PARP inhibitors for the treatment of advanced prostate cancer

Abstract

The present disclosure provides methods for detecting homozygous deletions in BRCA1 and / or BRCA2, in individuals with APC who may benefit from treatment comprising the administration of a drug effective in treating an HRD positive cancer, as well as methods, kits and reagents for detecting homozygous deletions in nucleic acid molecules and polypeptides. The disclosure also provides methods for evaluating, identifying, assessing, and / or treating an individual having a cancer.

Description

PARP INHIBITORS FOR THE TREATMENT OF ADVANCED PROSTATECANCERCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority benefit to U.S. Provisional Application No. 63 / 631,861, filed April 9, 2024, and U.S. Provisional Application No. 63 / 681,009 filed August 8, 2024, the entire contents of which are each incorporated herein by reference for all purposes.FIELD

[0002] The present disclosure methods of characterizing cancer, in particular metastatic prostate cancer for treatment comprising the administration of a drug effective at treating HRD positive cancers by detecting BRCA1 and / or BRCA2 homozygous deletions in the tumor.BACKGROUND

[0003] Cancer represents the phenotypic endpoint of multiple genetic lesions that endow cells with a full range of biological properties required for tumorigenesis. A hallmark genomic feature of many cancers is the presence of numerous complex chromosome structural aberrations, including translocations, intra-chromosomal inversions, point mutations, deletions, gene copy number changes, gene expression level changes, gene fusions, and germline mutations, among others.

[0004] Prostate cancer is a common solid tumor cancer among men. Advanced prostate cancers (APC) are advanced and potentially metastatic prostate cancers and include metastatic castration resistant prostate cancer (mCRPC). Genetic alterations in genes involved in DNA homologous repair (HRR) pathways, such as BRCA1 and BRCA2 are common in APC. Understanding the types of genetic alterations in HRR genes as well as the connection between the genetic alterations and cancer treatments will aid in improving treatment outcomes.

[0005] Previous studies suggest that HRR alterations are largely truncal and would be present in an archival prostate tissue biopsy (Zurita et al., Genomic Biomarkers and Genome-Wide Loss-of-Heterozygosity Scores in Metastatic Prostate Cancer Following Progression on Androgen-Targeting Therapies, JCO Precis Oncol, e2200195 (2022); Schweizer et al., Concordance of DNA Repair Gene Mutations in Paired Primary Prostate Cancer Samplesand Metastatic Tissue or Cell-Free DNA, JAMA Oncol 2021;7:1378-82. (2021)) suggesting that it would be valid to use archival specimens to guide decisions for PARPi use in metastatic settings. Further, detection of BRCA alterations were associated with improved overall survival for mCRPC patients compared to control in a post-hoc analysis of a mCRPC study (Tukachinsky et al., Genomic Analysis of Circulating Tumor DNA in 3,334 Patients with Advanced Prostate Cancer Identifies Targetable BRCA Alterations and AR Resistance Mechanisms, Clinical Cancer Research 2021;27:3094-105 (2021), Matsubara et al., Efficacy in Patients with Metastatic Castration-resistant Prostate Cancer and BRCA1, BRCA2, or ATM Alterations Identified by Testing Circulating Tumor, Clinical Cancer Research;29:92-9 (2023).BRIEF SUMMARY

[0006] Described herein are methods that can be used to identify individuals with cancer, e.g., advanced prostate cancer (APC), who may benefit from treatment with a drug effective at treating an HRD positive cancer. The methods comprise identifying BRCA1 and / or BRCA2 homozygous deletions (or loss) in patients with cancer, e.g., APC, and recommending treatment comprising the administration of a drug effective at treating an HRD positive cancer. Provided herein are methods that can be used to identify a BRCA1 and / or BRCA2 homozygous deletion (or loss) from a liquid biopsy taken from a patient with cancer, e.g., APC. Using the methods provided herein a tumor fraction (TF) may be determined. If the TF of the liquid biopsy sample is above about 20% the liquid biopsy sample may be reliable for identifying BRCA1 / BRCA2 homozygous deletions (loss) in patients with cancer who may benefit from treatment with a drug effective at treating an HRD positive cancer. If the TF of the liquid biopsy is equal to or greater than about 20% and a BRCA1 homozygous deletion (or loss) and / or a BRCA2 homozygous deletion (or loss) is not detected, the individual can be consider to not have BRCA1 homozygous deletion (or loss) and / or a BRCA2 homozygous deletion (or loss) and one who may not benefit from treatment with a drug effective at treating HRD positive cancer.

[0007] Also provided herein are methods that can be used to identify individuals with cancer, e.g., APC. who have BRCA1 homozygous deletion (or loss) and / or a BRCA2 homozygous deletion (or loss), who may benefit from treatment with a drug effective at treating an HRD positive cancer. The method comprises determining the TF of a liquid biopsy sample, and if the liquid biopsy sample has a TF of less than about 20% and a BRCA1 homologous deletion(or loss) and a BRCA2 homozygous deletion (or loss) are not detected in the liquid biopsy sample, further detecting the BRCA1 homologous deletion (or loss) and / or a BRCA2 homozygous deletion (or loss) in a tissue biopsy sample from the individual. If a BRCA1 and / or BRCA2 homozygous deletion (or loss) is identified in the tissue biopsy sample, the individual is identified as one who may benefit from treatment with a drug effective at treating an HRD positive cancer.

[0008] Also provided herein are methods that can be used to identify individuals with cancer, e.g., APC, who may benefit from treatment with a drug effective at treating an HRD positive cancer. The method comprises determining the TF of a liquid biopsy sample, and if the liquid biopsy sample has a TF of less than about 20%, and a BRCA1 homologous deletion (or loss) and a BRCA2 homozygous deletion (or loss) are not detected in the liquid biopsy sample, further detecting the BRCA1 homologous deletion (or loss) and / or a BRCA2 homozygous deletion (or loss) in a tissue biopsy sample from the individual. If a BRCA1 and / or BRCA2 homozygous deletion (or loss) is identified in the tissue biopsy sample, the individual is identified as one who may benefit from treatment with a drug effective at treating an HRD positive cancer.

[0009] Disclosed herein are method of identifying an individual having advanced prostate cancer (APC) that may benefit from a treatment of a drug effective in treating an HRD positive cancer to the individual, said method comprising detecting a BRCA1 homozygous deletion in a sample from the individual, wherein the presence of a one BRCA1 homozygous deletion in the sample identifies the individual as one who may benefit from the treatment.

[0010] Also provided herein, are methods of selecting treatment for an individual having advanced prostate cancer (APC), comprising acquiring knowledge of a BRCA1 homozygous deletion in a sample from an individual having APC, wherein responsive to the acquisition of said knowledge: (i) the individual is classified as a candidate to receive treatment with a drug effective in treating an HRD positive cancer and / or (ii) the individual is identified as likely to respond to a treatment with the drug effective in treating an HRD positive cancer.

[0011] Also provided herein, are methods of treating or delaying progression of advanced prostate cancer (APC) comprising, responsive to knowledge of a BRCA1 homozygous deletion in a sample from an individual, administering to the individual an effective amount of a drug effective in treating an HRD positive cancer.

[0012] Also provided herein, are methods of identifying one or more treatment options for an individual having advanced prostate cancer (APC), the methods comprising: (a) detecting a BRCA1 homozygous deletion in a sample from the individual; and (b) generating a reportcomprising one or more treatment options identified for the individual based at least in part on the presence of the BRCA1 homozygous deletion in the sample, wherein the one or more treatment options comprise a drug effective in treating an HRD positive cancer.

[0013] Also provided herein, are methods of predicting survival of an individual having advanced prostate cancer (APC) treated with a drug effective in treating an HRD positive cancer, comprising acquiring knowledge of a BRCA1 homozygous deletion in a sample from the individual, wherein responsive to the acquisition of said knowledge, the individual is predicted to have longer survival after treatment with the drug effective in treating an HRD positive cancer, as compared to an individual whose APC does not exhibit the BRCA1 homozygous deletion.

[0014] Also provided herein, are methods of diagnosing / assessing a BRCA1 homozygous deletion, the methods comprising: (a) detecting in a sample from an individual with advanced prostate cancer (APC); and (b) providing a diagnosis of the BRCA1 homozygous deletion.

[0015] In some instances, the methods further comprise recommending treatment options including administration of a HRR pathway drug.

[0016] In some instances, the BRCA1 homozygous deletion comprises a homozygous deletion of the full BRCA1 gene.

[0017] In some instances, the sample comprises a tissue biopsy sample or a liquid biopsy. In some instances, the sample is a tissue biopsy and compromises a tumor biopsy. In some instances, the sample is a liquid biopsy sample and comprises blood, serum, plasma, cerebrospinal fluid, sputum, stool, urine, or saliva.

