhibitor to the patient, wherein the patient has received a radiotherapy treatment.

[0041] Aspect 24 includes a method of sensitizing radiotherapy-resistant cancer cells to a radiotherapy, the method comprising delivering to the cells a POLQ inhibitor.299183876.1 - 6 -

[0042] Aspect 25 includes a method of measuring radiotherapy-resistance markers, the method comprising measuring at least one gene product of a radiotherapy-resistance marker in a biological sample obtained from a patient, wherein the radiotherapy-resistance marker is NCOA2, MAP3K1, PCMTD1, PIK3CB, LPP, IL7R, POLQ, SALL4, RBM38, FANCC, TNC, GNAQ, NF2, PTCH1, NUP214, EWSR1, GNAS, PIM1, MYH9, RBFOX2, EP300, or a combination thereof.

[0043] Aspect 26 depends upon Aspect 25, wherein the patient is a human patient.

[0044] Aspect 27 depends upon Aspect 25 or 26, wherein the patient has, is suspected of having, has been diagnosed with having, or has symptoms of prostate cancer.

[0045] Aspect 28 depends upon any one of Aspects 25 to 27, wherein the patient has received conventional fractionation radiotherapy.

[0046] Aspect 29 depends upon any one of Aspects 25 to 27, wherein the patient has received hypofractionation radiotherapy.

[0047] Aspect 30 includes a method of treating a prostate cancer patient, the method comprising administering a non-radiotherapy to the patient, wherein the patient is determined to have a POLQ amplification or increase in a POLQ gene product in cancer cells from the patient.

[0048] Aspect 31 depends upon Aspect 30, wherein the POLQ gene product is POLQ mRNA.

[0049] Aspect 32 depends upon Aspect 30 or 31, wherein the POLQ gene product is POLQ protein.

[0050] Aspect 33 includes a method of prognosing radiotherapy-resistance in a cancer patient, the method comprising measuring at least one gene product of a radiotherapy-resistance marker in a biological sample obtained from the patient, wherein the radiotherapy-resistance marker is NCOA2, MAP3K1, PCMTD1, PIK3CB, LPP, IL7R, POLQ, SALL4, RBM38, FANCC, TNC, GNAQ, NF2, PTCH1, NUP214, EWSR1, GNAS, PIM1, MYH9, RBFOX2, EP300, or a combination thereof.

[0051] Aspect 34 depends upon Aspect 33, wherein the patient is a human patient.

[0052] Aspect 35 depends upon Aspect 33 or 34, wherein the patient has, is suspected of having, has been diagnosed with having, or has symptoms of prostate cancer.

[0053] Aspect 36 depends upon any one of Aspects 33 to 35, wherein the patient has received conventional fractionation radiotherapy.

[0054] Aspect 37 depends upon any one of Aspects 33 to 35, wherein the patient has received hypofractionation radiotherapy.299183876.1 - 7 -

[0055] Aspect 38 includes a kit comprising one or more detection agents for determining levels of one or more radiotherapy-resistance markers, wherein the radiotherapy-resistance marker is NCOA2, MAP3K1, PCMTD1, PIK3CB, LPP, IL7R, POLQ, SALL4, RBM38, FANCC, TNC, GNAQ, NF2, PTCH1, NUP214, EWSR1, GNAS, PIM1, MYH9, RBFOX2, EP300, or a combination thereof.

[0056] Aspect 39 depends upon Aspect 38, wherein the kit further comprises one or more negative or positive control samples and / or control detection agents.

[0057] Aspect 40 depends upon Aspect 38 or 39, wherein the kit further comprises instructions for use.

[0058] Aspect 41 depends upon any one of Aspects 38 to 40, wherein the kit excludes reagents for detection of other biomarkers.

[0059] Aspect 42 depends upon any one of Aspects 38 to 41, wherein the detection agents comprise primers capable of hybridizing the one or more radiotherapy-resistance markers.

[0060] Aspect 43 depends upon any one of Aspects 38 to 42, wherein the detection agents comprise primers capable of amplifying the one or more radiotherapy-resistance markers. Aspect 44 depends upon any one of Aspects 38 to 43, wherein the detection agents comprise an antigen-binding protein capable of binding the one or more radiotherapy-resistance markers.

[0061] Throughout this application, the term “about” is used to indicate that a value includes the inherent variation of error for the measurement or quantitation method.

[0062] The use of the word “a” or “an” when used in conjunction with the term “comprising” may mean “one,” but it is also consistent with the meaning of “one or more,” “at least one,” and “one or more than one.”

[0063] The phrase “and / or” means “and” or “or”. To illustrate, A, B, and / or C includes: A alone, B alone, C alone, a combination of A and B, a combination of A and C, a combination of B and C, or a combination of A, B, and C. In other words, “and / or” operates as an inclusive or.

[0064] The words “comprising” (and any form of comprising, such as “comprise” and “comprises”), “having” (and any form of having, such as “have” and “has”), “including” (and any form of including, such as “includes” and “include”) or “containing” (and any form of containing, such as “contains” and “contain”) are inclusive or open-ended and do not exclude additional, unrecited elements or method steps.299183876.1 - 8 -

[0065] The compositions and methods for their use can “comprise,” “consist essentially of,” or “consist of” any of the ingredients or steps disclosed throughout the specification. Compositions and methods “consisting essentially of” any of the ingredients or steps disclosed limits the scope of the claim to the specified materials or steps which do not materially affect the basic and novel characteristic of the claimed invention.

[0066] It is contemplated that any aspect discussed in this specification can be implemented with respect to any method or composition of the invention, and vice versa. Furthermore, compositions of the invention can be used to achieve methods of the invention.

[0067] Other objects, features and advantages of the present invention will become apparent from the following detailed description. It should be understood, however, that the detailed description and the specific examples, while indicating specific aspects of the invention, are given by way of illustration only, since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description. BRIEF DESCRIPTION OF THE DRAWINGS

[0068] The following drawings form part of the present specification and are included to further demonstrate certain aspects of the present invention. The invention may be better understood by reference to one or more of these drawings in combination with the detailed description of specific aspects presented herein.

[0069] FIGS. 1A-1G show the genomic landscape of CF- and HF-resistant prostate cancer cells. FIG. 1A shows a schematic of experimental design and workflow. FIG. 1B shows somatic single nucleotide variant (SNV) count for CF- vs HF-resistant cells. CF-resistant cells gained twice more SNVs than HF (P = 0.1, Mann–Whitney U test). FIG. 1C shows a graphical representation of SNVs in cancer driver genes. Presented are all driver genes that are predicted to be strongly influenced by SNVs. Considered are SNVs that were identified in all three replicates for each cell type. Top panel, single-base substitution types. Bottom panel, the predicted annotation. FIG. 1D shows a graphical representation of mutational signatures. Gained SNVs converged on partly similar cancer mutational signatures. Most signatures of known etiology, irrespective of the treatment schedule, are associated with defective DNA mismatch repair. Signature etiologies: SBS5, unknown; SBS26 and SBS15, Defective DNA mismatch repair; SBS1, Spontaneous deamination of 5-methylcytosine; SBS14, Concurrent polymerase epsilon mutation and defective299183876.1 - 9 -DNA mismatch repair; SBS20, Concurrent POLD1 mutations and defective DNA mismatch repair; SBS44 Defective DNA mismatch repair. FIG. 1E shows somatic structural variant (SV) counts for CF- and HF-resistant cells. The number of somatic structural variants (sSVs) is similar between CF- and HF-resistant cells (P = 1, Mann–Whitney U test). FIG. 1F. shows distinct SVs in CF-resistant cells compared to HF across the genome. Considered are SVs that were identified in all three replicates for each cell type. Chromosome numbers are presented on the x-axis. The colored lines represent types of SVs: INV, inversion; DEL, deletion; DUP, duplication; TRA, translocation. FIG. 1G. shows the fusion transcripts that were identified in either CF- and / or HF- resistant cells. Purple, the fusion transcript was identified; white, the fusion transcript was not identified. The results are presented for three replicates for each cell line.

[0070] FIGS.2A-2F shows transcriptomic signatures of cancer-related genes in radioresistant cells. FIG.2A is a graphical representation on the differences in RNA abundances of driver cancer genes. The most abundant transcript out of all transcripts per gene was taken for visualization. FIGS. 2B – 2C show that the differences in RNA abundances of affected driver-gene, compared to their abundance differences at the protein level. FIG. 2B shows a graphical representation of the differences between CF-resistant cells and the parental cells. FIG. 2C shows a graphical representation of the differences between HF-resistant cells and the parental cells. FIG.2D shows a heatmap of RNA abundance in CF-resistant cells, HF-resistant cells, and parental cells. The RNA abundance profile of cancer hallmark genes in CF-resistant cells is distinct from HF-resistant cells and the parental profiles, which are relatively similar. The heatmap presents normalized counts of RNA isoforms that were significantly differentially abundant in CF-resistant cells compared to HF. Red, high abundance; blue, low abundance. For visualization, RNA abundances as a function of log10 were converted to z-scores. FIG. 2E shows a graphical representation of an enrichment analysis of hallmark gene sets. The dot size represents the enrichment score, and the dot color represents the directionality: for CF vs. PAR, HF vs. PAR, and HF vs. CF tests, orange represents upregulation toward CF-resistant cells, HF-resistant cells, and HF-resistant cells respectively. FIG. 2F shows a graphical representation of the differences in RNA abundances of the lncRNA UCA1, the miRNAs 200c-3P and 200b-3P, and ZEB2 (the target of the miRNAs 200c-3P and 200b-3P). For UCA1 transcripts, the small letters represent different isoforms. In FIGS. 2A and 2F, the dot size represents the log2(fold change) size, and the dot color represents the directionality: for CF vs. PAR, HF vs. PAR, and HF vs. CF tests, red represents upregulation toward CF-resistant299183876.1 - 10 -cells, HF-resistant cells, and HF-resistant cells respectively. For all figure panels, three RNA- sequencing replicates were used for the radioresistant cell lines and two for the parental line. In FIGS. 2B- 2C, three replicates were used for all cell types.

[0071] FIGS. 3A-3C shows fractionation-dependent protein profiles in radioresistant cells. FIG. 3A shows a heatmap depicting the difference in relationships of consensus-module eigengenes and cellular fractions, between CF- and HF-resistant cells. The colors represent the difference in correlations between consensus module eigengenes and a specific subcellular fraction of HF-resistant cells compared to CF: Blue, a higher correlation in CF-resistant cells; Red, a higher correlation in HF-resistant cells. At the top panel, each color represents a module, which is a detected group of positively correlated genes that are highly interconnected. FIG.3B depicts gene ontology enrichment of module genes for biological processes. The top two enrichments for each module are presented. FIG. 3C depicts the differences in protein abundances of driver, across different subcellular fractions and in whole cell lysates. Main panel, protein Cohen's d effect sizes of significant proteins across cell fractions. Only significant changes at the level of FDR ≤ 0.025 were plotted. The dot color is the directionality: magenta and green represent upregulation and downregulation, respectively, toward CF- or HF-resistant cells. Right panel, the consensus modules that each gene was assigned to. For all figure panels, three replicates were used for CF- and HF- resistant cells. For the parental cells, at least two replicates were used.

