Biomarkers in pre-operative prostate cancer

Abstract

Provided herein are biomarkers for calculating a risk score (e.g., a PORTOS score) and / or determining an expected response to treatment comprising dose-escalated radiation therapy in a subject with prostate cancer who has not undergone radical prostatectomy (e.g., intermediate risk prostate cancer), methods of using the biomarkers including methods of treatment based on the risk score and / or expected response to treatment comprising dose-escalated radiation therapy determined using the biomarkers, as well as kits for detecting the biomarkers.

Description

GENDX.036WO / 720 PCTBIOMARKERS IN PRE-OPERATIVE PROSTATE CANCERFIELD

[0001] The present disclosure pertains to the field of prostate cancer.BACKGROUND

[0002] Globally, prostate cancer accounts for a fifth of male cancer-related deaths. Definitive radiation therapy is one of the primary’ treatment modalities for localized prostate cancer. The radiation course historically consisted of a conventionally fractionated dose of around 70 Gy. However, numerous randomized controlled trials established an improvement in biochemical control with dose escalation to around 80 Gy. Based on these data, escalated dose has become the new standard of care for conventionally fractionated radiation. However, large meta-analyses have failed to demonstrate a metastasis-free survival (MFS) or overall survival (OS) benefit. No clinically validated biomarkers exist which can distinguish between the patients that truly derive a benefit from dose-escalation, and those that could be safely deescalated.

[0003] This background information is provided for the purpose of making known information believed by the applicant to be of possible relevance to the present disclosure. No admission is necessarily intended, nor should be construed, that any of the preceding information constitutes prior art against the present disclosure.SUMMARY

[0004] Provided herein are biomarkers for determining an expected response to treatment comprising dose-escalated radiation therapy in a subject with prostate cancer (e.g., intermediate risk prostate cancer) who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate), methods of using the biomarkers including methods of treatment based on the expected response to treatment comprising dose-escalated radiation therapy determined using the biomarkers, as well as kits for detecting the biomarkers.

[0005] Some embodiments provided herein are described by way of the following numbered embodiments:1. A method comprising:a. obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy, wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23; andb, calculating or having calculated a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and optionally determining a response to treatment comprising dose-escalated radiation therapy that the subject with prostate cancer who has not undergone radical prostatectomy is expected to have based on at least the risk score. A method comprising:a. obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy, wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of: KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23; andb. determining a response to treatment comprising dose-escalated radiation therapy that the subject with prostate cancer who has not undergone radical prostatectomy is expected to have based on at least the expression level of the one target or each of the plurality of targets in the sample in step (a. ).A method comprising:a. obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy, wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected fromthe group of genes consisting of: KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23;b. determining or having determined a response to treatment comprising dose-escalated radiation therapy that the subject with prostate cancer who has not undergone radical prostatectomy is expected to have based on at least the expression level of the one target or each of the plurality of targets in the sample in step (a.), optionally wherein the determining or having determined comprises calculating or having calculated a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets and the determining or having determined is based on the risk score; andc. administering or having administered to the subject with prostate cancer who has not undergone radical prostatectomy a treatment selected from:i. a treatment comprising dose-escalated radiation therapy; or ii. a treatment comprising non-dose-escalated radiation therapy, wherein the treatment is selected based on the expected response to treatment comprising dose-escalated radiation therapy determined in (b.). A method comprising:a. obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy, wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of: KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23;b. calculating or having calculated a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets of (a.); and c. administering or having administered to the subject with prostate cancer who has not undergone radical prostatectomy in a treatment selected from:i. a treatment comprising dose-escalated radiation therapy; or ii. a treatment comprising non-dose-escalated radiation therapy, wherein the treatment is selected based on the risk score of (b.).5. The method of any one of embodiment 1-3, wherein the response to treatment comprising dose-escalated radiation therapy comprises risk of one or more of: overall survival, distant metastasis, metastasis progression-free survival, metastasis-free survival, biochemical recurrence-free survival, failure-free survival, and prostate cancer specific mortality, optionally biochemical failure (BF), distant metastasis (DM) and receipt of salvage therapy, optionally wherein the risk is at 3, 5, 10 or 15 years post-treatment with the radiation therapy, optionally wherein the risk is at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 years, or a range defined by any two of the preceding values, post-treatment with the radiation therapy.6. The method of any one of embodiments 3-5, wherein the treatment (i.) comprising dose-escalated radiation therapy further comprises administration of an additional treatment for prostate cancer, and / or wherein the treatment (ii.) comprising non-dose-escalated radiation therapy further comprises an additional treatment for prostate cancer.7. The method of any one of embodiments 1-6, wherein the one target or plurality of targets comprises or consists of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23 or 24 target nucleic acid sequences, optionally wherein each target nucleic acid sequence is selected from a different gene selected from the group of genes consisting of: KRT14, EMEI, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23.8. The method of any one of embodiments 1-7, wherein the one target or plurality of targets comprises or consists of a nucleic acid sequence of each of:a. KRT14;b. KRT14, and EME1;c. KRT14, EME1, and DRAM1;d. KRT14, EME1, DRAM1, and ANLN;e. KRT14, EMEI, DRAM1, ANLN, and APEX2;f. KRT14, EMEI, DRAM1, ANLN, APEX2, and DTL;g. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, and ARHGAP15;h. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, and GNG11; i. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, and CDKN2AIP;j. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, and PTPN22;k. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, and RPS27A;l. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, and CDKN3;m. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, and IL1B;n. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, and NEK1;o. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, and UBA7;p. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, and MUM1; q. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, and BIN2;r. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, and TOP2A;s. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, and PLK2;t. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, and ZMAT3;u. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, and SULF2;v. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, and IL7R;w. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, and HCLS1; orx. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23.9. The method of any one of embodiments 1-3 and 5-8, wherein said determining or having determined comprises calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, optionally wherein said determining or having determined is based on the risk score.10. The method of any one of the preceding embodiments, wherein said determining or having determined comprises determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non¬ dose-escalated radiation therapy.11. The method of any one of embodiments 1-9, wherein said determining or having determined comprises determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy.12. The method of any one of embodiments 1 and 3-11, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and wherein said determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy:a. is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or greater than, 0, - 0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, - 0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, -0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.42 on a scale from -1 to 1; orb. is expected to respond no better or worse to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from - 1 to 1, the risk score being equal to or less than 0, -0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, -0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, -0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.41 on a scale from -1 to 1.The method of embodiment 12, further comprising:a. selecting for administration and / or administering, or having administered, to the subject expected to respond better to treatment comprising dose-escalated radiation therapy a treatment comprising dose-escalated radiation therapy; or b. selecting for administration and / or administering, or having administered, to the subject expected to respond no better or worse to treatment comprising dose-escalated radiation therapy a treatment comprising non-dose-escalated radiation therapy.14. The method of any one of embodiments 1-12, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and wherein the method comprises:a. selecting for administration and / or administering, or having administered, to the subject a treatment comprising dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; and ii. on a scale from -1 to 1, the risk score being equal to or greater than, 0, - 0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, - 0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, - 0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.42 on a scale from -1 to 1; orb. selecting for administration and / or administering, or having administered, to the subject a treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or less than 0, -0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, -0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, -0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.41 on a scale from -1 to 1.15. The method of any one of embodiments 1 -9, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the pluralityof targets, and wherein said determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy:a. is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or greater than, - 0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, - 0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, - 0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.67 on a scale from -1 to 1; orb. is expected to respond no better or worse to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or less than, -0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.68 on a scale from -1 to 1.16. The method of embodiment 15, further comprising:a. selecting for administration and / or administering, or having administered, to the subject expected to respond better to treatment comprising dose-escalated radiation therapy a treatment comprising dose-escalated radiation therapy; orb. selecting for administration and / or administering, or having administered, to the subject expected to respond no better or worse to treatment comprising dose- escalated radiation therapy a treatment comprising non-dose-escalated radiation therapy.17. The method of any one of embodiments 1 -9 or 15, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and wherein the method comprises:a. selecting for administration and / or administering, or having administered, to the subject a treatment comprising dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or greater than, - 0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, - 0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, - 0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.67 on a scale from -1 to 1; orb. selecting for administration and / or administering, or having administered, to the subject a treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or less than, -0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.68 on a scale from -1 to 1.18. The method of any one of embodiments 1 -9, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and wherein said determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy:a. is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; and ii. on a scale from -1 to 1, the risk score being equal to or greater than, 0, - 0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, - 0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, -0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.42 on a scale from -1 to 1; orb. is expected to respond no better or worse to treatment comprising dose- escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or less than, -0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.68 on a scale from -1 to 1.19. The method of embodiment 18, further comprising:a. selecting for administration and / or administering, or having administered, to the subject expected to respond better to treatment comprising dose-escalated radiation therapy a treatment comprising dose-escalated radiation therapy; or b. selecting for administration and / or administering, or having administered, to the subject expected to respond no better or worse to treatment comprising dose- escalated radiation therapy a treatment comprising non-dose-escalated radiation therapy.20. The method of any one of embodiments 1-9 or 18, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and wherein the method comprises:a. selecting for administration and / or administering, or having administered, to the subject a treatment comprising dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; and ii. on a scale from -1 to 1, the risk score being equal to or greater than, 0, - 0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, - 0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, - 0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.42 on a scale from -1 to 1; orb. selecting for administration and / or administering, or having administered, to the subject a treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or less than, -0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75,-0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.68 on a scale from -1 to 1.21. The method of any one of the preceding embodiments, comprising determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy:a. is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on at least the risk score being greater than 0 on a scale from - 1 to 1; or b. is expected to respond no better or worse to treatment comprising dose- escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on at least the risk score being less than or equal to 0 on a scale from -1 to 1,22. The method of any one of the preceding embodiments, wherein the method comprisesa. selecting for administration and / or administering, or having administered, to the subject a treatment comprising dose-escalated radiation therapy based on at least the risk score being greater than 0 on a scale from -1 to 1; or b. selecting for administration and / or administering, or having administered, to the subject a treatment comprising non-dose-escalated radiation therapy based at least on the risk score being equal to or less than 0 on a scale from -1 to 1. 23. The method of any one of embodiments 10-22, wherein the population of patients with prostate cancer who have not undergone radical prostatectomy comprises at least 10, 25, 50, 75 or 100 patients, optionally wherein the population of patients does not have metastatic prostate cancer, and / or the population of patients has intermediate-risk prostate cancer, optionally wherein the population patients have: histologically confirmed adenocarcinoma of the prostate, clinical stage Tla-T2b (stage I or II), Gleason score 2-6 and prostate-specific antigen (PSA) 10 to less than 20 ng / mL, or Gleason score 7 and PSA less than 15 ng / mL, and no distant metastases and no regional lymph node involvement.24. The method of any one of embodiments 1 -23, wherein the subject is determined to be expected to respond better to treatment comprising dose-escalated radiation therapy thanto treatment comprising non-dose-escalated radiation therapy, optionally wherein the subject is selected for administration and / or is administered a treatment comprising dose-escalated radiation therapy.25. The method of any one of embodiments 1 -23, wherein the subject is determined to be expected to respond no better or worse to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy, optionally wherein the subject is selected for administration and / or is administered a treatment comprising non- dose-escalated radiation therapy.26. The method of any one of embodiments 1-25, wherein the expected better response to treatment comprising dose-escalated radiation therapy is one or more of:a. a reduction in risk of BF of at least 5%, 10%, 20%, or 30% as compared to treatment comprising non-dose-escalated radiation therapy;b. a reduction in risk of DM of at least 5%, 10%, 20%, or 30% as compared to treatment comprising non-dose-escalated radiation therapy; and c. a reduction in risk of receipt of salvage therapy of at least 5%, 10%, 20%, or 30% as compared to treatment comprising non-dose-escalated radiation therapy;optionally wherein the risk is at 3, 5, 10 or 15 years post-treatment with the radiation therapy, optionally wherein the risk is at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 years, or a range defined by any two of the preceding values, post-treatment with the radiation therapy.27. The method of any one of embodiments 1 -26, wherein the expected no better or worse response to treatment comprising dose-escalated radiation therapy is one or more of:a. reduction in risk of BF of less than 5%, 0%, -5%, or -10% as compared to treatment comprising non-dose-escalated radiation therapy;b. reduction in risk of DM of less than 5%, 0%, -5%, or -10% as compared to treatment comprising non-dose-escalated radiation therapy; and c. reduction in risk of receipt of salvage therapy of less than 5%, 0%, -5%, or - 10% as compared to treatment comprising non-dose-escalated radiation therapy;optionally wherein the risk is at 3, 5, 10 or 15 years post-treatment with the radiation therapy, optionally wherein the risk is at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 years, or a range defined by any two of the preceding values, post-treatment with the radiation therapy.28. The method of any one of embodiments 5-27, wherein the BF is Pheonix BF.29. The method of any one of the preceding embodiments, wherein the subject does not have metastatic prostate cancer, and / or the subject has intermediate- risk prostate cancer, optionally wherein the subject has a PSA score of < 20 ng / ml, a clinical stage of T1 or T2, and / or a Gleason grade group of 1, 2 or 3.30. The method of any one of the preceding embodiments, wherein the subject satisfies the following criteria:a. histologically confirmed adenocarcinoma of the prostate,b. clinical stage Tl -T2b (stage I or II),c. Gleason score 2-6 and prostate-specific antigen (PSA) 10 to less than 20 ng / mL, or Gleason score 7 and PSA less than 15 ng / mL,d. no distant metastases, ande. no regional lymph node involvement.31. The method of any one of the preceding embodiments, wherein the subject has one or more of the following:a. intermediate risk prostate cancer as defined by the NCCN risk groups; b. low risk prostate cancer as defined by the NCCN risk groups;c. high or very high risk prostate cancer as defined by the NCCN risk groups; d. one or more of the following risk factors: clinical stage T2b- T2c, Grade Group 2 or 3 (Gleason 7), PSA 10-20;e. clinical stage cT1-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL; f. at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL; and / org. metastatic prostate cancer.32. The method of any one of the preceding embodiments, wherein the subject does not have one or more of the following:a. intermediate risk prostate cancer as defined by the NCCN risk groups; b. low risk prostate cancer as defined by the NCCN risk groups;c. high or very high risk prostate cancer as defined by the NCCN risk groups; d. one or more of the following risk factors: clinical stage T2b- T2c, Grade Group 2 or 3 (Gleason 7), PSA 10-20;e. clinical stage cT1-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL; f. at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL; and / org. metastatic prostate cancer.33. The method of any one of the preceding embodiments, wherein said determining or having determined the expected response to treatment comprising dose-escalated radiation therapy is in comparison to a treatment comprising non-dose-escalated radiation therapy.34. The method of any one of the preceding embodiments, wherein said determining or having determined the expected response of the subject with prostate cancer who has not undergone radical prostatectomy to treatment comprising dose-escalated radiation therapy is not based on prognostic clinical variables, optionally wherein the prognostic clinical variables are tumor stage, nodal stage, metastatic tumor volume categorization, a Gleason score and / or a serum PSA level of the subject.35. The method of any one of embodiments 1-33, wherein said determining or having determined the expected response of the subject with prostate cancer who has not undergone radical prostatectomy to treatment comprising dose-escalated radiation therapy is further based on one or more prognostic clinical variables, optionally wherein the prognostic clinical variables are tumor stage, nodal stage, metastatic tumor volume categorization, a Gleason score and / or a serum PSA level of the subject.36. The method of any one of the preceding embodiments, wherein said obtaining or having obtained the expression level of the one target or the plurality of targets comprises utilizing a microarray to assess one target or the plurality of targets, performing qPCR on the one target or the plurality of targets, and / or sequencing the one target or the plurality of targets.37. The method of any one of the preceding embodiments, wherein the sample is a resected specimen, optionally a transurethral resection of prostate (TURP) specimen, or a biopsy of the prostate cancer, optionally a needle biopsy.38. The method of any one of the preceding embodiments, wherein the sample is from a primary prostate tumor.39. The method of any one of the preceding embodiments, wherein the dose- escalated radiation therapy is at least 78 Gy, optionally at least 79 Gy or 80 Gy.40. The method of any one of the preceding embodiments, wherein the non-dose-escalated radiation therapy is not more than 71 Gy, optionally not more than 70 Gy.41. The method of any one of the preceding embodiments, wherein is selected from 3 -Dimensional Conformal Radiation Therapy (3D-CRT), Intensity -Modulated Radiation Therapy (IMRT), volumetric modulated arc therapy (VMAT), stereotactic body radiotherapy (SBRT or SABR), proton beam RT, and brachytherapy boost, optionally wherein it is 3D-CRT or IMRT.42. The method of any one of the preceding embodiments wherein the dose-escalated and / or non-dose-escalated radiation therapy is definitive radiation therapy.43. The method of any one of the preceding embodiments, wherein the dose-escalated and / or non-dose-escalated radiation therapy is not salvage radiation therapy.44. The method of any one of the preceding embodiments, wherein the risk factor score is a PORTOS score.45. The method of any one of the preceding embodiments, wherein the subject does not have NCCN unfavorable intermediate-risk prostate cancer.46. A kit for use in the method of any one of the preceding embodiments, the kit comprising reagents for obtaining the expression level of the one target or each of the plurality of targets.47. The kit of embodiment 46, wherein the reagents comprise primer(s) and / or probe(s) for obtaining the expression level of the one target or each of the plurality of targets.INCORPORATION BY REFERENCE

[0006] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference for the disclosures referenced herein and in their entireties to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference.BRIEF DESCRIPTION OF THE DRAWINGS

[0007] Figure 1 is an embodiment of a table of patient characteristics in NRG / RTOG 0126 by PORTOS tertile.

[0008] Figures 2A and 2B are embodiments of cumulative incidence curves for Biochemical Failure (BF) per (FIG. 2A) Phoenix and (FIG. 2B) ASTRO criteria comparing 79.2Gy vs. 70.2Gy stratified by PORTOS tertile in NRG / RTOG 0126.

[0009] Figures 3A and 3B are embodiments of cumulative incidence curves for (FIG. 3 A) Distant Metastasis and (FIG. 3B) Salvage Therapy comparing 79.2Gy vs. 70.2Gy stratified by PORTOS tertile in NRG / RTOG 0126

[0010] Figures 4A and 4B are embodiments of Biochemical Failure (BF) per Phoenix and ASTRO criteria forest plots showing PORTOS and clinicopathologic variables and the benefit of dose escalation in NRG / RTOG 0126.

[0011] Figure 5 is an embodiment of PORTOS scores in NCCN intermediate risk patients in commercial Decipher biopsy grouped by race, PSA, clinical stage, Gleason grade group, and favorable vs. unfavorable intermediate risk,

[0012] Figure 6 is an embodiment of DNA damage response (DDR), hypoxia, immune (Immune! 90) scores, and molecular subtypes in NCCN intermediate risk patients in commercial Decipher biopsy grouped by PORTOS tertile.

[0013] Figures 7A-7C are embodiments of Interaction plots for 5-year Biochemical Failure (BF) per (A) Phoenix and (B) ASTRO criteria, and (C) Receipt of Salvage Therapy comparing 79.2Gy vs. 70.2Gy stratified by PORTOS tertile in NRG / RTOG 0126.

[0014] Figures 8A and 8B are embodiments of Kaplan-Meier curves for Biochemical Failure (BF) per (A) Phoenix and (B) ASTRO criteria comparing 79.2Gy vs.70.2Gy stratified by PORTOS low vs. average / high tertile in NRG / RTOG 0126.

[0015] Figures 9A and 9B are embodiments of Interaction plots for (A) 5-year and (B) 10-year Biochemical Failure (BF) per Phoenix and ASTRO criteria comparing 79.2Gy vs.70.2Gy stratified by PORTOS low vs. average / high tertile in NRG / RTOG 0126.

[0016] Figures 10A and 10B are embodiments Kaplan-Meier curves for (A) Distant Metastasis and (B) Salvage Therapy comparing 79.2Gy vs. 70.2Gy stratified by PORTOS low vs. average / high tertile in NRG / RTOG 0126.

[0017] Figures 11A and 11B are embodiments of Biochemical Failure (BF) per Phoenix criteria forest plots showing subsets of PORTOS biomarkers and the benefit of dose escalation.DETAILED DESCRIPTION

[0018] Provided herein are biomarkers for calculating a risk score (e.g., a PORTOS score) and / or determining an expected response to treatment comprising dose-escalated radiation therapy in a subject with prostate cancer (e.g., intermediate risk prostate cancer per the modern NCCN risk groups) who has not undergone radical prostatectomy (or any other surgery’ to remove the cancerous prostate), methods of using the biomarkers including methods of treatment based on the risk score (e.g., a PORTOS score) and / or expected response to treatment comprising dose-escalated radiation therapy determined using the biomarkers, as well as kits for detecting the biomarkers.

[0019] Some embodiments disclosed herein are a method comprising: a.) obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate), wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of: KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23; and b.) calculating or having calculated a risk score (e.g., a PORTOS score) for the sample based on at least the expression level of the one target or each of the plurality of targets. In some embodiments, the method optionally includes determining a response to treatment comprising dose-escalated radiation therapy that the subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) is expected to have based on at least the risk score (e.g., a PORTOS score).

[0020] Some embodiments disclosed herein a method comprising: a.) obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery’ to remove the cancerous prostate), wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of: KRT14, EMEI, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEKI, UBA7, MUM1, BIN2, TOP2A,PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23; and b.) determining a response to treatment comprising dose-escalated radiation therapy that the subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) is expected to have based on at least the expression level of the one target or each of the plurality of targets in the sample in step (a.).

[0021] Some embodiments disclosed herein are a method comprising: a.) obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate), wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of: KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23; b.) determining or having determined a response to treatment comprising dose-escalated radiation therapy that the subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) is expected to have based on at least the expression level of the one target or each of the plurality of targets in the sample in step (a. ), optionally wherein the determining or having determined comprises calculating or having calculated a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets and the determining or having determined is based on the risk score; and c.) administering or having administered to the subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) a treatment selected from: i.) a treatment comprising dose-escalated radiation therapy; or ii.) a treatment comprising non-dose-escalated radiation (e.g., isodose) therapy, wherein the treatment is selected based on the expected response to treatment comprising dose-escalated radiation therapy determined in (b.).