[0018] In some instances, the sample comprises cells and / or nucleic acids from the APC. In some instances, the sample comprises mRNA, DNA, circulating tumor DNA (ctDNA), cell- free DNA, cell-free RNA from the cancer, or any combination thereof. In some instances, the sample is a liquid biopsy sample and comprises circulating tumor cells (CTCs). In some instances, the sample is a liquid biopsy sample and comprises cell-free DNA (cfDNA), circulating tumor DNA (ctDNA), or any combination thereof.

[0019] In some instances, acquiring knowledge of the BRCA1 homozygous deletion comprises detecting the BRCA1 homozygous deletion.

[0020] In some instances, the BRCA1 homozygous deletion is detected in the sample by one or more of: a nucleic acid hybridization assay, an amplification-based assay, a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay, real-time PCR, a screening analysis, fluorescence in situ hybridization (FISH), spectral karyotyping, multicolor FISH (mFISH), comparative genomic hybridization, in situ hybridization,sequence-specific priming (SSP) PCR, high-performance liquid chromatography (HPLC), mass-spectrometric genotyping, or sequencing.

[0021] In some instances, the advanced prostate cancer (APC) is metastatic castration resistant prostate cancer (mPRPC). In some instances, the advanced prostate cancer (APC) is hormone sensitive prostate cancer. In some instances, the advanced prostate cancer (APC) is de novo metastatic prostate cancer.

[0022] Also provided herein are methods of identifying if an individual having advanced prostate cancer (APC) that may benefit from a treatment with a drug effective in treating an HRD positive cancer, said methods comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to about 20% or less than about 20% and a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion is detected in the liquid biopsy sample, identifying the individual as one who may benefit from the treatment of a drug effective in treating an HRD positive cancer.

[0023] Also provided herein are methods of identifying if an individual having advanced prostate cancer (APC) that may benefit from a treatment with a drug effective in treating an HRD positive cancer, said methods comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to about 20% and a BRCA1 homozygous deletion and a BRCA2 homozygous deletion is not detected in the liquid biopsy sample, identifying the individual as one who may not receive the most benefit from the treatment with a drug effective in treating an HRD positive cancer.

[0024] Also provided herein are methods of selecting treatment for an individual having advanced prostate cancer (APC), said methods comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to 20% or less than about 20% and a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion is detected in the liquid biopsy sample from the individual, classifying the individual as a candidate to receive treatment with a drug effective in treating an HRD positive cancer.

[0025] Also provided herein, are methods of selecting treatment for an individual having advanced prostate cancer (APC), said methods comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater thanor equal to 20% and BRCA1 homozygous deletion and BRCA2 homozygous deletion is not detected in the liquid biopsy sample, classifying the individual as one who may not receive the most benefit from treatment with a drug effective in treating an HRD positive cancer.

[0026] Also provided herein are methods of treating or delaying progression of advanced prostate cancer (APC) in an individual said methods comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to 20% or less than about 20%, and a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion is detected in the liquid biopsy sample from the individual, administering to the individual an effective amount of a drug effective in treating an HRD positive cancer.

[0027] Also provided herein are methods of treating or delaying progression of advanced prostate cancer (APC) in an individual said methods comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to 20% and a BRCA1 homozygous deletion and a BRCA2 homozygous deletion is not detected in the liquid biopsy sample from the individual, administering to the individual an effective amount of a drug other than a drug effective in treating an HRD positive cancer.

[0028] Also provided herein are methods of predicting survival of an individual having advanced prostate cancer (APC) treated with a drug effective in treating an HRD positive cancer, said methods comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to about 20% or less than about 20%, and a BRCA1 homologous deletion and / or a BRCA2 homozygous deletion is detected in the liquid biopsy sample from the individual, the individual is predicted to have longer survival after treatment with a drug effective in treating an HRD positive cancer, as compared to an individual whose APC does not exhibit the BRCA1 homozygous deletion and / or BRCA2 homozygous deletion.

[0029] Also provided herein are methods of predicting survival of an individual having advanced prostate cancer (APC) treated with administration of a drug effective in treating an HRD positive cancer, said methods comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than orequal to about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual, the individual is predicted to have shorter survival after treatment with the drug effective in treating an HRD positive cancer, as compared to an individual whose APC does exhibit the BRCA1 homozygous deletion and / or BRCA2 homozygous deletion.

[0030] Also provided herein are methods of identifying if an individual having cancer that may benefit from a treatment with a drug effective in treating an HRD positive cancer, said methods comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; if the TF in the liquid biopsy sample is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detecting in the liquid biopsy sample from the individual, further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if BRCA1 homozygous deletion and / or BRCA2 homozygous deletion are detected in the tissue biopsy sample from the individual, identifying the individual as one who may benefit from the treatment with a drug effective in treating an HRD positive cancer.

[0031] Also provided herein are methods of identifying if an individual having cancer that may benefit from a treatment with a drug effective in treating an HRD positive cancer, said methods comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample from the individual, if the TF in the liquid biopsy sample is less than about 20% and BRCA1 homozygous deletion and BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual; further detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if a BRCA1 homozygous deletion and a BRCA2 homozygous deletion are not detected in the tissue biopsy sample, identifying the individual as one who may not benefit from the treatment with a drug effective in treating an HRD positive cancer; or if a BRCA1 homozygous deletion or a BRCA2 homozygous deletion is detected in the tissue biopsy sample, identifying the individual as one who may benefit from the treatment with a drug effective in treating an HRD positive cancer.

[0032] Also provided herein are methods of selecting treatment for an individual having cancer, said methods comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual; further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if BRCA1 homozygous deletion and / or BRCA2 homozygous deletion are detected in the tissue biopsy sample from the individual, classifying the individual as a candidate to receive treatment with a drug effective in treating an HRD positive cancer.

[0033] Also provided herein are methods of selecting treatment for an individual having cancer, said methods comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual; further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if a BRCA1 homozygous deletion and a BRCA2 homozygous deletion are not detected in the tissue biopsy sample, classifying the individual as not a candidate to receive treatment with a drug effective in treating an HRD positive cancer, or if a BRCA1 homozygous deletion or a BRCA2 homozygous deletion is detected in the tissue biopsy sample, classifying the individual as a candidate to receive treatment with a drug effective in treating an HRD positive cancer.

[0034] Also provided herein are methods of treating or delaying progression of advanced cancer in an individual said methods comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual; further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual, and if a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion are detected in the tissue biopsy sample from the individual, administering to the individual an effective amount of a drug effective in treating an HRD positive cancer.

[0035] Also provided herein are methods of treating or delaying progression of cancer in an individual said methods comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are notdetected in the liquid biopsy sample from the individual, further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if a BRCA1 homozygous deletion and a BRCA2 homozygous deletion are not detected in the tissue biopsy sample, administering to the individual an effective amount of a drug other than a drug effective in treating an HRD positive cancer, or if a BRCA1 homozygous deletion or a BRCA2 homozygous deletion is detected in the tissue biopsy sample, administering to the individual a drug effective in treating an HRD positive cancer.

[0036] Also provided herein are methods of predicting survival of an individual having cancer treated with administration of a drug effective in treating an HRD positive cancer, said methods comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual, further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion are detected in the tissue biopsy sample the individual is predicted to have longer survival after treatment with the administration of the drug effective in treating an HRD positive cancer, as compared to an individual whose cancer does not exhibit the BRCA1 homozygous deletion and / or BRCA2 homozygous deletion.

[0037] Also provided herein are methods of predicting survival of an individual having cancer treated with administration of a drug effective in treating an HRD positive cancer, said methods comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual, further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if a BRCA1 homozygous deletion and a BRCA2 homozygous deletion are not detected in the tissue biopsy sample the individual is not predicted to have longer survival after treatment with the administration of the drug effective in treating an HRD positive cancer, as compared to an individual whose cancer does not exhibit the BRCA1 homozygous deletion and / or BRCA2 homozygous deletion, or if a BRCA1 homozygous deletion or a BRCA2 homozygous deletion is detected in the tissue biopsy sample, the individual is predicted to have longersurvival after treatment with the administration of the drug effective in treating an HRD positive cancer, as compared to an individual whose cancer does not exhibit the BRCA1 homozygous deletion and / or BRCA2 homozygous deletion.

[0038] In some instances, the liquid biopsy sample comprises blood, serum, plasma, cerebrospinal fluid, sputum, stool, urine, or saliva. In some instances, the tissue sample is a tissue biopsy and comprises a tumor biopsy. In some instances, the liquid biopsy sample comprises mRNA, DNA, circulating tumor DNA (ctDNA), cell-free DNA, cell-free RNA from the cancer, or any combination thereof. In some instances, the liquid biopsy sample comprises circulating tumor cells (CTCs). In some instances, the sample comprises cell-free DNA (cfDNA), circulating tumor DNA (ctDNA), or any combination thereof.