[0072] FIGS. 4A-4H shows the characteristics of radioresistance modulator candidates in primary prostate tumors. FIG. 4A shows significant associations (FDR ≤ 0.05) between CNAs in candidate genes and biochemical recurrence, as shown by fitting Cox proportional hazard models in ICGC. FIG.4B shows that POLQ amplification is associated with biochemical relapse in ICGC (Cox proportional hazard model, FDR = 7.78 x 10-3). FIG. 4C shows that high POLQ RNA abundance is associated with BCR following an RT treatment in NCCS (Cox proportional hazard model, P = 0.06). FIG. 4D shows that increased RNA abundance of POLQ is associated with a high mutation ratio in clinically relevant candidate genes based on linear regression in ICGC. Padjusted levels after FDR correction are presented on the right. FIG. 4E shows significant correlations between RNA abundances of clinically relevant candidates and POLQ RNA abundance. FIGS.4F-4G show strong correlation between POLQ and BRCA2 RNA abundance in ICGC (FIG.4F) and NCCS (FIG.4G). FIG. 4H shows that increased RNA abundance of POLQ299183876.1 - 11 -is associated with high Gleason grades. In FIGS. 4F-4H, ρ: Spearman correlation. Pρ: the Padjustedafter FDR

[0073] FIGS. 5A-5F shows the proteome signature upon POLQ inhibition. FIG. 5A shows a schematic of the experimental design. FIG. 5B depicts results from genetic POLQ inhibition. The parental, CF-resistant and HF-resistant cells were treated with either POLQ siRNA or scramble siRNA as the control. Significant radiosensitization was observed in all cell lines upon POLQ knockdown (P = 0.01, 0.002 and 0.02 for the parental, CF-resistant and HF-resistant cells, respectively; Paired T-test). Three biological replicas of 4000 cells per well, and three of 6000 cells per well were considered for each sample. FIG. 5C depicts results from pharmacologic POLQ inhibition. CF-resistant, HF-resistant, and the parental cells were treated with 100μM Novobiocin to achieve POLQ inhibition (Nvb 100μM). Significant radiosensitization was observed in all DU145 cell lines upon POLQ inhibition (P = 0.04, 0.02 and 0.01, for the parental, CF-resistant and HF-resistant cells; Paired T-test). In FIGS. 5B – 5C both the control and the treated cells were irradiated with 0Gy or 4Gy in 2 fractions. The surviving fraction of the treated cells (POLQ siRNA or Nvb 100μM) was normalized to the surviving fraction of the corresponding control of each cell line (black dots). FIG.5D depicts the enrichment score of the top ten activated and suppressed biological processes at the protein level following POLQ depletion, in CF-resistant cells and the parental. The dot size represents the enrichment score, and the dot color is the directionality: orange shows upregulation toward POLQ-treated cells. FIG. 5E shows that POLQ genetic inhibition creates a proteomic signature, involving twelve affected genes in CF-resistant cells. Three replicates from each cell type were used for the analysis. Left panel: signature genes whose protein abundances were changed between POLQ-depleted cells and the control, in CF- resistant cells and the parental cells. The dot size represents the Cohen’s d effect size, and the dot color is the directionality: magenta, upregulation, and green, downregulation toward POLQ- treated cells. Middle panel: changes in protein abundances of signature genes, between CF- resistant cells and the parental. The dot size represents the Cohen’s d effect size, and the dot color is the directionality: magenta, upregulation, and green, downregulation toward CF-resistant cells. Right panel: changes in RNA abundances of signature genes between CF-resistant cells and the parental. The dot size represents the log2(fold change) values. the dot color represents the directionality: red shows upregulation toward CF-resistant cells. FIG. 5F shows results from investigating signature genes (that were affected by POLQ inhibition in cell lines) in primary299183876.1 - 12 -patient data. Left: RNA-RNA Pearson correlations between POLQ and signature genes. Middle: Pearson correlation between the abundances of POLQ RNA and proteins of signature genes. Right: Abundance of signature genes in normal vs. tumor samples at the RNA and protein levels. For all panels, datasets refer to the cohort names used for analysis, and the data type for the type of molecule tested for signature genes (i.e. RNA or protein).

[0074] FIGS. 6A-6C. FIG. 6A shows the density of SNVs induced by radiation. For each sample, the number of SNVs that were not present in the corresponding parental cells, and were common to all replicates was divided by the number of genome sites covered by at least 20 reads in all replicates. FIG. 6B shows the mutational signatures in 22Rv1 based on SNVs induced by radiation. Signature etiologies: SBS1, Spontaneous deamination of 5-methylcytosine; SBS5, unknown; SBS44 Defective DNA mismatch repair. FIG. 6C shows the gene annotations of SVs and the effect on the RNA and protein abundances. Left panel, differences in transcript abundance of SVs distrusted genes, using DEseq2 with the default parameter lfcThreshold = 0. The dot size represents the log2(fold change) size, and the dot color represents the directionality: for CF vs. PAR, HF vs. PAR, and HF vs. CF tests, red represents upregulation toward CF-, HF-, and HF- resistant cells respectively. For each gene, the most abundant transcript was taken. Right panel, differences in the protein levels of sSVs disrupted genes across different cell fractions. The dot size represents the Cohen's d effect size, and the dot color represents the directionality: magenta and green represent upregulation and downregulation, respectively, toward CF- or HF-resistant cells. In both the left and right panels, X represents ‘non-detected’.

[0075] FIG. 7 shows the difference in RNA abundance of a PCNA isoform (ENST00000379143.10) and POLD1 isoforms (ENST00000440232.7 and ENST00000596648.1). The dot size represents the log2(fold change) size, and the dot color represents the directionality: for CF vs. PAR, HF vs. PAR, and HF vs. CF tests, red represents upregulation toward CF-, HF-, and HF-resistant cells respectively.

[0076] FIGS.8A-8K show supporting data for differential RNA abundance analysis. FIG.8A shows the normalized transcript count distribution for each replicate. The median is shown as a solid line. FIG. 8B shows the normalized transcript counts for all replicates. Red, high counts; blue, low counts. For visualization, transcript counts as a function of log10 were converted to z- scores. In FIGS. 8A-8B, a value of 1 was added to all values in the count matrix before operating log. FIGS. 8C-8E show V=volcano plots following differential RNA-abundance analysis, using299183876.1 - 13 -DEseq2 with lfcThreshold = 0.5. In all panels, nsignificant refers to the number of significant transcripts (FDR £ 0.05). FIG.8C shows a volcano plot for CF vs. PAR. FIG.8D shows a volcano plot for HF vs. PAR. FIG. 8E shows a volcano plot for HF vs. CF. FIG. 8F shows a pie chart depicting the overlap between significant differentially abundant transcripts in CF- and HF- resistant cells compared to the parental, following a strict test. FIG.8G shows a pie chart depicting the overlaps between CF-differentially abundant genes, HF-differentially abundant genes and the GSEA hallmark cancer gene set. For this plot, transcript names from FIG.8F were converted into gene names, and each gene was counted only once (even if there were more than one significant transcript per gene) in CF- and HF-resistant cells. FIG. 8H-8I depicts the normalized miRNA counts. Red, high counts; blue, low counts. For visualization, transcript counts as a function of log2 were converted to z-scores. FIG. 8H depicts counts for CF and Parental groups. FIG. 8I depicts counts for HF and Parental groups. A value of 1 was added to all values in the count matrix before operating log. FIG. 8J shows a pie chart depicting the overlap between significant differentially abundant miRNAs in CF- and HFresistant cells compared to the parental. FIG. 8K shows a pie chart depicting the overlap between significant differentially abundant genes (listed in FIG. 8G) in CF- or HF-resistant cells compared to the parental, that were identified as targets for significant miRNAs (listed in FIG. 8J) for CF or HF, correspondingly.

[0077] FIGS. 9A-9N show volcano plots following differential protein-abundance analysis. In all panels, significant proteins are plotted in red. Significance is defined as targets with Padjusted ≤ 0.05 following an FDR correction. FIG. 9A shows a volcano plot of whole cell CF vs. PAR. FIG. 9B shows a volcano plot of whole cell HF vs. PAR. FIG. 9C shows a volcano plot of mitochondria CF vs. PAR. FIG. 9D shows a volcano plot of mitochondria HF vs. PAR. FIG. 9E shows a volcano plot of microsome CF vs PAR. FIG. 9F shows a microsome HF vs. PAR. FIG. 9G shows a volcano plot of nuclear pellet CF vs PAR. FIG. 9H shows a volcano plot of nuclear pellet HF vs PAR. FIG.9I shows a volcano plot of cytosol vs PAR. FIG.9J shows a volcano plot of cytosol HF vs. PAR. FIG. 9K shows a volcano plot of nuclear soluble CF vs PAR. FIG. 9L shows a volcano plot of nuclear soluble HF vs PAR. FIG. 9M shows a volcano plot of plasma membrane CF vs PAR. FIG. 9N plasma membrane HF vs PAR.

[0078] FIGS. 10A-10D show RNA-protein relationships. FIG. 10A depicts differences in RNA abundance of CD44 isoforms. The dot size represents the log2(fold change) size, and the dot color represents the directionality: for CF vs. PAR, HF vs. PAR, and HF vs. CF tests, red represents299183876.1 - 14 -upregulation toward CF-, HF-, and HF-resistant cells respectively. FIG.10B shows a volcano plot of depicts differences in protein abundance of CD44 protein. Right panel: protein abundances across different subcellular fractions and cell types. Red, high abundance; blue, low abundance. For visualization, imputed protein intensities in a log2- space were converted to z-scores. Right panel: differences in protein abundance of CD44 proteins across different subcellular fractions. The dot size represents the Cohen’s d effect size, and the dot color represents the directionality: for CF vs. PAR, HF vs. PAR, and HF vs. CF tests, magenta represents upregulation toward CF-, HF-, and HF-resistant cells respectively. FIG. 10C depicts Pearson correlations between the median abundance of RNA and proteins for all detected genes (left panel) or only hallmark cancer genes (right panel). Targets for which the median value was zero, were removed from correlation calculations. Protein intensities are stratified by subcellular localization. FIG. 10D depicts differences in protein levels of driver genes that were affected at the RNA level in the same direction, across different subcellular fractions and in whole cell lysates. Left panel, the consensus modules that each gene was assigned to. Middle panel, protein abundance changes across cell fractions. Right panel, differences in the abundance of corresponding genes at the RNA levels, between CF- or HF-resistant cells and the parental cells. The dot size represents the log2(fold change) size, and the dot color represents the directionality: red represents upregulation toward CF- and HF-resistant cells.