[0022] Some embodiments disclosed herein are a method comprising: a.) obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate), wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from thegroup of genes consisting of: KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23; b.) calculating or having calculated a risk score (e.g., a PORTOS score) for the sample based on at least the expression level of the one target or each of the plurality of targets of (a.); and c.) administering or having administered to the subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) in a treatment selected from: i.) a treatment comprising dose-escalated radiation therapy; or ii.) a treatment comprising nondose-escalated radiation therapy, wherein the treatment is selected based on the risk score (e.g., a PORTOS score) of (b.),

[0023] In some embodiments, the dose-escalated radiation therapy is at least 78 Gy, optionally at least 79 Gy or 80 Gy. In some embodiments, the non-dose-escalated radiation therapy is not more than 71 Gy, optionally not more than 70 Gy. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is selected from 3-Dimensional Conformal Radiation Therapy (3D-CRT), Intensity-Modulated Radiation Therapy (IMRT), volumetric modulated arc therapy (VMAT), stereotactic body radiotherapy (SBRT or SABR), proton beam RT, and brachytherapy boost In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is 3D-CRT or IMRT. In some embodiments, the dose- escalated and / or non-dose-escalated radiation therapy is definitive radiation therapy. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is not salvage radiation therapy. In some embodiments, the subject has intermediate-risk prostate cancer. In some embodiments, the subject has a PSA score of < 20 ng / ml, a clinical stage of T1 or T2, and / or a Gleason grade group of 1, 2 or 3. In some embodiments, the subject satisfies the following: histologically confirmed adenocarcinoma of the prostate, clinical stage Tla-T2b (stage I or II), Gleason score 2-6 and prostate-specific antigen (PSA) 10 to less than 20 ng / mL or Gleason score 7 and PSA less than 15 ng / mL, no distant metastases, and no regional lymph node involvement. In some embodiments, the subject has intermediate risk prostate cancer as defined by the NCCN risk groups disclosed in Spratt, D. E. et al. Genomic Classifier Performance in Intermediate-Risk Prostate Cancer: Results From NRG Oncology / RTOG 0126 Randomized Phase 3 Trial. IntJ Radiat Oncol Biol Phys 117, 370-377 (2023), which is herein incorporated by reference in its entirety. In some embodiments, the subject does not haveNCCN unfavorable intermediate-risk prostate cancer. In some embodiments, the subject has one or more intermediate risk factors: clinical stage T2b- T2c, Grade Group 2 or 3 (Gleason 7), PSA 10-20. In some embodiments, the subject has low-risk prostate cancer. In some embodiments, the subject has clinical stage cTl-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL. In some embodiments, the subject has high / very high-risk prostate cancer. In some embodiments, the subject has at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL. In some embodiments, the subject has metastatic prostate cancer. In some embodiments, the subject does not have low-risk prostate cancer. In some embodiments, the subject does not have clinical stage cTl-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL. In some embodiments, the subject does not have high / very high-risk prostate cancer. In some embodiments, the subject does not have at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL. In some embodiments, the subject does not have metastatic prostate cancer. In some embodiments, the risk level is determined according to NCCN risk groups disclosed m Spratt et al. (2023).Biomark ers / Genes for predicting response to dose-escalated radiation therapy

[0024] In some embodiments, Table 1 is a list of biomarkers / genes for use in calculating a risk score (e.g., a PORTOS score) and / or predicting the response of prostate cancer (e.g., prostate cancer in a subject who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate)) to treatment comprising dose-escalated radiation therapy. The coefficient can be used to calculate a risk score (e.g., a PORTOS score). The importance of each gene to the risk score is shown in Table 1, with the least important gene having a score of 24.Table 1: Prostate Cancer Biomarkers / GenesImportanceGene CoefficientRankKRT14 -0.847 1EME1 1.39 2DRAM1 -0.102 3ANLN -1.233 4APEX2 0.671 5DTL 1.161 6ARHGAP15 -1.114 7GNG11 0.41 8CDKN2AIP 0.466 9PTPN22 -1.029 10RPS27A 0.364 11CDKN3 -0.848 12IL1B -1.502 13NEK1 0.678 14UBA7 0.291 15MUM1 0.444 16BIN2 0.529 17TOP2A 1.378 18PLK2 -1.294 19ZMAT3 0.118 20SULF2 -0.288 21IL7R 0.135 22HCLS1 -0.008 23KIF23 1.01 24

[0025] In some embodiments, biomarkers / genes listed in Table 1 are combined into a risk score. In some embodiments, the coefficients of this model can have positive or negative weights. In some embodiments, an elevated (e.g., relative to matched healthy tissue or low grade prostate cancer) expression level of the biomarkers / genes in Table 1 with a positive coefficient value indicates that a subject is more likely to respond more to treatment comprising dose-escalated radiation therapy, and an elevated expression level of the target selected from a biomarker / gene with a negative coefficient value indicates that a subject is likely to experience less benefit from treatment comprising dose-escalated radiation therapy. The greater the absolute Importance Rank value of the biomarker / gene, the greater the relative predictive value of the biomarker / gene to the overall model. In some embodiments the risk score is a PORTOS score. In some embodiments, a nucleic acid expression-based signature, (e.g., a signature including genes / biomarkers selected from Table I, a risk score (e.g. PORTOS), or a signature or risk score provided below and elsewhere herein) provide clinically-important information in prostate cancer in a subject who has not undergone radical prostatectomy independent of prognostic clinical variables (e.g., Gleason score, PSA, tumor grade, etc.). In some embodiments, the nucleic acid expression-based signature (e.g., risk score, PORTOS score) provides a basis for selecting patients for dose-escalated radiation therapy. In some embodiments, a nucleic acid expression-based signature, (e.g., risk score, PORTOS score) measured at diagnosis from routinely-acquired primary tumors can classify prostate cancer in a subject who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) into clinically-useful groups that predict dose-escalatedradiation therapy benefit. In some embodiments, the signature (e.g., the selected targets or risk score) is a signature disclosed in PCT / US2017 / 048486, which is herein incorporated by reference in its entirety. In some embodiments, the signature (e.g., the selected targets or risk score) comprises or consists of those used in PORTOS, disclosed in Zhao, S. G. et al. Development and validation of a 24-gene predictor of response to postoperative radiotherapy in prostate cancer: a matched, retrospective analysis. Lancet Oncol (2016) Nov;17(ll):1612-1620, which is herein incorporated by reference in its entirety. In some embodiments, the signature (e.g., the selected targets) comprises or consists of, or of at least, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23 or 24 of those used in PORTOS. In some embodiments, the signature is PORTOS as disclosed in Zhao, S. G. et al. Development and validation of a 24-gene predictor of response to postoperative radiotherapy in prostate cancer: a matched, retrospective analysis. Lancet Oncol (2016) Nov;17(11):1612-1620. In some embodiments, risk score is calculated using targets comprising or consisting of, or of at least, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23 or 24 of those used in PORTOS, and the risk score is calculated in the manner disclosed for the PORTOS score in Zhao, S. G. et al. Development and validation of a 24-gene predictor of response to postoperative radiotherapy in prostate cancer: a matched, retrospective analysis. Lancet Oncol (2016) Nov;17(ll):1612-1620 using the at least, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23 or 24 targets. As used herein, a “PORTOS” score is a score calculated using the method disclosed in Zhao, S. G. et al. Lancet Oncol (2016) Nov; 17(11): 1612-1620. Where the risk score uses fewer than all of the genes / markers used by Zhao, S. G. et al., the “PORTOS” score is the score calculated using the method disclosed in Zhao, S. G. et al. applied to the selected genes / markers.

[0026] In some embodiments, the method comprises: a.) obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate), wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of: KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23; and b.) calculating or having calculated a risk score (e.g.,a PORTOS score) for the sample based on at least the expression level of the one target or each of the plurality of targets. In some embodiments, the method optionally includes determining a response to treatment comprising dose-escalated radiation therapy that the subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) is expected to have based on at least the risk score (e.g., a PORTOS score). In some embodiments, the method comprises: a.) obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate), wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of: KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23; and b.) determining a response to treatment comprising dose-escalated radiation therapy that the subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) is expected to have based on at least the expression level of the one target or each of the plurality of targets in the sample in step (a.). In some embodiments, the method comprises: a.) obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate), wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of: KRT14, EMEI, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23; b.) determining or having determined a response to treatment comprising dose-escalated radiation therapy that the subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) is expected to have based on at least the expression level of the one target or each of the plurality of targets in the sample in step (a.), optionally wherein the determining or having determined comprises calculating or having calculated a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets and the determining or having determined is based on the risk score; and c.) administering or having administeredto the subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) a treatment selected from: i.) a treatment comprising dose-escalated radiation therapy; or ii.) a treatment comprising non-dose-escalated radiation therapy, wherein the treatment is selected based on the expected response to treatment comprising dose-escalated radiation therapy determined in (b.). In some embodiments, the method comprises: a.) obtaining or having obtained an expression level of one target or a plurality' of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate), wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of: KRT14, EMEI, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23; b.) calculating or having calculated a risk score (e.g., a PORTOS score) for the sample based on at least the expression level of the one target or each of the plurality of targets of (a.); and c.) administering or having administered to the subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) in a treatment selected from: i.) a treatment comprising dose-escalated radiation therapy; or ii. ) a treatment comprising non-dose-escalated radiation therapy, wherein the treatment is selected based on the risk score (e.g., a PORTOS score) of (b.). In some embodiments, the response to treatment comprising dose-escalated radiation therapy' comprises risk of one or more of: biochemical failure (BF), distant metastasis (DM) and receipt of salvage therapy. In some embodiments, the response to treatment comprising dose-escalated radiation therapy comprises overall survival, metastasis-free survival, progression-free survival, and / or metastatic progression-free survival. In some embodiments, the risk is at 3, 5, 10 or 15 years or a range defined by any two of the preceding values, post-treatment with the radiation therapy. In some embodiments, the risk is at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 years, or a range defined by any two of the preceding values, post-treatment with the radiation therapy. In some embodiments, the expected better response to treatment comprising dose- escalated radiation therapy is one or more of: a reduction in risk of BF of, or of at least 5%, 10%, 20%, or 30% or a range defined by any two of the preceding values, as compared to treatment comprising non-dose-escalated radiation therapy; a reduction in risk of DM of, or ofat least 5%, 10%, 20%, or 30% or a range defined by any two of the preceding values, as compared to treatment comprising non-dose-escalated radiation therapy; and a reduction in risk of receipt of salvage therapy of or of at least 5%, 10%, 20%, or 30% or a range defined by any two of the preceding values, as compared to treatment comprising non-dose-escalated radiation therapy. In some embodiments, the expected better response to treatment comprising dose-escalated radiation therapy is one or more of: an increase in overall survival, metastasis- free survival, progression-free survival, and / or metastatic progression-free survival of, or of at least 5%, 10%, 20%, or 30% or a range defined by any two of the preceding values, as compared to treatment comprising non-dose-escalated radiation therapy. In some embodiments, the risk is at 3, 5, 10 or 15 years or a range defined by any two of the preceding values, post-treatment with the radiation therapy. In some embodiments, the risk is at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 years, or a range defined by any two of the preceding values, post-treatment with the radiation therapy. In some embodiments, the expected no better or worse response to treatment comprising dose-escalated radiation therapy is one or more of: reduction in risk of BF of, or of less than 5%, 0%, -5%, or -10% or a range defined by any two of the preceding values, as compared to treatment comprising non-dose-escalated radiation therapy; reduction in risk of DM of, or of less than 5%, 0%, -5%, or -10% or a range defined by any two of the preceding values, as compared to treatment comprising non-dose-escalated radiation therapy; and reduction in risk of receipt of salvage therapy of, or of less than 5%, 0%, -5%, or -10% or a range defined by any two of the preceding values, as compared to treatment comprising non-dose-escalated radiation therapy. In some embodiments, the expected no better or worse response to treatment comprising dose-escalated radiation therapy is one or more of: increase in overall survival, metastasis-free survival, progression-free survival, and / or metastatic progression-free survival of, or of less than 5%, 0%, -5%, or -10% or a range defined by any two of the preceding values, as compared to treatment comprising non-dose-escalated radiation therapy. In some embodiments, the risk is at 3, 5, 10 or 15 years or a range defined by any two of the preceding values, post-treatment with the radiation therapy. In some embodiments, the risk is at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 years, or a range defined by any two of the preceding values, post-treatment with the radiation therapy. In some embodiments, the BF is Pheonix BF. In some embodiments, the dose-escalated radiation therapy is at least 78 Gy, optionally at least 79 Gy or 80 Gy. In some embodiments, the non-dose-escalated radiation therapy is not more than 71 Gy, optionally not more than 70 Gy. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is selected from 3-Dimensional Conformal Radiation Therapy (3D-CRT), Intensity-Modulated Radiation Therapy (IMRT), volumetric modulated arc therapy (VMAT), stereotactic body radiotherapy (SBRT or SABR), proton beam RT, and brachytherapy boost. In some embodiments, the dose- escalated and / or non-dose-escalated radiation therapy is 3D-CRT or IMRT. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is definitive radiation therapy. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is not salvage radiation therapy. In some embodiments, the subject has intermediate-risk prostate cancer. In some embodiments, the subject has a PSA score of < 20 ng / ml, a clinical stage of T1 or T2, and / or a Gleason grade group of 1, 2 or 3, In some embodiments, the subject satisfies the following: histologically confirmed adenocarcinoma of the prostate, clinical stage Tla-T2b (stage I or II), Gleason score 2-6 and prostate-specific antigen (PSA) 10 to less than 20 ng / mL or Gleason score 7 and PSA less than 15 ng / mL, no distant metastases, and no regional lymph node involvement. In some embodiments, the subject has intermediate risk prostate cancer as defined by the NCCN risk groups disclosed in Spratt, D. E. et al. Int J Radiat Oncol Biol Phys 117, 370-377 (2023). In some embodiments, the subject does not have NCCN unfavorable intermediate-risk prostate cancer. In some embodiments, the subject has one or more intermediate risk factors: clinical stage T2b- T2c, Grade Group 2 or 3 (Gleason 7), PSA 10-20. In some embodiments, the subject has low-risk prostate cancer. In some embodiments, the subject has clinical stage cTl-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL. In some embodiments, the subject has high / very high-risk prostate cancer. In some embodiments, the subject has at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL. In some embodiments, the subject has metastatic prostate cancer. In some embodiments, the subject does not have low-risk prostate cancer. In some embodiments, the subject does not have clinical stage cTl-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL. In some embodiments, the subject does not have high / very high-risk prostate cancer. In some embodiments, the subject does not have at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL. In some embodiments, the subject does not have metastatic prostate cancer. In some embodiments, the risk level is determined according to NCCN risk groups disclosed in Spratt et al. (2023). In some embodiments, the treatment (i.) comprising dose-escalatedradiation therapy further comprises administration of an additional treatment for prostate cancer. In some embodiments, the treatment (ii.) comprising non-dose-escalated radiation therapy further comprises an additional treatment for prostate cancer.

[0027] In some embodiments, the one target or plurality of targets comprises or consists of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23 or 24 target nucleic acid sequences, or a range defined by any two of the preceding values, for example, 1-24, 1-15, 1-13, 13-24, 5-24, 5-15, 10-24, 10-15, 14-24, or 18-24 target nucleic acid sequences. In some embodiments, each target nucleic acid sequence is selected from a different gene (biomarker). In some embodiments, not more than 1, 2, 3, 4, or 5 target nucleic acid sequences are from the same gene. In some embodiments, not more than 1, 2, 3, 4 or 5 genes have more than 1 target nucleic acid sequences. In some embodiments, each target nucleic acid sequence is selected from a different gene selected from the group of genes consisting of: KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23. In some embodiments, the targets are selected from biomarkers / genes in Table 1 that have coefficient values that are positive. In some embodiments, the targets are selected from biomarkers / genes in Table 1 that have coefficient values that are negative. In some embodiments, the targets are selected from biomarkers / genes in Table 1 that have coefficient values that are positive and negative. In some embodiments, the targets are selected from biomarkers / genes in Table 1 that have an absolute coefficient value that is, or is at least, 0.005, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, or 1.5, or a range defined by any two of the preceding values. In some embodiments, the targets are selected from biomarkers / genes in Table 1 that have an absolute coefficient value that is, or is at least, 0.4. In some embodiments, the targets are selected from biomarkers / genes in Table 1 that have an absolute coefficient value that is, or is at least, 1.0. In some embodiments, the targets are selected from biomarkers / genes in Table 1 that have an Importance Rank of at least 24, 23, 22, 21, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1. In some embodiments, the targets comprise or consist of at least one target nucleic acid from each of the biomarkers / genes in Table 1.

[0028] As explained in Example 2, Table 2 provides groups (subsets) of biomarkers / genes made by removing genes, starting with the least important as ranked in Table1. The Spearman correlation coefficient value for each group in Table 2 is calculated by comparing the performance of a risk score calculated using the markers in the group relative to the risk score using the markers in group x (the PORTOS score disclosed in Zhao, S. G et al. Lancet Oncol (2016) Nov;17(ll):1612-1620). Figures 11A and 11B depicts of Biochemical Failure (BF) per Phoenix criteria forest plots showing the subsets of PORTOS biomarkers listed in Table 2 and the benefit of dose escalation.Table 2: Groups (subsets) of Biomarkers / GenesSpearman Group Descriptions Genes Correlation PORTOSa -23 genes KRT14; 0.395 b -22 genes KRT14, and EMEI; 0.499 c -21 genes KRT14, EMEI, and DRAM1; 0.545 d -20 genes KRT14, EMEI, DRAM1, and ANLN; 0.698 e -19 genes KRT14, EMEI, DRAM1, ANLN, and APEX2; 0.710KRT14, EMEI, DRAM1, ANLN, APEX2, andf -18 genes 0.720DTL;KRT14, EMEI, DRAM1, ANLN, APEX2, DTL,g -17 genes 0.778and ARHGAP15;KRT14, EMEI, DRAM1, ANLN, APEX2, DTL,h -16 genes 0.799ARHGAP15, and GNG11;KRT14, EMEI, DRAM1, ANLN, APEX2, DTL,i -15 genes 0.831ARHGAP15, GNG11, and CDKN2AIP;KRT14, EMEI, DRAM1, ANLN, APEX2, DTL,J -14 genes ARHGAP15, GNG11, CDKN2AIP, and 0.870PTPN22;KRT14, EMEI, DRAM1, ANLN, APEX2, DTL,k -13 genes ARHGAP15, GNG11, CDKN2AIP, PTPN22, 0.877and RPS27A;KRT14, EME1, DRAM1, ANLN, APEX2, DTL,1 -12 genes ARHGAP15, GNG11, CDKN2AIP, PTPN22, 0.909RPS27A, and CDKN3;KRT14, EMEI, DRAM1, ANLN, APEX2, DTL,m -11 genes ARHGAP15, GNG11, CDKN2AIP, PTPN22, 0.916RPS27A, CDKN3, and IL1B;KRT14, EMEI, DR AMI, ANLN, APEX2, DTL,n -10 genes ARHGAP15, GNG11, CDKN2AIP, PTPN22, 0.931RPS27A, CDKN3, IL1B, andNEKl;KRT14, EMEI, DRAM1, ANLN, APEX2, DTL,o -9 genes ARHGAP15, GNG11, CDKN2AIP, PTPN22, 0.934RPS27A, CDKN3, IL1B, NEK1, and UBA7;Spearman Group Descriptions Genes Correlation PORTOS KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22,P -8 genes 0.934RPS27A, CDKN3, IL1B, NEK1, UBA7, and MUM1;KRT14. EMEL DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22,q -7 genes 0.935RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1,and BIN2;KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22,r -6 genes 0.960RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1,BIN2, and TOP2A;KRT14, EMEI, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22,s -5 genes 0.968RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1,BIN2, TOP2A, and PLK2;KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22,t -4 genes 0.984RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1,BIN2, TOP2A, PLK2, and ZMAT3;KRT14, EMEI, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22,u -3 genes 0.992RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1,BIN2, TOP2A, PLK2, ZMAT3, and SULF2;KRT14, EMEI, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22,V -2 genes RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, 0.997BIN2, TOP2A, PLK2, ZMAT3, SULF2, andIL7R;KRT14, EMEI, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22,w -1 genes RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, 0.997BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, and HCLS1; orKRT14, EMEI, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22,X -0 genes RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, 1.000BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R,HCLS1, and KIF23.

[0029] In some embodiments, the one target or plurality of targets comprises or consists of a nucleic acid sequence of each of the biomarkers / genes in a group selected from: groups a-x, groups b-x, groups c-x, groups d-x, groups e-x, groups f-x, groups g-x, groups h-x, groups i-x, groups j-x, groups k-x, groups 1-x, groups m-x, groups n-x, groups o-x, groups p-x, groups q-x, groups r-x, groups s-x, groups t-x, groups u-x, groups v-x, groups w-x or group x, of Table 2. In some embodiments, the one target or plurality of targets comprises or consists of a nucleic acid sequence of each of the biomarkers / genes in a group selected from: groups 1-x, groups m-x, groups n-x, groups o-x, groups p-x, groups q-x, groups r-x, groups s-x, groups t-x, groups u-x, groups v-x, groups w-x or group x, of Table 2, In some embodiments, the one target or plurality of targets comprises or consists of a nucleic acid sequence of each of the biomarkers / genes in a group selected from groups of Table 2 having a Spearman correlation value of at least 0.7, 0.75, 0.8, 0.85, 0.9, or 0.95. In some embodiments, the one target or plurality of targets comprises or consists of a nucleic acid sequence of each of the biomarkers / genes in a group selected from groups of Table 2 having a Spearman correlation value of at least 0.85, 0.9, or 0.95. In some embodiments, the one target or plurality of targets comprises or consists of a nucleic acid sequence of each of the biomarkers / genes in a group selected from groups of Table 2 having a Spearman correlation value of at least 0.9, or 0.95.