[0039] In some instances, the BRCA1 homozygous deletion and / or the or BRCA2 homozygous deletion is detected in the liquid biopsy sample by one or more of: a nucleic acid hybridization assay, an amplification-based assay, a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay, real-time PCR, a screening analysis, fluorescence in situ hybridization (FISH), spectral karyotyping, multicolor FISH (mFISH), comparative genomic hybridization, in situ hybridization, sequence-specific priming (SSP) PCR, high-performance liquid chromatography (HPLC), mass-spectrometric genotyping, or sequencing.

[0040] In some instances, the BRCA1 homozygous deletion and / or the or BRCA2 homozygous deletion is detected in the tissue biopsy sample by one or more of: a nucleic acid hybridization assay, an amplification-based assay, a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay, real-time PCR, a screening analysis, fluorescence in situ hybridization (FISH), spectral karyotyping, multicolor FISH (mFISH), comparative genomic hybridization, in situ hybridization, sequence-specific priming (SSP) PCR, high-performance liquid chromatography (HPLC), mass-spectrometric genotyping, or sequencing.

[0041] In some instances, the cancer is advanced prostate cancer (APC). In some instances, the advanced prostate cancer (APC) is metastatic castration resistant prostate cancer (mPRPC). In some instances, the advanced prostate cancer (APC) is hormone sensitive prostate cancer. In some instances, the advanced prostate cancer (APC) is de novo metastatic prostate cancer.

[0042] In some instances, the drug effective in treating an HRD positive cancer is a Poly ADP-ribose polymerase inhibitor (PARPi). In some instances, the PARPi is selected from a group consisting of niraparib, olaparib, talazoparib, or rucaparib. In some instances, the drugeffective in treating an HRD positive cancer is a platinum agent. In some instances, the drug effective in treating an HRD positive cancer is a drug that targets double strand break repair.

[0043] In some instances, the treatment comprises administration of a hormonal therapy. In some instances, the hormonal therapy is an androgen receptor blocker. In some instances, the hormonal therapy comprises one or more of abiraterone, enzalutamide, apalutamide, and darolutamide, or any combination thereof.

[0044] In some instances, the method of any of claims 1-54, wherein the individual has received a prior anti-cancer treatment or is being treated with an anti-cancer treatment. In some instances,, the prior anti-cancer treatment comprises one or more of a small molecule inhibitor, a chemotherapeutic agent, a cancer immunotherapy, an antibody, a cellular therapy, a nucleic acid, a surgery, a radiotherapy, an anti- angiogenic therapy, an anti-DNA repair therapy, an anti-inflammatory therapy, an anti-neoplastic agent, a growth inhibitory agent, a cytotoxic agent, a vaccine, a small molecule agonist, a virus-based therapy, an antibody-drug conjugate, a recombinant protein, a fusion protein, a natural compound, a peptide, a PROteolysis-Targeting Chimera (PROTAC), or any combination thereof. In some instances, the individual has not received a prior anti-cancer treatment.

[0045] In some instances, survival is an overall survival, a progression-free survival, a disease-free survival, an objective response rate, a time to tumor progression, a time to treatment failure, a durable complete response, a time to death, or a time to next treatment. In some instances, the objective response rate is a change in prostate specific antigen (PSA) from baseline.

[0046] In some instances, the methods further comprise obtaining the sample from the individual. In some instances, the methods comprise obtaining the liquid biopsy sample from the individual. In some instances, the methods comprise obtaining the tissue biopsy sample from the individual.

[0047] In some instances, the sequencing comprises a massively parallel sequencing (MPS) technique, whole genome sequencing (WGS), whole exome sequencing, targeted sequencing, direct sequencing, or a Sanger sequencing technique; an optionally wherein the massively parallel sequencing (MPS) technique comprises next- generation sequencing. In some instances, wherein the sequencing comprises: (a) providing a plurality of nucleic acid molecules obtained from the sample, wherein the plurality of nucleic acid molecules comprises a mixture of tumor nucleic acid molecules and non-tumor nucleic acid molecules; (b) optionally, ligating one or more adapters onto one or more nucleic acid molecules from the plurality of nucleic acid molecules; (c) amplifying nucleic acid molecules from theplurality of nucleic acid molecules; (d) optionally, capturing nucleic acid molecules from the amplified nucleic acid molecules, wherein the captured nucleic acid molecules are captured from the amplified nucleic acid molecules by hybridization to one or more bait molecules; and (e) sequencing, by a sequencer, the captured nucleic acid molecules to obtain a plurality of sequence reads corresponding to one or more genomic loci within a subgenomic interval in the sample. In some instance, the sequencing comprises: (a) providing a plurality of nucleic acid molecules obtained from the liquid biopsy sample, wherein the plurality of nucleic acid molecules comprises a mixture of tumor nucleic acid molecules and non-tumor nucleic acid molecules; (b) optionally, ligating one or more adapters onto one or more nucleic acid molecules from the plurality of nucleic acid molecules; (c) amplifying nucleic acid molecules from the plurality of nucleic acid molecules; (d) optionally, capturing nucleic acid molecules from the amplified nucleic acid molecules, wherein the captured nucleic acid molecules are captured from the amplified nucleic acid molecules by hybridization to one or more bait molecules; and (e) sequencing, by a sequencer, the captured nucleic acid molecules to obtain a plurality of sequence reads corresponding to one or more genomic loci within a subgenomic interval in the liquid biopsy sample. In some instances, the sequencing comprises: (a) providing a plurality of nucleic acid molecules obtained from the tissue biopsy sample, wherein the plurality of nucleic acid molecules comprises a mixture of tumor nucleic acid molecules and non-tumor nucleic acid molecules; (b) optionally, ligating one or more adapters onto one or more nucleic acid molecules from the plurality of nucleic acid molecules; (c) amplifying nucleic acid molecules from the plurality of nucleic acid molecules; (d) optionally, capturing nucleic acid molecules from the amplified nucleic acid molecules, wherein the captured nucleic acid molecules are captured from the amplified nucleic acid molecules by hybridization to one or more bait molecules; and (e) sequencing, by a sequencer, the captured nucleic acid molecules to obtain a plurality of sequence reads corresponding to one or more genomic loci within a subgenomic interval in the tissue biopsy sample. In some isntances, the adapters comprise one or more of amplification primer sequences, flow cell adapter hybridization sequences, unique molecular identifier sequences, substrate adapter sequences, or sample index sequences. In some instances, amplifying nucleic acid molecules comprises performing a polymerase chain reaction (PCR) technique, a non-PCR amplification technique, or an isothermal amplification technique. In some instances, the one or more bait molecules comprise one or more nucleic acid molecules, each comprising a region that is complementary to a region of a captured nucleic acid molecule. Insome instances, the one or more bait molecules each comprise a capture moiety. In some instances, the capture moiety is biotin.

[0048] In some instances, the individual is a patient. In some instances, the individual is a human.

[0049] Also provided herein are drugs effective in treating an HRD positive cancer for use in a method of treating or delaying progression of advanced prostate cancer (APC), wherein the method comprises administering the drug effective in treating an HRD positive cancer to an individual, wherein a BRCA1 homozygous deletion is detected in a sample obtained from the individual. Also provided herein are drugs effective in treating an HRD positive cancer for use in the manufacture of a medicament for treating or delaying progression of advanced prostate cancer (APC), wherein the medicament is to be administered to an individual, wherein a BRCA1 homozygous deletion has been detected in a sample obtained from the individual. Also provided herein are drugs effective in treating an HRD positive cancer for use in a method of treating or delaying progression of clinically advanced prostate cancer (APC), wherein the method comprises administering the drug effective in treating an HRD positive cancer to an individual, wherein a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion is detected in a liquid biopsy sample obtained from the individual; and wherein the same is a liquid biopsy has a TF of greater than or equal to about 20%. Also provided herein are drugs effective in treating an HRD positive cancer for use in a method of treating or delaying progression of clinically advanced prostate cancer (APC), wherein the method comprises administering the drug effective in treating an HRD positive cancer to an individual, wherein a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion is detected in a tissue biopsy sample obtained from the individual; wherein a liquid biopsy sample has been collected from the individual and the liquid biopsy sample has a TF of less than about 20%. Also provided herein are drugs effective in treating an HRD positive cancer for use in the manufacture of a medicament for treating or delaying progression of advanced prostate cancer (APC), wherein the medicament is to be administered to an individual, wherein a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion has been detected in a sample obtained from the individual; and wherein the same is a liquid biopsy has a TF of greater than or equal to about 20%. Also provided herein are drugs effective in treating an HRD positive cancer for use in the manufacture of a medicament for treating or delaying progression of advanced prostate cancer (APC), wherein the medicament is to be administered to an individual, wherein a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion is detected in a tissue biopsy sample obtained from the individual;wherein a liquid biopsy sample has been collected from the individual and the liquid biopsy sample has a TF of less than about 20%.BRIEF DESCRIPTION OF THE DRAWINGS

[0050] Various aspects of the disclosed methods, devices, and systems are set forth with particularity in the appended claims. A better understanding of the features and advantages of the disclosed methods, devices, and systems will be obtained by reference to the following detailed description of illustrative embodiments and the accompanying drawings, of which:

[0051] FIG. 1 provides a non-limiting example of a cohort selection diagram and treatment and outcome overview diagram.