[0079] FIGS. 11A-11D shows validation of the association of POLQ with prostate cancer aggressiveness. FIG. 11A shows that high POLQ RNA abundance is associated with BCR based on a Cox proportional hazard model. The tested dataset was downloaded from GEO, accession number GSE70770. FIG. 11B shows RNA abundance of POLQ in normal vs. tumor samples in 51 TCGA patients with matched normal and tumor samples. P represents the P-value following a paired T-test. FIG. 11C shows a positive association between POLQ RNA abundance and T- categories in TCGA. FIG.11D shows a positive association between POLQ RNA abundance and ISUP grade groups in TCGA.

[0080] FIG.12 shows the association between CNA events in seven candidate genes and BCR, using a log-rank test. P represents the Padjusted after FDR correction.

[0081] FIGS. 13A-13I shows supporting data for POLQ inhibition effects. FIGS. 13A-13C shows that real-time PCR following POLQ siRNA treatments confirmed a successful POLQ siRNA depletion. FIG. 13A shows comparison to samples treated with scramble siRNA in CF-299183876.1 - 15 -resistant cells. FIG. 13B shows comparison to samples treated with sample siRNA in the parental cells. FIG. 13C shows comparison to samples treated with sample siRNA in HF-resistant cells. On the y-axis, RQ is the relative quantification. FIG. 13D depicts the abundances of all detected proteins following POLQ knockdown FIG. 13E depicts the abundances of significantly affected proteins in CF-resistant cells, following POLQ knockdown. In FIGS. 13D-13E, Red, high intensity; blue, low intensity. For visualization, protein intensities as a function of log2 were converted to z-scores. FIGS. 13F-13G. FIG. 13F shows a volcano plot for differential protein- abundance analysis, following POLQ knockdown in CF-resistant cells. FIG.13G shows a volcano plot for differential protein-abundance analysis, following POLQ knockdown in the parental cells. In FIGS. 13F-13G, red dots represent significant changes in protein abundance (FDR £ 0.05). FIG.13H depicts the association between BCR and CNA events in the signature gene, PSAT1, by fitting a Cox proportional hazard model for PSAT1. FIG.13I depicts the association between BCR and CNA events in the signature gene, CLPX, by using a Heinze log-rank test for CLPX. P represents the Padjusted after FDR correction. detailed description of the invention DETAILED DESCRIPTION OF THE INVENTION I. Sample Preparation

[0082] In certain aspects, methods involve obtaining a sample from a subject. The methods of obtaining provided herein may include methods of biopsy such as fine needle aspiration, core needle biopsy, vacuum assisted biopsy, incisional biopsy, excisional biopsy, punch biopsy, shave biopsy or skin biopsy. In certain aspects, the sample is obtained from a biopsy from prostate tissue by any of the biopsy methods previously mentioned. In other aspects, the sample may be obtained from any of the tissues provided herein that include but are not limited to non-cancerous or cancerous tissue and non-cancerous or cancerous tissue from the serum, gall bladder, mucosal, skin, heart, lung, breast, pancreas, blood, liver, muscle, kidney, smooth muscle, bladder, colon, intestine, brain, prostate, esophagus, or thyroid tissue. Alternatively, the sample may be obtained from any other source including but not limited to urine, blood, sweat, hair follicle, buccal tissue, tears, menses, feces, or saliva. In certain aspects of the current methods, any medical professional such as a doctor, nurse or medical technician may obtain a biological sample for testing. Yet further, the biological sample can be obtained without the assistance of a medical professional.299183876.1 - 16 -

[0083] A sample may include but is not limited to, tissue, cells, or biological material from cells or derived from cells of a subject. The biological sample may be a heterogeneous or homogeneous population of cells or tissues. The biological sample may be obtained using any method known to the art that can provide a sample suitable for the analytical methods described herein. The sample may be obtained by non-invasive methods including but not limited to: scraping of the skin or cervix, swabbing of the cheek, saliva collection, urine collection, feces collection, collection of menses, tears, or semen.

[0084] The sample may be obtained by methods known in the art. In certain aspects, the samples are obtained by biopsy. In other aspects, the sample is obtained by swabbing, endoscopy, scraping, phlebotomy, or any other methods known in the art. In some cases, the sample may be obtained, stored, or transported using components of a kit of the present methods. In some cases, multiple samples, such as multiple prostate samples may be obtained for diagnosis by the methods described herein. In other cases, multiple samples, such as one or more samples from one tissue type (for example prostate) and one or more samples from another specimen (for example serum) may be obtained for diagnosis by the methods. In some cases, multiple samples such as one or more samples from one tissue type (e.g. prostate) and one or more samples from another specimen (e.g. serum) may be obtained at the same or different times. Samples may be obtained at different times are stored and / or analyzed by different methods. For example, a sample may be obtained and analyzed by routine staining methods or any other cytological analysis methods.

[0085] In some aspects, the biological sample may be obtained by a physician, nurse, or other medical professional such as a medical technician, endocrinologist, cytologist, phlebotomist, radiologist, or a pulmonologist. The medical professional may indicate the appropriate test or assay to perform on the sample. In certain aspects a molecular profiling business may consult on which assays or tests are most appropriately indicated. In further aspects of the current methods, the patient or subject may obtain a biological sample for testing without the assistance of a medical professional, such as obtaining a whole blood sample, a urine sample, a fecal sample, a buccal sample, or a saliva sample.

[0086] In other cases, the sample is obtained by an invasive procedure including but not limited to: biopsy, needle aspiration, endoscopy, or phlebotomy. The method of needle aspiration may further include fine needle aspiration, core needle biopsy, vacuum assisted biopsy, or large core299183876.1 - 17 -biopsy. In some aspects, multiple samples may be obtained by the methods herein to ensure a sufficient amount of biological material.

[0087] General methods for obtaining biological samples are also known in the art. Publications such as Ramzy, Ibrahim Clinical Cytopathology and Aspiration Biopsy 2001, which is herein incorporated by reference in its entirety, describes general methods for biopsy and cytological methods. In one aspect, the sample is a fine needle aspirate of a esophageal or a suspected esophageal tumor or neoplasm. In some cases, the fine needle aspirate sampling procedure may be guided by the use of an ultrasound, X-ray, or other imaging device.

[0088] In some aspects, of the present methods, the molecular profiling business may obtain the biological sample from a subject directly, from a medical professional, from a third party, or from a kit provided by a molecular profiling business or a third party. In some cases, the biological sample may be obtained by the molecular profiling business after the subject, a medical professional, or a third party acquires and sends the biological sample to the molecular profiling business. In some cases, the molecular profiling business may provide suitable containers, and excipients for storage and transport of the biological sample to the molecular profiling business.

[0089] In some aspects, of the methods described herein, a medical professional need not be involved in the initial diagnosis or sample acquisition. An individual may alternatively obtain a sample through the use of an over the counter (OTC) kit. An OTC kit may contain a means for obtaining said sample as described herein, a means for storing said sample for inspection, and instructions for proper use of the kit. In some cases, molecular profiling services are included in the price for purchase of the kit. In other cases, the molecular profiling services are billed separately. A sample suitable for use by the molecular profiling business may be any material containing tissues, cells, nucleic acids, genes, gene fragments, expression products, gene expression products, or gene expression product fragments of an individual to be tested. Methods for determining sample suitability and / or adequacy are provided.

[0090] In some aspects, the subject may be referred to a specialist such as an oncologist, surgeon, or endocrinologist. The specialist may likewise obtain a biological sample for testing or refer the individual to a testing center or laboratory for submission of the biological sample. In some cases the medical professional may refer the subject to a testing center or laboratory for submission of the biological sample. In other cases, the subject may provide the sample. In some cases, a molecular profiling business may obtain the sample.299183876.1 - 18 -II. Detection and Measurement of Markers

[0091] Certain aspects herein relate to the detection and / or measurement of markers, including radiotherapy-resistance markers such as those described herein. In some aspects, the markers are detected by sequencing nucleic acids from a sample. In some aspects, the markers are detected by hybridizing a probe to nucleic acids from a sample. In some aspects, the markers are detected by amplifying nucleic acids from a sample. The amplified nucleic acids may then be further detected or measured. In some aspects, the markers are detected by contacting proteins and / or nucleic acids from a sample. The sample may be a biological sample obtained from a patient.

[0092] In certain aspects, the detected and / or measured markers are quantified. For example, the detected and / or measured markers may be quantified by total read count and / or abundance, such as from a cell and / or from a sample. The detected and / or measured markers may be quantified by comparing the level to a reference marker, such as a housekeeping gene. In certain aspects, the detected and / or measured markers are compared to a control. The control may be the level of the markers present in a healthy individual. The control may be the level of the markers present in a population of individuals, including a population of healthy individuals. The control may be a baseline level found in a patient before or after diagnosis of a disease, such as a cancer described herein. The control may be a baseline level found in a patient before, during, or after treatment of a disease, such as a cancer described herein. The control may be a standard determined by skilled artisans, such as clinicians, oncologists, and / or epidemiologists. The standard may be determined after one or more clinical trials to determine the level above which indicates a patient for or against a certain treatment, such as a radiotherapy. A. Hybridization

[0093] Methods of the present disclosure may use nucleic acids that hybridize to other nucleic acids under particular hybridization conditions. Various methods may be used for hybridizing nucleic acids. See, e.g., Current Protocols in Molecular Biology, John Wiley and Sons, N.Y. (1989), 6.3.1-6.3.6, which is incorporated by reference herein in its entirety. Methods of the present disclosure may use a moderately stringent hybridization condition using a prewashing solution containing 5× sodium chloride / sodium citrate (SSC), 0.5% SDS, 1.0 mM EDTA (pH 8.0), hybridization buffer of about 50% formamide, 6×SSC, and a hybridization temperature of 55° C.299183876.1 - 19 -(or other similar hybridization solutions, such as one containing about 50% formamide, with a hybridization temperature of 42° C), and washing conditions of 60° C. in 0.5×SSC, 0.1% SDS. A stringent hybridization condition hybridizes in 6×SSC at 45° C., followed by one or more washes in 0.1×SSC, 0.2% SDS at 68° C. Furthermore, one can manipulate the hybridization and / or washing conditions to increase or decrease the stringency of hybridization such that nucleic acids comprising nucleotide sequence that are at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identical to each other may remain hybridized to each other.

[0094] The parameters affecting the choice of hybridization conditions and guidance for devising suitable conditions may be described by, for example, Sambrook, Fritsch, and Maniatis (Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., chapters 9 and 11 (1989); Current Protocols in Molecular Biology, Ausubel et al., eds., John Wiley and Sons, Inc., sections 2.10 and 6.3-6.4 (1995), each of which is herein incorporated by reference in their entirety) and can be readily determined based on, for example, the length and / or base composition of the DNA. B. Probes In another aspect, nucleic acid molecules are suitable for use as primers or hybridization probes for the detection or purification of nucleic acid sequences.