[0030] In some embodiments, the method comprises calculating a risk score (e.g., a PORTOS score) for the sample based on at least the expression level of the one target or each of the plurality of targets. In some embodiments, the determining or having determined comprises calculating a risk score (e.g., a PORTOS score) for the sample based on at least the expression level of the one target or each of the plurality of targets. In some embodiments, the determining or having determined comprises determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising nondose-escalated radiation therapy. In some embodiments, the determining or having determined comprises determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy is expected to respond better to treatment comprising dose- escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy. In some embodiments, the risk score (e.g., a PORTOS score) is on a scale from -1 to 1. In some embodiments, the risk score is a PORTOS score calculated using the method disclosedin disclosed in Zhao, S. G et al. Lancet Oncol (2016) Nov;17(11):1612-1620, applied to the one target or each of the plurality of targets. In some embodiments, the risk score (e.g., a PORTOS score) on a scale from -1 to 1 is divided into three groups, e.g., a Lower, Middle and Higher group. In some embodiments, the cutoff for the risk score between the Higher and Middle group on a scale from -1 to 1 is equal to or greater than, 0, -0.05, -0.10, -0.15, -0.20, - 0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, -0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, -0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, for example -0.32 to -0.52. In some embodiments, the cutoff for the risk score between the Higher and Middle group is equal to or greater than -0.42 on a scale from -1 to 1. In some embodiments, the risk score for the Higher group is equal to or greater than -0.42 on a scale from -1 to 1, In some embodiments, the risk score for the Middle and Lower group is equal to or less than -0.41 on a scale from -1 to 1. In some embodiments, the cutoff for the risk score between the Middle and Lower group on a scale from -1 to 1 is equal to or greater than, -0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, for example, -0.57 to -0.77. In some embodiments, the cutoff for the risk score between the Middle and Lower group is equal to or greater than -0.67 on a scale from -1 to 1. In some embodiments, the risk score for the Higher and Middle group is equal to or greater than -0.66 on a scale from -1 to 1. In some embodiments, the risk score for the Lower group is equal to or less than -0.67 on a scale from -1 to 1. In some embodiments, the risk score for the Higher group is greater than about -0.41, the risk score for the Middle group less than or equal to about -0.41 and greater than about - 0.67, and the risk score for the Lower group is less than or equal to about -0.67. In some embodiments, the subject is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on having a risk score (e.g., a PORTOS score) in the Higher group, or in the Middle or Higher group. In some embodiments, the method comprises selecting for administration and / or administering, or having administered, to the subject expected to respond better to treatment comprising dose-escalated radiation therapy a treatment comprising dose-escalated radiation therapy. In some embodiments, the subject is expected to respond no better or worse to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based having a risk score (e.g., a PORTOS score) in the Lower group, or in the Lower and Middle group. In some embodiments, the risk score (e.g., a PORTOS score) on a scale from -1 to 1 is divided into two groups, e g., a Low and High group. In some embodiments, the cutoff for the risk score between the Low and High group on a scale from -1 to 1 is equal to or less than, 0. In some embodiments, the method comprises selecting for administration and / or administering, or having administered, to the subject expected to respond no better or worse to treatment comprising dose-escalated radiation therapy a treatment comprising non-dose-escalated radiation therapy. In some embodiments, the dose-escalated radiation therapy is at least 78 Gy, optionally at least 79 Gy or 80 Gy. In some embodiments, the non-dose-escalated radiation therapy is not more than 71 Gy, optionally not more than 70 Gy, In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is selected from 3-Dimensional Conformal Radiation Therapy (3D-CRT), Intensity -Modulated Radiation Therapy (IMRT), volumetric modulated arc therapy (VMAT), stereotactic body radiotherapy (SBRT or SABR), proton beam RT, and brachytherapy boost. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is 3D-CRT or IMRT. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is definitive radiation therapy. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is not salvage radiation therapy. In some embodiments, the subject has intermediate-risk prostate cancer. In some embodiments, the subject has a PSA score of < 20 ng / ml, a clinical stage of T1 or T2, and / or a Gleason grade group of 1, 2 or 3. In some embodiments, the subject satisfies the following: histologically confirmed adenocarcinoma of the prostate, clinical stage Tla-T2b (stage I or 11), Gleason score 2-6 and prostate-specific antigen (PSA) 10 to less than 20 ng / mL or Gleason score 7 and PSA less than 15 ng / mL, no distant metastases, and no regional lymph node involvement. In some embodiments, the subject has intermediate risk prostate cancer as defined by the NCCN risk groups disclosed in Spratt, D. E. et al. IntJ Radiat Oncol Biol Phys 117, 370-377 (2023). In some embodiments, the subject does not have NCCN unfavorable intermediate-risk prostate cancer. In some embodiments, the subject has one or more intermediate risk factors: clinical stage T2b- T2c, Grade Group 2 or 3 (Gleason 7), PSA 10-20. In some embodiments, the subject has low-risk prostate cancer. In some embodiments, the subject has clinical stage cTl-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL. In some embodiments, the subjecthas high / very high-risk prostate cancer. In some embodiments, the subject has at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL. In some embodiments, the subject has metastatic prostate cancer. In some embodiments, the subject does not have low-risk prostate cancer. In some embodiments, the subject does not have clinical stage cTl-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL. In some embodiments, the subject does not have high / very high-risk prostate cancer. In some embodiments, the subject does not have at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL. In some embodiments, the subject does not have metastatic prostate cancer. In some embodiments, the risk level is determined according to NCCN risk groups disclosed in Spratt et al. (2023). Embodiments utilizing the Higher / Middle / Lower grouping described above and elsewhere herein, include the following non-limiting embodiments.

[0031] Higher v, Middle / Lower: In some embodiments, the method comprises calculating a risk score (e g., a PORTOS score) for the sample based on at least the expression level of the one target or each of the plurality of targets, and the determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate):a. is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate); and ii. on a scale from -1 to 1, the risk score being equal to or greater than, 0, -0.05, -0.10, - 0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, -0.36, -0.37, -0.38, -0.39, - 0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, -0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.42 on a scale from -1 to 1; orb. is expected to respond no better or worse to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate); and ii. on a scale from -1 to 1, the risk score being equal to or less than 0, -0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, -0.36, -0.37, -0.38, -0.39, -0.40, - 0.41, -0.42, -0.43, -0.44, -0.45, -0.46, -0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0,41 on a scale from -1 to 1.

[0032] Higher / Middle v. Lower: In some embodiments, the method comprises calculating a risk score (e g., a PORTOS score) for the sample based on at least the expression level of the one target or each of the plurality of targets, and the determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate):a. is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate); andii. on a scale from -1 to 1, the risk score being equal to or greater than, - 0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.67 on a scale from -1 to 1; orb. is expected to respond no better or worse to treatment comprising dose- escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower third of the risk scores observed in a population of patients with prostate cancer who have notundergone radical prostatectomy (or any other surgery to remove the cancerous prostate); andii. on a scale from -1 to 1, the risk score being equal to or less than, -0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.68 on a scale from -1 to 1.

[0033] Hi gher vs. Lower: In some embodiments, the method comprises calculating a risk score (e.g., a PORTOS score) for the sample based on at least the expression level of the one target or each of the plurality of targets, and the determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate):a. is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate); and ii. on a scale from -1 to 1, the risk score being equal to or greater than, 0, -0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, -0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, -0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.42 on a scale from -1 to 1; or b. is expected to respond no better or worse to treatment comprising dose- escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower third of the risk scores observed in a population of patients with prostate cancer who have notundergone radical prostatectomy (or any other surgery to remove the cancerous prostate); andii. on a scale from -1 to 1, the risk score being equal to or less than, -0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.68 on a scale from -1 to 1.

[0034] In some embodiments, including but not limited to the three embodiments above, the population of patients with prostate cancer who have not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) comprises at least 10, 25, 50, 75 or 100 patients, or a range defined by any two of the preceding values, for example, 10-100, 75-100, 10-50, or 50-100. In some embodiments, the population of patients does not have metastatic prostate cancer, and / or the population of patients has intermediate-risk prostate cancer. In some embodiments, including but not limited to the three embodiments above, the method of further comprises: a. selecting for administration and / or administering, or having administered, to the subject expected to respond better to treatment comprising dose-escalated radiation therapy a treatment comprising dose-escalated radiation therapy; or b. selecting for administration and / or administering, or having administered, to the subject expected to respond no better or worse to treatment comprising dose-escalated radiation therapy a treatment comprising non-dose-escalated radiation therapy. In some embodiments, the dose-escalated radiation therapy is at least 78 Gy, optionally at least 79 Gy or 80 Gy. In some embodiments, the non-dose-escalated radiation therapy is not more than 71 Gy, optionally not more than 70 Gy. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is selected from 3 -Dimensional Conformal Radiation Therapy (3D-CRT), Intensity -Modulated Radiation Therapy (IMRT), volumetric modulated arc therapy (VMAT), stereotactic body radiotherapy (SBRT or SABR), proton beam RT, and brachytherapy boost. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is 3D-CRT or IMRT. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is definitive radiation therapy. In some embodiments, the dose-escalated and / or non-dose- escalated radiation therapy is not salvage radiation therapy. In some embodiments, the subjecthas intermediate-risk prostate cancer. In some embodiments, the subject has a PSA score of < 20 ng / ml, a clinical stage of T1 or T2, and / or a Gleason grade group of 1, 2 or 3. In some embodiments, the subject satisfies the following: histologically confirmed adenocarcinoma of the prostate, clinical stage Tla-T2b (stage I or II), Gleason score 2-6 and prostate-specific antigen (PSA) 10 to less than 20 ng / mL or Gleason score 7 and PSA less than 15 ng / mL, no distant metastases, and no regional lymph node involvement. In some embodiments, the subject has intermediate risk prostate cancer as defined by the NCCN risk groups disclosed in Spratt, D. E. et al. Int J Radiat Oncol Biol Phys 117, 370-377 (2023). In some embodiments, the subject does not have NCCN unfavorable intermediate-risk prostate cancer. In some embodiments, the subject has one or more intermediate risk factors: clinical stage T2b- T2c, Grade Group 2 or 3 (Gleason 7), PSA 10-20. In some embodiments, the subject has low-risk prostate cancer. In some embodiments, the subject has clinical stage cTl-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL. In some embodiments, the subject has high / very high-risk prostate cancer. In some embodiments, the subject has at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL. In some embodiments, the subject has metastatic prostate cancer. In some embodiments, the subject does not have low-risk prostate cancer. In some embodiments, the subject does not have clinical stage cTl-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL. In some embodiments, the subject does not have high / very high-risk prostate cancer. In some embodiments, the subject does not have at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL. In some embodiments, the subject does not have metastatic prostate cancer. In some embodiments, the risk level is determined according to NCCN risk groups disclosed in Spratt et al. (2023).

[0035] In some embodiments utilizing the Higher / Middle / Lower groups described above and elsewhere herein, the method comprises calculating a risk score (e.g., a PORTOS score) for the sample based on at least the expression level of the one target or each of the plurality of targets, but does not require (but optionally can include) determining the expected response to dose-escalated radiation therapy. Embodiments include the following non-limiting embodiments.

[0036] Higher vs. Middle / Lower: In some embodiments, the method comprises calculating a risk score (e.g., a PORTOS score) for the sample based on at least the expression level of the one target or each of the plurality of targets, and the method comprises:a. selecting for administration and / or administering, or having administered, to the subject a treatment comprising dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate); andii. on a scale from -1 to 1, the risk score being equal to or greater than, 0, -0.05, - 0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, -0.36, -0.37, - 0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, -0.47, -0.48, -0.49, - 0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.42 on a scale from -1 to 1; orb. selecting for administration and / or administering, or having administered, to the subject a treatment comprising non-dose-escalated radiation therapy based on either or both ofi. the risk score being within the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate); and ii. on a scale from -1 to 1, the risk score being equal to or less than 0, -0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, -0.36, -0.37, -0.38, - 0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, -0.47, -0.48, -0.49, -0.50, - 0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.41 on a scale from -1 to 1.

[0037] Higher / Middle vs. Lower: In some embodiments, the method comprises calculating a risk score (e g., a PORTOS score) for the sample based on at least the expression level of the one target or each of the plurality of targets, and the method comprises:a. selecting for administration and / or administering, or having administered, to the subject a treatment comprising dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate); andii. on a scale from -1 to 1, the risk score being equal to or greater than, -0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, - 0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.67 on a scale from -1 to 1; orb. selecting for administration and / or administering, or having administered, to the subject a treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate); andii. on a scale from -1 to 1, the risk score being equal to or less than, -0.53, -0.54, - 0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, - 0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.68 on a scale from -1 to 1.

[0038] Higher vs. Lower: In some embodiments, the method comprises calculating a risk score (e.g., a PORTOS score) for the sample based on at least the expression level of the one target or each of the plurality of targets, and the method comprises:a. selecting for administration and / or administering, or having administered, to the subject a treatment comprising dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate); andii. on a scale from -1 to 1, the risk score being equal to or greater than, 0, -0.05, - 0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, -0.36, -0.37, - 0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, -0.47, -0.48, -0.49, - 0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.42 on a scale from -1 to 1; orb. selecting for administration and / or administering, or having administered, to the subject a treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate); and u. on a scale from -1 to 1, the risk score being equal to or less than, -0.53, -0.54, - 0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.68 on a scale from -1 to 1.

[0039] In some embodiments, including but not limited to the embodiments above, the method comprises determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy:a. is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on at least the risk score being greater than 0 on a scale from -1 to 1; or b. is expected to respond no better or worse to treatment comprising dose- escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on at least the risk score being less than or equal to 0 on a scale from -1 to 1.

[0040] In some embodiments, including but not limited to the embodiments above, the method comprises:a. selecting for administration and / or administering, or having administered, to the subject a treatment comprising dose-escalated radiation therapy based on at least the risk score being greater than 0 on a scale from - 1 to 1; or b. selecting for administration and / or administering, or having administered, to the subject a treatment comprising non-dose-escalated radiation therapy based at least on the risk score being equal to or less than 0 on a scale from -1 to 1.

[0041] In some embodiments, including but not limited to the embodiments above, the population of patients with prostate cancer who have not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) comprises at least 10, 25, 50, 75 or 100 patients, or a range defined by any two of the preceding values, for example, 10-100, 75-100, 10-50, or 50-100, In some embodiments, the population of patients does not have metastatic prostate cancer, and / or the population of patients has intermediate- risk prostate cancer. In some embodiments, the dose-escalated radiation therapy is at least 78 Gy, optionally at least 79 Gy or 80 Gy. In some embodiments, the non-dose-escalated radiation therapy is not more than 71 Gy, optionally not more than 70 Gy. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is selected from 3-Dimensional Conformal Radiation Therapy (3D-CRT), Intensity -Modulated Radiation Therapy (IMRT), volumetric modulated arc therapy (VMAT), stereotactic body radiotherapy (SBRT or SABR), proton beam RT, and brachytherapy boost. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is 3D-CRT or IMRT. In some embodiments, the dose-escalated and / or non- dose-escalated radiation therapy is definitive radiation therapy. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is not salvage radiation therapy. In some embodiments, the subject has intermediate-risk prostate cancer. In some embodiments, the subject has a PSA score of < 20 ng / ml, a clinical stage of T1 or T2, and / or a Gleason grade group of 1, 2 or 3. In some embodiments, the subject satisfies the following: histologically confirmed adenocarcinoma of the prostate, clinical stage Tla-T2b (stage I or II), Gleason score 2-6 and prostate-specific antigen (PSA) 10 to less than 20 ng / mL or Gleason score 7 and PSA less than 15 ng / mL, no distant metastases, and no regional lymph node involvement. In some embodiments, the subject has intermediate risk prostate cancer as defined by the NCCN risk groups disclosed in Spratt, D. E. et al. Int J Radiat Oncol Biol Phys 117, 370-377 (2023). In some embodiments, the subject does not have NCCN unfavorable intermediate-risk prostatecancer. In some embodiments, the subject has one or more intermediate risk factors: clinical stage T2b- T2c, Grade Group 2 or 3 (Gleason 7), PSA 10-20. In some embodiments, the subject has low-risk prostate cancer. In some embodiments, the subject has clinical stage cTl-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL. In some embodiments, the subject has high / very high-risk prostate cancer. In some embodiments, the subject has at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL. In some embodiments, the subject has metastatic prostate cancer. In some embodiments, the subject does not have low-risk prostate cancer. In some embodiments, the subject does not have clinical stage cTl-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL. In some embodiments, the subject does not have high / very high-risk prostate cancer. In some embodiments, the subject does not have at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL, In some embodiments, the subject does not have metastatic prostate cancer. In some embodiments, the risk level is determined according to NCCN risk groups disclosed in Spratt et al. (2023).

[0042] In some embodiments, including but not l imited to the embodiments above that utilize the Higher / Middle / Lower groups, the response to treatment comprising dose- escalated radiation therapy comprises risk of one or more of: biochemical failure (BF), distant metastasis (DM) and receipt of salvage therapy. In some embodiments, the risk is at 3, 5, 10 or 15 years or a range defined by any two of the preceding values, post-treatment with the radiation therapy. In some embodiments, the risk is at I, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 years, or a range defined by any two of the preceding values, post- treatment with the radiation therapy. In some embodiments, the expected better response to treatment comprising dose-escalated radiation therapy is one or more of: a reduction in risk of BF of, or of at least 5%, 10%, 20%, or 30% or a range defined by any two of the preceding values, as compared to treatment comprising non-dose-escalated radiation therapy; a reduction in risk of DM of, or of at least 5%, 10%, 20%, or 30% or a range defined by any two of the preceding values, as compared to treatment comprising non-dose-escalated radiation therapy; and a reduction in risk of receipt of salvage therapy of or of at least 5%, 10%, 20%, or 30% or a range defined by any two of the preceding values, as compared to treatment comprising non-dose-escalated radiation therapy. In some embodiments, the risk is at 3, 5, 10 or 15 years or a range defined by any two of the preceding values, post-treatment with the radiation therapy. In some embodiments, the risk is at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 years, or a rangedefined by any two of the preceding values, post-treatment with the radiation therapy. In some embodiments, the expected no better or worse response to treatment comprising dose-escalated radiation therapy is one or more of: reduction in risk of BF of, or of less than 5%, 0%, -5%, or -10% or a range defined by any two of the preceding values, as compared to treatment comprising non-dose-escalated radiation therapy; reduction in risk of DM of, or of less than 5%, 0%, -5%, or -10% or a range defined by any two of the preceding values, as compared to treatment comprising non-dose-escalated radiation therapy; and reduction in risk of receipt of salvage therapy of, or of less than 5%, 0%, -5%, or -10% or a range defined by any two of the preceding values, as compared to treatment comprising non-dose-escalated radiation therapy. In some embodiments, the risk is at 3, 5, 10 or 15 years or a range defined by any two of the preceding values, post-treatment with the radiation therapy. In some embodiments, the risk is at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 years, or a range defined by any two of the preceding values, post-treatment with the radiation therapy. In some embodiments, the BF is Pheonix BF.

[0043] In some embodiments, the determining or having determined the expected response to treatment comprising dose-escalated radiation therapy is in comparison to a treatment comprising non-dose-escalated radiation therapy.

[0044] In some embodiments, the determining or having determined the expected response of the subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) to treatment comprising dose-escalated radiation therapy is not based on prognostic clinical variables. In some embodiments, the determining or having determined the expected response of the subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) to treatment comprising dose-escalated radiation therapy is further based on one or more prognostic clinical variables. In some embodiments, the prognostic clinical variables are tumor stage, nodal stage, metastatic tumor volume categorization, a Gleason score and a serum PSA level of the subject. In some embodiments, the prognostic clinical variables are tumor stage, nodal stage, metastatic tumor volume categorization, and a Gleason score. In some embodiments, the prognostic clinical variable is a serum PSA level of the subject.

[0045] In some embodiments, the obtaining or having obtained the expression level of the one target or the plurality of targets comprises utilizing a microarray to assess on theplurality of targets, performing qPCR on the plurality of targets, and / or sequencing the plurality of targets. In some embodiments, the sample is from a resected specimen. In some embodiments, the sample is from a transurethral resection of prostate (TURP) specimen. In some embodiments, the sample is from a biopsy of the prostate cancer. In some embodiments, the biopsy is a needle biopsy. In some embodiments, the sample is from a primary prostate tumor.

[0046] In some embodiments, the population of patients with prostate cancer who have not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) comprises, or comprises at least 10, 25, 50, 75 or 100 patients, or a range defined by any two of the preceding values, for example, 10-100, 75-100, 10-50, or 50-100. In some embodiments, the population of patients with prostate cancer who have not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) comprises at least 10, 25, 50, 75 or 100 patients.

[0047] In some embodiments, the dose-escalated radiation therapy is at least 78 Gy, optionally at least 79 Gy or 80 Gy. In some embodiments, the non-dose-escalated radiation therapy is not more than 71 Gy, optionally not more than 70 Gy. In some embodiments, he dose-escalated and / or non-dose-escalated radiation therapy is selected from 3-Dimensional Conformal Radiation Therapy (3D-CRT), Intensity-Modulated Radiation Therapy (IMRT), volumetric modulated arc therapy (VMAT), stereotactic body radiotherapy (SBRT or SABR), proton beam RT, and brachytherapy boost. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is 3D-CRT or IMRT. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is definitive radiation therapy. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is not salvage radiation therapy. In some embodiments, the subject has intermediate-risk prostate cancer. In some embodiments, the subject has a PSA score of < 20 ng / ml, a clinical stage of T1 or T2, and / or a Gleason grade group of 1, 2 or 3. In some embodiments, the subject satisfies the following: histologically confirmed adenocarcinoma of the prostate, clinical stage Tla-T2b (stage I or II), Gleason score 2-6 and prostate-specific antigen (PSA) 10 to less than 20 ng / mL or Gleason score 7 and PSA less than 15 ng / mL, no distant metastases, and no regional lymph node involvement. In some embodiments, the subject has intermediate risk prostate cancer as defined by the NCCN risk groups disclosed in Spratt, D. E. et al. Int J Radiat Oncol Biol Phys117, 370-377 (2023). In some embodiments, the subject does not have NCCN unfavorable intermediate-risk prostate cancer. In some embodiments, the subject has one or more intermediate risk factors: clinical stage T2b- T2c, Grade Group 2 or 3 (Gleason 7), PSA 10-20. In some embodiments, the subject has low-risk prostate cancer. In some embodiments, the subject has clinical stage cTl-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL. In some embodiments, the subject has high / very high-risk prostate cancer. In some embodiments, the subject has at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL. In some embodiments, the subject has metastatic prostate cancer. In some embodiments, the subject does not have low-risk prostate cancer. In some embodiments, the subject does not have clinical stage cTl-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL. In some embodiments, the subject does not have high / very high-risk prostate cancer. In some embodiments, the subject does not have at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL. In some embodiments, the subject does not have metastatic prostate cancer. In some embodiments, the risk level is determined according to NCCN risk groups disclosed in Spratt et al. (2023).Kits

[0048] In some embodiments, a kit for use in determining a response to treatment according to any one of the methods herein is provided. In some embodiments, the kit comprises reagents for obtaining the expression level of biomarkers comprising or consisting of the one or each of the plurality of targets. In some embodiments, the reagents comprise primer(s) and / or probe(s) for obtaining the expression level of the one or plurality of targets. In some embodiments, the kit comprises a container or housing for holding the components of the kit. In some embodiments, the reagents include those useful for performing a method disclosed herein, for example, amplification reagents, primers, probes, etc. In some embodiments, the reagents include one or more probes, primers or primer pairs, enzymes (including polymerases and ligases), intercalating dyes, labeled probes, and labels that can be incorporated into amplification products.

[0049] In some embodiments, the kit comprises primers (or primer pairs) specific for those subsets and combinations of target sequences described herein. In some embodiments, at least two, three, four or five primers (or pairs) of primers suitable for selectively amplifying the same number of target sequence-specific polynucleotides can beprovided in kit form. In some embodiments, the kit comprises from about five to one hundred primers or pairs of primers suitable for amplifying the same number of target sequence¬ representative polynucleotides of interest.

[0050] In some embodiments, the primers (or primer pairs) of the kit, when used in an amplification reaction, specifically amplify a non-coding target, coding target, or non-exonic target described herein, for example a nucleic acid sequence corresponding to a target selected from the biomarkers from Table 1 or from a subset of Table 2, an RNA form thereof, or a complement to either thereof. In some embodiments, the kit may include a plurality of such primers or primer pairs which can specifically amplify a corresponding plurality of different amplify a non-codmg target, coding target, or non-exonic transcript described herein, a nucleic acid sequence corresponding to a target selected from the biomarkers from Table 1 or from a subset of Table 2, RNA forms thereof, or complements thereto. In some embodiments, at least two, three, four or five primers or pairs of primers suitable for selectively amplifying the same number of target sequence-specific polynucleotides can be provided in kit form. In some embodiments, the kit comprises from five to one hundred primers (or pairs of primers) suitable for amplifying the same number of target sequence-representative polynucleotides of interest.

[0051] In some embodiments, the reagents may independently be provided in liquid or solid form. In some embodiments, the reagents are provided in mixtures. In some embodiments, control samples and / or nucleic acids are provided in the kit. In some embodiments, control samples may include tissue and / or nucleic acids obtained from or representative of tumor samples from patients showing no evidence of disease. In some embodiments, control samples may include tissue and / or nucleic acids obtained from or representative of tumor samples from patients that have and / or develop cancer.