[0052] FIGS. 2A-2G provides a non-limiting example of Swimmer’s plots of time to next treatment (TTNT) per patient receiving single-agent PARPi and genomic profiling. FIG.2A shows TNTT for samples with BRCA1 / 2 mutations detected in tissue biopsy. FIG. 2B shows TNTT for samples with ATM or other homologous recombination repair (HRR) gene alterations detected in tissue biopsy. FIG. 2C shows TNTT for samples with no HRR alterations detected in tissue biopsy. FIG. 2D shows TNTT for samples with BRCA1 / 2 homozygous deletions detected in liquid biopsy. FIG. 2E shows TNTT for samples with ATM detected in liquid biopsy. FIG. 2F shows TNTT for samples with other HRR alterations in liquid biopsy FIG. 2G shows TNTT for samples with no HRR alterations detected in liquid biopsy. As used in FIGS. 2A-2G, ECOG = Eastern Cooperative Oncology Group (ECOG) performance score; LOT = line of therapy in mCRPC setting; PSA = pre- PARPi prostate specific antigen response (PSA) level; sv = short variant mutation; del = deletion; re = rearrangement.

[0053] FIGS. 3A-3D provides a non-limiting example of Multivariable Cox PH models comparing outcomes by HRR mutation status in tissue and liquid biopsy samples. FIG. 3A shows Multivariable Cox PH models comparing TNTT for samples with and without HRR mutations in tissue biopsy samples, all results are adjusted for baseline prognostic factors. FIG. 3B shows Multivariable Cox PH models comparing real-world overall survival (rwOS)for samples with and without HRR mutations in tissue biopsy samples, all results are adjusted for baseline prognostic factors. FIG. 3C shows Multivariable Cox PH models comparing TNTT for samples with and without HRR mutations, all results adjusted for baseline prognostic factors. FIG. 3D shows Multivariable COX PH models comparing rwOS for samples with and without HRR mutations, all results adjusted for baseline prognosticfactors. As used in FIGS. 3A-3D, ECOG = ECOG performance score; PSA = pre-PARPi PSA level; NHT = novel hormonal therapy.

[0054] FIGS. 4A-4B provides a non-limiting example of Multivariable Cox PH models. FIG. 6A shows Multivariable Cox PH models comparing time to death (TTD) between samples with and without HRR mutations in tissue biopsy samples, adjusted for baseline prognostic factors. FIG. 4B shows Multivariable Cox PH models comparing time to death (TTD) between samples with and without HRR mutations in liquid biopsy samples, adjusted for baseline prognostic factors. As used in FIGS. 4A-4B, ECOG = ECOG performance score; PSA = pre-PARPi PSA level; NHT = novel hormonal therapy.

[0055] FIGS. 5A-5F provides a non-limiting example of unadjusted Kaplan-Meier plots stratified by HRR group status. FIG. 5A shows unadjusted Kaplan-Meier plots stratified by HRR group status for TNTT in tissue biopsy samples (TBx). FIG. 5B shows unadjusted Kaplan-Meier plots stratified by HRR group status for TTD in TBx. FIG. 5C shows unadjusted Kaplan-Meier plots stratified by HRR group status for overall survival (OS) in TBx. FIG. 5D shows unadjusted Kaplan-Meier plots stratified by HRR group status for TNTT in liquid biopsy samples (LBx). FIG. 5E shows unadjusted Kaplan-Meier plots stratified by HRR group status for TTD in LBx. FIG. 5F shows unadjusted Kaplan-Meier plots stratified by HRR group status for OS in LBx.

[0056] FIGS. 6A-6I provides a non-limiting example of outcomes shown by HRDsig status detected in tissue biopsies from patients on PARPi. FIG. 6A shows TTNT segregated by HRDsig status for all patients. FIG. 6B shows TNTT segregated by HRDsig for patients that are positive for BRCA1 / 2 alterations. FIG. 6C shows TNTT segregated by patients that are negative for BRCA1 / 2 alterations FIG. 6D shows TTD segregated by HRDsig status for all patients. FIG. 6E shows TTD segregated by HRDsig for patients that are positive for BRCA1 / 2 alterations. FIG. 6F shows TTD segregated by HRDsig for patients that are negative for BRCA1 / 2 alterations. FIG 6G shows OS segregated by HRDsig status for all patients. FIG. 6H shows OS segregated by HRDsig for patients that are positive for BRCA1 / 2 alterations. FIG. 61 shows OS segregated by HRDsig for patients that are negative for BRCA1 / 2 alterations.

[0057] FIGS. 7A-7B provides a non-limiting example of plots demonstrating the change in prostate specific antigen response (PSA) from baseline to on-therapy. FIG. 7A shows change in PSA for patients with evaluable baseline and on-therapy PSA with alterations identified via tissue biopsy. FIG. 7B shows change in PSA for patients with evaluable baseline and on-therapy PSA with alterations identified via liquid biopsy. As shown in FIG. 7A-7B, dashed horizontal lines represent 30% and 50% PSA declines from baseline.

[0058] FIGS. 8A-8D provides a non-limiting example of plots showing the outcomes associated with BRCA 1 / 2 vs other alterations among the subset of patients with BRCA 1 / 2 alterations. FIG. 8A shows TNTT outcomes associated with BRCA 1 / 2 homozygous deletions / loss vs. other alterations identified in tissue biopsies. FIG. 8B shows OS outcomes associated with BRCA 1 / 2 homozygous deletions / loss versus other alterations identified in tissue biopsies. FIG. 8C shows the prevalence of BRCA 1 / 2 homozygous deletions / loss among tissue biopsies and liquid biopsies included in the database. FIG. 8D shows the distribution of ctDNA tumor fraction (TF) associated with detection of BRCA alterations in liquid biopsy.

[0059] FIG. 9 provides a non-limiting example of a plot of TDD in patients with BRCA 1 / 2 homozygous deletions / loss vs. other alterations in with tissue biopsies.

[0060] FIGS. 10A- 10C provides a non-limiting example of a plot showing the outcomes associated with BRCA 1 / 2 homozygous deletions / loss vs. other alterations. FIG. 10A shows TNTT associated with BRCA 1 / 2 homozygous deletions / loss versus other alterations. FIG. 10B shows TDD associated with BRCA 1 / 2 homozygous deletions / loss vs. other alterations. FIG. 10C shows OS associated with BRCA 1 / 2 homozygous deletions / loss versus other alterations. As shows in FIGS. 10A- 10D, all results are adjusted for baseline prognostic factors among the subset of patients with tissue biopsy and BRCA 1 / 2 alterations.DETAILED DESCRIPTION

[0061] The effectiveness of treatment regimens for individuals with cancer, e.g., advanced prostate cancer (APC) is associated with mutations in homologous recombination repair (HRR) genes, such as BRCA1 and BRCA2. As described herein, it was discovered that individuals with cancer, e.g., APC, with homozygous deletions in BRCA1 and / or BRCA2 or BRCA 1 / 2 loss may respond more effectively to treatment comprising the administration of a drug effective at treating HRD positive cancers than those with other HRR mutations. As such, it is important for physicians to characterize cancers, e.g., APC according to homozygous deletions in BRCA1 and / or BRCA2. In addition, early characterization of cancers, e.g., APC from liquid biopsies with low tumor fractions will facilitate efficient treatment of patients with APC with drugs likely to be effective.

[0062] Certain types of alterations, such as copy number loss, are computationally more challenging to reliably detect than other types of alterations e.g., mutations, and when assessed with liquid biopsy, some genes can additionally require a higher level of ctDNA in the sample (Husain et al. Tumor Fraction Correlates With Detection of Actionable Variants Across > 23,000 Circulating Tumor DNA Samples. JCO Precis Oncol: e2200261. (2022)) to confidently detect (Herberts et al. Technical and biological constraints on ctDNA-based genotyping. Trends Cancer. 7:995-1009. (2021)). Accordingly, detection of a BRCA1 and / or BRCA2 homozygous deletion (or loss) in liquid biopsy samples with low tumor fraction may confer less reliability in detecting BRCA1 and / or BRCA2 homozygous deletion (or loss). Thus, it was also discovered that it may be important to also analyze a tissue biopsy sample if the tumor fraction in the liquid sample is below a particular threshold and BRCA1 and / or BRCA2 homozygous deletions are clinically relevant to the individuals cancer. The present exemplary embodiments show that a reflexive confirmatory tissue sample is necessary if the tumor fraction is less than about 20%. Determining TF and detecting a BRCA1 and / or BRCA2 homozygous deletion in a tissue sample if the TF of the liquid biopsy is less than 20% may prevent false negative or misclassifications of individuals as candidates to receive a benefit from drugs effective at treating a HRD positive cancer. The methods of determining tumor fraction and proceeding to characterize a BRCA1 and / or BRCA2 homozygous deletion in a tissue sample if the tumor fraction is less than about 20% may be useful for any cancer.