[0095] Probes based on the desired sequence of a nucleic acid can be used to detect the nucleic acid or similar nucleic acids, for example, transcripts encoding a polypeptide of interest. The probe can comprise a label group, e.g., a radioisotope, a fluorescent compound, an enzyme, or an enzyme co-factor. Such probes can be used to isolate or purify certain nucleic acids. C. Sequencing

[0096] DNA, including bisulfite-converted DNA, may be used for the amplification of the region of interest followed by sequencing. Primers can be designed around the CpG island and used for PCR amplification of bisulfite-converted DNA. The resulting PCR products may be cloned and sequenced. Accordingly, aspects of the disclosure may include sequencing nucleic acids to detect methylation of nucleic acids and / or biomarkers. In some aspects, the methods of the disclosure include a sequencing method.299183876.1 - 20 -D. Additional Assay Methods

[0097] In some aspects, methods involve amplifying and / or sequencing one or more target genomic regions or transcripts, including any described herein, using at least one pair of primers specific to the target genomic regions or transcripts. In certain aspects, the primers are 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or more (or any range derivable therein) nucleotides. In other aspects, enzymes are added such as primases or primase / polymerase combination enzyme to the amplification step to synthesize primers.

[0098] In some aspects, arrays can be used to detect nucleic acids of the disclosure. An array comprises a solid support with nucleic acid probes attached to the support. Arrays may comprise a plurality of different nucleic acid probes that are coupled to a surface of a substrate in different, known locations. These arrays, also described as “microarrays” or colloquially "chips", may be described by, for example, U.S. Pat. Nos. 5,143,854, 5,445,934, 5,744,305, 5,677,195, 6,040,193, 5,424,186, and Fodor et al., 1991), each of which is incorporated by reference in its entirety. Techniques for the synthesis of these arrays using mechanical synthesis methods are described in, e.g., U.S. Pat. No. 5,384,261, incorporated herein by reference in its entirety. Although a planar array surface is used in certain aspects, the array may be fabricated on a surface of virtually any shape or even a multiplicity of surfaces. Arrays may be nucleic acids on beads, gels, polymeric surfaces, fibers such as fiber optics, glass or any other appropriate substrate, see U.S. Pat. Nos. 5,770,358, 5,789,162, 5,708,153, 6,040,193 and 5,800,992, each of which is incorporated by reference herein in their entirety.

[0099] In addition to the use of arrays and microarrays, it is contemplated that a number of difference assays may be employed to analyze nucleic acids. Such assays include, but are not limited to, nucleic amplification, polymerase chain reaction, quantitative PCR, RT-PCR, in situ hybridization, digital PCR, dd PCR (digital droplet PCR), nCounter (nanoString), BEAMing (Beads, Emulsions, Amplifications, and Magnetics) (Inostics), ARMS (Amplification Refractory Mutation Systems), RNA-Seq, TAm-Seg (Tagged-Amplicon deep sequencing), PAP (Pyrophosphorolysis-activation polymerization), next generation RNA sequencing, northern hybridization, hybridization protection assay (HPA)(GenProbe), branched DNA (bDNA) assay (Chiron), rolling circle amplification (RCA), single molecule hybridization detection (US Genomics), Invader assay (ThirdWave Technologies), and / or Bridge Litigation Assay (Genaco).299183876.1 - 21 -

[0100] Amplification primers or hybridization probes can be prepared to be complementary to a genomic region, biomarker, probe, or oligo described herein. The term "primer" or “probe” as used herein, is meant to encompass any nucleic acid that is capable of priming the synthesis of a nascent nucleic acid in a template-dependent process and / or pairing with a single strand of an oligo of the disclosure, or portion thereof. Primers may be oligonucleotides from ten to twenty and / or thirty nucleic acids in length, but longer sequences can be employed. Primers may be provided in double-stranded and / or single-stranded form.

[0101] The use of a probe or primer of between 13 and 100 nucleotides, particularly between 17 and 100 nucleotides in length, or in some aspects up to 1-2 kilobases or more in length, allows the formation of a duplex molecule that is both stable and selective. Molecules having complementary sequences over contiguous stretches greater than 20 bases in length may be used to increase stability and / or selectivity of the hybrid molecules obtained. One may design nucleic acid molecules for hybridization having one or more complementary sequences of 20 to 30 nucleotides, or even longer where desired. Such fragments may be readily prepared, for example, by directly synthesizing the fragment by chemical approaches or by introducing selected sequences into recombinant vectors for recombinant production.

[0102] In one aspect, each probe / primer comprises at least 15 nucleotides. For instance, each probe can comprise at least or at most 20, 25, 50, 75, 100, 125, 150, 175, 200, 225, 250, 275, 300, 325, 350, 400 or more nucleotides (or any range derivable therein). They may have these lengths and have a sequence that is identical or complementary to a gene described herein. Particularly, each probe / primer has relatively high sequence complexity and does not have any ambiguous residue (undetermined "n" residues). The probes / primers can hybridize to the target gene, including its RNA transcripts, under stringent or highly stringent conditions. It is contemplated that probes or primers may have inosine or other design implementations that accommodate recognition of more than one human sequence for a particular biomarker.

[0103] For applications requiring high selectivity, one may desire to employ relatively high stringency conditions to form the hybrids. For example, relatively low salt and / or high temperature conditions, such as provided by about 0.02 M to about 0.10 M NaCl at temperatures of about 50°C to about 70°C. Such high stringency conditions tolerate little, if any, mismatch between the probe or primers and the template or target strand and may be particularly suitable for isolating specific299183876.1 - 22 -genes or for detecting specific mRNA transcripts. It is generally appreciated that conditions can be rendered more stringent by the addition of increasing amounts of formamide.

[0104] In one aspect, quantitative RT-PCR (such as TaqMan, ABI) is used for detecting and comparing the levels or abundance of nucleic acids in samples. The concentration of the target DNA in the linear portion of the PCR process is proportional to the starting concentration of the target before the PCR was begun. By determining the concentration of the PCR products of the target DNA in PCR reactions that have completed the same number of cycles and are in their linear ranges, it is possible to determine the relative concentrations of the specific target sequence in the original DNA mixture. This direct proportionality between the concentration of the PCR products and the relative abundances in the starting material is true in the linear range portion of the PCR reaction. The final concentration of the target DNA in the plateau portion of the curve is determined by the availability of reagents in the reaction mix and is independent of the original concentration of target DNA. Therefore, the sampling and quantifying of the amplified PCR products may be carried out when the PCR reactions are in the linear portion of their curves. In addition, relative concentrations of the amplifiable DNAs may be normalized to some independent standard / control, which may be based on either internally existing DNA species or externally introduced DNA species. The abundance of a particular DNA species may also be determined relative to the average abundance of all DNA species in the sample.

[0105] In one aspect, the PCR amplification utilizes one or more internal PCR standards. The internal standard may be an abundant housekeeping gene in the cell or it can specifically be GAPDH, GUSB and β-2 microglobulin. These standards may be used to normalize expression levels so that the expression levels of different gene products can be compared directly. An internal standard may be used to normalize expression levels. III. Cancer Therapy

[0106] Certain methods disclosed herein comprise administering a therapy, including a cancer therapy to the patient. The cancer therapy may be chosen based on copy number, RNA, or protein level measurements, alone or in combination with a calculated score for the patient. In some aspects, the cancer therapy comprises a local cancer therapy. In some aspects, the cancer therapy excludes a systemic cancer therapy. In some aspects, the cancer therapy excludes a local therapy. In some aspects, the cancer therapy comprises a local cancer therapy without the administration of299183876.1 - 23 -a system cancer therapy. In some aspects, the cancer therapy comprises an immunotherapy, which may be an immune checkpoint therapy. Any of these cancer therapies may also be excluded. Combinations of these therapies may also be administered.

[0107] The term “cancer,” as used herein, may be used to describe a solid tumor, metastatic cancer, or non-metastatic cancer. In certain aspects, the cancer may originate in the bladder, blood, bone, bone marrow, brain, breast, colon, esophagus, duodenum, small intestine, large intestine, colon, rectum, anus, gum, head, kidney, liver, lung, nasopharynx, neck, ovary, pancreas, prostate, skin, stomach, testis, tongue, or uterus. In some aspects, the cancer is recurrent cancer. In some aspects, the cancer is Stage I cancer. In some aspects, the cancer is Stage II cancer. In some aspects, the cancer is Stage III cancer. In some aspects, the cancer is Stage IV cancer.

[0108] The cancer may specifically be of the following histological type, though it is not limited to these: neoplasm, malignant; carcinoma; carcinoma, undifferentiated; giant and spindle cell carcinoma; small cell carcinoma; papillary carcinoma; squamous cell carcinoma; lymphoepithelial carcinoma; basal cell carcinoma; pilomatrix carcinoma; transitional cell carcinoma; papillary transitional cell carcinoma; adenocarcinoma; gastrinoma, malignant; cholangiocarcinoma; hepatocellular carcinoma; combined hepatocellular carcinoma and cholangiocarcinoma; trabecular adenocarcinoma; adenoid cystic carcinoma; adenocarcinoma in adenomatous polyp; adenocarcinoma, familial polyposis coli; solid carcinoma; carcinoid tumor, malignant; branchiolo-alveolar adenocarcinoma; papillary adenocarcinoma; chromophobe carcinoma; acidophil carcinoma; oxyphilic adenocarcinoma; basophil carcinoma; clear cell adenocarcinoma; granular cell carcinoma; follicular adenocarcinoma; papillary and follicular adenocarcinoma; nonencapsulating sclerosing carcinoma; adrenal cortical carcinoma; endometroid carcinoma; skin appendage carcinoma; apocrine adenocarcinoma; sebaceous adenocarcinoma; ceruminous adenocarcinoma; mucoepidermoid carcinoma; cystadenocarcinoma; papillary cystadenocarcinoma; papillary serous cystadenocarcinoma; mucinous cystadenocarcinoma; mucinous adenocarcinoma; signet ring cell carcinoma; infiltrating duct carcinoma; medullary carcinoma; lobular carcinoma; inflammatory carcinoma; paget’s disease, mammary; acinar cell carcinoma; adenosquamous carcinoma; adenocarcinoma w / squamous metaplasia; thymoma, malignant; ovarian stromal tumor, malignant; thecoma, malignant; granulosa cell tumor, malignant; androblastoma, malignant; sertoli cell carcinoma; leydig cell tumor, malignant; lipid cell tumor, malignant; paraganglioma, malignant; extra-mammary299183876.1 - 24 -paraganglioma, malignant; pheochromocytoma; glomangiosarcoma; malignant melanoma; amelanotic melanoma; superficial spreading melanoma; malignant melanoma in giant pigmented nevus; epithelioid cell melanoma; blue nevus, malignant; sarcoma; fibrosarcoma; fibrous histiocytoma, malignant; myxosarcoma; liposarcoma; leiomyosarcoma; rhabdomyosarcoma; embryonal rhabdomyosarcoma; alveolar rhabdomyosarcoma; stromal sarcoma; mixed tumor, malignant; mullerian mixed tumor; nephroblastoma; hepatoblastoma; carcinosarcoma; mesenchymoma, malignant; brenner tumor, malignant; phyllodes tumor, malignant; synovial sarcoma; mesothelioma, malignant; dysgerminoma; embryonal carcinoma; teratoma, malignant; struma ovarii, malignant; choriocarcinoma; mesonephroma, malignant; hemangiosarcoma; hemangioendothelioma, malignant; kaposi’s sarcoma; hemangiopericytoma, malignant; lymphangiosarcoma; osteosarcoma; juxtacortical osteosarcoma; chondrosarcoma; chondroblastoma, malignant; mesenchymal chondrosarcoma; giant cell tumor of bone; ewing's sarcoma; odontogenic tumor, malignant; ameloblastic odontosarcoma; ameloblastoma, malignant; ameloblastic fibrosarcoma; pinealoma, malignant; chordoma; glioma, malignant; ependymoma; astrocytoma; protoplasmic astrocytoma; fibrillary astrocytoma; astroblastoma; glioblastoma; oligodendroglioma; oligodendroblastoma; primitive neuroectodermal; cerebellar sarcoma; ganglioneuroblastoma; neuroblastoma; retinoblastoma; olfactory neurogenic tumor; meningioma, malignant; neurofibrosarcoma; neurilemmoma, malignant; granular cell tumor, malignant; malignant lymphoma; hodgkin’s disease; hodgkin’s; paragranuloma; malignant lymphoma, small lymphocytic; malignant lymphoma, large cell, diffuse; malignant lymphoma, follicular; mycosis fungoides; other specified non-hodgkin’s lymphomas; malignant histiocytosis; multiple myeloma; mast cell sarcoma; immunoproliferative small intestinal disease; leukemia; lymphoid leukemia; plasma cell leukemia; erythroleukemia; lymphosarcoma cell leukemia; myeloid leukemia; basophilic leukemia; eosinophilic leukemia; monocytic leukemia; mast cell leukemia; megakaryoblastic leukemia; myeloid sarcoma; and hairy cell leukemia. A. Immunotherapies