[0052] In some embodiments, instructions for using the kit to perform a method disclosed herein is provided with the container, and can be provided in any suitable fixed medium. The instructions may be located inside or outside the container or housing, and / or may be printed on the interior or exterior of any suitable surface thereof. A kit may be in multiplex form for concurrently detecting and / or quantitating one or more different target polynucleotides representing the expressed target sequences.

[0053] Disclosed herein are kits for use in detecting a response to treatment according to any one of the preceding methods, or methods disclosed elsewhere herein. In some embodiments, the kit comprises reagents for obtaining the expression level of biomarkers comprising or consisting of the one or each of the plurality of targets. In some embodiments, the reagents comprise primer(s) and / or probe(s) for obtaining the expression level of each of the plurality of targets.Coding and Non-coding Targets

[0054] In some embodiments, the methods disclosed herein comprise assaying the expression level of one target or a plurality of targets (e.g., target nucleic acid sequences). In some embodiments, the targets are referred to as biomarkers. In some embodiments, the targets may comprise coding targets and / or non-coding targets. In some embodiments, the target or plurality of targets may comprise coding targets and / or non-coding targets of a protein-coding gene or a non protein-coding gene. A protein-coding gene structure may comprise an exon and an intron. The exon may further comprise a coding sequence (CDS) and an untranslated region (UTR). The protein-coding gene may be transcribed to produce a pre-mRNA and the pre-mRNA may be processed to produce a mature mRNA. The mature mRNA may be translated to produce a protein.

[0055] In some embodiments, a non protein-coding gene structure may comprise an exon and intron. In some embodiments, the exon region of a non protein-coding gene primarily contains a UTR. The non protein-coding gene may be transcribed to produce a pre-mRNA and the pre-mRNA may be processed to produce a non-coding RNA (ncRNA).

[0056] In some embodiments, a coding target may comprise a coding sequence of an exon. In some embodiments, a non-coding target may comprise a UTR sequence of an exon, intron sequence, intergenic sequence, promoter sequence, non-coding transcript, CDS antisense, intronic antisense, UTR antisense, or non-coding transcript antisense. In some embodiments, a non-coding transcript may comprise a non-coding RNA (ncRNA).

[0057] In some embodiments, the plurality of targets are differentially expressed. In some embodiments, a plurality of probe selection regions (PSRs) is differentially expressed.

[0058] In some embodiments, the plurality of targets comprises target nucleic acid sequences from biomarkers selected from Table 1 or from a subset of biomarkers of Table 2. In some embodiments, the plurality of targets comprises target nucleic acid sequences from atleast 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, or at least 10 biomarkers selected from Table 1 or from a subset of Table 2. In some embodiments, the plurality of targets comprises target nucleic acid sequences from at least 12, at least 15, at least 17, at least 20, at least 22, or at least 24 biomarkers selected from Table 1 or from a subset of Table 2. In some embodiments, the targets are target nucleic acid sequences from biomarkers selected from Table 1. In some embodiments, the targets are target nucleic acid sequences from biomarkers from a subset of biomarkers of Table 2. In some embodiments, the plurality of targets comprises one or more targets selected from Table 1. In some embodiments, the plurality of targets comprises at least about 2, at least about 3, at least about 4, at least about 5, at least about 6, at least about 7, at least about 8, at least about 9, or at least about 10 targets selected from Table 1, In some embodiments, the plurality of targets comprises at least about 12, at least about 15, at least about 17, at least about 20, at least about 22, or at least about 24 targets selected from Table 1. In some embodiments, the targets are selected from Table 1. In some embodiments, the plurality of targets comprises a coding target, non-coding target, or any combination thereof. In some embodiments, the coding target comprises an exonic sequence. In some embodiments, the non-coding target comprises a non-exonic sequence. In some embodiments, a non-coding target comprises a UTR sequence, an intronic sequence, or a non-coding RNA transcript. In some embodiments, a non-coding target comprises sequences which partially overlap with a UTR sequence or an intronic sequence. A non-coding target also includes non-exonic transcripts. Exonic sequences may comprise regions on a protein-coding gene, such as an exon, U TR, or a portion thereof. Non-exonic sequences may comprise regions on a protein-coding, non protein-coding gene, or a portion thereof. For example, non-exonic sequences may comprise intronic regions, promoter regions, intergenic regions, a non-coding transcript, an exon anti-sense region, an intronic anti-sense region, UTR anti-sense region, non¬ coding transcript anti-sense region, or a portion thereof. In some embodiments, the plurality of targets comprises a non-coding RNA transcript.Probes / Primers

[0059] The present disclosure provides for a probe set (e g., in a kit) for predicting a response to treatment in a subject with prostate cancer who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) comprising a plurality of probes. In some embodiments, the kit comprises reagents for obtaining the expression levelof biomarkers (e.g., those comprising or consisting of each of the plurality of targets). In some embodiments, the reagents comprise primer(s) and / or probe(s) for obtaining the expression level of each of the plurality of targets. In some embodiments, the probe set may comprise one or more polynucleotide probes. In some embodiments, individual polynucleotide probes comprise a nucleotide sequence derived from the nucleotide sequence of the target sequences or complementary sequences thereof. In some embodiments, the nucleotide sequence of the polynucleotide probe is designed such that it corresponds to, or is complementary to the target sequences (sequences of the plurality of targets). In some embodiments, the polynucleotide probe can hybridize under either stringent or lowered stringency hybridization conditions to a region of the target sequences (sequences of the plurality of targets), to the complement thereof, or to a nucleic acid sequence (such as a cDNA) derived therefrom. In some embodiments, the selection of the polynucleotide probe sequences and determination of their uniqueness is carried out in silico using suitable techniques known in the art, for example, based on a BLASTN search of the polynucleotide sequence in question against gene sequence databases, such as the Human Genome Sequence, UniGene, dbEST or the non-redundant database at NCBI. In some embodiments, the polynucleotide probe is complementary to a region of a target mRNA derived from a target sequence (sequence of the plurality of targets) in the probe set. In some embodiments, computer programs can also be employed to select probe sequences that may not cross hybridize or may not hybridize non-specifically.

[0060] One skilled in the art understands that the nucleotide sequence of the polynucleotide probe need not be fully complementary to its target sequence in order to hybridize thereto, and can determine the required level complementarity needed for an expression assay. In some embodiments, the polynucleotide probes comprise a nucleotide sequence that is at least about 65% complementary to a region of the coding target or non¬ coding target of a biomarker selected from Table 1. In some embodiments, the nucleotide sequence of the polynucleotide probe is at least about 70% complementary to a region of the coding target or non-coding target of a biomarker from Table 1. In some embodiments, the nucleotide sequence of the polynucleotide probe is at least about 75% complementary to a region of the coding target or non-coding target of a biomarker from Table 1. In some embodiments, the nucleotide sequence of the polynucleotide probe is at least about 80% complementary to a region of the coding target or non-coding target of a biomarker from Table1. In some embodiments, the nucleotide sequence of the polynucleotide probe is at least about 85% complementary to a region of the coding target or non-coding target of a biomarker from Table 1. In some embodiments, the nucleotide sequence of the polynucleotide probe is at least about 90% complementary to a region of the coding target or non-coding target of a biomarker from Table 1. In some embodiments, the nucleotide sequence of the polynucleotide probe is at least about 95% complementary to a region of the coding target or non-coding target of a biomarker from Table 1. In some embodiments, the nucleotide sequence of the polynucleotide probe is 100% complementary to a region of the coding target or non-coding target of a biomarker from Table 1,

[0061] In some embodiments, the probe set comprises a plurality of probe sequences that hybridize to at least about 5, at least about 10, at least about 15, or at least about 20 coding targets and / or non-coding targets selected from biomarkers from Table 1 or a subset of Table 2.

[0062] In some embodiments, the system of the present disclosure further provides for primers and primer pairs capable of amplifying target sequences defined by the probe set, or fragments or subsequences or complements thereof. The nucleotide sequences of the probe set may be provided in computer-readable media for in silico applications and as a basis for the design of appropriate primers for amplification of one or more target sequences of the probe set. In some embodiments, the primer(s) are capable of amplifying (e.g., assisting in amplification) the plurality of target nucleic acid sequences, optionally as defined by the probe(s), or fragments or subsequences or complements of the plurality of target nucleic acid sequences. The nucleotide sequences of the plurality of target nucleic acid sequences, or of the probe(s) for detecting the target nucleic acid sequences, may be provided in computer-readable media for in silico applications and as a basis for the design of appropriate primers for amplification of one or more target sequences of the plurality of target nucleic acids or of the corresponding probe(s).

[0063] In some embodiments, the method or kits comprise primers and primer pairs capable of amplifying the plurality of target nucleic acid sequences, optionally as defined by the probe set, or fragments or subsequences or complements of the plurality of target nucleic acid sequences. The nucleotide sequences of the plurality of target nucleic acid sequences, or of the probe set for detecting the target nucleic acid sequences, may be provided in computer-readable media for in silico applications and as a basis for the design of appropriate primers for amplification of one or more target sequences of the plurality of target nucleic acids or of the corresponding probe set.

[0064] In some embodiments, primers based on the nucleotide sequences of target sequences (plurality of target sequences) are designed for use in amplification of the target sequences (plurality of target sequences). In some embodiments, for use in amplification reactions such as PCR, a pair of primers are used. In some embodiments, the primers may hybridize to specific sequences of the probe set under stringent conditions. In some embodiments, the primers may hybridize to specific sequences of the plurality of target nucleic acid sequences under stringent conditions. In some embodiments, these primers are used in combination with probes of the probe set, such as molecular beacons in amplifications using real-time PCR,

[0065] In some embodiments, the primers or primer pairs, when used in an amplification reaction. In some embodiments, the primers or primer pairs amplify at least a portion of a nucleic acid sequence of a target selected from any of Table 1 (or subgroups thereof as set forth herein), an RNA form thereof, or a complement to either thereof. In some embodiments, the targets are selected from Table 1. In some embodiments, the primers or primer pairs, when used in an amplification reaction amplify at least a portion of a target nucleic acid sequence in the plurality of targets selected from the biomarkers of Tables 1 or a subset of Table 2, an RNA form thereof, or a complement to either thereof.

[0066] In some embodiments, the nucleotide sequence of the entire length of the polynucleotide probe or primer does not need to be derived from the target sequence. In some embodiments, for example, the polynucleotide probe may comprise nucleotide sequences at the 5' and / or 3' termini that are not derived from the target sequences. In some embodiments, nucleotide sequences which are not derived from the nucleotide sequence of the target sequence may provide additional functionality to the polynucleotide probe. In some embodiments, for example, they may provide a restriction enzyme recognition sequence or a "tag" that facilitates detection, isolation, purification or immobilization onto a solid support. In some embodiments, the additional nucleotides may provide a self-complementary sequence that allows the primer / probe to adopt a hairpin configuration. Such configurations arenecessary for certain probes, for example, molecular beacon and Scorpion probes, which can be used in solution hybridization techniques.

[0067] In some embodiments, the polynucleotide probes or primers incorporate moieties useful in detection, isolation, purification, or immobilization, if desired. Such moieties are well-known in the art (see, for example, Ausubel et al., (1997 & updates) Current Protocols in Molecular Biology, Wiley & Sons, New York) and are chosen such that the ability of the probe to hybridize with its target sequence is not affected. In some embodiments, examples of suitable moieties are detectable labels, such as radioisotopes, fluorophores, chemiluminophores, enzymes, colloidal particles, and fluorescent microparticles, as well as antigens, antibodies, haptens, avidin / streptavidin, biotin, haptens, enzyme cofactors / substrates, enzymes, and the like. In some embodiments, a label is attached to or incorporated into a probe or primer polynucleotide to allow detection and / or quantitation of a target polynucleotide representing the target sequence of interest. In some embodiments, the target polynucleotide is the expressed target sequence RNA itself, a cDNA copy thereof, or an amplification product derived therefrom, and may be the positive or negative strand, so long as it can be specifically detected in the assay being used. In some embodiments, an antibody is labeled.

[0068] In some embodiments, polynucleotides of the disclosure comprise at least 20 consecutive bases of the nucleic acid sequence of a target selected from any of Table 1 or a complement thereto. The polynucleotides may comprise at least 21, 22, 23, 24, 25, 27, 30, 32, 35 or more consecutive bases of the nucleic acids sequence of a target selected from any of Table 1, as applicable. In some embodiments, the targets are selected from Table 1. In some embodiments, the polynucleotides comprise at least 20 consecutive bases of the nucleic acid sequence of a target selected from biomarkers from Table 1 or a subset from Table 2, or a complement thereto. The polynucleotides may comprise at least 21, 22, 23, 24, 25, 27, 30, 32, 35 or more consecutive bases of the nucleic acids sequence of a target selected from biomarkers Table 1 or a subset of Table 2, as applicable

[0069] In some embodiments, the polynucleotides are provided in a variety of formats, including as solids, in solution, or in an array. In some embodiments, the polynucleotides comprise one or more labels, which may be chemically and / or enzymatically incorporated into the polynucleotide.

[0070] In some embodiments, one or more polynucleotides provided herein are provided on a substrate. In some embodiments, the substrate can take the form of an array, a photodiode, an optoelectronic sensor such as an optoelectronic semiconductor chip or optoelectronic thin-film semiconductor, or a biochip. The location(s) of probe(s) on the substrate can be addressable; this can be done in highly dense formats, and the location(s) can be microaddressable or nanoaddressable.Samples

[0071] In some embodiments, samples for use in the present disclosure comprise nucleic acids suitable for providing RNA expression information. In some embodiments, the biological sample from which the expressed RNA is obtained and analyzed for target sequence expression can be any material suspected of comprising cancer tissue or cells. In some embodiments, the sample is a biological sample used directly in a method of the disclosure. In some embodiments, the sample is a sample prepared from a biological sample. In some embodiments, samples comprise nucleic acids suitable for providing information on the expression levels of targets (e.g., nucleic acid expression). In some embodiments, samples comprise nucleic acids suitable for providing RNA expression information. In some embodiments, the sample can comprise any suitable material suspected of comprising cancer tissue and / or cells. In some embodiments, the sample is used directly (e.g., without further preparation, processing, purification, etc.) once derived from the subject in an embodiment of a method disclosed herein. In some embodiments, the sample (once derived from a subject) is further prepared, processed, purified, and / or otherwise altered prior to use in an embodiment of a method disclosed herein.

[0072] In some embodiments, the sample or portion of the sample comprising or suspected of comprising cancer tissue or cells is any source of biological material, including cells, tissue or fluid, including bodily fluids. Non-limiting examples of the source of the sample include an aspirate, a needle biopsy, a cytology pellet, a bulk tissue preparation or a section thereof obtained for example by surgery or autopsy, lymph fluid, blood, plasma, serum, tumors, and organs. In some embodiments, the sample is from urine. In some embodiments, the sample is from blood, plasma or serum. In some embodiments, the sample is from saliva. In some embodiments, the sample or portion of the sample comprises or is suspected to comprise cancer tissue or cells. In some embodiments, the sample comprises any suitable source of biologicalmaterial. In some embodiments, the sample comprises cells, tissue, and / or fluid (e.g., including bodily fluids). Non-limiting examples of the source of the sample inclu de an aspirate, a needle biopsy, a cytology pellet, a bulk tissue preparation or a section thereof obtained for example by surgery or biopsy, lymph fluid, blood, plasma, serum, tumors, and organs. In some embodiments, the sample is from urine. In some embodiments, the sample is from blood, plasma or serum. In some embodiments, the sample is from saliva. In some embodiments, the sample is a fine needle aspiration.

[0073] In some embodiments, the samples are archival samples, having a known and documented medical outcome, or are samples from current patients whose ultimate medical outcome is not yet known.

[0074] In some embodiments, the sample are dissected prior to molecular analysis. The sample may be prepared via macrodissection of a bulk tumor specimen or portion thereof, or may be treated via microdissection, for example via Laser Capture Microdissection (LCM),

[0075] In some embodiments, the sample may initially be provided in a variety of states, as fresh tissue, fresh frozen tissue, fine needle aspirates, and may be fixed or unfixed. In some embodiments, medical laboratories routinely prepare medical samples in a fixed state, which facilitates tissue storage. A variety of fixatives can be used to fix tissue to stabilize the morphology of cells, and may be used alone or in combination with other agents. In some embodiments, fixatives include crosslinking agents, alcohols, acetone, Bouin's solution, Zenker solution, Hely solution, osmic acid solution and Carnoy solution.

[0076] In some embodiments, the biological sample, which may be fixed or unfixed, is embedded in an embedding medium. In some embodiments, the embedding media used in histology is paraffin, Tissue- Tek® V. I. P. TM, Paramat, Paramat Extra, Paraplast, Paraplast X-tra, Paraplast Plus, Peel Away Paraffin Embedding Wax, Polyester Wax, Carbowax Polyethylene Glycol, PolyfinTM, Tissue Freezing Medium TFM™, Cryo-Gel™, and OCT Compound (Electron Microscopy Sciences, Hatfield, PA). Prior to molecular analysis, the embedding material may be removed via any suitable techniques, as known in the art. For example, where the sample is embedded in wax, the embedding material may be removed by extraction with organic solvent(s), for example xylenes. Kits are commercially available for removing embedding media from tissues. Samples or sections thereof may besubjected to further processing steps as needed, for example serial hydration or dehydration steps.

[0077] In some embodiments, the sample is a fixed, wax-embedded biological sample. Frequently, samples from medical laboratories are provided as fixed, wax-embedded samples, most commonly as formalin-fixed, paraffin embedded (FFPE) tissues.

[0078] In some embodiments, the target polynucleotide that is assayed is prepared synthetically (e.g. in the case of control sequences). In some embodiments, the target polynucleotide that is assayed is purified from the biological source and subjected to one or more preparative steps. The RNA may be purified to remove or diminish one or more undesired components from the biological sample or to concentrate it. In some embodiments, where the RNA is too concentrated for the particular assay, it may be diluted.Nucleic Acid Extraction

[0079] In some embodiments, a nucleic acid, for example RNA, is extracted and purified from samples using any suitable technique. In some embodiments, RNA is extracted and purified from samples (e.g., biological samples) using any suitable technique. A number of suitable techniques are known in the art, and several are commercially available (e.g., FormaPure nucleic acid extraction kit, Agencourt Biosciences, Beverly MA, High Pure FFPE RNA Micro Kit, Roche Applied Science, Indianapolis, IN). In some embodiments, RNA can be extracted from frozen tissue sections using TRIzol (Invitrogen, Carlsbad, CA) and purified using RNeasy Protect kit (Qiagen, Valencia, CA). In some embodiments, RNA can be further purified using DNAse I treatment (Ambion, Austin, TX) to eliminate any contaminating DNA. RNA concentrations can be made using a Nanodrop ND- 1000 spectrophotometer (Nanodrop Technologies, Rockland, DE). RNA can be further purified to eliminate contaminants that interfere with cDNA synthesis by cold sodium acetate precipitation. In some embodiments, RNA integrity can be evaluated by running electropherograms, and RNA integrity number (RIN, a correlative measure that indicates intactness of mRNA) can be determined using the RNA 6000 PicoAssay for the Bioanalyzer 2100 (Agilent Technologies, Santa Clara, CA). Amplification and Hybridization

[0080] In some embodiments, following sample collection and nucleic acid extraction, the nucleic acid portion of the sample comprising RNA that is or can be used to prepare the target polynucleotide(s) of interest can be subjected to one or more preparativereactions. These preparative reactions can include in vitro transcription (IVT), labeling, fragmentation, amplification and other reactions. mRNA can first be treated with reverse transcriptase and a primer to create cDNA prior to detection, quantitation and / or amplification; this can be done in vitro with purified mRNA or in situ, e.g., in cells or tissues affixed to a slide.Detection and / or Quantification of Target Sequences

[0081] In some embodiments, any suitable method of detecting and / or quantitating the expression of the encoded target sequences is used. In some embodiments, the expressed target sequences is directly detected and / or quantitated, or may be copied and / or amplified to allow detection of amplified copies of the expressed target sequences or its complement. In some embodiments, a suitable method of obtaining the expression of the target (e.g., target sequences) is used. In some embodiments, the expressed targets (e.g., target sequences) are directly detected and / or quantitated, or may be copied and / or amplified to allow detection of amplified copies of the expressed target sequences or their complements,

[0082] In some embodiments, methods for detecting and / or quantifying a target can include Northern blotting, sequencing, array or microarray hybridization, by enzymatic cleavage of specific structures (e.g., an Invader® assay, Third Wave Technologies, e.g. as described in U. S. Pat. Nos. 5,846,717, 6,090,543; 6,001,567; 5,985,557; and 5,994,069) and amplification methods, e.g. RT-PCR, including in a TaqMan® assay (PE Biosystems, Foster City, Calif., e.g. as described in U. S. Pat. Nos. 5,962,233 and 5,538,848), and may be quantitative or semi- quantitative, and may vary depending on the origin, amount and condition of the available biological sample. In some embodiments, combinations of these methods may also be used. In some embodiments, for example, nucleic acids are amplified, labeled and subjected to microarray analysis.

[0083] In some embodiments, target sequences are detected by sequencing. In some embodiments, sequencing methods comprise whole genome sequencing or exome sequencing. In some embodiments, sequencing methods such as Maxim-Gilbert, chain¬ termination, or high-throughput systems are used. In some embodiments, suitable sequencing techniques include classic dideoxy sequencing reactions (e.g., Sanger method) using labeled terminators or primers and gel separation in slab or capillary, sequencing by synthesis using reversibly terminated labeled nucleotides, pyrosequencing, 454 sequencing, allele specifichybridization to a library of labeled oligonucleotide probes, sequencing by synthesis using allele specific hybridization to a library of labeled clones that is followed by ligation, real time monitoring of the incorporation of labeled nucleotides during a polymerization step, and SOLiD sequencing.

[0084] In some embodiments, methods for detecting and / or quantifying a target include single-molecule sequencing (e.g., Helicos, PacBio), sequencing by synthesis (e.g., Illumina, Ion Torrent), sequencing by ligation (e.g., ABI SOLID), sequencing by hybridization (e.g., Complete Genomics), in situ hybridization, bead-array technologies (e.g., Luminex xMAP, Illumina BeadChips), branched DNA technology (e.g., Panomics, Genisphere). In some embodiments, sequencing methods use fluorescent (e.g., Illumina) or electronic (e.g., Ion Torrent, Oxford Nanopore) methods of detecting nucleotides.Reverse Transcription for QRT-PCR Analysis

[0085] In some embodiments, reverse transcription is be performed by any suitable method known in the art. In some embodiments, for example, reverse transcription is performed using the Omniscript kit (Qiagen, Valencia, CA), Superscript III kit (Invitrogen, Carlsbad, CA), for RT-PCR. In some embodiments, target-specific priming is performed in order to increase the sensitivity of detection of target sequences and generate target-specific cDNA.TaqMan® Gene Expression Analysis

[0086] In some embodiments, TaqMan®RT-PCR is performed using Applied Biosystems Prism (ABI) 7900 HT instruments in a 5 µl volume with target sequence-specific cDNA equivalent to 1 ng total RNA.