[0063] The methods of the present application provide for the classification of cancers, e.g., APC, as those with homozygous deletions in BRCA1 and / or BRCA2 and that may respond to a drug effective at treating HRD positive cancers. In some aspects, the methods comprise determining a tumor fraction of a liquid biopsy sample and characterization BRCA1 and / or BRCA2 homozygous deletion in APC using the liquid biopsy sample. If the tumor fraction is above about 20%, detection of a homozygous deletion of BRCA1 and / or BRCA2 can be informative for treating the individual with cancer, e.g., APC, with a drug effective at treating an HR positive cancer. If the tumor fraction is less than 20% and a homozygous deletion in BRCA1 and / or BRCA2 is detected in the liquid biopsy sample, the detection of the homozygous deletion in BRCA1 and / or BRCA2 is similarly informative. When the tumor fraction is less than about 20% and no homozygous deletion in BRCA1 and / or BRCA2 is detected, a tissue biopsy sample can be used to detect a BRCA1 and / or BRCA2 homozygous deletion in the individual with cancer, e.g., APC, and characterize the individual as one who may benefit from a drug effective at treating an HRD positive cancer. When a TF in a liquid biopsy is less than about 20%, the methods described herein may be used to identify anindividual with any cancer as one who may benefit from a drug effective at treating an HRD positive cancer.

[0064] Described herein are methods for characterizing cancers, e.g., APC, as being likely responsive to a treatment that includes the administration of a drug effective in treating an HRD-positive cancer (such as a PARP inhibitor (PARPi)), based on a homozygous deletion of or loss of the BRCA1 and / or BRCA2 gene. Also described are methods of selecting a treatment option for an individual having such APC, as well as methods of treating such APC in a subject. In some embodiments, the methods comprise determining a tumor fraction of a liquid biopsy from the individual and detecting the BRCA1 and / or BRCA2 homozygous deletion in either the liquid biopsy sample or in a tissue biopsy sample from the individual then selecting a treatment option accordingly.

[0065] As further described herein, homozygous deletions of BRCA1 and / or BRCA2 are associated with the effectiveness of treatment regimens that include the administration of a drug generally used to treat HRD-positive cancers, such as PARPi. In some embodiments particularly, for treating APC. The disclosure is additionally based, in part, on the discovery identifying a tumor fraction value in which the lack of detection of a BRCA1 and / or BRCA2 homozygous deletion in a liquid biopsy sample that informs pursuing methods for identifying a BRCA1 and / or BRCA2 homozygous deletion in a tissue biopsy sample. In turn, provided herein are uses of the identified biomarker in either the liquid biopsy sample or tissue biopsy sample to increase effectiveness in treating cancer, such as APC, with treatment with a drug effective at treating HRD positive cancer, such as PARPi.Definitions

[0066] Unless otherwise defined, technical terms used herein have the same meaning as commonly understood by one of ordinary skill in the art in the field to which this disclosure belongs.

[0067] As used in this specification and the appended claims, the singular forms “a”, “an” and “the” include plural referents unless the content clearly dictates otherwise. Thus, for example, reference to “a molecule” optionally includes a combination of two or more such molecules, and the like.

[0068] The term “or” is used herein to mean, and is used interchangeably with, the term “and / or”, unless context clearly indicates otherwise.

[0069] The terms “about” or “approximately” as used herein refer to the usual error range for the respective value readily known to the skilled person in this technical field, for example,an acceptable degree of error or deviation for the quantity measured given the nature or precision of the measurements. Reference to “about” or “approximately” a value or parameter herein includes (and describes) embodiments that are directed to that value or parameter per se.

[0070] The term “isolated” in the context of a nucleic acid molecule or a polypeptide refers to a nucleic acid molecule or polypeptide being separated from other nucleic acid molecules or polypeptides that are present in the natural source of the nucleic acid molecule or polypeptide. In some certain embodiments, the isolated nucleic acid molecule or polypeptide is free of or substantially free of other cellular material or culture medium when produced by recombinant techniques, or free of or substantially free of chemical precursors or other chemicals when chemically synthesized.

[0071] An “individual” or “subject” is a mammal. Mammals include, but are not limited to, domesticated animals (e.g., cows, sheep, cats, dogs, and horses), primates (e.g., humans and non-human primates such as monkeys), rabbits, and rodents (e.g., mice and rats). In certain embodiments, the individual or subject is a human. In some embodiments, the individual is human patient, e.g., a human patient having cancer (e.g, an APC tumor) as described herein, and / or a homozygous deletion in BRCA1 or BRCA2.

[0072] An “effective amount” or a “therapeutically effective amount” of an agent, e.g., an anti-cancer agent, or a pharmaceutical formulation, refers to an amount effective, at dosages and for periods of time necessary, to achieve the desired therapeutic or prophylactic result, e.g., in the treatment or management of a cancer, for example, delaying or minimizing one or more symptoms associated with the cancer. In some embodiments, an effective amount or a therapeutically effective amount of an agent refers to an amount of the agent at dosages and for periods of time necessary, alone or in combination with other therapeutic agents, which provides a therapeutic benefit in the treatment or management of a disease such as a cancer. In some embodiments, an effective amount or a therapeutically effective amount of an agent enhances the therapeutic efficacy of another therapeutic agent or another therapeutic modality.

[0073] As used herein, “treatment” (and grammatical variations thereof such as “treat” or “treating”) refers to clinical intervention in an attempt to alter the natural course of the individual being treated, and can be performed during the course of clinical pathology. Desirable effects of treatment include, but are not limited to, preventing occurrence or recurrence of disease, delaying progression of disease, alleviation of symptoms, diminishment of any direct or indirect pathological consequences of the disease, decreasingthe rate of disease progression, amelioration or palliation of the disease state, and remission or improved prognosis. In some embodiments, the terms “treatment,” “treat,” or “treating” include preventing a disease, such as cancer, e.g., before an individual begins to suffer from symptoms of a cancer or from re-growth or recurrence of the cancer. In some embodiments, the terms “treatment,” “treat,” or “treating” include inhibiting or reducing the severity of a disease such as cancer (e.g, an APC tumor).

[0074] “Likely to” or “increased likelihood,” as used herein, refer to an increased probability that an event, item, object, thing or person will occur. Thus, in one example, an individual that is likely to respond to treatment with an anti-cancer therapy, e.g., an anti-cancer therapy provided herein, alone or in combination, has an increased probability of responding to treatment with the anti-cancer therapy alone or in combination, relative to a reference individual or group of individuals. “Unlikely to” refers to a decreased probability that an event, item, object, thing or person will occur relative to a reference individual or group of individuals. Thus, an individual that is unlikely to respond to treatment with an anti-cancer therapy, e.g., an anti-cancer therapy provided herein, alone or in combination, has a decreased probability of responding to treatment with the anti-cancer therapy, alone or in combination, relative to a reference individual or group of individuals.

[0075] “Sample,” as used herein, refers to a biological sample obtained or derived from a source of interest, as described herein.

[0076] “Liquid biopsy” as used herein, refers to sample from an individual with cancer, e.g., APC, and may comprise circulating tumor cells (CTCs), cell free DNA (cfDNA), circulating tumor DNA (ctDNA), or a combination thereof.

[0077] “Homozygous Deletion “ as used herein refers to a total loss of a gene on both chromosomes of two homologous chromosomes. A homozygous deletion as used herein may also be referred to as a homozygous loss.

[0078] When a range of values is provided, it is to be understood that each intervening value between the upper and lower limit of that range, and any other stated or intervening value in that states range, is encompassed within the scope of the present disclosure. Where the stated range includes upper or lower limits, ranges excluding either of those included limits are also included in the present disclosure.

[0079] Some of the analytical methods described herein include mapping sequences to a reference sequence, determining sequence information, and / or analyzing sequence information. It is well understood in the art that complementary sequences can be readilydetermined and / or analyzed, and that the description provided herein encompasses analytical methods performed in reference to a complementary sequence.

[0080] The section headings used herein are for organization purposes only and are not to be construed as limiting the subject matter described. The description is presented to enable one of ordinary skill in the art to make and use the invention and is provided in the context of a patent application and its requirements. Various modifications to the described embodiments will be readily apparent to those persons skilled in the art and the generic principles herein may be applied to other embodiments. Thus, the present invention is not intended to be limited to the embodiment shown but is to be accorded the widest scope consistent with the principles and features described herein.

[0081] The figures illustrate processes according to various embodiments. In the exemplary processes, some blocks are, optionally, combined, the order of some blocks is, optionally, changed, and some blocks are, optionally, omitted. In some examples, additional steps may be performed in combination with the exemplary processes. Accordingly, the operations as illustrated (and described in greater detail below) are exemplary by nature and, as such, should not be viewed as limiting.