[0109] In some aspects, the therapeutic treatment comprises a cancer immunotherapy. Cancer immunotherapy (sometimes called immuno-oncology, abbreviated IO) is the use of the immune system to treat cancer. Immunotherapies can be categorized as active, passive or hybrid (active and passive). These approaches exploit the fact that cancer cells often have molecules on their surface that can be detected by the immune system, known as tumour-associated antigens (TAAs);299183876.1 - 25 -they are often proteins or other macromolecules (e.g. carbohydrates). Active immunotherapy directs the immune system to attack tumor cells by targeting TAAs. Passive immunotherapies enhance existing anti-tumor responses and include the use of monoclonal antibodies, lymphocytes and cytokines. Immumotherapies are known in the art, and some are described below. B. Chemotherapies

[0110] In some aspects, the therapeutic treatment comprises a chemotherapy. Suitable classes of chemotherapeutic agents include (a) Alkylating Agents, such as nitrogen mustards (e.g., mechlorethamine, cylophosphamide, ifosfamide, melphalan, chlorambucil), ethylenimines and methylmelamines (e.g., hexamethylmelamine, thiotepa), alkyl sulfonates (e.g., busulfan), nitrosoureas (e.g., carmustine, lomustine, chlorozoticin, streptozocin) and triazines (e.g., dicarbazine), (b) Antimetabolites, such as folic acid analogs (e.g., methotrexate), pyrimidine analogs (e.g., 5-fluorouracil, floxuridine, cytarabine, azauridine) and purine analogs and related materials (e.g., 6-mercaptopurine, 6-thioguanine, pentostatin), (c) Natural Products, such as vinca alkaloids (e.g., vinblastine, vincristine), epipodophylotoxins (e.g., etoposide, teniposide), antibiotics (e.g., dactinomycin, daunorubicin, doxorubicin, bleomycin, plicamycin and mitoxanthrone), enzymes (e.g., L-asparaginase), and biological response modifiers (e.g., Interferon-α), and (d) Miscellaneous Agents, such as platinum coordination complexes (e.g., cisplatin, carboplatin), substituted ureas (e.g., hydroxyurea), methylhydiazine derivatives (e.g., procarbazine), and adreocortical suppressants (e.g., taxol and mitotane). In some aspects, cisplatin is a particularly suitable chemotherapeutic agent.

[0111] Cisplatin has been widely used to treat cancers such as, for example, metastatic testicular or ovarian carcinoma, advanced bladder cancer, head or neck cancer, cervical cancer, lung cancer or other tumors. Cisplatin is not absorbed orally and must therefore be delivered via other routes such as, for example, intravenous, subcutaneous, intratumoral or intraperitoneal injection. Cisplatin can be used alone or in combination with other agents, with efficacious doses used in clinical applications including about 15 mg / m2 to about 20 mg / m2 for 5 days every three weeks for a total of three courses being contemplated in certain aspects. In some aspects, the amount of cisplatin delivered to the cell and / or subject in conjunction with the construct comprising an Egr-1 promoter operably linked to a polynucleotide encoding the therapeutic polypeptide is less than the amount that would be delivered when using cisplatin alone.299183876.1 - 26 -

[0112] Other suitable chemotherapeutic agents include antimicrotubule agents, e.g., Paclitaxel (“Taxol”) and doxorubicin hydrochloride (“doxorubicin”). The combination of an Egr-1 promoter / TNFα construct delivered via an adenoviral vector and doxorubicin was determined to be effective in overcoming resistance to chemotherapy and / or TNF-α, which suggests that combination treatment with the construct and doxorubicin overcomes resistance to both doxorubicin and TNF-α.

[0113] Doxorubicin is absorbed poorly and is preferably administered intravenously. In certain aspects, appropriate intravenous doses for an adult include about 60 mg / m2 to about 75 mg / m2 at about 21-day intervals or about 25 mg / m2 to about 30 mg / m2 on each of 2 or 3 successive days repeated at about 3 week to about 4 week intervals or about 20 mg / m2 once a week. The lowest dose should be used in elderly patients, when there is prior bone-marrow depression caused by prior chemotherapy or neoplastic marrow invasion, or when the drug is combined with other myelopoietic suppressant drugs.

[0114] Nitrogen mustards are another suitable chemotherapeutic agent useful in the methods of the disclosure. A nitrogen mustard may include, but is not limited to, mechlorethamine (HN2), cyclophosphamide and / or ifosfamide, melphalan (L-sarcolysin), and chlorambucil. Cyclophosphamide (CYTOXAN®) is available from Mead Johnson and NEOSTAR® is available from Adria), is another suitable chemotherapeutic agent. Suitable oral doses for adults include, for example, about 1 mg / kg / day to about 5 mg / kg / day, intravenous doses include, for example, initially about 40 mg / kg to about 50 mg / kg in divided doses over a period of about 2 days to about 5 days or about 10 mg / kg to about 15 mg / kg about every 7 days to about 10 days or about 3 mg / kg to about 5 mg / kg twice a week or about 1.5 mg / kg / day to about 3 mg / kg / day. Because of adverse gastrointestinal effects, the intravenous route is preferred. The drug also sometimes is administered intramuscularly, by infiltration or into body cavities.

[0115] Additional suitable chemotherapeutic agents include pyrimidine analogs, such as cytarabine (cytosine arabinoside), 5-fluorouracil (fluouracil; 5-FU) and floxuridine (fluorode- oxyuridine; FudR).5-FU may be administered to a subject in a dosage of anywhere between about 7.5 to about 1000 mg / m2. Further, 5-FU dosing schedules may be for a variety of time periods, for example up to six weeks, or as determined by one of ordinary skill in the art to which this disclosure pertains.299183876.1 - 27 -

[0116] Gemcitabine diphosphate (GEMZAR®, Eli Lilly & Co., “gemcitabine”), another suitable chemotherapeutic agent, is recommended for treatment of advanced and metastatic pancreatic cancer, and will therefore be useful in the present disclosure for these cancers as well.

[0117] The amount of the chemotherapeutic agent delivered to the patient may be variable. In one suitable aspect, the chemotherapeutic agent may be administered in an amount effective to cause arrest or regression of the cancer in a host, when the chemotherapy is administered with the construct. In other aspects, the chemotherapeutic agent may be administered in an amount that is anywhere between 2 to 10,000 fold less than the chemotherapeutic effective dose of the chemotherapeutic agent. For example, the chemotherapeutic agent may be administered in an amount that is about 20 fold less, about 500 fold less or even about 5000 fold less than the chemotherapeutic effective dose of the chemotherapeutic agent. The chemotherapeutics of the disclosure can be tested in vivo for the desired therapeutic activity in combination with the construct, as well as for determination of effective dosages. For example, such compounds can be tested in suitable animal model systems prior to testing in humans, including, but not limited to, rats, mice, chicken, cows, monkeys, rabbits, etc. In vitro testing may also be used to determine suitable combinations and dosages, as described in the examples. C. Radiotherapy

[0118] In some aspects, the therapeutic treatment comprises radiation, such as ionizing radiation. As used herein, “ionizing radiation” means radiation comprising particles or photons that have sufficient energy or can produce sufficient energy via nuclear interactions to produce ionization (gain or loss of electrons). An exemplary and preferred ionizing radiation is an x- radiation. Means for delivering x-radiation to a target tissue or cell are well known in the art.

[0119] In some aspects, the amount of ionizing radiation is greater than 20 Gy and is administered in one dose. In some aspects, the amount of ionizing radiation is 18 Gy and is administered in three doses. In some aspects, the amount of ionizing radiation is at least, at most, or exactly 2, 4, 6, 8, 10, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 18, 19, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 40 Gy (or any derivable range therein). In some aspects, the ionizing radiation is administered in at least, at most, or exactly 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 does (or any derivable range therein). When more than one dose is administered, the does may be about 1, 4, 8, 12, or 24 hours or 1, 2, 3, 4, 5, 6, 7, or 8 days or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 14, or 16 weeks apart, or any derivable range therein.299183876.1 - 28 -

[0120] In some aspects, the amount of IR may be presented as a total dose of IR, which is then administered in fractionated doses. For example, in some aspects, the total dose is 50 Gy administered in 10 fractionated doses of 5 Gy each. In some aspects, the total dose is 50-90 Gy, administered in 20-60 fractionated doses of 2-3 Gy each. In some aspects, the total dose of IR is at least, at most, or about 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40,41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 125, 130, 135, 140, or 150 (or any derivable range therein). In some aspects, the total dose is administered in fractionated doses of at least, at most, or exactly 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 14, 15, 20, 25, 30, 35, 40, 45, or 50 Gy (or any derivable range therein. In some aspects, at least, at most, or exactly 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40,41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100 fractionated doses are administered (or any derivable range therein). In some aspects, at least, at most, or exactly 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 (or any derivable range therein) fractionated doses are administered per day. In some aspects, at least, at most, or exactly 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 (or any derivable range therein) fractionated doses are administered per week. D. Surgery

[0121] Approximately 60% of persons with cancer will undergo surgery of some type, which includes preventative, diagnostic or staging, curative, and palliative surgery. Curative surgery includes resection in which all or part of cancerous tissue is physically removed, excised, and / or destroyed and may be used in conjunction with other therapies, such as the treatment of the present aspects, chemotherapy, radiotherapy, hormonal therapy, gene therapy, immunotherapy, and / or alternative therapies. Tumor resection refers to physical removal of at least part of a tumor. In addition to tumor resection, treatment by surgery includes laser surgery, cryosurgery, electrosurgery, and microscopically-controlled surgery (Mohs’ surgery).