[0087] In some embodiments, primer and probe concentrations for TaqMan analysis are added to amplify fluorescent amplicons using PCR cycling conditions such as 95°C for 10 minutes for one cycle, 95°C for 20 seconds, and 60°C for 45 seconds for 40 cycles. In some embodiments, a reference sample is assayed to ensure reagent and process stability. In some embodiments, negative controls (e.g., no template) are assayed to monitor any exogenous nucleic acid contamination.Classification Arrays

[0088] In some embodiments, a probe set or probes derived therefrom is provided in an array format. In the context of the present disclosure, an "array" has its plain and ordinarymeaning as understood in light of the specification, and refers to a spatially or logically organized collection of polynucleotide probes. In some embodiments, an array comprising probes specific for a coding target, non-coding target, or a combination thereof is used. In some embodiments, an array comprising probes specific for two or more of transcripts of a target selected from any of Table 1 or a product derived thereof is used. In some embodiments, an array is specific for 5, 10, 15, 20, 25, 30, 50, 75, 100, 150, 200 or more of transcripts of a target selected from any of Table 1. In some embodiments, the targets are selected from Table 1. In some embodiments, expression of these sequences is detected alone or in combination with other transcripts. In some embodiments, an array is used which comprises a wide range of sensor probes for prostate- specific expression products, along with appropriate control sequences. In some embodiments, the array may comprise the Human Exon 1.0 ST Array (HuEx 1.0 ST, Affymetrix, Inc., Santa Clara, CA.)

[0089] In some embodiments, the polynucleotide probes are attached to a solid substrate and are ordered so that the location (e.g., on the substrate) and the identity of each are known. In some embodiments, the polynucleotide probes are attached to one of a variety of solid substrates capable of withstanding the reagents and conditions necessary for use of the array. In some embodiments, examples include, but are not limited to, polymers, such as (poly)tetrafluoroethylene, (poly)vinylidenedifluoride, polystyrene, polycarbonate, polypropylene and polystyrene; ceramic; silicon; silicon dioxide; modified silicon; (fused) silica, quartz or glass; functionalized glass; paper, such as filter paper; diazotized cellulose; nitrocellulose filter; nylon membrane; and polyacrylamide gel pad. In some embodiments, substrates that are transparent to light are useful for arrays that are used in an assay that involves optical detection. In some embodiments, examples of array formats include membrane or filter arrays (for example, nitrocellulose, nylon arrays), plate arrays (for example, multi-well, such as a 24-, 96-, 256-, 384-, 864- or 1536-well, microtitre plate arrays), pin arrays, and bead arrays (for example, in a liquid "slurry"). In some embodiments, arrays on substrates such as glass or ceramic slides, often referred to as chip arrays or "chips," are well known in the art and are used in the assays disclosed herein.Data Analysis

[0090] In some embodiments, one or more pattern recognition methods is used in analyzing the expression level of target sequences. In some embodiments, the patternrecognition method comprises a linear combination of expression levels, or a nonlinear combination of expression levels. In some embodiments, expression measurements for RNA transcripts or combinations of RNA transcript levels are formulated into linear or non-linear models or algorithms (e.g., an 'expression signature') and converted into a likelihood score. In some embodiments, this likelihood score indicates the probability that a biological sample is from a patient who will respond to treatment with dose-escalated radiation therapy. In some embodiments, the likelihood score is used to distinguish these disease states. In some embodiments, the models and / or algorithms are provided in machine readable format, and may be used to correlate expression levels or an expression profile with a response to treatment with dose-escalated radiation therapy for a patient or class of patients. In some embodiments, the models and / or algorithms are provided in machine readable format, and may be used to correlate expression levels or an expression profile with a disease state, for example cancer risk, for a patient or class of patients,

[0091] In some embodiments, assaying the expression level for a plurality of targets comprises the use of an algorithm or classifier, or a signature. Array data can be managed, classified, and analyzed using suitable techniques known in the art. In some embodiments, assaying the expression level for a plurality of targets comprises probe set modeling and data pre-processing. In some embodiments, probe set modeling and data pre¬ processing is derived using the Robust Multi-Array (RMA) algorithm or variants GC-RMA, fRMA, Probe Logarithmic Intensity Error (PLIER) algorithm or variant iterPLIER. In some embodiments, variance or intensity filters are applied to pre-process data using the RMA algorithm, for example by removing target sequences with a standard deviation of < 10 or a mean intensity of < 100 intensity units of a normalized data range, respectively.

[0092] In some embodiments, assaying the expression level for a plurality of targets may comprise the use of a machine learning algorithm. In some embodiments, the machine learning algorithm comprises a supervised learning algorithm. In some embodiments, examples of supervised learning algorithms include Average One-Dependence Estimators (AODE), Artificial neural network (e.g., Backpropagation), Bayesian statistics (e.g., Naive Bayes classifier, Bayesian network, Bayesian knowledge base), Case-based reasoning, Decision trees, Inductive logic programming, Gaussian process regression, Group method of data handling (GMDH), Learning Automata, Learning Vector Quantization, Minimummessage length (e.g., decision trees, decision graphs, etc.), Lazy learning, Instance-based learning Nearest Neighbor Algorithm, Analogical modeling, Probably approximately correct learning (PAC) learning, Ripple down rules, a knowledge acquisition methodology, Symbolic machine learning algorithms, Subsymbolic machine learning algorithms, Support vector machines, Random Forests, Ensembles of classifiers, Bootstrap aggregating (bagging), and Boosting. Supervised learning may comprise ordinal classification such as regression analysis and Information fuzzy networks (IFN). In some embodiments, supervised learning methods comprise statistical classification, such as AODE, Linear classifiers (e.g., Fisher's linear discriminant. Logistic regression, Naive Bayes classifier, Perceptron, and Support vector machine), quadratic classifiers, k-nearest neighbor, Boosting, Decision trees (e.g., C4.5, Random forests), Bayesian networks, and Hidden Markov models. In some embodiments, supervised learning methods compri e prediction analysis of microarrays methods that uses shrunken centroids to classify samples, as described in Tibshirani et al., Proc Natl Acad Sci U S A. 2002 May 14;99(10):6567-72, which is incorporated herein by reference,

[0093] In some embodiments, the machine learning algorithms also comprise an unsupervised learning algorithm. In some embodiments, examples of unsupervised learning algorithms include artificial neural network, Data clustering, Expectation-maximization algorithm, Self-organizing map, Radial basis function network, Vector Quantization, Generative topographic map, Information bottleneck method, and IBSEAD. In some embodiments, unsupervised learning comprises association rule learning algorithms such as Apriori algorithm, Eclat algorithm and FP-growth algorithm. Hierarchical clustering, such as Single-linkage clustering and Conceptual clustering, and is also used. In some embodiments, unsupervised learning comprises partitional clustering such as K-means algorithm and Fuzzy clustering.

[0094] In some embodiments, the machine learning algorithms comprise a reinforcement learning algorithm. Examples of reinforcement learning algorithms include, but are not limited to, temporal difference learning, Q-learning and Learning Automata. In some embodiments, the machine learning algorithm comprises Data Pre-processing.

[0095] In some embodiments, the machine learning algorithms includes, but are not limited to, Average One-Dependence Estimators (AODE), Fisher's linear discriminant, Logistic regression,, Perceptron, Multilayer Perceptron, Artificial Neural Networks, Supportvector machines, Quadratic classifiers, Boosting, Decision trees, C4.5, Bayesian networks, Hidden Markov models, High-Dimensional Discriminant Analysis, and Gaussian Mixture Models. In some embodiments, the machine learning algorithm comprises support vector machines, Naive Bayes classifier, k-nearest neighbor, high-dimensional discriminant analysis, or Gaussian mixture models. In some embodiments, the machine learning algorithm comprises Random Forests.Additional Techniques and Tests

[0096] In some embodiments, suitable factors known in the art for diagnosing and / or suggesting, selecting, designating, recommending or otherwise determining a course of treatment for a patient or class of patients having or suspected of having prostate cancer can be employed in combination with measurements of the target sequence expression. In some embodiments, the methods disclosed herein may include additional techniques such as cytology, histology, ultrasound analysis, MRI results, CT scan results, and measurements of PSA levels and / or cancer marker levels.

[0097] In some embodiments, certified tests for classifying disease status and / or designating treatment modalities are also used in diagnosing, predicting, and / or monitoring the status or outcome of a cancer in a subject. In some embodiments, a certified test comprises a means for characterizing the expression levels of one or more of the target sequences of interest, and a certification from a government regulatory agency endorsing use of the test for classifying the disease status of a biological sample, e.g., response to treatment with dose- escalated radiation therapy.

[0098] In some embodiments, the certified test comprises reagents for amplification reactions used to detect and / or quantitate expression of the target sequences to be characterized in the test. In some embodiments, an array of probe nucleic acids is used, with or without prior target amplification, for use in measuring target sequence expression.

[0099] In some embodiments, the test is submitted to an agency having authority to certify the test for use in distinguishing disease status and / or outcome. In some embodiments, results of detection of expression levels of the target sequences used in the test and correlation with disease status and / or outcome are submitted to the agency. In some embodiments, a certification authorizing the diagnostic and / or prognostic use of the test is obtained.

[0100] In some embodiments, also provided are portfolios of expression levels comprising a plurality of normalized expression levels of the targets selected from any of Table 1. In some embodiments, the targets are selected from Table 1. In some embodiments, also provided are portfolios of expression levels comprising a plurality of normalized expression levels of the targets selected from any of the biomarkers of Table 1 or the subsets of Table 2. In some embodiments, the targets are selected from biomarkers selected from Table 1, or from subsets of Table 2. Such portfolios may be provided by performing the methods described herein to obtain expression levels from an individual patient or from a group of patients. In some embodiments, the expression levels are normalized by any suitable method known in the art. In some embodiments, normalization methods that are used include Robust Multichip Average (RMA), probe logarithmic intensity error estimation (PLIER), non-linear fit (NLFIT) quantile-based and nonlinear normalization, and combinations thereof. In some embodiments, background correction can also be performed on the expression data. In some embodiments, techniques useful for background correction include mode of intensities, normalized using median polish probe modeling and sketch-normalization.

[0101] In some embodiments, portfolios are established such that the combination of targets in the portfolio exhibit improved sensitivity and specificity relative to known methods. In some embodiments, a group of targets are selected for inclusion in a portfolio based on a small standard deviation in expression measurements correlating with greater specificity. In some embodiments, other measurements of variation such as correlation coefficients can also be used in this capacity. In some embodiments, the expression level determines the status or outcome (e.g., response to treatment with dose-escalated radiation therapy) of a prostate cancer m the subject with at least about 45% specificity, at least about 50% specificity, at least about 55% specificity, at least about 60% specificity, at least about 65% specificity, at least about 70% specificity, at least about 75% specificity, at least about 80% specificity. In some embodiments, t the expression level determines the status or outcome of a cancer in the subject with at least about 85% specificity, at least about 90% specificity, or at least about 95% specificity.

[0102] In some embodiments of the methods disclosed herein, the accuracy of diagnosing, monitoring, and / or predicting a status or outcome (e.g., response to treatment with dose-escalated radiation therapy) of a prostate cancer is at least about 45%, at least about 50%,at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, or at least about 95%.

[0103] In some embodiments, the accuracy of a classifier or biomarker (e g., signature) is determined by the 95% confidence interval (CI). In some embodiments, a classifier or biomarker or signature is considered to have good accuracy if the 95% CI does not overlap 1. In some embodiments, the 95% CI of a classifier or biomarker or signature is at least about 1.08, 1.10, 1.12, 1.14, 1.15, 1.16, 1.17, 1.18, 1.19, 1.20, 1.21, 1.22, 1.23, 1.24, 1.25, 1.26, 1,27, 1.28, 1.29, 1,30, 1.31, 1.32, 1,33, 1.34, or 1.35 or more. In some embodiments, the 95% CI of a classifier or biomarker or signature is at least about 1.14, 1.15, 1.16, 1.20, 1.21, 1.26, or 1.28, In some embodiments, the 95% CI of a classifier or biomarker or signature is less than about 1.75, 1.74, 1.73, 1.72, 1.71, 1.70, 1.69, 1.68, 1.67, 1.66, 1.65, 1.64, 1.63, 1.62, 1.61, 1.60, 1.59, 1.58, 1.57, 1.56, 1.55, 1.54, 1.53, 1.52, 1.51, 1.50 or less. In some embodiments, the 95% CI of a classifier or biomarker or signature is less than about 1.61, 1.60, 1.59, 1.58, 1.56, 1.55, or 1,53. In some embodiments, the 95% CI of a classifier or biomarker or signature is between about 1.10 to 1.70, between about 1.12 to about 1.68, between about 1.14 to about 1.62, between about 1.15 to about 1.61, between about 1.15 to about 1.59, between about 1.16 to about 1.160, between about 1.19 to about 1.55, between about 1.20 to about 1.54, between about 1.21 to about 1.53, between about 1.26 to about 1.63, between about 1.27 to about 1.61, or between about 1.28 to about 1.60.

[0104] In some embodiments, the accuracy of a biomarker or classifier or signature is dependent on the difference in range of the 95% Cl (e.g., difference in the high value and low value of the 95% CI interval). In some embodiments, biomarkers or classifiers or signatures with large differences m the range of the 95% CI interval have greater variability and are considered less accurate than biomarkers or classifiers or signatures with small differences in the range of the 95% CI intervals. In some embodiments, a biomarker or classifier or signature is considered more accurate if the difference in the range of the 95% Cl is less than about 0.60, 0.55, 0.50, 0.49, 0.48, 0.47, 0.46, 0.45, 0.44, 0.43, 0.42, 0.41, 0.40, 0.39, 0.38, 0.37, 0.36, 0.35, 0.34, 0.33, 0.32, 0.31, 0.30, 0.29, 0.28, 0.27, 0.26, 0.25 or less. In some embodiments, the difference in the range of the 95% CI of a biomarker or classifier or signature is less than about 0.48, 0.45, 0.44, 0.42, 0.40, 0.37, 0.35, 0.33, or 0.32. In some embodiments, the difference in the range of the 95% CI for a biomarker or classifier orsignature is between about 0.25 to about 0.50, between about 0.27 to about 0.47, or between about 0.30 to about 0.45.

[0105] In some embodiments of the methods disclosed herein, the sensitivity is at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, or at least about 95%.

[0106] In some embodiments, the classifiers or biomarkers, or signatures disclosed herein are clinically significant. In some embodiments, the clinical significance of the classifiers or biomarkers or signature is determined by the AUC value. In order to be clinically significant, the AUC value is at least about 0.5, 0.55, 0.6, 0.65, 0.7, 0.75, 0.8, 0.85, 0.9, or 0.95. In some embodiments, the clinical significance of the classifiers or biomarkers or signature is determined by the percent accuracy. For example, in some embodiments, a classifier or biomarker or signature is determined to be clinically significant if the accuracy of the classifier or biomarker or signature is at least about 50%, 55%, 60%, 65%, 70%, 72%, 75%, 77%, 80%, 82%, 84%, 86%, 88%, 90%, 92%, 94%, 96%, or 98%.

[0107] In some embodiments, the clinical significance of the classifiers or biomarkers or signature is determined by the median fold difference (MDF) value. In some embodiments, m order to be clinically significant, the MDF value is at least about 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.9, or 2.0. In some embodiments, the MDF value is greater than or equal to 1.1. In some embodiments, the MDF value is greater than or equal to 1.2. In some embodiments, alternatively, or additionally, the clinical significance of the classifiers or biomarkers or signature is determined by the t-test P-value. In some embodiments, in order to be clinically significant, the t-test P-value is less than about 0.070, 0.065, 0.060, 0.055, 0.050, 0.045, 0.040, 0.035, 0.030, 0.025, 0.020, 0.015, 0.010, 0.005, 0.004, or 0.003. The t-test P-value can be less than about 0.050. In some embodiments, the t-test P-value is less than about 0.010.

[0108] In some embodiments, the clinical significance of the classifiers or biomarkers or signature is determined by the clinical outcome. For example, in some embodiments, different clinical outcomes can have different minimum or maximum thresholds for AUC values, MDF values, t-test P-values, and accuracy values that determine whether the classifier or biomarker or signature is clinically significant. In some embodiments, a classifieror biomarker or signature is considered clinically significant if the P-value of the t-test is less than about 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02, 0.01, 0.005, 0.004, 0.003, 0.002, or 0.001. In some embodiments, the P-value is based on any of the following comparisons: BCR vs non¬ BCR, CP vs non-CP, PCSM vs non-PCSM. For example, in some embodiments, a classifier or biomarker or signature is determined to be clinically significant if the P-values of the differences between the KM curves for BCR vs non-BCR, CP vs non-CP, PCSM vs non-PCSM is lower than about 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02, 0.01, 0.005, 0.004, 0.003, 0.002, or 0.001, For example, in some embodiments, a signature is determined to be clinically significant if the P-values of the differences between the KM curves for progression tree survival / disease recurrence or overall survival is lower than about 0.08, 0,07, 0.06, 0,05, 0.04, 0.03, 0.02, 0.01, 0.005, 0.004, 0.003, 0.002, or 0.001.

[0109] In some embodiments, the performance of the classifier or biomarker or signature is based on the odds ratio. In some embodiments, a classifier or biomarker or signature is considered to have good performance if the odds ratio is at least about 1.30, 1.31, 1.32, 1.33, 1.34, 1.35, 1.36, 1.37, 1.38, 1.39, 1.40, 1.41, 1.42, 1.43, 1.44, 1.45, 1.46, 1.47, 1.48, 1.49, 1.50, 1.52, 1.55, 1.57, 1.60, 1.62, 1.65, 1.67, 1.70 or more. In some embodiments, the odds ratio of a classifier or biomarker or signature is at least about 1.33.

[0110] In some embodiments, the clinical significance of the classifiers and / or biomarkers or signature is based on Univariable Analysis Odds Ratio P-value (uvaORPval ). In some embodiments, the Univariable Analysis Odds Ratio P-value (uvaORPval ) of the classifier and / or biomarker or signature is between about 0-0.4. In some embodiments, the Univariable Analysis Odds Ratio P-value (uvaORPval ) of the classifier and / or biomarker or signature is between about 0-0.3. In some embodiments, the Univariable Analysis Odds Ratio P-value (uvaORPval ) of the classifier and / or biomarker or signature is between about 0-0.2. In some embodiments, the Univariable Analysis Odds Ratio P-value (uvaORPval ) of the classifier and / or biomarker or signature is less than or equal to 0.25, 0.22, 0.21, 0.20, 0.19, 0.18, 0.17, 0.16, 0.15, 0.14, 0.13, 0.12, 0.11. In some embodiments, the Univariable Analysis Odds Ratio P-value (uvaORPval ) of the classifier and / or biomarker or signature is less than or equal to 0.10, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02, 0.01. In some embodiments, the Univariable Analysis Odds Ratio P-value (uvaORPval ) of the classifier and / or biomarker or signature is less than or equal to 0.009, 0.008, 0.007, 0.006, 0.005, 0.004, 0.003, 0.002, 0.001.

[0111] In some embodiments, the clinical significance of the classifiers and / or biomarkers (e.g., signature) is based on multivariable analysis Odds Ratio P-value (mvaORPval ). In some embodiments, the multivariable analysis Odds Ratio P-value (mvaORPval ) of the classifier and / or biomarker or signature is between about 0-1. In some embodiments, the multivariable analysis Odds Ratio P-value (mvaORPval ) of the classifier and / or biomarker or signature is between about 0-0.9. In some embodiments, the multivariable analysis Odds Ratio P-value (mvaORPval ) of the classifier and / or biomarker or signature is between about 0-0.8. In some embodiments, the multivariable analysis Odds Ratio P-value (mvaORPval ) of the classifier and / or biomarker or signature is less than or equal to 0,90, 0.88, 0.86, 0.84, 0.82, 0.80, In some embodiments, the multivariable analysis Odds Ratio P-value (mvaORPval ) of the classifier and / or biomarker or signature is less than or equal to 0,78, 0.76, 0.74, 0.72, 0.70, 0.68, 0.66, 0.64, 0.62, 0.60, 0.58, 0.56, 0.54, 0.52, 0.50. In some embodiments, the multivariable analysis Odds Ratio P-value (mvaORPval ) of the classifier and / or biomarker or signature is less than or equal to 0.48, 0.46, 0.44, 0.42, 0.40, 0.38, 0.36, 0,34, 0,32, 0.30, 0.28, 0.26, 0.25, 0.22, 0.21, 0.20, 0.19, 0.18, 0.17, 0.16, 0.15, 0.14, 0.13, 0.12, 0.11. In some embodiments, the multivariable analysis Odds Ratio P-value (mvaORPval ) of the classifier and / or biomarker or signature is less than or equal to 0.10, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02, 0.01. In some embodiments, the multivariable analysis Odds Ratio P-value (mvaORPval ) of the classifier and / or biomarker or signature is less than or equal to 0.009, 0.008, 0.007, 0.006, 0.005, 0.004, 0.003, 0.002, 0.001.

[0112] In some embodiments, the clinical significance of the classifiers and / or biomarkers or signature is based on the Kaplan Meier P-value (KM P-value). In some embodiments, the Kaplan Meier P-value (KM P-value) of the classifier and / or biomarker or signature is between about 0-0.8. In some embodiments, the Kaplan Meier P-value (KM P-value) of the classifier and / or biomarker or signature is between about 0-0.7. In some embodiments, the Kaplan Meier P-value (KM P-value) of the classifier and / or biomarker or signature is less than or equal to 0.80, 0.78, 0.76, 0.74, 0.72, 0.70, 0.68, 0.66, 0.64, 0.62, 0.60, 0.58, 0.56, 0.54, 0.52, 0.50. In some embodiments, the Kaplan Meier P-value (KM P-value) of the classifier and / or biomarker or signature is less than or equal to 0.48, 0.46, 0.44, 0.42, 0.40, 0.38, 0.36, 0.34, 0.32, 0.30, 0.28, 0.26, 0.25, 0.22, 0.21, 0.20, 0.19, 0.18, 0.17, 0.16, 0.15, 0.14, 0.13, 0.12, 0.11. In some embodiments, the Kaplan Meier P-value (KM P-value) of theclassifier and / or biomarker or signature is less than or equal to 0.10, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02, 0.01. In some embodiments, the Kaplan Meier P-value (KM P- value) of the classifier and / or biomarker or signature is less than or equal to 0.009, 0.008, 0.007, 0.006, 0.005, 0.004, 0.003, 0.002, 0.001.