[0082] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference in their entirety to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference in its entirety. In the event of a conflict between a term herein and a term in an incorporated reference, the term herein controls.Characterization of APC with homozygous deletion in BRCA1

[0083] Provided herein are methods that can be used to characterize APC tumors with homozygous deletions in BRCA1 as potentially responsive to drugs effective at treating HRD positive cancers. The methods comprise identifying a homozygous deletion of BRCA1 in a sample collected from a subject having APC using the methods described herein. Once the homozygous deletion in BRCA1 has been detected, the APC can be classified as one likely to respond to a drug effective at treating an HRD positive cancer, such as a PARP inhibitor. In some respects, the APC is metastatic castration resistant prostate cancer (mCRPC). The characterization of APC according to the methods described herein can be use in a variety of applications such as, but not limited to, diagnosing a patient with APC, selecting a treatment, treating a patient with APC, and generating reports about the APC for the patient or clinical provider. In some embodiments, the sample is a sample from a subject having APC, such as atissue biopsy sample or a liquid biopsy sample as described herein. In some aspects, the sample is a tissue biopsy sample or liquid biopsy sample comprising cells from a tumor. In some aspects, the sample comprises cells and / or nucleic acids from the APC.

[0084] Provided herein are methods that can be used to detect a homozygous deletion in BRCA1 in a sample from an individual with APC in order to identify the individual as a candidate for treatment comprising the administration of a drug effective at treating an HRD positive cancer. Detecting a homozygous deletion in BRCA1 may be accomplished using any of the methods known in the art to detect a deletion in a gene, such as but not limited to the in situ hybridization, array-based, amplification based, sequencing based, and protein detection based methods described herein. The sample from an individual with APC may be a tumor tissue sample or a liquid biopsy sample as described herein. Once a homozygous deletion in BRCA1 is detected in the sample from an individual with APC, they may be identified as a candidate for treatment comprising a drug effective at treating an HRD positive cancer. In some embodiments, the absence of a BRCA1 homozygous deletion in a sample may indicate the individual may not receive the most benefit from a treatment with a drug effective at treating an HRD positive cancer. In some embodiments, the absence of a BRCA1 homozygous deletion in a sample may indicate the individual may be less likely to benefit from a treatment with a drug effective at treating an HRD positive cancer. For example, the individual may be less likely to benefit from treatment with a drug effective at treating an HRD positive cancer than an individual wherein a BRCA1 homozygous deletion was detected in the same. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent.

[0085] Provided here are methods that can be used to identify an individual having advanced prostate cancer that may benefit from a treatment with a drug effective in treating an HRD positive cancer to the individual may have a benefit from the treatment, comprising detecting a BRCA1 homozygous deletion in a sample from the individual, wherein the presence of the a BRCA1 homozygous deletion in the sample identifies the individual as one who may benefit, or may have a benefit from the treatment.

[0086] Provided here are methods that can be used to identify if an individual having advanced prostate cancer that may receive a benefit from a treatment a drug effective in treating an HRD positive cancer to the individual, comprising detecting a BRCA1 homozygous deletion in a sample from the individual, wherein the presence of a BRCA1 homozygous deletion in the sample identifies the individual as one who may receive a benefitfrom a treatment comprising a drug effective at treating an HRD positive cancer and wherein the absence of a BRCA1 homozygous deletion in the sample identifies the individual as one who may not receive the most benefit from a treatment comprising a drug effective at treating an HRD positive cancer. In some embodiments, a drug effective at treating an HRD positive cancer may be more effective at treating APC when the sample from the individual has a BRCA1 homozygous deletion than when the sample from the individual does not have BRCA1 homozygous deletion.

[0087] Provided herein are methods that can be used to identify individuals who may benefit from treatment with a drug effective at treating an HRD positive cancer using knowledge of a homozygous deletion of BRCA1 in a sample from the individuals. In some aspects, the knowledge may be the result of employing one of the methods for detecting a homozygous deletion of BRCA1 in a sample from an individual described herein. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent. In some embodiments, the methods may for identifying individuals who may receive a benefit from treatments comprising a drug effective at treating an HRD positive cancer. In some embodiment, the methods may comprise identifying individuals who may not receive the most benefit from treatment with a drug effective at treating an HRD positive cancer using knowledge of the absence of a homozygous deletion in BRCA1 in a sample from the individuals.

[0088] Provided herein are methods that can be used to predict survival for an individual with APC with a homozygous deletion in BRCA1, in response to knowledge of the BRCA1 homozygous deletion in a sample from the individual. In some aspects, predicting survival may comprise predicting that the individual is likely to survive longer after treatment with a drug effective at treating an HRD positive cancer than an individual with APC that does not have a BRCA1 homozygous deletion. In some aspects, survival may be measured using any survival measurement described herein, for example, but not limited to real- world overall survival (rwOS), time to next treatment (TTNT), or time to treatment discontinuation (TTD). In some aspects, knowledge of a BRCA1 homozygous deletion in a sample may be obtained using the methods of detecting a BRCA1 homozygous deletion described herein. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent.

[0089] In some embodiments, provided herein is a method of predicting survival of an individual having APC treated with a drug effective in treating an HRD positive cancer, comprising acquiring knowledge of a BRCA1 homozygous deletion in a sample from the individual, wherein responsive to the acquisition of said knowledge, the individual is predicted to have longer survival after treatment with the drug effective in treating an HRD positive cancer, as compared to an individual whose APC does not exhibit the BRCA1 homozygous deletion.

[0090] Provided herein are methods that can be used to select a treatment comprising a drug effective at treating an HRD positive cancer for an individual having APC in response to information about a BRCA1 homozygous deletion detected in a sample from the individual. Selecting the treatment for an individual having APC may be in response to identifying the individual as someone likely to benefit from treatment comprising the administration of a drug effective at treating an HRD positive cancer. Selecting the treatment for an individual having APC may be in response to identifying the individual as someone likely to receive benefit from treatment comprising the administration of a drug effective at treating an HRD positive cancer. Selecting the treatment may comprise selecting one or more treatment options for the individual with APC wherein one of the treatments comprises the administration of a drug effective at treating an HRD positive cancer. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent. The method may further comprise generating a report comprising the one or more treatment options identified for the individual based in part on the characterization of a homozygous deletion in BRCA1 in the sample from the individual.

[0091] Provided herein are methods that can be used to select a treatment comprising a drug effective at treating an HRD positive cancer for an individual having APC in response to information about a BRCA1 homozygous deletion detected in a sample from the individual. Selecting the treatment for an individual having APC may be in response to identifying the individual as someone likely to benefit, or not likely to have the most benefit from treatment comprising the drug effective at treating an HRD positive cancer. Selecting the treatment may comprise selecting one or more treatment options for the individual with APC wherein one of the treatments comprises the administration of a drug effective at treating an HRD positive cancer. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent. The method may further comprise generating a report comprisingthe one or more treatment options identified for the individual based in part on the characterization of a homozygous deletion in BRCA1 in the sample from the individual. In some aspects, the methods further comprise administering the drug effective at treating an HRD positive cancer to the individual.

[0092] In some embodiments, provided herein is a method of selecting treatment for an individual having advanced prostate cancer (APC), comprising acquiring knowledge of a BRCA1 homozygous deletion in a sample from an individual having APC, wherein responsive to the acquisition of said knowledge: (i) the individual is classified as a candidate to receive treatment with administration of a drug effective in treating an HRD positive cancer and / or (ii) the individual is identified as likely to respond to a treatment that comprises administration of the drug effective in treating an HRD positive cancer.

[0093] In some embodiments, provided herein is a method of identifying one or more treatment options for an individual having advanced prostate cancer (APC), the method comprising: (a) detecting a BRCA1 homozygous deletion in a sample from the individual; and (b) generating a report comprising one or more treatment options identified for the individual based at least in part on the presence of the BRCA1 homozygous deletion in the sample, wherein the one or more treatment options comprise a drug effective in treating an HRD positive cancer.

[0094] Provided herein are methods that can be used to treat an individual with APC with a homozygous deletion in BRCA1 by administering a drug effective at treating an HRD positive cancer. The treatment may delay progression of the APC in the individual. In some aspects, the treatment may be in response to knowledge that the individual is likely to respond to treatment with a drug effective at treating an HRD positive cancer. In some aspects, the treatment may be response to the knowledge of a BRCA1 homozygous deletion in a sample from the individual. In some aspects, knowledge of a BRCA1 homozygous deletion in a sample may be obtained using the methods of detecting a BRCA1 homozygous deletion described herein. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent.