[0122] Upon excision of part or all of cancerous cells, tissue, or tumor, a cavity may be formed in the body. Treatment may be accomplished by perfusion, direct injection, or local application of299183876.1 - 29 -the area with an additional anti-cancer therapy. Such treatment may be repeated, for example, every 1, 2, 3, 4, 5, 6, or 7 days, or every 1, 2, 3, 4, and 5 weeks or every 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months. These treatments may be of varying dosages as well. IV. Administration of Therapeutic Compositions

[0123] In some aspects, a therapeutic composition is administered to a patient before, during, or after the detection and / or measurement of one or more markers, including any marker described herein such as the radiotherapy-resistance markers. In certain aspects, a therapeutic composition is administered to a patient when the patient is determined to have abnormal expression of the one or more markers. Abnormal expression may be an increase or decrease of the marker(s) relative to a control, including any control described herein.

[0124] The therapy provided herein may comprise administration of a combination of therapeutic agents, such as a first therapy and a second therapy. The therapies may be administered in any suitable manner known in the art. For example, the first and second treatment may be administered sequentially (at different times) or concurrently (at the same time). In some aspects, the first and second treatments are administered in a separate composition. In some aspects, the first and second treatments are in the same composition.

[0125] Aspects of the disclosure relate to compositions and methods comprising therapeutic compositions. The different therapies may be administered in one composition or in more than one composition, such as 2 compositions, 3 compositions, or 4 compositions. Various combinations of the agents may be employed.

[0126] The therapeutic agents of the disclosure may be administered by the same route of administration or by different routes of administration. In some aspects, the therapy is administered intravenously, intramuscularly, subcutaneously, topically, orally, transdermally, intraperitoneally, intraorbitally, by implantation, by inhalation, intrathecally, intraventricularly, or intranasally. In some aspects, the antibiotic is administered intravenously, intramuscularly, subcutaneously, topically, orally, transdermally, intraperitoneally, intraorbitally, by implantation, by inhalation, intrathecally, intraventricularly, or intranasally. The appropriate dosage may be determined based on the type of disease to be treated, severity and course of the disease, the clinical condition of the individual, the individual's clinical history and response to the treatment, and the discretion of the attending physician.299183876.1 - 30 -

[0127] The treatments may include various “unit doses.” Unit dose is defined as containing a predetermined-quantity of the therapeutic composition. The quantity to be administered, and the particular route and formulation, is within the skill of determination of those in the clinical arts. A unit dose need not be administered as a single injection but may comprise continuous infusion over a set period of time. In some aspects, a unit dose comprises a single administrable dose.

[0128] Precise amounts of the therapeutic composition also depend on the judgment of the practitioner and are peculiar to each individual. Factors affecting dose include physical and clinical state of the patient, the route of administration, the intended goal of treatment (alleviation of symptoms versus cure) and the potency, stability and toxicity of the particular therapeutic substance or other therapies a subject may be undergoing. V. Pharmaceutical Compositions

[0129] In certain aspects, the compositions or agents for use in the methods, such as therapeutic agents for inhibiting POLQ, are suitably contained in a pharmaceutically acceptable carrier. The carrier is non-toxic, biocompatible and is selected so as not to detrimentally affect the biological activity of the agent. The agents in some aspects of the disclosure may be formulated into preparations for local delivery (i.e. to a specific location of the body, such as skeletal muscle or other tissue) or systemic delivery, in solid, semi-solid, gel, liquid or gaseous forms such as tablets, capsules, powders, granules, ointments, solutions, depositories, inhalants and injections allowing for oral, parenteral or surgical administration. Certain aspects of the disclosure also contemplate local administration of the compositions by coating medical devices and the like.

[0130] Suitable carriers for parenteral delivery via injectable, infusion or irrigation and topical delivery include distilled water, physiological phosphate-buffered saline, normal or lactated Ringer's solutions, dextrose solution, Hank's solution, or propanediol. In addition, sterile, fixed oils may be employed as a solvent or suspending medium. For this purpose any biocompatible oil may be employed including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid find use in the preparation of injectables. The carrier and agent may be compounded as a liquid, suspension, polymerizable or non-polymerizable gel, paste or salve.

[0131] The carrier may also comprise a delivery vehicle to sustain (i.e., extend, delay or regulate) the delivery of the agent(s) or to enhance the delivery, uptake, stability or pharmacokinetics of the therapeutic agent(s). Such a delivery vehicle may include, by way of non-299183876.1 - 31 -limiting examples, microparticles, microspheres, nanospheres or nanoparticles composed of proteins, liposomes, carbohydrates, synthetic organic compounds, inorganic compounds, polymeric or copolymeric hydrogels and polymeric micelles.

[0132] In certain aspects, the actual dosage amount of a composition administered to a patient or subject can be determined by physical and physiological factors such as body weight, severity of condition, the type of disease being treated, previous or concurrent therapeutic interventions, idiopathy of the patient and on the route of administration. The practitioner responsible for administration will, in any event, determine the concentration of active ingredient(s) in a composition and appropriate dose(s) for the individual subject.

[0133] Solutions of pharmaceutical compositions can be prepared in water suitably mixed with a surfactant, such as hydroxypropylcellulose. Dispersions also can be prepared in glycerol, liquid polyethylene glycols, mixtures thereof and in oils. Under ordinary conditions of storage and use, these preparations contain a preservative to prevent the growth of microorganisms.

[0134] In certain aspects, the pharmaceutical compositions are advantageously administered in the form of injectable compositions either as liquid solutions or suspensions; solid forms suitable or solution in, or suspension in, liquid prior to injection may also be prepared. These preparations also may be emulsified. A typical composition for such purpose comprises a pharmaceutically acceptable carrier. For instance, the composition may contain 10 mg or less, 25 mg, 50 mg or up to about 100 mg of human serum albumin per milliliter of phosphate buffered saline. Other pharmaceutically acceptable carriers include aqueous solutions, non-toxic excipients, including salts, preservatives, buffers and the like.

[0135] Examples of non-aqueous solvents are propylene glycol, polyethylene glycol, vegetable oil and injectable organic esters such as ethyloleate. Aqueous carriers include water, alcoholic / aqueous solutions, saline solutions, parenteral vehicles such as sodium chloride, Ringer's dextrose, etc. Intravenous vehicles include fluid and nutrient replenishers. Preservatives include antimicrobial agents, antgifungal agents, anti-oxidants, chelating agents and inert gases. The pH and exact concentration of the various components the pharmaceutical composition are adjusted according to well-known parameters.

[0136] Additional formulations are suitable for oral administration. Oral formulations include such typical excipients as, for example, pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharine, cellulose, magnesium carbonate and the like. The299183876.1 - 32 -compositions take the form of solutions, suspensions, tablets, pills, capsules, sustained release formulations or powders.

[0137] In further aspects, the pharmaceutical compositions may include classic pharmaceutical preparations. Administration of pharmaceutical compositions according to certain aspects may be via any common route so long as the target tissue is available via that route. This may include oral, nasal, buccal, rectal, vaginal or topical. Alternatively, administration may be by orthotopic, intradermal, subcutaneous, intramuscular, intraperitoneal or intravenous injection. Such compositions would normally be administered as pharmaceutically acceptable compositions that include physiologically acceptable carriers, buffers or other excipients. For treatment of conditions of the lungs, aerosol delivery can be used. Volume of the aerosol is between about 0.01 ml and 0.5 ml.

[0138] An effective amount of the pharmaceutical composition is determined based on the intended goal. The term “unit dose” or “dosage” refers to physically discrete units suitable for use in a subject, each unit containing a predetermined-quantity of the pharmaceutical composition calculated to produce the desired responses discussed above in association with its administration, i.e., the appropriate route and treatment regimen. The quantity to be administered, both according to number of treatments and unit dose, depends on the protection or effect desired.

[0139] Precise amounts of the pharmaceutical composition also depend on the judgment of the practitioner and are peculiar to each individual. Factors affecting the dose include the physical and clinical state of the patient, the route of administration, the intended goal of treatment (e.g., alleviation of symptoms versus cure) and the potency, stability and toxicity of the particular therapeutic substance. VII. Kits

[0140] Certain aspects of the present invention also concern kits containing compositions of the invention or compositions to implement methods of the invention. In some aspects, kits can be used to evaluate one or more biomarkers. In certain aspects, a kit contains, contains at least or contains at most 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 100, 500, 1,000 or more probes, primers or primer sets, synthetic molecules, antibodies, or inhibitors,299183876.1 - 33 -or any value or range and combination derivable therein. In some aspects, there are kits for evaluating biomarker activity or level in a cell.

[0141] Kits may comprise components, which may be individually packaged or placed in a container, such as a tube, bottle, vial, syringe, or other suitable container means.

[0142] Individual components may also be provided in a kit in concentrated amounts; in some aspects, a component is provided individually in the same concentration as it would be in a solution with other components. Concentrations of components may be provided as 1x, 2x, 5x, 10x, or 20x or more.

[0143] Kits for using probes, antibodies, synthetic nucleic acids, nonsynthetic nucleic acids, and / or inhibitors of the disclosure for prognostic or diagnostic applications are included as part of the disclosure. Specifically contemplated are any such molecules corresponding to any biomarker identified herein, which includes antibodies that bind to such biomarkers as well as nucleic acid primers / primer sets and probes that are identical to or complementary to all or part of a biomarker, which may include noncoding sequences of the biomarker, as well as coding sequences of the biomarker.

[0144] In certain aspects, negative and / or positive control nucleic acids, antibodies, probes, and inhibitors are included in some kit aspects. In addition, a kit may include a sample that is a negative or positive control for methylation of one or more biomarkers.