[0113] In some embodiments, the clinical significance of the classifiers and / or biomarkers or signature is based on the survival AUC value (survAUC). In some embodiments, the survival AUC value (survAUC) of the classifier and / or biomarker or signature is between about 0-1. In some embodiments, the survival AUC value (surv AUC) of the classifier and / or biomarker or signature is between about 0-0.9. In some embodiments, the survival AUC value (survAUC) of the classifier and / or biomarker or signature is less than or equal to 1, 0.98, 0.96, 0.94, 0.92, 0.90, 0.88, 0.86, 0.84, 0.82, 0.80, In some embodiments, the survival AUC value (survAUC) of the classifier and / or biomarker or signature is less than or equal to 0.80, 0.78, 0.76, 0.74, 0.72, 0.70, 0.68, 0.66, 0.64, 0.62, 0.60, 0.58, 0.56, 0.54, 0.52, 0.50. In some embodiments, the survival AUC value (survAUC) of the classifier and / or biomarker or signature is less than or equal to 0.48, 0.46, 0.44, 0.42, 0.40, 0.38, 0.36, 0.34, 0.32, 0.30, 0.28, 0.26, 0.25, 0.22, 0.21, 0.20, 0.19, 0.18, 0.17, 0.16, 0.15, 0.14, 0.13, 0.12, 0.11. In some embodiments, the survival AUC value (survAUC) of the classifier and / or biomarker or signature is less than or equal to 0.10, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02, 0.01. In some embodiments, the survival AUC value (survAUC) of the classifier and / or biomarker or signature is less than or equal to 0.009, 0.008, 0.007, 0.006, 0.005, 0.004, 0.003, 0.002, 0.001.

[0114] In some embodiments, the clinical significance of the classifiers and / or biomarkers or signature is based on the Univariable Analysis Hazard Ratio P-value (uvaHRI’val). In some embodiments, the Univariable Analysis Hazard Ratio P-value (uvaHRPval) of the classifier and / or biomarker or signature is between about 0-0.4. In some embodiments, the Univariable Analysis Hazard Ratio P-value (uvaHRI’val) of the classifier and / or biomarker or signature is between about 0-0.3. In some embodiments, the Univariable Analysis Hazard Ratio P-value (uvaHRPval) of the classifier and / or biomarker or signature is less than or equal to 0.40, 0.38, 0.36, 0.34, 0.32. In some embodiments, the Univariable Analysis Hazard Ratio P-value (uvaHRPval) of the classifier and / or biomarker or signature is less than or equal to 0.30, 0.29, 0.28, 0.27, 0.26, 0.25, 0.24, 0.23, 0.22, 0.21, 0.20. In some embodiments, the Univariable Analysis Hazard Ratio P-value (uvaHRPval) of the classifierand / or biomarker or signature is less than or equal to 0.19, 0.18, 0.17, 0.16, 0.15, 0.14, 0.13, 0.12, 0.11. In some embodiments, the Univariable Analysis Hazard Ratio P-value (uvaHRPval) of the classifier and / or biomarker or signature is less than or equal to 0.10, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02, 0.01. In some embodiments, the Univariable Analysis Hazard Ratio P-value (uvaHRPval) of the classifier and / or biomarker or signature is less than or equal to 0.009, 0.008, 0.007, 0.006, 0.005, 0.004, 0.003, 0.002, 0.001.

[0115] In some embodiments, the clinical significance of the classifiers and / or biomarkers or signature is based on the Multivariable Analysis Hazard Ratio P-value (mvaHRPval)mva HRPval. In some embodiments, the Multivariable Analysis Hazard Ratio P-value (mvaHRPval)mva HRPval of the classifier and / or biomarker or signature is between about 0-1. In some embodiments, the Multivariable Analysis Hazard Ratio P-value (mvaHRPval )mva HRPval of the classifier and / or biomarker or signature is between about 0-0.9. In some embodiments, the Multivariable Analysis Hazard Ratio P-value (mvaHRPval )mva HRPval of the classifier and / or biomarker or signature is less than or equal to 1, 0.98, 0.96, 0.94, 0.92, 0.90, 0.88, 0.86, 0.84, 0.82, 0.80. In some embodiments, the Multivariable Analysis Hazard Ratio P-value (mvaHRPval)mva HRPval of the classifier and / or biomarker or signature is less than or equal to 0.80, 0.78, 0.76, 0.74, 0.72, 0.70, 0.68, 0.66, 0.64, 0.62, 0.60, 0.58, 0.56, 0.54, 0.52, 0.50. In some embodiments, the Multivariable Analysis Hazard Ratio P-value (mvaHRPval)mva HRPval of the classifier and / or biomarker or signature is less than or equal to 0.48, 0.46, 0.44, 0.42, 0.40, 0.38, 0.36, 0.34, 0.32, 0.30, 0.28, 0.26, 0.25, 0.22, 0.21, 0.20, 0.19, 0.18, 0.17, 0.16, 0.15, 0.14, 0.13, 0.12, 0.11. In some embodiments, the Multivariable Analysis Hazard Ratio P-value (mvaHRPval)mva HRPval of the classifier and / or biomarker or signature is less than or equal to 0.10, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02, 0.01. In some embodiments, the Multivariable Analysis Hazard Ratio P-value (mvaHRI’val)mva HRPval of the classifier and / or biomarker or signature is less than or equal to 0.009, 0.008, 0.007, 0.006, 0.005, 0.004, 0.003, 0.002, 0.001.

[0116] In some embodiments, the clinical significance of the classifiers and / or biomarkers or signature is based on the Multivariable Analysis Hazard Ratio P-value (mvaHRPval). In some embodiments, the Multivariable Analysis Hazard Ratio P-value (mvaHRPval) of the classifier and / or biomarker or signature is between about 0 to about 0.60. significance of the classifier and / or biomarker or signature is based on the MultivariableAnalysis Hazard Ratio P-value (mvaHRPval). In some embodiments, the Multivariable Analysis Hazard Ratio P-value (mvaHRPval) of the classifier and / or biomarker or signature is between about 0 to about 0.50. significance of the classifier and / or biomarker or signature is based on the Multivariable Analysis Hazard Ratio P-value (mvaHRPval). In some embodiments, the Multivariable Analysis Hazard Ratio P-value (mvaHRPval) of the classifier and / or biomarker or signature is less than or equal to 0.50, 0.47, 0.45, 0.43, 0.40, 0.38, 0.35, 0.33, 0.30, 0.28, 0.25, 0.22, 0.20, 0.18, 0.16, 0.15, 0.14, 0.13, 0.12, 0.11, 0.10. In some embodiments, the Multivariable Analysis Hazard Ratio P-value (mvaHRPval) of the classifier and / or biomarker or signature is less than or equal to 0,10, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02, 0.01. In some embodiments, the Multivariable Analysis Hazard Ratio P-value (mvaHRPval) of the classifier and / or biomarker or signature is less than or equal to 0,01, 0.009, 0.008, 0.007, 0.006, 0.005, 0.004, 0.003, 0.002, 0.001.

[0117] In some embodiments, the classifiers and / or biomarkers or signature disclosed herein outperform current classifiers or clinical variables or signatures in providing clinically relevant analysis of a sample from a subject. In some embodiments, the classifiers or biomarkers or signature more accurately predict a clinical outcome or status as compared to current classifiers or clinical variables or signatures. For example, m some embodiments, a classifier or biomarker or signature may more accurately predict response of prostate cancer in a subject who has not undergone radical prostatectomy (or any other surgery to remove the cancerous prostate) to dose-escalated radiation therapy. In some embodiments, the performance of a classifier or biomarker or signature disclosed herein is based on the AUC value, odds ratio, 95% CI, difference in range of the 95% CI, p-value or any combination thereof.

[0118] In some embodiments, the performance of the classifiers and / or biomarkers or signature disclosed herein is determined by AUC values and an improvement in performance is determined by the difference in the AUC value of the classifier or biomarker or signature disclosed herein and the AUC value of current classifiers or clinical variables. In some embodiments, a classifier and / or biomarker or signature disclosed herein outperforms current classifiers or clinical variables or signatures when the AUC value of the classifier and / or or biomarker or signature disclosed herein is greater than the AUC value of the current classifiers or clinical variables or signatures by at least about 0.05, 0.06, 0.07, 0.08, 0.09, 0.10,0.11, 0.12, 0.13, 0.14, 0.15, 0.16, 0.17, 0.18, 0.19, 0.20, 0.022, 0.25, 0.27, 0.30, 0.32, 0.35, 0.37, 0.40, 0.42, 0.45, 0.47, 0.50 or more. In some embodiments, the AUC value of the classifier and / or or biomarker or signature disclosed herein is greater than the AUC value of the current classifiers or clinical variables or signatures by at least about 0.10. In some embodiments, the AUC value of the classifier and / or or biomarker or signature disclosed herein is greater than the AUC value of the current classifiers or clinical variables or signatures by at least about 0.13. In some embodiments, the AUC value of the classifier and / or or biomarker or signature disclosed herein is greater than the AUC value of the current classifiers or clinical variables or signatures by at least about 0, 18.

[0119] In some embodiments, the performance of the classifiers and / or biomarkers or signature disclosed herein is determined by the odds ratios and an improvement in performance may be determined by comparing the odds ratio of the classifier or biomarker or signature disclosed herein and the odds ratio of current classifiers or clinical variables. In some embodiments, comparison of the performance of two or more classifiers, biomarkers, and / or clinical variables or signatures is based on the comparison of the absolute value of (1-odds ratio) of a first classifier, biomarker or clinical variable or signature to the absolute value of (1-odds ratio) of a second classifier, biomarker or clinical variable or signature. In some embodiments, the classifier, biomarker or clinical variable or signature with the greater absolute value of (1-odds ratio) is considered to have better performance as compared to the classifier, biomarker or clinical variable or signature with a smaller absolute value of (1-odds ratio).

[0120] In some embodiments, the performance of a classifier, biomarker or clinical variable or signature is based on the comparison of the odds ratio and the 95% confidence interval (CI). In some embodiments, for example, a first classifier, biomarker or clinical variable or signature may have a greater absolute value of (1-odds ratio) than a second classifier, biomarker or clinical variable or signature, however, the 95% CI of the first classifier, biomarker or clinical variable or signature may overlap 1 (e.g., poor accuracy), whereas the 95% CI of the second classifier, biomarker or clinical variable or signature does not overlap 1. In this instance, the second classifier, biomarker or clinical variable or signature is considered to outperform the first classifier, biomarker or clinical variable or signature because the accuracy of the first classifier, biomarker or clinical variable or signature is lessthan the accuracy of the second classifier, biomarker or clinical variable or signature. In another example, a first classifier, biomarker or clinical variable or signature may outperform a second classifier, biomarker or clinical variable or signature based on a comparison of the odds ratio; however, the difference in the 95% CI of the first classifier, biomarker or clinical variable or signature is at least about 2 times greater than the 95% CI of the second classifier, biomarker or clinical variable or signature. In this instance, the second classifier, biomarker or clinical variable or signature is considered to outperform the first classifier or signature.

[0121] In some embodiments, a classifier or biomarker or signature disclosed herein is more accurate than a current classifier or clinical variable or signature. In some embodiments, the classifier or biomarker or signature disclosed herein is more accurate than a current classifier or clinical variable or signature if the range of 95% CI of the classifier or biomarker or signature disclosed herein does not span or overlap 1 and the range of the 95% CI of the current classifier or clinical variable or signature spans or overlaps 1.

[0122] In some embodiments, a classifier or biomarker or signature disclosed herein is more accurate than a current classifier or clinical variable or signature. In some embodiments, the classifier or biomarker or signature disclosed herein is more accurate than a current classifier or clinical variable or signature when difference in range of the 95% CI of the classifier or biomarker or signature disclosed herein is about 0.70, 0.60, 0.50, 0.40, 0.30, 0.20, 0.15, 0.14, 0.13, 0.12, 0.10, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02 times less than the difference in range of the 95% CI of the current classifier or clinical variable or signature. In some embodiments, the classifier or biomarker or signature disclosed herein is more accurate than a current classifier or clinical variable or signature when difference in range of the 95% CI of the classifier or biomarker or signature disclosed herein between about 0.20 to about 0.04 times less than the difference in range of the 95% CI of the current classifier or clinical variable or signature.

[0123] In some embodiments, the methods disclosed herein may comprise the use of a genomic classifier (GC) model. In some embodiments, a general method for developing a GC model may comprise (a) providing a sample from a subject suffering from a cancer; (b) assaying the expression level for a plurality of targets; (c) generating a model (e.g., signature) by using a machine learning algorithm. In some embodiments, the machine learning algorithm comprises Random Forests, or those described above and elsewhere herein. In someembodiments, a GC model (or signature) is developed by using a machine learning algorithm to analyze and rank genomic features. In some embodiments, analyzing the genomic features comprises classifying one or more genomic features. In some embodiments, the method further comprise validating the classifier or signature and / or refining the classifier or signature by using a machine learning algorithm.

[0124] In some embodiments, a general method for developing a model comprises (a) providing a sample from a subject suffering from a cancer; (b) assaying the expression level for a plurality of targets; (c) generating a model (e.g., signature) by using a machine learning algorithm. In some embodiments, the machine learning algorithm comprises Random Forests, In some embodiments, the methods disclosed herein may comprise the use of a genomic-clinical classifier (GCC) model. In some embodiments, a general method for developing a GCC model comprises (a) providing a sample from a subject suffering from a cancer; (b) assaying the expression level for a plurality of targets; (c) generating a model by using a machine learning algorithm. In some embodiments, the machine learning algorithm comprises Random Forests.Terms

[0125] Unless defined otherwise or the context clearly dictates otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs when read m light of the current disclosure.

[0126] " Biomarker" has its plain and ordinary meaning as understood in light of this disclosure. In some embodiments biomarker refers to an measurable characteristic or indicator, comprising a nucleic acid, gene, its transcript (e.g., RNA expressed from the gene), protein, or metabolite, found in blood, other body fluids, or tissues, that signals a normal or abnormal biological process, a disease state, or a person’s response to an exposure or treatment. In some embodiments, the biomarker is a nucleic acid, optionally a gene, optionally a transcript (e.g., an RNA).

[0127] A "target sequence" has its plain and ordinary meaning as understood in light of the specification In some embodiments, target nucleic acid sequence refers to the sequence of a nucleic acid that is utilized to determine the expression level of a biomarker (e.g., a nucleic acid biomarker). In some embodiments, target nucleic acid sequence can be a regionof genome and / or specific gene. In some embodiments, it may be selected from the biomarkers disclosed in Table 1 or a subset of Table 2. In some embodiments, one or more probes or primers can be designed against such biomarkers, e.g., for detecting, quantifying, amplifying or sequencing. In some embodiments, target nucleic acid sequence comprises a coding sequence or a non-coding sequence. In some embodiments, target nucleic acid sequence comprise regions on a protein- coding gene or a non-protein coding gene. In some embodiments, a target nucleic acid sequence comprises exonic and / or non-exonic sequences. In some embodiments, exonic sequences may comprise an exon, UTR, or a portion thereof (e.g., on a protein-coding gene). In some embodiments, non-exonic sequences may comprise regions on a protein-coding gene, non-protein-coding gene, or a portion thereof.. In some embodiments, non-exonic sequences may comprise regions on a protein-coding gene, non-protein-coding gene, or a portion thereof. In some embodiments, for example, non-exonic sequences may comprise intronic regions, promoter regions, intergenic regions, a non-coding transcript, an exon anti-sense region, an intronic anti-sense region, UTR anti-sense region, noncoding transcript anti-sense region, or a portion thereof.

[0128] In some embodiments, a probe is any polynucleotide capable of selectively hybridizing to a target sequence or its complement, or to an RNA version of either. A probe may comprise ribonucleotides, deoxyribonucleotides, peptide nucleic acids, and combinations thereof. In some embodiments, a probe may comprise one or more labels. In some embodiments, a probe is used to amplify one or both strands of a target sequence or an RNA form thereof, acting as a sole primer in an amplification reaction or as a member of a set of primers.

[0129] Unless stated otherwise or it is clear from the context, as used herein, the term "about" is + / - 10% of a given value.

[0130] Use of the singular forms "a," "an," and "the" include plural references unless the context clearly dictates otherwise. Thus, for example, reference to "a polynucleotide" includes a plurality of polynucleotides, reference to "a target" includes a plurality of such targets, reference to "a normalization method" includes a plurality of such methods, and the like. Additionally, use of specific plural references, such as "two," "three," etc., read on larger numbers of the same subject, unless the context clearly dictates otherwise.

[0131] Where a range of values is recited, it is to be understood that each intervening integer value, and each fraction thereof, between the recited upper and lower limits of that range is also disclosed, along with each subrange between such values. In some embodiments, the upper and lower limits of any range can independently be included m or excluded from the range, and each range where either, neither or both limits are included is also encompassed within the disclosure. Where a value being discussed has inherent limits, for example, where a component can be present at a concentration of from 0 to 100%, or where the pH of an aqueous solution can range from 1 to 14, those inherent limits are disclosed. Where a value is explicitly recited, it is to be understood that values, which are about the same quantity or amount as the recited value, are also within the scope of the disclosure, as are ranges based thereon. In some embodiments, where a combination is disclosed, each sub-combination of the elements of that combination is also specifically disclosed and is within the scope of the disclosure. In some embodiments, where different elements or groups of elements are disclosed, combinations thereof are also disclosed. In some embodiments where any element of a disclosure is disclosed as having a plurality of alternatives, examples of that disclosure in which each alternative is excluded singly or in any combination with the other alternatives are also hereby disclosed; more than one element of an disclosure can have such exclusions, and all combinations of elements having such exclusions are hereby disclosed.Embodiments