[0095] In some embodiments, provided herein is a method of treating or delaying progression of advanced prostate cancer (APC) comprising, responsive to knowledge of a BRCA1 homozygous deletion in a sample from an individual, administering to the individual an effective amount of a drug effective in treating an HRD positive cancer.Provided herein are methods that can be used to generate a report about a about an individual with APC with a homozygous deletion in BRCAl.The report may provide disclosure about the homozygous deletion of BRCA1 in a sample obtained from an individual with APC. In some aspects, the disclosure may be in response to knowledge of a BRCA1 homozygous deletion in a sample from the individual. In some aspects, knowledge of a BRCA1 homozygous deletion in a sample may be obtained using the methods of detecting a BRCA1 homozygous deletion described herein. The report may further provide disclosure identifying the individual as an individual with APC who is likely to respond to treatment comprising the administration of a drug effective at treating an HRD positive cancer. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent. In some aspects, the report may be about an individual with APC wherein no homozygous deletion in BRCA1 was obtained. In such aspects, the report may provide information that the individual may not receive the most benefit from treatment with a drug effective in treating an HRD positive cancer. In some embodiments, the report may be generated and provided to a reporting party according to the methods described herein.Characterization of APC with homozygous deletion in BRCA1 and / or BRCA2 from a liquid biopsy sample

[0096] In additional aspects, provided herein are methods that can be used to characterize an APC with a homozygous deletion in BRCA1 and / or BRCA2 as potentially responsive to drugs effecting at treating HRD positive cancers using a liquid biopsy. In some aspects, the ability to characterize an APC with a homozygous deletion in BRCA1 and / or BRCA2 from a liquid biopsy sample may depend on the percent of circulating tumor DNA (ctDNA) in the cell-free DNA (cfDNA) of a liquid biopsy sample, referred to as a tumor fraction.

[0097] The methods described herein, may comprise determining a tumor fraction (TF) in a liquid biopsy sample from the individual, detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample, and if the TF is greater than or equal to about 20% or less than about 20% characterizing the APC having a BRCA1 and / or BRCA2 homozygous deletion.

[0098] In such aspects the methods may comprise, detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to about 20% and a BRCA1 homozygous deletion and a BRCA2 homozygous deletion is not detected in the liquid biopsy sample, characterizing the APC as not containingBRCA1 and BRCA2 homozygous deletion. In some aspects, the methods may comprise detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to about 25%, or about 30% or about 35%, or about 45% or about 50% and a BRCA1 homozygous deletion and a BRCA2 homozygous deletion is not detected in the liquid biopsy sample, characterizing the APC as not containing BRCA1 and BRCA2 homozygous deletion.

[0099] The methods may comprise determining a tumor fraction in the liquid biopsy sample collected from an individual using the methods described herein. The liquid biopsy sample may be a liquid biopsy sample as described herein, and may have less than 40% TF, less than 35% TF, less than 30% TF, less than 25% TF, less than 20% TF, less than 15% TF, less than 10% TF, or less than 5% TF and greater than or equal to about 1% TF. The liquid biopsy sample may be a liquid biopsy sample as described herein, and may have greater than or equal to 20% TF, greater than or equal to 25% TF, greater than or equal to 30% TF, greater than or equal to 35% TF, greater than or equal to 40% TF, greater than or equal to 45% or greater than or equal to 50%.

[0100] Detecting a homozygous deletion in BRCA1 and / or BRCA2 in the liquid biopsy sample from the individual with APC may be accomplished with using any of the methods known in the art to detect a deletion in a gene, such as but not limited to the in situ hybridization, array -based, amplification based, sequencing based, and protein detection based methods described herein.

[0101] In some aspects, if the TF is greater than or equal to about 20% or less than about 20% and a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion is detected in the liquid biopsy sample, the APC may be classified as having a BRCA1 and / or homozygous deletion. In some aspects, if the TF is greater than or equal to about 20%, greater than or equal to 25% TF, greater than or equal to 30% TF, greater than or equal to 35% TF, greater than or equal to 40% TF, greater than or equal to 45% or greater than or equal to 50% and a BRCA1 homozygous deletion and a BRCA2 homozygous deletion is not detected in the liquid biopsy sample, the APC may be classified as not having a BRCA1 and BRCA2.

[0102] The characterization of APC according to the methods described herein can be use in a variety of applications such as, but not limited to, diagnosing a patient with APC, selecting a treatment, treating a patient with APC, and generating reports about the APC for the patient or clinical provider. The patient may be identified as one likely to respond to a drug effective at treating an HRD positive cancer, such as a PARPi. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer 1described herein, such as but not limited to a PARPi or a platinum agent. In some respects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0103] Provided herein are methods that can be used to identify if an individual having APC who may benefit from a treatment with a drug effective at treating an HRD positive cancer, said method comprising: determining a TF in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in the liquid biopsy sample if the TF is greater than or equal to about 20% or less than about 20% and a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion is detected in the liquid biopsy sample, identifying the individual as one who may benefit from the treatment of a drug effective in treating an HRD positive cancer. In some embodiments, the method may comprise: determining a TF in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in the liquid biopsy sample if the TF is greater than or equal to about 20 and a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion is detected in the liquid biopsy sample, identifying the individual as one who may benefit from the treatment of a drug effective in treating an HRD positive cancer. In some embodiments, the APC is mPRPC. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent. In some embodiments, the benefit may be the most benefit. In some embodiments, the benefit may be a higher benefit than a benefit from a treatment other than a drug effective in treating an HRD positive cancer.

[0104] In some embodiments, provided herein is a method of identifying if an individual having advanced prostate cancer (APC) that may benefit from a treatment with a drug effective in treating an HRD positive cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to about 20% and a BRCA1 homozygous deletion and a BRCA2 homozygous deletion is not detected in the liquid biopsy sample, identifying the individual as one who may not receive the most benefit from the treatment with a drug effective in treating an HRD positive cancer. In some embodiments, the individual may be identified as one who may receive less benefit from the treatment with a drug effective in treating an HRD positive cancer. In some embodiments, the individual may be identified as one who may receive less of a benefit from the treatment with a drug effective in treating an HRD positive cancer than an individual identified as having an APC with a homozygous deletion in BRCA1 and / orBRCA2. In some embodiments, the APC is mPRPC. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent.

[0105] Provided herein are methods that can be used select treatment for an individual having APC, said method comprising: determining a TF in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in the liquid biopsy sample if the TF is greater than or equal to 20% or less than about 20% and a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion is detected in the liquid biopsy sample from the individual, classifying the individual as a candidate to receive treatment with a drug effective in treating an HRD positive cancer. In some embodiments, the method may comprise: determining a TF in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in the liquid biopsy sample if the TF is greater than or equal to 20% and a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion is detected in the liquid biopsy sample from the individual, classifying the individual as a candidate to receive treatment with a drug effective in treating an HRD positive cancer. In some embodiments, the APC is mPRPC. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent.

[0106] In some embodiments, the method for selecting treatment for an individual with APC may comprise: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; if the TF is greater than or equal to 20% and BRCA1 homozygous deletion and BRCA2 homozygous deletion is not detected in the liquid biopsy sample, classifying the individual as not a candidate to receive a benefit from treatment with a drug effective in treating an HRD positive cancer. In some embodiments, the individual may be classified as a candidate that will receive less of a benefit from treatment with a drug effective in treating an HRD positive cancer. In some embodiments, the individual may be classified as a candidate that will receive less of a benefit from treatment with a drug effective in treating an HRD positive cancer than an individual that is classified as a candidate to receive a benefit from the treatment. In some embodiments, the APC is mPRPC. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent.

[0107] Provided herein are methods that can be used to treat or delay progression of advanced prostate cancer (APC) in an individual, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to 20% or less than about 20%, and a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion is detected in the liquid biopsy sample from the individual, administering to the individual an effective amount of a drug effective in treating an HRD positive cancer. In some embodiments, the methods may comprise: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to 20% and a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion is detected in the liquid biopsy sample from the individual, administering to the individual an effective amount of a drug effective in treating an HRD positive cancer. In some embodiments, the APC is mPRPC. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent.

[0108] In some embodiments, the methods that can be used to treat or delay progression of an APC comprise: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to 20% and a BRCA1 homozygous deletion and a BRCA2 homozygous deletion is not detected in the liquid biopsy sample from the individual, administering to the individual an effective amount of a drug other than a drug effective in treating an HRD positive cancer. In some embodiments, the methods that can be used to treat or delay progression of an APC comprise: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to 20% and a BRCA1 homozygous deletion and a BRCA2 homozygous deletion is not detected in the liquid biopsy sample from the individual, administering to the individual an effective amount of a drug a drug effective in treating an HRD positive cancer, wherein the individual may not receive the most benefit from the treatment. In some embodiments, the treatment may not be as effective for treating the individual. In some embodiments, the treatment may not be the most effective for treating the individual. In some embodiments, the APC is mPRPC. The drug effective at treating anHRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent.