[0145] It is contemplated that any method or composition described herein can be implemented with respect to any other method or composition described herein and that different aspects may be combined. The claims originally filed are contemplated to cover claims that are multiply dependent on any filed claim or combination of filed claims. Examples

[0146] The following examples are included to demonstrate certain aspects of the disclosure. It should be appreciated by those of skill in the art that the techniques disclosed in the examples which follow represent techniques discovered by the inventor to function well in the practice of the invention, and thus can be considered to constitute preferred modes for its practice. However, those of skill in the art should, in light of the present disclosure, appreciate that many changes can be made in the specific embodiments which are disclosed and still obtain a like or similar result without departing from the spirit and scope of the invention.299183876.1 - 34 -Example 1. Radioresistance induces widespread genomic instability.

[0147] To characterize the molecular determinants of radioresistance in prostate cancer, the inventors used isogenic DU145 radioresistant cells created by mimicking CF and HF treatment schedules18,78. The inventors performed DNA whole-genome sequencing (WGS), RNA sequencing and whole-cell and organellar proteomics (FIG. 1A) to characterize the molecular response to radiation. The inventors first evaluated the number of mutations induced by radiation across the entire genome. CF-resistant cells acquired ~60,000 somatic single nucleotide variants (SNVs) per biological replicate after radiotherapy – mutations absent in the parental cells (FIG. 1B). In contrast, HF-resistant cells acquired fewer, at ~30,000 new SNVs (P = 0.1, Mann–Whitney U test; FIG. 1B).

[0148] Relative to the typical few thousand SNVs in newly diagnosed localized prostate cancers61,79the number of radiotherapy-associated point mutations was very large. To validate this result, the inventors next created CF-radioresistant 22Rv1 cells, and again performed DNA sequencing. The inventors saw a comparable increase in SNV mutations / Mbp of genomic DNA in this second model system (FIG. 6A). CF-resistant cells experienced widespread thymine to cytosine mutations, whereas cytosine to thymine predominated in HF-resistant cells (Supplementary Table 5). Both CF- and HF-resistant cells contained multiple driver-affecting SNVs, including two separate point mutations in MTOR in CF-resistant cells (FIG. 1C).

[0149] Independent of fractionation, mutations occurring in radioresistant cells showed strong signatures of defective DNA mismatch repair, but there were fractionation-dependent differences in the signature types: SBS20 and SBS26 for CF-resistant cells and SBS14, SBS15 and SBS44 for HF (FIG. 1D). Upregulation of proliferating cell nuclear antigen (PCNA) transcript, and dysregulation of polymerase delta (POLD1) isoforms were detected in CF-resistant cells (FIG.7), consistent with the association of the CF-exclusive signature SBS20 with POLD180–82. The inventors again confirmed these results in CF-radioresistant 22Rv1 cells (FIG. 6B). SBS5 was conserved between CF- and HF (FIG. 1D).

[0150] Radioresistance was also associated with increased genomic instability in double- stranded breaks, with significant additional structural variation (SV; FIG. 1E). While the total number of SVs was similar between CF- and HF-resistant cells, there were no common SV regions. (FIG.1F). No SVs appeared to directly perturb known cancer driver genes, although a subset was associated with gene expression changes (FIG.6C). For example, a chromosome 10 inversion that299183876.1 - 35 -disrupted MYOF and GRK5 in HF-resistant cells led to the presence of MYOF-GRK5 fusion transcripts solely in these cells (FIG. 1G; Supplementary Table 6). Example 2. Extensive transcriptional and post-transcriptional responses to radiotherapy

[0151] We next considered global RNA abundance profiling, where similar numbers of transcripts were detected in all groups (Supplementary Table 7; FIG. 8A) with similar global patterns between groups (FIG. 8B). Hundreds to thousands of specific transcripts were differentially abundant between parental and radioresistant groups (FIGS. 8C-8E). Consistent with DNA sequencing, the transcriptome of HF-resistant cells was less perturbed than that of CF- resistant cells (271 vs.1,416 significant transcripts at the level of FDR ≤ 0.05; FIG.8F). This was particularly evident in cancer-driver genes, in which almost all changes occurred in CF-resistant but not HF-resistant cells (FIG. 2A; Supplementary Table 1). Most of these RNA changes translated to differential protein abundance in HF (FIG. 2B) or CF (FIG. 2C) cells. Most notable of these was CDH1, a classic prostate cancer driver gene that its loss has been previously associated with radioresistance83. Intriguingly CDH1 was down-regulated at the RNA level only in CF-resistant cells but not in HF-resistant cells (FIG.2A). By contrast, it was significantly down- regulated in protein for both fractionation schedules, suggesting differing mechanisms (FIGS.2B- C). Supporting this, cancer hallmark genes were preferentially changed in radioresistant cells (FIG. 8G), and distinguished fractionation schedules (FIG. 2D) much more clearly than did the global transcriptome (FIG. 8B). Multiple pathways were specifically dysregulated in radioresistant cells (FIG. 2E), with CF-resistant cells being more disrupted, consistent with genomic and univariate transcriptomic data. Similar numbers of significant RNA abundance changes in drivers occurred in CF-resistant 22Rv1 cells. Table 1. Prostate cancer cohort characteristics ICGC NCCSmedian (range) Age at treatment 66 (42 - 83) 71 (66 – 75)6 65 6299183876.1 - 36 -7 305 109 8 8 34es theed with CF radioresistance17, and consistently here the inventors identified four UCA1 isoforms heavily upregulated in CF-resistant cells, but none in HF-resistant cells (FIG. 2F). The RNA counts of the oncogenic lncRNA THOR84, were very low in all samples and did not yield significant changes following radiotherapy (FIG. 2F). The lncRNA SCHLAP1, which promotes aggressive prostate cancer85, was not detected in any of the samples.

[0153] To further characterize post-transcriptional signaling responses to radiation, the inventors then quantified global microRNAs (miRNA) abundance in each condition. Similar miRNA abundance patterns were observed between groups (FIGS. 8H-8I). The inventors299183876.1 - 37 -identified 23 differentially abundant miRNAs (FIG. 8J; Supplementary Table 8). CF and HF- exclusive miRNAs targeted 14 and 11 percent of the exclusive CF and HF dysregulated genes, respectively (FIG. 8K), suggesting that RNA abundance differences in CF vs. HF are at least in part caused by differential regulation of miRNA “master regulators”. The inventors highlighted the miRNA target ZEB2, which promotes radioresistance via recombination-dependent DNA repair86, and its regulation by miRNA200b-3p and miRNA 200c-3p (FIG. 2F). Taken together, these data demonstrate wide-spread, fraction-specific modes of radioresistance that are significantly shaped by post-transcriptional regulatory processes. Example 3. Protein subcellular responses to radiotherapy are fractionation dependent

[0154] To better evaluate post-transcriptional regulation in radioresistance, the inventors performed subcellular fractionation to enrich for distinct organelles47, which were independently analyzed by proteomics (Supplementary Tables 9-10). Using network analysis, the inventors identified 28 gene modules (groups of co-expressed genes among two data sets) with disparate abundance patterns across fractions (FIG. 3A). Most were more abundant in the cytosols of HF- resistant cells, but conversely were more abundant in other subcellular compartments of CF- resistant cells. Several of these fraction-biased networks were preferentially associated with specific pathways (FIG.3B). For example, genes involved in RNA splicing ( “dark grey” module) were preferentially cytosolic in HF-resistant cells but nuclear in CF-resistant cells.

[0155] To understand how specific proteins were associated with radioresistance, the inventors performed differential proteomic abundance analysis in each fraction and in whole cell lysates. Consistent with the DNA and RNA findings, CF-resistant cells were significantly more perturbed in whole cell lysates (FIGS. 9A-9B). This difference was driven by all subcellular fractions (besides the mitochondria, where no dysregulation was observed for both cell types), and especially by the nucleus and the plasma membrane (FIGS. 9A-9N). Across fractions, a total of 132 cancer driver genes showed proteomic dysregulation (FIG. 3C; Supplementary Table 3), larger than at either the DNA or RNA levels (FIG. 10C). As a specific example, CD44 has been proposed as a potential driver of radioresistance18. Its transcriptional dysregulation was restricted to five isoforms identified in CF-resistant cells, and four in HF-resistant cells (FIG. 10A); these isoforms have distinctive functional roles87. Similarly, its protein dysregulation was exclusively299183876.1 - 38 -reflected in the nuclear soluble fraction (FIG. 10B). This highlights the isoform- and subcellular compartment-specific changes induced by radioresistance.

[0156] Consistent with the presence of significant post-transcriptional and translational components of radioresistance, RNA and protein abundances were only weakly correlated (FIG. 10C; left panel). The median correlation was 0.15, lower than the typical 0.3 observed in primary prostate cancers67. By contrast, these correlations strengthened significantly when only cancer hallmark genes were considered, most prominently in CF cells (FIG. 10C; right panel). This suggests that transcriptional regulation was an important mode of radioresistance for cancer- related genes. Indeed, a large group of driver genes that were transcriptomically dysregulated were proteomically dysregulated in one or more specific subcellular compartments (FIG. 10D). Fully 28% of drivers that showed RNA and protein changes in CF cells were members of the green module (FIG. 10D), suggesting that specific network modules may reflect specific regulatory patterns. Taken together, these data demonstrate that radioresistance significantly reshapes post- transcriptional, translational and post-translational processes. Example 4. Primary patient data highlights POLQ as a mediator of radioresistance

[0157] To determine which genes associated with radioresistance in pre-clinical model systems might influence primary patient phenotypes, the inventors interrogated the 382 patient ICGC PRAD-CA dataset. This cohort includes patients treated with curative intent, either by surgery or radiotherapy (Table 1). Of 291 pre-clinical candidates selected based on the cell line investigations (Methods; Supplementary Table 11), 28 were prognostic of biochemical recurrence (BCR) (FIG. 4A). Out of these 28 genes, the inventors chose to focus on POLQ, due to its well-known role in double-strand DNA (dsDNA) break repair88. In cell lines, POLQ transcripts were upregulated strongly in CF resistant cells (FIG. 2A), and modestly in HF (Supplementary Table 2). Amplification of POLQ was associated with significantly worse prognosis in treatment-naïve prostate cancer (FIG. 4B; HR [Hazard Ratio]= 2.49, FDR = 7.78 x 10-3, CI [confidence interval] = 1.54-4.02). The inventors validated this finding by demonstrating an increased association of POLQ RNA abundance with BCR in two independent cohorts: 1) the NCCS cohort of 185 treatment-naïve prostate cancers treated with radiotherapy (Table 1; FIG. 4C; HR = 2.4, P = 0.06, CI = 0.96-5.71) 2) a cohort by Ross-Adams et al. with 93 patients treated with RP (FIG. 11A; HR = 1.9, P = 0.06, CI = 0.93-3.70).299183876.1 - 39 -

[0158] To understand the molecular consequences of POLQ dysregulation, the inventors evaluated the74association of POLQ RNA abundance with known prostate cancer drivers and prognostic biomarkers in ICGC PRAD-CA61,62,66,67. High POLQ RNA abundance was associated with increased mutation rates in many pre-clinical candidates and known drivers, including MYC and RAD21 (Linear regression, FDR £ 0.05; FIG. 4D; FIG. 12). Increased POLQ RNA abundances were also associated with amplifications in POLQ itself, suggesting that the dysregulation in POLQ at the RNA level is driven, at least partly, by POLQ amplification. Positive RNA-RNA correlations were wide-spread between POLQ and multiple cancer driver genes (FIG. 4E), most notably BRCA2 (FIG. 4F). The inventors validated this finding in the same 185 patient independent NCCS cohort (FIG. 4G). POLQ RNA abundances were also significantly higher in tumor vs. normal samples (FIG. 11B), linked to pathologic T-categories (FIG. 11C) and closely associated with grade in two independent cohorts (FIG. 4H; Supplementary Table 12; FIG. 11D). Collectively, the inventors revealed a widespread association of POLQ with somatic and clinical features of prostate tumors that marked POLQ as the top radioresistance modulator candidate.