[0132] Some embodiments provided herein are described by way of the following numbered embodiments:1. A method comprising:a. obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy, wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of: KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23; andb. calculating or having calculated a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and optionally determining a response to treatment comprising dose-escalated radiation therapy that the subject with prostate cancer who has not undergone radical prostatectomy is expected to have based on at least the risk score. A method comprising:a. obtaining or having obtained an expression level of one target or a plurality of targets m a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy, wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of: KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23; andb. determining a response to treatment comprising dose-escalated radiation therapy that the subject with prostate cancer who has not undergone radical prostatectomy is expected to have based on at least the expression level of the one target or each of the plurality of targets in the sample in step (a. ).A method comprising:a. obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy, wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of: KRT14, EMEI, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23;b. determining or having determined a response to treatment comprising dose- escalated radiation therapy that the subject with prostate cancer who has not undergone radical prostatectomy is expected to have based on at least the expression level of the one target or each of the plurality of targets in the samplein step (a.), optionally wherein the determining or having determined comprises calculating or having calculated a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets and the determining or having determined is based on the risk score; and c. administering or having administered to the subject with prostate cancer who has not undergone radical prostatectomy a treatment selected from:i. a treatment comprising dose-escalated radiation therapy; or ii, a treatment comprising non-dose-escalated radiation therapy, wherein the treatment is selected based on the expected response to treatment comprising dose-escalated radiation therapy determined in (b.). 4. A method comprising:a. obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy, wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of: KRT14, EMEI, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3. SULF2, IL7R, HCLS1, and KIF23;b. calculating or having calculated a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets of (a. ); and c. administering or having administered to the subject with prostate cancer who has not undergone radical prostatectomy in a treatment selected from:i. a treatment comprising dose-escalated radiation therapy; or ii. a treatment comprising non-dose-escalated radiation therapy, wherein the treatment is selected based on the risk score of (b. ).5. The method of any one of embodiment 1-3, wherein the response to treatment comprising dose-escalated radiation therapy comprises risk of one or more of: biochemical failure (BF), distant metastasis (DM) and receipt of salvage therapy, optionally wherein the risk is at 3, 5, 10 or 15 years post-treatment with the radiation therapy.6. The method of any one of embodiments 3-5, wherein the treatment (i.) comprising dose-escalated radiation therapy further comprises administration of another treatment for prostate cancer, and / or wherein the treatment (li.) comprising non-dose-escalated radiation therapy further comprises another treatment for prostate cancer.7. The method of any one of embodiments 1-6, wherein the one target or plurality of targets comprises or consists of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23 or 24 target nucleic acid sequences, optionally wherein each target nucleic acid sequence is selected from a different gene selected from the group of genes consisting of: KRT14, EMEI, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23.8. The method of any one of embodiments 1-7, wherein the one target or plurality of targets comprises or consists of a nucleic acid sequence of each of:a. KRT14;b. KRT14, and EME1;c. KRT14, EME1, and DRAM1;d. KRT14, EME1, DRAM1, and ANLN;e. KRT14, EME1, DRAM1, ANLN, and APEX2;f. KRT14, EMEI, DRAM1, ANLN, APEX2, and DTL;g. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, and ARHGAP15; h. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, and GNG11; i. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, and CDKN2AIP;j. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, and PTPN22;k. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, and RPS27A;l. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, and CDKN3;m. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, and IL1B;il. KRT14, EMEI, DRAMI, ANLN, APEX2, DTL, ARHGAP15, GNGII, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, and NEK1;o. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, and UBA7;p. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, and MUM1; q. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, and BIN2;r. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, and TOP2A;s. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, and PLK2;t. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, and ZMAT3;u. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, and SULF2;v. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, and IL7R;w. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, and HCLS1; orx. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23.-so9. The method of any one of embodiments 1-3 and 5-8, wherein said determining or having determined comprises calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, optionally wherein said determining or having determined is based on the risk score.10. The method of any one of embodiments 1 and 3-9, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and wherein said determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy:a. is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; and ii. on a scale from -1 to 1, the risk score being equal to or greater than, 0, - 0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, - 0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, - 0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.42 on a scale from -1 to 1; orb. is expected to respond no better or worse to treatment comprising dose- escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or less than 0, -0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, -0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, -0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two ofthe preceding values, optionally the risk score being equal to or less than -0.41 on a scale from -1 to 1.11. The method of embodiment 10, further comprising:a. selecting for administration and / or administering, or having administered, to the subject expected to respond better to treatment comprising dose-escalated radiation therapy a treatment comprising dose-escalated radiation therapy; or b. selecting for administration and / or administering, or having administered, to the subject expected to respond no better or worse to treatment comprising dose- escalated radiation therapy a treatment comprising non-dose-escalated radiation therapy,12. The method of any one of embodiments 1-10, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and wherein the method comprises:a. selecting for administration and / or administering, or having administered, to the subject a treatment comprising dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; and ii. on a scale from -1 to 1, the risk score being equal to or greater than, 0, - 0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, - 0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, - 0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.42 on a scale from -1 to 1; orb. selecting for administration and / or administering, or having administered, to the subject a treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or less than 0, -0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, -0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, -0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.41 on a scale from -1 to 1.13. The method of any one of embodiments 1 -9, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and wherein said determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy:a. is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or greater than, - 0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, - 0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, - 0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.67 on a scale from -1 to 1; orb. is expected to respond no better or worse to treatment comprising dose- escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or less than, -0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75,-0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.68 on a scale from -1 to 1.14. The method of embodiment 13, further comprising:a. selecting for administration and / or administering, or having administered, to the subject expected to respond better to treatment comprising dose-escalated radiation therapy a treatment comprising dose-escalated radiation therapy; or b. selecting for administration and / or administering, or having administered, to the subject expected to respond no better or worse to treatment comprising dose- escalated radiation therapy a treatment comprising non-dose-escalated radiation therapy.15. The method of any one of embodiments 1-9 or 13, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and wherein the method comprises:a. selecting for administration and / or administering, or having administered, to the subject a treatment comprising dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or greater than, - 0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, - 0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, - 0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.67 on a scale from -1 to 1; orb. selecting for administration and / or administering, or having administered, to the subject a treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or less than, -0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.68 on a scale from -1 to 1.16. The method of any one of embodiments 1 -9, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and wherein said determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy:a. is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; and ii. on a scale from -1 to 1, the risk score being equal to or greater than, 0, - 0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, - 0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, - 0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.42 on a scale from -1 to 1; orb. is expected to respond no better or worse to treatment comprising dose- escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or less than, -0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.68 on a scale from -1 to 1.17. The method of embodiment 16, further comprising:a. selecting for administration and / or administering, or having administered, to the subject expected to respond better to treatment comprising dose-escalated radiation therapy a treatment comprising dose-escalated radiation therapy; or b. selecting for administration and / or administering, or having administered, to the subject expected to respond no better or worse to treatment comprising dose- escalated radiation therapy a treatment comprising non-dose-escalated radiation therapy.18. The method of any one of embodiments 1-9 or 16, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and wherein the method comprises:a. selecting for administration and / or administering, or having administered, to the subject a treatment comprising dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; and ii. on a scale from -1 to 1, the risk score being equal to or greater than, 0, - 0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, - 0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, - 0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.42 on a scale from -1 to 1; orb. selecting for administration and / or administering, or having administered, to the subject a treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or less than, -0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.68 on a scale from -1 to 1.19. The method of any one of embodiments 10-18, wherein the population of patients with prostate cancer who have not undergone radical prostatectomy comprises at least 10, 25, 50, 75 or 100 patients, optionally wherein the population of patients does not have metastatic prostate cancer, and / or the population of patients has intermediate-risk prostate cancer, optionally wherein the population patients have: histologically confirmed adenocarcinoma of the prostate, clinical stage Tla-T2b (stage I or II), Gleason score 2-6 and prostate-specific antigen (PSA) 10 to less than 20 ng / mL, or Gleason score 7 and PSA less than 15 ng / mL, and no distant metastases and no regional lymph node involvement.20. The method of any one of embodiments 1-19, wherein the subj ect is determined to be expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy, optionally wherein the subject is selected for administration and / or is administered a treatment comprising dose-escalated radiation therapy.21. The method of any one of embodiments 1-19, wherein the subj ect is determined to be expected to respond no better or worse to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy, optionally wherein the subject is selected for administration and / or is administered a treatment comprising non- dose-escalated radiation therapy.22. The method of any one of embodiments 1-21, wherein the expected better response to treatment comprising dose-escalated radiation therapy is one or more of:a. a reduction in risk of BF of at least 5%, 10%, 20%, or 30% as compared to treatment comprising non-dose-escalated radiation therapy;b. a reduction in risk of DM of at least 5%, 10%, 20%, or 30% as compared to treatment comprising non-dose-escalated radiation therapy; and c. a reduction in risk of receipt of salvage therapy of at least 5%, 10%, 20%, or 30% as compared to treatment comprising non-dose-escalated radiation therapy;optionally wherein the risk is at 3, 5, 10 or 15 years post-treatment with the radiation therapy,23. The method of any one of embodiments 1-22, wherein the expected no better or worse response to treatment comprising dose-escalated radiation therapy is one or more of:a. reduction in risk of BF of less than 5%, 0%, -5%, or -10% as compared to treatment comprising non-dose-escalated radiation therapy;b. reduction in risk of DM of less than 5%, 0%, -5%, or -10% as compared to treatment comprising non-dose-escalated radiation therapy; and c. reduction in risk of receipt of salvage therapy of less than 5%, 0%, -5%, or - 10% as compared to treatment comprising non-dose-escalated radiation therapy;optionally wherein the risk is at 3, 5, 10 or 15 years post-treatment with the radiation therapy.24. The method of any one of embodiments 5-23, wherein the BF is Pheonix BF.25. The method of any one of the preceding embodiments, wherein the subject does not have metastatic prostate cancer, and / or the subject has intermediate-risk prostate cancer, optionally wherein the subject has a PSA score of < 20 ng / ml, a clinical stage of T1 or T2, and / or a Gleason grade group of 1, 2 or 3.26. The method of any one of the preceding embodiments, wherein the subject satisfies the following criteria:a. histologically confirmed adenocarcinoma of the prostate,b. clinical stage Tla-T2b (stage I or II),c. Gleason score 2-6 and prostate- specific antigen (PSA) 10 to less than 20 ng / mL, or Gleason score 7 and PSA less than 15 ng / mL,d. no distant metastases, ande. no regional lymph node involvement.27. The method of any one of the preceding embodiments, wherein the subject has one or more of the following:a. intermediate risk prostate cancer as defined by the NCCN risk groups; b. low risk prostate cancer as defined by the NCCN risk groups;c. high or very high risk prostate cancer as defined by the NCCN risk groups; d. one or more of the following risk factors: clinical stage T2b- T2c, Grade Group 2 or 3 (Gleason 7), PSA 10-20;e. clinical stage cT1-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL; f. at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL; and / org. metastatic prostate cancer.28. The method of any one of the preceding embodiments, wherein the subject does not have one or more of the following:a. intermediate risk prostate cancer as defined by the NCCN risk groups; b. low risk prostate cancer as defined by the NCCN risk groups;c. high or very high risk prostate cancer as defined by the NCCN risk groups; d. one or more of the following risk factors: clinical stage T2b- T2c, Grade Group 2 or 3 (Gleason 7), PSA 10-20;e. clinical stage cT1-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL; f. at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL; and / org. metastatic prostate cancer.29. The method of any one of the preceding embodiments, wherein said determining or having determined the expected response to treatment comprising dose- escalated radiation therapy is in comparison to a treatment comprising non-dose-escalated radiation therapy.30. The method of any one of the preceding embodiments, wherein said determining or having determined the expected response of the subject with prostate cancer who has not undergone radical prostatectomy to treatment comprising dose-escalated radiationtherapy is not based on prognostic clinical variables, optionally wherein the prognostic clinical variables are tumor stage, nodal stage, metastatic tumor volume categorization, a Gleason score and / or a serum PSA level of the subject.31. The method of any one of the preceding embodiments, wherein said obtaining or having obtained the expression level of the one target or the plurality of targets comprises utilizing a microarray to assess one target or the plurality of targets, performing qPCR on the one target or the plurality of targets, and / or sequencing the one target or the plurality of targets.32. The method of any one of the preceding embodiments, wherein the sample is a resected specimen, optionally a transurethral resection of prostate (TURP) specimen, or a biopsy of the prostate cancer, optionally a needle biopsy,33. The method of any one of the preceding embodiments, wherein the dose-escalated radiation therapy is at least 78 Gy, optionally at least 79 Gy or 80 Gy,34. The method of any one of the preceding embodiments, wherein the non-dose-escalated radiation therapy is not more than 71 Gy, optionally not more than 70 Gy,35. The method of any one of the preceding embodiments, wherein is selected from 3-Dimensional Conformal Radiation Therapy (3D-CRT), Intensity-Modulated Radiation Therapy (IMRT), volumetric modulated arc therapy (VMAT), stereotactic body radiotherapy (SBRT or SABR), proton beam RT, and brachytherapy boost, optionally wherein it is 3D-CRT or IMRT.36. The method of any one of the preceding embodiments wherein the dose- escalated and / or non-dose-escalated radiation therapy is definitive radiation therapy. In some embodiments, the dose-escalated and / or non-dose-escalated radiation therapy is not salvage radiation therapy.37. The method of any one of the preceding embodiments, wherein the risk factor score is a PORTOS score.38. The method of any one of the preceding embodiments, wherein the subject does not have NCCN unfavorable intermediate-risk prostate cancer.39. A kit for use in the method of any one of the preceding embodiments, the kit comprising reagents for obtaining the expression level of the one target or each of the plurality of targets.40. The kit of embodiment 39, wherein the reagents comprise primer(s) and / or probe(s) for obtaining the expression level of the one target or each of the plurality of targets.ReferencesThe following references are incorporated herein by reference in their entirety.1. 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). https: / / doi.org / 10.1056 / NEJMoa16062202. Kishan, A. U. et al. High-dose Radiotherapy or Androgen Deprivation Therapy (HEAT) as Treatment Intensification for Localized Prostate Cancer: An Individual Patient-data Network Meta-analysis from the MARCAP Consortium. Eur Urol 82, 106-114 (2022). https: / / doi.org / 10.1016 / j.eururo.2022.04.0033. Zhao, S. G. et al. Development and validation of a 24-gene predictor of response to postoperative radiotherapy in prostate cancer: a matched, retrospective analysis. Lancet Oncol (2016). https: / / doi.org / 10.1016 / S1470-2045(16)30491-04. Dal Pra, A. et al. in ASTRO Annual Meeting.5. Michalski, J. M. et al. Effect of Standard vs Dose-Escalated Radiation Therapy for Patients With Intermediate-Risk Prostate Cancer: The NRG Oncology RTOG 0126 Randomized Clinical Trial. JAMA Oncol 4, el 80039 (2018). https: / / doi.org / 10.1001 / jamaoncol.2018.00396. Spratt, D. E. et al. Genomic Classifier Performance in Intermediate-Risk Prostate Cancer: Results From NRG Oncology / RTOG 0126 Randomized Phase 3 Trial. Int J Radiat Oncol Biol Phys 117, 370-377 (2023). https: / / doi.org / 10.1016 / j.ijrobp.2023.04.0107. Roach, M., 3rd et al. Defining biochemical failure following radiotherapy with or without hormonal therapy in men with clinically localized prostate cancer: recommendations of the RTOG- ASTRO Phoenix Consensus Conference. Int J Radiat Oncol Biol Phys 65, 965-974 (2006). https: / / doi.org / 10.1016 / j.ijrobp.2006.04.0298. Andrade, C. Mean Difference, Standardized Mean Difference (SMD), and Their Use in Meta- Analysis: As Simple as It Gets. J Clin Psychiatry 81 (2020). https: / / doi.org / 10.4088 / JCP.20f13681Milosevic, M. et al. Tumor hypoxia predicts biochemical failure following radiotherapy for clinically localized prostate cancer. Clin Cancer Res 18, 2108-2114 (2012). https: / / doi.org / 10.1158 / 1078-0432.CCR-11-2711Bristow, R. G, Berlin, A. & Dal Pra, A. An arranged marriage for precision medicine: hypoxia and genomic assays in localized prostate cancer radiotherapy. Br J Radiol 87, 20130753 (2014). https: / / doi.org / 10.1259 / bjr.20130753Zhao, S. G. et al. Associations of Luminal and Basal Subtyping of Prostate Cancer With Prognosis and Response to Androgen Deprivation Therapy, JAMA Oncol 3, 1663-1672 (2017). https: / / doi.org / 10.1001 / jamaoncol.2017.0751Zhao, S. G. et al. Clinical and Genomic Implications of Luminal and Basal Subtypes Across Carcinomas. Clin Cancer Res 25, 2450-2457 (2019). https: / / doi.org / 10.1158 / 1078-0432.CCR-18-3121Aggarwal, R. et al. Prognosis Associated With Luminal and Basal Subtypes of Metastatic Prostate Cancer. JAMA Oncol (2021). https: / / doi.org / 10.1001 / jamaoncol.2021.3987Weiner, A. B. et al. A novel prostate cancer subtyping classifier based on luminal and basal phenotypes. Cancer 129, 2169-2178 (2023). https: / / doi.org / 10.1002 / cncr.34790 Ngwa, W. et al. Using immunotherapy to boost the abscopal effect. Nat Rev Cancer 18, 313-322 (2018). https: / / doi.org / 10.1038 / nrc.2018.6Kerkmeijer, L. G. W. et al. Focal Boost to the Intraprostatic Tumor in External Beam Radiotherapy for Patients With Localized Prostate Cancer: Results From the FLAME Randomized Phase Ill Trial. J Clin Oncol 39, 787-796 (2021). https: / / doi.org / 10.1200 / JCO.20.02873Michalski, J. M. et al. Effect of Brachytherapy With External Beam Radiation Therapy Versus Brachytherapy Alone for Intermediate-Risk Prostate Cancer: NRG Oncology RTOG 0232 Randomized Clinical Trial. J Clin Oncol 41, 4035-4044 (2023). https: / / doi.org / 10.1200 / JCO.22.01856Simon, R. M., Paik, S. & Hayes, D. F. Use of archived specimens in evaluation of prognostic and predictive biomarkers. J Natl Cancer Inst 101, 1446-1452 (2009). https: / / doi.org / 10.1093 / jnci / djp335.EXAMPLESExample 1:

[0133] The following is a summary of the example described in detail below:

[0134] Background: NRG / RTOG 0126 randomized intermediate risk prostate cancer patients to 70.2Gy vs. 79.2Gy and found improvement in biochemical failure rates with dose escalation, similar to other trials. It was hypothesized that a previously developed post-operative radiation therapy outcomes score (PORTOS) could distinguish patients who did not benefit from dose escalation in the definitive setting.

[0135] Methods: PORTOS scores were calculated on prostate biopsy samples and divided into three equal tertiles (lower, average / middle, and higher). The primary objective of this study was to evaluate PORTOS as a predictive biomarker for the benefit of RT dose escalation on biochemical failure (BF) via the contemporary Phoenix criteria, though the historical ASTRO criteria are reported as this was a protocol endpoint (N=215). Also examined was distant metastasis (DM) and receipt of salvage therapy as additional secondary endpoints. In addition, also investigated was clinical and molecular correlates in a large real-world dataset of 31,107 prostate biopsy samples from men with intermediate risk prostate cancer.

[0136] Results: In NRG / RTOG 0126, in patients with lower tertile PORTOS scores, there was no difference in Phoenix BF (sIIR 1.14 [0.54-2.40], P=0.73). However, for patients in the average and higher tertile PORTOS score range, there was a significant benefit for RT DE for BF Phoenix (average PORTOS: sHR 0.45 [0.22-0.90], P=0.02; higher PORTOS: sHR 0.30 [0.12-0.75], P=0.009). An interaction test indicated a significant difference in benefit for DE between higher and lower PORTOS groups (P=0.048). Similar trends were observed for BF AS TRO, DM, and receipt of salvage therapy. Interestingly, PORTOS was not consistently associated with clinicopathologic variables in the NRG / RTOG 0126 trial or the large real-world dataset. In the latter, PORTOS was modestly associated with hypoxia signatures, and strongly associated with immune signatures and subtypes.

[0137] Conclusion: In the NRG / RTOG 0126 randomized controlled trial, this study validated that PORTOS can identify a subset of patients who do not benefit from dose escalation from 70.2Gy to 79.2Gy, a subgroup that cannot be identified using clinicopathologic or prognostic variables. Validation in a randomized trial with a significant interaction providesstrong evidence that PORTOS is a predictive biomarker for definitive radiation therapy response.INTRODUCTION:

[0138] Definitive radiation therapy is one of the primary treatment modalities for localized prostate cancer. The ProtecT trial established the equivalency in clinical outcomes compared to radical prostatectomy. The radiation course historically consisted of a conventionally fractionated dose of around 70 Gy. However, numerous randomized controlled trials established an improvement in biochemical control with dose escalation to around 80 Gy. Based on these data, escalated dose has become the new standard of care for conventionally fractionated radiation. However, large meta-analyses have failed to demonstrate a metastasis-free survival (MFS) or overall survival (OS) benefit. No clinically validated biomarkers exist which can distinguish between the patients that truly derive a benefit from dose-escalation, and those that could be safely de-escalated.

[0139] A post-operative radiation outcomes score (PORTOS) has been developed, which has been validated to predict patients who benefit from dose escalation in the post¬ operative setting in SAKK 09 / 104. The present study determined that PORTOS can predict which patients may benefit from dose-escalation in the definitive setting, by validating PORTOS in NRG / RTOG 01265, a randomized controlled trial in men with primarily intermediate-risk prostate cancer randomized to 79.2Gy vs 70.2Gy. This trial showed an improvement in biochemical control and distant metastasis with dose-escalation, but not OS, at the cost of increased toxicity.METHODS NRG / RTOG 0126

[0140] The NRG / RTOG 0126 randomized phase III trial tested whether dose- escalated definitive radiation (79.2Gy) was superior to conventional dose definitive radiation (70.2Gy). The primary endpoints were biochemical failure (BF) by the ASTRO / Phoenix criteria. More details on study exclusion and inclusion criteria are available at ClinicalTrials.gov (NCT00033631) which is herein incorporated by reference in its entirety. Briefly, the subjects satisfied the following: histologically confirmed adenocarcinoma of the prostate, clinical stage Tla-T2b (stage I or II), Gleason score 2-6 and prostate-specific antigen (PSA) 10 to less than 20 ng / mL or Gleason score 7 and PSA less than 15 ng / mL, no distantmetastases, and no regional lymph node involvement, among other criteria. Details on the translational research project investigating genomic markers have been previously described. Genomic signatures were generated using the whole transcriptome Decipher assay (Veracyte, Inc., San Francisco, CA). Only 215 patients had tissue available and passed quality control for inclusion in the final analysis as reported previously. All patients in the final analysis had intermediate-risk prostate cancer per the modern NCCN risk groups.Decipher GRID

[0141] Prospectively obtained transcriptomic data from prostate cancer biopsy specimens for men with intermediate-risk prostate cancer (mirroring the NRG / RTOG 0126 population) were obtained from the Genomic Resource for Intelligent Discovery (GRID) database (NCT02609269). A total of 31,107 specimens tested between May 2016 and February 2022 were included to investigate associations between PORTOS and clinicopathologic and genomic variables in a large scale, real-world clinical dataset. These patients’ data were deidentified in accordance with the Safe Harbor method described in the HIPAA Privacy Rule 45 CFR 164.514(b) and (c) (Veracyte, Inc., San Diego, CA) prior to analysis.PORTOS calculation

[0142] PORTOS was derived on the same clinical-grade microarray platform as Decipher, and the score was calculated as previously published. However, the cutoff for the PORTOS high vs. low score was originally derived on radical prostatectomy specimens, not biopsy specimens as was tested. Furthermore, the study evaluated definitive radiation, not salvage therapy. Therefore, PORTOS was investigated in tertiles, balancing subgroup size with discrimination across the range of PORTOS scores. The PORTOS cutoff for Lower / Average is between -0.679 and -0.673, and the cutoff for Average and Higher is between -0.419 and -0.416.Statistical methods

[0143] Descriptive statistics were provided for 215 patients from the NRG / RTOG 0126 randomized phase III trial, including median and interquartile range (IQR) for continuous variables, and frequency and proportion for categorical variables. Comparisons between PORTOS subgroups, divided into three equal tertiles (lower, average, and higher) were conducted using Fisher's exact test for categorical variables and the Kruskal-Wallis test for continuous variables. The primary endpoints of this study were time to biochemical failure(BF) per the Phoenix and ASTRO criteria. The objective of this study was to evaluate PORTOS' potential as a moderator for the benefit of definitive radiation dose escalation on BF. Distant metastasis (DM) and receipt of salvage therapy were also considered as the secondary endpoints in this study.

[0144] Univariable analysis (UVA) of Fine-Gray models, treating death without events as a competing risk, were constructed for all endpoints. Cumulative incidence curves were plotted for each PORTOS subgroup by randomization arms (79.2Gy vs. 70.2Gy) to evaluate the effect of definitive radiation dose escalation. UV As were conducted to investigate the association between definitive radiation dose escalation and endpoints within each PORTOS tertile and to compare PORTOS’ performance with other clinicopathologic variables. Furthermore, standardized mean differences (SMD) were utilized to evaluate the relationship between PORTOS and clinicopathologic variables in a real-world clinical dataset comprising 31,107 NCCN intermediate- risk prostate cancer patients who were de-identified in the GRID database. All statistical tests were two-sided, with p-values less than 0.05 considered statistically significant. All subdistribution hazard ratios (sHRs) and landmark outcomes at five years were reported with 95% confidence intervals (Cis). Analyses were conducted using R, version 4.3.1 (R Foundation for Statistical Computing).RESULTSValidation of PORTOS in NRG / RTOG 0126

[0145] In total, 215 intermediate risk prostate cancer patients from the NRG / RTOG 0126 trial had evaluable PORTOS scores. Since the post-operative cutoffs for high vs. low PORTOS likely do not apply to the definitive setting, we instead divided patients into three equal tertiles for lower, average, and higher PORTOS scores. We first examined associations of PORTOS and clinicopathologic variables (Figure 1). There were no associations that were both statistically significant and consistently ordered (i.e. increasing or decreasing consistently from PORTOS low to medium to high).

[0146] We next examined PORTOS for prognosis as well as potentially identifying patients who benefit from definitive dose-escalation for BF endpoints. We primarily focused on the Phoenix definition as the more contemporary and clinically used definition, but the historical ASTRO definition is reported as it was part of the original protocol. PORTOS score was not prognostic overall for either endpoint (BF ASTRO P=0.15; BF Phoenix P=0.23).Regardmg the predictive value for dose-escalation (Figure 2), in patients with lower PORTOS scores, there was no difference in BF (Phoenix sHR 1.14 [0.54-2.40], P=0.73; ASTRO sHR 1.03 [95% CI: 0.45-2.36], P=0.94). However, for patients with average PORTOS scores, there was a significant benefit for dose-escalation for BF per the Phoenix criteria (sHR 0.45 [0.22- 0.90], P=0.02), with a trend for BF per the ASTRO criteria (sHR 0.60 [0.32-1.14], P=0.12). For patients with higher PORTOS scores, this benefit was even more pronounced for a BF benefit (Phoenix sHR0.30 [0.12-0.75], P=0.009; ASTRO sHR 0.46 [0.23-0.92], P=0.03), with interaction tests indicating a significant difference in benefit for dose escalation between higher and lower PORTOS tertiles for BF Phoenix (p = 0.048; p = 0.09 for ASTRO; Figure 7A-B), When we group patients with average / higher PORTOS scores together (Figures 8A-8B), the interaction P- value was 0.06 for BF Phoenix (Figures 9A-9B) and 0.1 for BF ASTRO (Figure 8B). These results indicate that patients with higher PORTOS scores appear to be benefitting the most from dose-escalated radiation therapy, with minimal benefit in the lower fertile of PORTOS scores,

[0147] We also examined the secondary endpoints of distant metastasis (DM) and receipt of salvage therapy. Like BF, in patients with lower PORTOS scores, there was no difference in DM (sHR 1.73 [0.16-18.43], P=0.65). However, for patients with average PORTOS scores, there was a trend for benefit for dose-escalation for DM (sHR 0.14 [0.02-1.11], P=0.06). In patients with high PORTOS scores, there were no DM events in the dose- escalated arm, only in the standard dose arm, and thus a model did not converge (Figure 3A). For receipt of salvage therapy (Figure 3B), both lower (sHR 0.91 [0.37-2.21], P=0.83) and average (sHR 0.73 [0.30-1.77], P=0.48) PORTOS groups did not show a difference, but the higher PORTOS group demonstrated a significant benefit for dose-escalation (sHR 0.12 [0.03-0.53], P=0.005), with interaction tests indicating a significant difference in benefit for dose escalation between higher and lower PORTOS tertiles for (P=0.026; Figure 7C). The differences in DM and salvage therapy were both statistically significant when comparing lower vs. average / higher PORTOS score (Figures 10A-10B).

[0148] We next evaluated other clinicopathologic variables and compared them to PORTOS in predicting the benefit of dose escalation. Of all the variables tested, no other clinicopathologic variables were consistently significant across both Phoenix and ASTRO BF in identifying patients that benefitted from dose escalation (Figure 4). In total, these datasuggest that PORTOS is predictive of benefit from dose escalation in this population, identifying a subset of patients who have improved outcomes from 79.2Gy vs. 70.2Gy definitive radiation that could not be determined solely from clinicopathologic variables. Evaluation of PORTOS in a real-world clinical dataset

[0149] We next evaluated the association of PORTOS with clinicopathologic and molecular variables in a real- world clinical dataset of 31, 107 intermediate-risk prostate cancer patients with pre-treatment biopsy testing with Decipher (GRID cohort). Because of the size of the dataset, all P- values are highly statistically significant, and so we instead report a standardized median difference (SMD) to compare effect sizes. SMD values of >0.2, >0.5, and >0.8 could be considered small, medium, and large differences. We first examined the association with PORTOS and clinicopathologic variables across this large dataset (Figure 5). Interestingly, there was a modest association of PORTOS with race (SMD = 0.18), However, PSA, clinical T-stage, Gleason grade group, and NCCN risk group all had SMDs <0.1. These results suggest that PORTOS is conveying independent information from clinicopathologic variables in the biopsy setting.

[0150] We next evaluated the association of PORTOS with biological variables (Figure 6). PORTOS was not especially associated with the expression of DNA Damage Response (DDR) gene signature (SMD=0.14). Tumor hypoxia is known to be associated with radiation resistance and we also observed a modest association with higher PORTOS (deriving more benefit from dose-escalation) and a hypoxia signature (SMD=0.29). The strongest association was with an immune signature (SMD=0.81). This was further confirmed when comparing with published subtypes of localized prostate cancer, where there was a very strong enrichment of basal-immune tumors in the higher PORTOS score tumors, with 60.8% of samples falling in this subtype compared to 29.7% of the average PORTOS tumors and 12.7% of the lower PORTOS score tumors (SMD=0.82). These data provide intriguing insights into the underlying biological mechanisms underpinning the differences in radiation response of PORTOS.DISCUSSION

[0151] PORTOS has been validated in predicting a benefit for dose escalation in both the post-operative randomized trial SAKK 09 / 104 and now here in NRG / RTOG 0126. To our knowledge, this is the first molecular signature that has been shown to predict the benefitfor biochemical failure of dose escalation of definitive radiation. The top two thirds of PORTOS patients demonstrated some benefit from dose-escalation, which is most pronounced in the highest tertile across endpoints, consistent with the overall results of NRG / RTOG 0126. However, a significant minority (the bottom third of PORTOS patients) derived no benefit. Since dose escalation comes at the cost of increased toxicity, PORTOS could be used to identify men in which to safely de-escalate radiation dose and could be particularly helpful in situations where dose constraints are proving difficult to meet, or even to stop treatment early in patients who are tolerating treatment poorly.

[0152] Our analysis of a large real-world cohort of molecularly profiled prostate biopsy samples sheds further light into the biological underpinnings of PORTOS, and mirrors the results found in a large real-world radical prostatectomy cohort. There were not strong associations of PORTOS and clinicopathologic variables, suggesting that PORTOS is providing orthogonal information not captured by Gleason, PSA, or T-stage. There was a modest association with expression of hypoxia genes. Hypoxia is known to increase radio resistance in tumors, and thus it makes sense that more hypoxic tumors might have higher PORTOS scores and thus require increased doses of radiation to adequately treat. However, the strongest biological association was with expression of immune pathways, and enrichment of the basal-immune subtype as PORTOS scores increased. Previous studies have established a luminal / basal axis in prostate cancer, and four further subtypes have been identified: luminal- differentiated (LD), luminal-proliferating (LP), basal-immune (BI) and basal-neuroendocrine (BN). Interactions between radiation and the immune system continue to be areas of intense investigation. These data provide some insight into the biological underpinnings of the PORTOS score.

[0153] Dose-escalation in prostate cancer has also not stopped at 80 Gy. Further dose-escalation using simultaneous integrated boosts (SIBs) m randomized trials has been explored with conventionally fractionated doses in the FLAME trial, showing a further biochemical control benefit. Patients predicted to benefit from dose-escalation by PORTOS, especially those with the highest scores, may also benefit from further dose escalation using SIB, as they have tumors which benefit from higher radiation doses. In intermediate-risk patients receiving brachytherapy, EBRT boost is another method of dose escalation. In NRG / RTOG 0232, no overall benefit was identified for combination EBRT and brachytherapycompared with brachytherapy alone, but there was a trend suggesting a subset of patients might benefit. Investigation of PORTOS in the context of these additional modalities could help establish additional clinical scenarios where PORTOS may be useful in identifying the best candidates for dose-escalation.

[0154] Patients in the NRG / RTOG 0126 were not treated with ADT, which would be considered as standard of care today for patients with NCCN unfavorable intermediate-risk prostate cancer. However, these patients made up the minority of patients in our study. The NRG / RTOG 0126 study was also conducted prior to the widespread adoption of PSMA PET scans. However, metastasis rates were overall low, suggesting that this would likely not have impacted our results. These results provide evidence that PORTOS is a predictive biomarker for definitive radiation therapy response.Example 2:

[0155] This study identified subsets of the PORTOS genes that can still predict DE (dose escalation) response for the Higher and Average groups when evaluated against the BF Phoenix endpoint. Once the genes are ranked by importance to the PORTOS score, we generated subsets by removing the least important gene to create a 23 gene subset, the 2 least important to create a 22 gene subset, etc. The resulting subsets are shown in Table 2. we evaluated the correlation between the subset PORTOS and the original PORTOS by removing varying numbers of the least important genes. The results of the correlation is shown in Table 2. Additionally, we assessed the hazard ratio between the two doses of radiation therapy in both the Higher and Average groups to ensure that the p-values remain significant for both groups. The results are shown in Figures 11A and 11B, which are a Biochemical Failure (BF) per Phoenix criteria forest plot summarizing the hazard ratios for the different subsets of PORTOS genes. The p-values remain significant for the Higher and Average groups even after removing large numbers of the least important genes from the subsets.

Claims

WHAT IS CLAIMED IS:

1. A method comprising:a. obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy, wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of: KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23; andb, calculating or having calculated a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and optionally determining a response to treatment comprising dose-escalated radiation therapy that the subject with prostate cancer who has not undergone radical prostatectomy is expected to have based on at least the risk score.

2. A method comprising:a. obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy, wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of: KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23; andb. determining a response to treatment comprising dose-escalated radiation therapy that the subject with prostate cancer who has not undergone radical prostatectomy is expected to have based on at least the expression level of the one target or each of the plurality of targets in the sample in step (a.).

3. A method comprising:a. obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has notundergone radical prostatectomy, wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23;b. determining or having determined a response to treatment comprising dose- escalated radiation therapy that the subject with prostate cancer who has not undergone radical prostatectomy is expected to have based on at least the expression level of the one target or each of the plurality of targets in the sample in step (a.), optionally wherein the determining or having determined comprises calculating or having calculated a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets and the determining or having determined is based on the risk score; andc. administering or having administered to the subject with prostate cancer who has not undergone radical prostatectomy a treatment selected from:i. a treatment comprising dose-escalated radiation therapy; or ii. a treatment comprising non-dose-escalated radiation therapy, wherein the treatment is selected based on the expected response to treatment comprising dose-escalated radiation therapy determined in (b.).

4. A method comprising:a. obtaining or having obtained an expression level of one target or a plurality of targets in a sample obtained from a subject with prostate cancer who has not undergone radical prostatectomy, wherein the one target or plurality of targets comprises or consists of at least one target nucleic acid sequence selected from the group of genes consisting of: KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23;b. calculating or having calculated a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets of (a.); andc. administering or having administered to the subject with prostate cancer who has not undergone radical prostatectomy in a treatment selected from:i. a treatment comprising dose-escalated radiation therapy; or ii. a treatment comprising non-dose-escalated radiation therapy, wherein the treatment is selected based on the risk score of (b.).

5. The method of any one of claim 1-3, wherein the response to treatment comprising dose-escalated radiation therapy comprises risk of one or more of: overall survival, distant metastasis, metastasis progression-free survival, metastasi -free survival, biochemical recurrence-free survival, failure-free survival, and prostate cancer specific mortality, optionally biochemical failure (BF), distant metastasis (DM) and receipt of salvage therapy, optionally wherein the risk is at 3, 5, 10 or 15 years post-treatment with the radiation therapy, optionally wherein the risk is at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 years, or a range defined by any two of the preceding values, post-treatment with the radiation therapy.

6. The method of any one of claims 3-5, wherein the treatment (i.) comprising dose-escalated radiation therapy further comprises administration of an additional treatment for prostate cancer, and / or wherein the treatment (ii.) comprising non-dose-escalated radiation therapy further comprises an additional treatment for prostate cancer.

7. The method of any one of claims 1-6, wherein the one target or plurality of targets comprises or consists of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23 or 24 target nucleic acid sequences, optionally wherein each target nucleic acid sequence is selected from a different gene selected from the group of genes consisting of: KRT14, EMEI, DRAM1, ANLN, APEX2, DLL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23.

8. The method of any one of claims 1-7, wherein the one target or plurality of targets comprises or consists of a nucleic acid sequence of each of:a. KRT14;b. KRT14, and EME1;c. KRT14, EME1, and DRAM1;d. KRT14, EME1, DRAM1, and ANLN;e. KRT14, EME1, DRAM1, ANLN, and APEX2;f. KRT14, EME1, DRAM1, ANLN, APEX2, and DTL;g. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, and ARHGAP15;h. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, and GNG11; i. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, and CDKN2AIP;j. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, and PTPN22;k. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, and RPS27A;l. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, and CDKN3;m. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, and IL1B;n. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, and NEK1;o. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, and UBA7;p. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, and MUM1; q. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, and BIN2;r. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, and TOP2A;s. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, and PLK2;t. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, and ZMAT3;u. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, and SULF2;v. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, and IL7R;w. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, and HCLS1; orx. KRT14, EME1, DRAM1, ANLN, APEX2, DTL, ARHGAP15, GNG11, CDKN2AIP, PTPN22, RPS27A, CDKN3, IL1B, NEK1, UBA7, MUM1, BIN2, TOP2A, PLK2, ZMAT3, SULF2, IL7R, HCLS1, and KIF23.

9. The method of any one of claims 1-3 and 5-8, wherein said determining or having determined comprises calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, optionally wherein said determining or having determined is based on the risk score.

10. The method of any one of the preceding claims, wherein said determining or having determined comprises determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy.

11. The method of any one of claims 1-9, wherein said determining or having determined comprises determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy.

12. The method of any one of claims 1 and 3-11, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and wherein said determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy:a. is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or greater than, 0, - 0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, - 0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, -0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.42 on a scale from -1 to 1; orb. is expected to respond no better or worse to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from - 1 to 1, the risk score being equal to or less than 0, -0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, -0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, -0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.41 on a scale from -1 to 1.

13. The method of claim 12, further comprising:a. selecting for administration and / or administering, or having administered, to the subject expected to respond better to treatment comprising dose-escalated radiation therapy a treatment comprising dose-escalated radiation therapy; or b. selecting for administration and / or administering, or having administered, to the subject expected to respond no better or worse to treatment comprising dose-escalated radiation therapy a treatment comprising non-dose-escalated radiation therapy.

14. The method of any one of claims 1-12, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and wherein the method comprises:a. selecting for administration and / or administering, or having administered, to the subject a treatment comprising dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; and ii. on a scale from -1 to 1, the risk score being equal to or greater than, 0, - 0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, - 0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, - 0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.42 on a scale from -1 to 1; orb. selecting for administration and / or administering, or having administered, to the subject a treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or less than 0, -0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, -0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, -0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.41 on a scale from -1 to 1.

15. The method of any one of claims 1-9, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality oftargets, and wherein said determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy:a. is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or greater than, - 0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, - 0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, - 0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.67 on a scale from -1 to 1; orb. is expected to respond no better or worse to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or less than, -0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.68 on a scale from -1 to 1.

16. The method of claim 15, further comprising:a. selecting for administration and / or administering, or having administered, to the subject expected to respond better to treatment comprising dose-escalated radiation therapy a treatment comprising dose-escalated radiation therapy; orb. selecting for administration and / or administering, or having administered, to the subject expected to respond no better or worse to treatment comprising dose- escalated radiation therapy a treatment comprising non-dose-escalated radiation therapy.

17. The method of any one of claims 1-9 or 15, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and wherein the method comprises:a. selecting for administration and / or administering, or having administered, to the subject a treatment comprising dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or greater than, - 0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, - 0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, - 0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.67 on a scale from -1 to 1; orb. selecting for administration and / or administering, or having administered, to the subject a treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or less than, -0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.68 on a scale from -1 to 1.

18. The method of any one of claims 1-9, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and wherein said determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy:a. is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; and ii. on a scale from -1 to 1, the risk score being equal to or greater than, 0, - 0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, - 0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, -0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.42 on a scale from -1 to 1; orb. is expected to respond no better or worse to treatment comprising dose- escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or less than, -0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75, -0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.68 on a scale from -1 to 1.

19. The method of claim 18, further comprising:-no-a. selecting for administration and / or administering, or having administered, to the subject expected to respond better to treatment comprising dose-escalated radiation therapy a treatment comprising dose-escalated radiation therapy; or b. selecting for administration and / or administering, or having administered, to the subject expected to respond no better or worse to treatment comprising dose- escalated radiation therapy a treatment comprising non-dose-escalated radiation therapy.

20. The method of any one of claims 1-9 or 18, comprising calculating a risk score for the sample based on at least the expression level of the one target or each of the plurality of targets, and wherein the method comprises:a. selecting for administration and / or administering, or having administered, to the subject a treatment comprising dose-escalated radiation therapy based on either or both of:i. the risk score for the sample being greater than the lower two thirds of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; and ii. on a scale from -1 to 1, the risk score being equal to or greater than, 0, - 0.05, -0.10, -0.15, -0.20, -0.25, -0.30, -0.31, -0.32, -0.33, -0.34, -0.35, - 0.36, -0.37, -0.38, -0.39, -0.40, -0.41, -0.42, -0.43, -0.44, -0.45, -0.46, - 0.47, -0.48, -0.49, -0.50, -0.51, or -0.52, or in a range defined by any two of the preceding values, optionally the risk score being equal to or greater than -0.42 on a scale from -1 to 1; orb. selecting for administration and / or administering, or having administered, to the subject a treatment comprising non-dose-escalated radiation therapy based on either or both of:i. the risk score being within the lower third of the risk scores observed in a population of patients with prostate cancer who have not undergone radical prostatectomy; andii. on a scale from -1 to 1, the risk score being equal to or less than, -0.53, -0.54, -0.55, -0.56, -0.57, -0.58, -0.59, -0.60, -0.61, -0.62, -0.63, -0.64, -0.65, -0.66, -0.67, -0.68, -0.69, -0.70, -0.71, -0.72, -0.73, -0.74, -0.75,-0.76, or -0.77, or in a range defined by any two of the preceding values, optionally the risk score being equal to or less than -0.68 on a scale from -1 to 1.

21. The method of any one of the preceding claims, comprising determining or having determined that the subject with prostate cancer who has not undergone radical prostatectomy:a. is expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on at least the risk score being greater than 0 on a scale from - 1 to 1; or b. is expected to respond no better or worse to treatment comprising dose- escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy based on at least the risk score being less than or equal to 0 on a scale from -1 to 1,22. The method of any one of the preceding claims, wherein the method comprises a. selecting for administration and / or administering, or having administered, to the subject a treatment comprising dose-escalated radiation therapy based on at least the risk score being greater than 0 on a scale from -1 to 1; or b. selecting for administration and / or administering, or having administered, to the subject a treatment comprising non-dose-escalated radiation therapy based at least on the risk score being equal to or less than 0 on a scale from -1 to 1.

23. The method of any one of claims 10-22, wherein the population of patients with prostate cancer who have not undergone radical prostatectomy comprises at least 10, 25, 50, 75 or 100 patients, optionally wherein the population of patients does not have metastatic prostate cancer, and / or the population of patients has intermediate-risk prostate cancer, optionally wherein the population patients have: histologically confirmed adenocarcinoma of the prostate, clinical stage Tla-T2b (stage 1 or II), Gleason score 2-6 and prostate-specific antigen (PSA) 10 to less than 20 ng / mL, or Gleason score 7 and PSA less than 15 ng / mL, and no distant metastases and no regional lymph node involvement.

24. The method of any one of claims 1-23, wherein the subject is determined to be expected to respond better to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy, optionally wherein the subject isselected for administration and / or is administered a treatment comprising dose-escalated radiation therapy.

25. The method of any one of claims 1-23, wherein the subject is determined to be expected to respond no better or worse to treatment comprising dose-escalated radiation therapy than to treatment comprising non-dose-escalated radiation therapy, optionally wherein the subject is selected for administration and / or is administered a treatment comprising non- dose-escalated radiation therapy.

26. The method of any one of claims 1-25, wherein the expected beter response to treatment comprising dose-escalated radiation therapy is one or more of:a. a reduction in risk of BF of at least 5%, 10%, 20%, or 30% as compared to treatment comprising non-dose-escalated radiation therapy;b. a reduction in risk of DM of at least 5%, 10%, 20%, or 30% as compared to treatment comprising non-dose-escalated radiation therapy; and c. a reduction in risk of receipt of salvage therapy of at least 5%, 10%, 20%, or 30% as compared to treatment comprising non-dose-escalated radiation therapy;optionally wherein the risk is at 3, 5, 10 or 15 years post-treatment with the radiation therapy, optionally wherein the risk is at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 years, or a range defined by any two of the preceding values, post- treatment with the radiation therapy.

27. The method of any one of claims 1 -26, wherein the expected no better or worse response to treatment comprising dose-escalated radiation therapy is one or more of:a. reduction in risk of BF of less than 5%, 0%, -5%, or -10% as compared to treatment comprising non-dose-escalated radiation therapy;b. reduction in risk of DM of less than 5%, 0%, -5%, or -10% as compared to treatment comprising non-dose-escalated radiation therapy; and c. reduction in risk of receipt of salvage therapy of less than 5%, 0%, -5%, or - 10% as compared to treatment comprising non-dose-escalated radiation therapy;optionally wherein the risk is at 3, 5, 10 or 15 years post-treatment with the radiation therapy, optionally wherein the risk is at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 years,or a range defined by any two of the preceding values, post-treatment with the radiation therapy.

28. The method of any one of claims 5-27, wherein the BF is Pheonix BF.

29. The method of any one of the preceding claims, wherein the subject does not have metastatic prostate cancer, and / or the subject has intermediate-risk prostate cancer, optionally wherein the subject has a PSA score of < 20 ng / ml, a clinical stage of T1 or T2, and / or a Gleason grade group of 1, 2 or 3.

30. The method of any one of the preceding claims, wherein the subject satisfies the following criteria:a. histologically confirmed adenocarcinoma of the prostate,b. clinical stage Tla-T2b (stage I or II),c. Gleason score 2-6 and prostate-specific antigen (PSA) 10 to less than 20 ng / mL, or Gleason score 7 and PSA less than 15 ng / mL,d. no distant metastases, ande. no regional lymph node involvement.

31. The method of any one of the preceding claims, wherein the subject has one or more of the following:a. intermediate risk prostate cancer as defined by the NCCN risk groups; b. low risk prostate cancer as defined by the NCCN risk groups;c. high or very high risk prostate cancer as defined by the NCCN risk groups; d. one or more of the following risk factors: clinical stage T2b- T2c, Grade Group 2 or 3 (Gleason 7), PSA 10-20;e. clinical stage cT1-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL; f. at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL; and / org. metastatic prostate cancer.

32. The method of any one of the preceding claims, wherein the subject does not have one or more of the following:a. intermediate risk prostate cancer as defined by the NCCN risk groups; b. low risk prostate cancer as defined by the NCCN risk groups;c. high or very high risk prostate cancer as defined by the NCCN risk groups;d. one or more of the following risk factors: clinical stage T2b- T2c, Grade Group 2 or 3 (Gleason 7), PSA 10-20;e. clinical stage cT1-cT2a and Grade Group 1 (Gleason 6) and PSA <10 ng / mL; f. at least two of: cT3-cT4, GG4-5, PSA>20 ng / mL; and / org. metastatic prostate cancer.

33. The method of any one of the preceding claims, wherein said determining or having determined the expected response to treatment comprising dose-escalated radiation therapy is in comparison to a treatment comprising non-dose-escalated radiation therapy.

34. The method of any one of the preceding claims, wherein said determining or having determined the expected response of the subject with prostate cancer who has not undergone radical prostatectomy to treatment comprising dose-escalated radiation therapy is not based on prognostic clinical variables, optionally wherein the prognostic clinical variables are tumor stage, nodal stage, metastatic tumor volume categorization, a Gleason score and / or a serum PSA level of the subject.

35. The method of any one of claims 1-33, wherein said determining or having determined the expected response of the subject with prostate cancer who has not undergone radical prostatectomy to treatment comprising dose-escalated radiation therapy is further based on one or more prognostic clinical variables, optionally wherein the prognostic clinical variables are tumor stage, nodal stage, metastatic tumor volume categorization, a Gleason score and / or a serum PSA level of the subject.

36. The method of any one of the preceding claims, wherein said obtaining or having obtained the expression level of the one target or the plurality of targets comprises utilizing a microarray to assess one target or the plurality of targets, performing qPCR on the one target or the plurality of targets, and / or sequencing the one target or the plurality of targets.

37. The method of any one of the preceding claims, wherein the sample is a resected specimen, optionally a transurethral resection of prostate (TORI’) specimen, or a biopsy of the prostate cancer, optionally a needle biopsy.

38. The method of any one of the preceding claims, wherein the sample is from a primary prostate tumor.

39. The method of any one of the preceding claims, wherein the dose-escalated radiation therapy is at least 78 Gy, optionally at least 79 Gy or 80 Gy.

40. The method of any one of the preceding claims, wherein the non-dose-escalated radiation therapy is not more than 71 Gy, optionally not more than 70 Gy.

41. The method of any one of the preceding claims, wherein is selected from 3- Dimensional Conformal Radiation Therapy (3D-CRT), Intensity-Modulated Radiation Therapy (IMRT), volumetric modulated arc therapy (VMAT), stereotactic body radiotherapy (SBRT or SABR), proton beam RT, and brachytherapy boost, optionally wherein it is 3D-CRT or IMRT.

42. The method of any one of the preceding claims wherein the dose-escalated and / or non-dose-escalated radiation therapy is definitive radiation therapy.

43. The method of any one of the preceding claims, wherein the dose-escalated and / or non-dose-escalated radiation therapy is not salvage radiation therapy.

44. The method of any one of the preceding claims, wherein the risk factor score is a PORTOS score.

45. The method of any one of the preceding claims, wherein the subject does not have NCCN unfavorable intermediate-risk prostate cancer.

46. A kit for use in the method of any one of the preceding claims, the kit comprising reagents for obtaining the expression level of the one target or each of the plurality of targets.

47. The kit of claim 46, wherein the reagents comprise primer(s) and / or probe(s) for obtaining the expression level of the one target or each of the plurality of targets.

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