[0159] Next, to functionally validate the role of POLQ in radioresistance, the inventors knocked it down in parental, CF- and HF-resistant cells using siRNA, followed by irradiation (FIG. 5A; FIGS. 13A-13C). POLQ knockdown significantly radiosensitized all three cell lines (FIG. 5B). POLQ pharmacologic inhibition with the inhibitor novobiocin89, yielded similar radiosensitization (FIG. 5C). Proteomic profiling of cells following siRNA mediated POLQ knockdown revealed widespread changes (FIG. 13D), and dysregulation of multiple signaling pathways (FIG. 5D; Supplementary Table 13). A signature of 12 individual strongly dysregulated proteins (FDR ≤ 0.05) defined POLQ-inactivation (FIG. 5E; FIGS. 13E-13G). Opposite dysregulation of these genes in untreated CF-resistant cells suggests their involvement in radioresistance, while POLQ depletion reverses their dysregulation, leading to radiosensitization (FIG. 5E, right two panels).

[0160] To validate the 12-gene POLQ inhibition signature (FIG. 5D), the inventors used transcriptomic66,68,70and proteomic67,69profiling of primary prostate tumors. The inventors first calculated the RNA abundance correlations between the signature genes and POLQ. Significant positive correlations between all signature genes and POLQ were identified in TCGA, with a mix of positive and negative correlations in the other small datasets used (Supplementary Table 14;299183876.1 - 40 -FIG. 5F). All 12 signature genes showed transcriptomic proteomic differential abundance (FIG. 5F, right panel), while two were associated individually with BCR: PSAT1 and CLPX deletion (FIGS. 13H-13I). Taken together, these data establish that POLQ is associated with radioresistance in pre-clinical model systems, that its pharmacologic or genetic inhibition reverses this phenotype, and that it is associated with a gene expression signature that predicts aggressive cancer * * *

[0161] All of the methods disclosed and claimed herein can be made and executed without undue experimentation in light of the present disclosure. While the compositions and methods of this invention have been described in terms of preferred embodiments, it will be apparent to those of skill in the art that variations may be applied to the methods and in the steps or in the sequence of steps of the method described herein without departing from the concept, spirit and scope of the invention. More specifically, it will be apparent that certain agents which are both chemically and physiologically related may be substituted for the agents described herein while the same or similar results would be achieved. All such similar substitutes and modifications apparent to those skilled in the art are deemed to be within the spirit, scope and concept of the invention as defined by the appended claims.299183876.1 - 41 -REFERENCES The references provided herein, to the extent that they provide exemplary procedural or other details supplementary to those set forth herein, are specifically incorporated herein by reference. 1. Siegel, R. L., Miller, K. D., Fuchs, H. E. & Jemal, A. Cancer statistics, 2022. CA. Cancer J. Clin. 72, 7–33 (2022). 2. Hamdy, F. C. et al. 10-Year Outcomes after Monitoring, Surgery, or Radiotherapy for Localized Prostate Cancer. N. Engl. J. Med. 375, 1415–1424 (2016). 3. Schymura, M. J. et al. Factors associated with initial treatment and survival for clinically localized prostate cancer: results from the CDC-NPCR Patterns of Care Study (PoC1). BMC Cancer 10, 152 (2010). 4. De Ruysscher, D. et al. Radiotherapy toxicity. Nat. Rev. Dis. Primer 5, 13 (2019). 5. Reisz, J. 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Claims

WHAT IS CLAIMED IS:

1. A method of treating cancer in a patient, the method comprising administering a non- radiotherapy treatment to the patient, wherein the patient has been determined to have a change in at least one radiotherapy-resistance marker, wherein the radiotherapy-resistance marker comprises NCOA2, MAP3K1, PCMTD1, PIK3CB, LPP, IL7R, POLQ, SALL4, RBM38, FANCC, TNC, GNAQ, NF2, PTCH1, NUP214, EWSR1, GNAS, PIM1, MYH9, RBFOX2, EP300, or a combination thereof.

2. The method of claim 1, wherein radiotherapy-resistance marker further comprises MYC, RAD21, or a combination thereof.

3. The method of claim 1 or 2, wherein the patient is a human patient.

4. The method of any one of claims 1 to 3, wherein the patient has, is suspected of having, has been diagnosed with having, or has symptoms of prostate cancer.

5. The method of any one of claims 1 to 4, wherein the patient has received conventional fractionation radiotherapy.

6. The method of any one of claims 1 to 4, wherein the patient has received hypofractionation radiotherapy.

7. The method of any one of claims 1 to 6, wherein the radiotherapy-resistance marker is POLQ.

8. The method of any one of claims 1 to 7, wherein the change comprises a genetic amplification of the radiotherapy-resistance marker.

9. The method of any one of claims 1 to 8, wherein the change comprises an increase in RNA of the radiotherapy-resistance marker.

10. The method of any one of claims 1 to 9, wherein the change comprises an increase in protein of the radiotherapy-resistance marker.

11. The method of any one of claims 1 to 10, wherein the change comprises an amplification of POLQ.

12. The method of any one of claims 1 to 11, wherein the change comprises an increase in POLQ protein.299183876.1 - 50 -13. A method of treating cancer in a patient, the method comprising administering a POLQ inhibitor to the patient.

14. The method of claim 13, wherein the patient has received a radiotherapy treatment.

15. The method of claim 13 or 14, wherein the patient receives a radiotherapy treatment after the administering of the POLQ inhibitor.

16. The method of any one of claims 13 to 15, further comprising administering a radiotherapy treatment to the patient after administering the POLQ inhibitor to the patient.

17. The method of any one of claims 13 to 16, wherein the radiotherapy treatment comprises conventional fractionation radiotherapy.

18. The method of any one of claims 13 to 16, wherein the radiotherapy treatment comprises hypofractionation radiotherapy.

19. The method of any one of claims 13 to 18, wherein patient is a human patient.

20. The method of any one of claims 13 to 19, wherein the patient has, is suspected of having, has been diagnosed with having, or has symptoms of prostate cancer.

21. The method of any one of claims 13 to 20, wherein the POLQ inhibitor comprises an RNAi inhibitor.

22. The method of any one of claims 13 to 21, wherein the POLQ inhibitor comprises novobiocin and / or ART4215.

23. A method of treating a radiotherapy-resistant cancer in a patient, the method comprising administering a POLQ inhibitor to the patient, wherein the patient has received a radiotherapy treatment.

24. A method of sensitizing radiotherapy-resistant cancer cells to a radiotherapy, the method comprising delivering a POLQ inhibitor to the cells.

25. A method of measuring radiotherapy-resistance markers, the method comprising measuring at least one gene product of a radiotherapy-resistance marker in a biological sample obtained from a patient, wherein the radiotherapy-resistance marker is NCOA2, MAP3K1, PCMTD1, PIK3CB, LPP, IL7R, POLQ, SALL4, RBM38, FANCC, TNC, GNAQ, NF2, PTCH1, NUP214, EWSR1, GNAS, PIM1, MYH9, RBFOX2, EP300, or a combination thereof.299183876.1 - 51 -26. The method of claim 25, wherein the patient is a human patient.

27. The method of claim 25 or 26, wherein the patient has, is suspected of having, has been diagnosed with having, or has symptoms of prostate cancer.

28. The method of any one of claims 25 to 27, wherein the patient has received conventional fractionation radiotherapy.

29. The method of any one of claims 25 to 27, wherein the patient has received hypofractionation radiotherapy.

30. A method of treating a prostate cancer patient, the method comprising administering a non- radiotherapy to the patient, wherein the patient is determined to have a POLQ amplification or increase in a POLQ gene product in cancer cells from the patient.

31. The method of claim 30, wherein the POLQ gene product is POLQ mRNA.

32. The method of claim 30 or 31, wherein the POLQ gene product is POLQ protein.

33. A method of prognosing radiotherapy-resistance in a cancer patient, the method comprising measuring at least one gene product of a radiotherapy-resistance marker in a biological sample obtained from the patient, wherein the radiotherapy-resistance marker is NCOA2, MAP3K1, PCMTD1, PIK3CB, LPP, IL7R, POLQ, SALL4, RBM38, FANCC, TNC, GNAQ, NF2, PTCH1, NUP214, EWSR1, GNAS, PIM1, MYH9, RBFOX2, EP300, or a combination thereof.

34. The method of claim 33, wherein the patient is a human patient.

35. The method of claim 33 or 34, wherein the patient has, is suspected of having, has been diagnosed with having, or has symptoms of prostate cancer.

36. The method of any one of claims 33 to 35, wherein the patient has received conventional fractionation radiotherapy.

37. The method of any one of claims 33 to 35, wherein the patient has received hypofractionation radiotherapy.

38. A kit comprising one or more detection agents for determining levels of one or more radiotherapy-resistance markers, wherein the radiotherapy-resistance marker is NCOA2, MAP3K1, PCMTD1, PIK3CB, LPP, IL7R, POLQ, SALL4, RBM38, FANCC, TNC, GNAQ,299183876.1 - 52 -NF2, PTCH1, NUP214, EWSR1, GNAS, PIM1, MYH9, RBFOX2, EP300, or a combination thereof.

39. The kit of claim 38, wherein the kit further comprises one or more negative or positive control samples and / or control detection agents.

40. The kit of claim 38 or 39, wherein the kit further comprises instructions for use.

41. The kit of any one of claims 38 to 40, wherein the kit excludes reagents for detection of other biomarkers.

42. The kit of any one of claims 38 to 41, wherein the detection agents comprise primers capable of hybridizing the one or more radiotherapy-resistance markers.

43. The kit of any one of claims 38 to 42, wherein the detection agents comprise primers capable of amplifying the one or more radiotherapy-resistance markers.

44. The kit of any one of claims 38 to 43, wherein the detection agents comprise an antigen-binding protein capable of binding the one or more radiotherapy-resistance markers.299183876.1 - 53 -

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