[0109] Provided herein are methods that can be used to predict survival for an individual with APC treated with a drug effective at treating an HRD positive cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to about 20% or less than about 20%, and a BRCA1 homologous deletion and / or a BRCA2 homozygous deletion is detected in the liquid biopsy sample from the individual, the individual is predicted to have longer survival after treatment with a drug effective in treating an HRD positive cancer, as compared to an individual whose APC does not exhibit the BRCA1 homozygous deletion and / or BRCA2 homozygous deletion. In some embodiments, the methods may comprise :detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to about 20% and a BRCA1 homologous deletion and / or a BRCA2 homozygous deletion is detected in the liquid biopsy sample from the individual, the individual is predicted to have longer survival after treatment with a drug effective in treating an HRD positive cancer, as compared to an individual whose APC does not exhibit the BRCA1 homozygous deletion and / or BRCA2 homozygous deletion. In some aspects, survival may be measured using any survival measurement described herein, for example, but not limited to real-world overall survival (rwOS), time to next treatment (TTNT), or time to treatment discontinuation (TTD). The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent.

[0110] Provided herein are methods that can be used to predict survival for an individual with APC treated with a drug effective at treating an HRD positive cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual, the individual is predicted to have shorter survival after treatment with the drug effective in treating an HRD positive cancer, as compared to an individual whose APC does exhibit the BRCA1 homozygous deletion and / or BRCA2 homozygous deletion. In some aspects, survival may be measured using any survival measurement described herein, for example, but not limited to real- world overall survival(rwOS), time to next treatment (TTNT), or time to treatment discontinuation (TTD). The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent.

[0111] Provided herein are methods that can be used to generate a report an individual with APC with a homozygous deletion in BRCA1 and / or BRCA2. The report may provide disclosure about the homozygous deletion of BRCA1 and / or BRCA2 identified in a liquid biopsy sample obtained from an individual with APC. The liquid biopsy sample may be a liquid biopsy sample as described herein and may have a TF greater than or equal to 20%, greater than or equal to 25%, greater than or equal to 30%, greater than or equal to 40%, greater than or equal to 45% or greater than or equal to 50%. The liquid biopsy sample may be a liquid biopsy sample as described herein and may have a TF less than 50%, less than 45%, less than 35%, less than 30%, less than 25% or less than 20%. Generating the report may comprise determining the TF in a liquid biopsy sample from the individual and detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample. If the TF is greater than or equal to 20% or less than about 20% and a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion is not detected in the liquid biopsy sample from the individual, the report may provide disclosure identifying the individual as an individual with APC who is likely to respond to treatment with a drug effective at treating an HRD positive cancer. If the TF is greater than or equal to 20% and a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion is not detected in the liquid biopsy sample from the individual, the report may provide disclosure identifying the individual as an individual with APC who is likely to respond to treatment with a drug effective at treating an HRD positive cancer. If the TF is greater than or equal to about 20% and a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion is not detected in the liquid biopsy sample from the individual, the report may provide disclosure identifying the individual as an individual with APC who is less likely to respond to treatment with of a drug effective at treating an HRD positive cancer, is likely to response to treatment comprising a drug effective at treating an HRD positive cancer, or may not receive the most benefit from treatment with a drug effective at treating an HRD positive cancer. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent. In some embodiments, the report may be generated and provided to a reporting party according to the methods described herein.Characterization of cancers with homozygous deletion in BRCA1 and / or BRCA2 from a liquid biopsy sample and a tissue biopsy sample

[0112] Provided herein are methods that can be used to characterize a cancer with homozygous deletion in BRCA1 and / or BRCA2 as potentially likely to respond to treatment with a drug effective at treating an HRD positive cancer. In some aspects, the ability to characterize a cancer with a homozygous deletion in BRCA1 and / or BRCA2 from a liquid biopsy sample may depend on the percent of circulating tumor DNA (ctDNA) in the cell-free DNA (cfDNA) of a liquid biopsy sample, referend to as a tumor fraction.

[0113] In some aspects, the methods comprise determining a tumor fraction (TF) in a liquid biopsy sample from an individual with a cancer; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; if the TF in the liquid biopsy sample is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detecting in the liquid biopsy sample from the individual, further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual. In some embodiments, the cancer is APC.

[0114] In some embodiments, a BRCA1 and a BRCA2 homozygous deletion is not detected in a liquid biopsy sample with a TF of less than 20%, but a BRCA1 and / or BRCA2 homozygous deletion is detected in a tumor biopsy sample from the same individual. In such cases, the cancer should be characterized as a cancer with a homozygous deletion in BRCA1 and / or BRCA2. In some aspects, detecting the BRCA1 and / or BRCA2 homozygous deletion in the tissue biopsy sample prevents mischaracterization of the cancer when the liquid biopsy sample has a tumor fraction of less than about 20%. In some aspects, tissue biopsy samples allow for detection of a BRCA1 and / or BRCA2 mutation in a cancer that was not detected in a liquid biopsy sample with a TF of less than about 20%. In some embodiments, the cancer is APC.

[0115] In some aspect, the method may comprise determining a tumor fraction in in the liquid biopsy sample collected from an individual using the methods described herein. The liquid biopsy sample may be a liquid biopsy sample as described herein, and may have less than 40% TF, less than 35% TF, less than 30% TF, less than 25% TF, less than 20% TF, less than 15% TF, less than 10% TF, or less than 5% TF and greater than or equal to about 1% TF. The liquid biopsy sample may be a liquid biopsy sample as described herein, and may have greater than or equal to 20% TF, greater than or equal to 25% TF, greater than or equalto 30% TF, greater than or equal to 35% TF, greater than or equal to 40% TF, greater than or equal to 45% or greater than or equal to 50%.

[0116] Detecting a homozygous deletion in BRCA1 and / or BRCA2 in the liquid biopsy sample from the individual with cancer may be accomplished with using any of the methods known in the art to detect a deletion in a gene, such as but not limited to the in situ hybridization, array -based, amplification based, sequencing based, and protein detection based methods described herein.

[0117] In some aspects, if the TF in the liquid biopsy sample is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detecting in the liquid biopsy sample from the individual, the methods further comprise detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual. Detecting a homozygous deletion in BRCA1 and / or BRCA2 in the tissue biopsy sample from the individual with cancer may be accomplished with using any of the methods known in the art to detect a deletion in a gene, such as but not limited to the in situ hybridization, array -based, amplification based, sequencing based, and protein detection based methods described herein.

[0118] The characterization of cancer according to the methods described herein can be use in a variety of applications such as, but not limited to, diagnosing a patient with cancer, selecting a treatment, treating a patient with cancer, and generating reports about the cancer for the patient or clinical provider. In some aspects, the characterization of the cancer as a cancer with a homozygous deletion in BRCA1 and / or BRCA2 can be used to classify the cancer as likely to respond to a drug effective at treating an HRD positive cancer. The patient may be identified as one likely to respond to a drug effective at treating an HRD positive cancer, such as a PARPi. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0119] Provided herein are methods that can be used to identifying if an individual having cancer that may benefit from a treatment with a drug effective in treating an HRD positive cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; if the TF in the liquid biopsy sample is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detecting in the liquid biopsy sample from the individual, further detecting a BRCA1homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if BRCA1 homozygous deletion and / or BRCA2 homozygous deletion are detected in the tissue biopsy sample from the individual, identifying the individual as one who may benefit from the treatment with a drug effective in treating an HRD positive cancer. In some aspects, the cancer is APC. In some embodiments, the APC is mPRPC. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent. In some embodiments, the benefit may be the most benefit. In some embodiments, the benefit may be a higher benefit than a benefit from a treatment other than a drug effective in treating an HRD positive cancer.

[0120] In some embodiments, provided herein is a method of identifying if an individual having cancer that may benefit from a treatment with a drug effective in treating an HRD positive cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample from the individual, if the TF in the liquid biopsy sample is less than about 20% and BRCA1 homozygous deletion and BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual; further detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if a BRCA1 homozygous deletion and a BRCA2 homozygous deletion are not detected in the tissue biopsy sample, identifying the individual as one who may not benefit from the treatment with a drug effective in treating an HRD positive cancer; or if a BRCA1 homozygous deletion or a BRCA2 homozygous deletion is detected in the tissue biopsy sample, identifying the individual as one who may benefit from the treatment with a drug effective in treating an HRD positive cancer. In some embodiments, if a BRCA1 homozygous deletion and a BRCA2 homozygous deletion are not detected in the tissue biopsy sample, the individual may be identified as an individual who may receive a less benefit from the treatment with a drug effective in treating an HRD positive cancer. In some embodiments, if a BRCA1 homozygous deletion and a BRCA2 homozygous deletion are not detected in the tissue biopsy sample, the individual may be identified as one who may receive less of a benefit from the treatment with a drug effective in treating an HRD positive cancer than an individual identified as having a cancer with a homozygous deletion in BRCA1 and / or BRCA2. In some aspects, the cancer is APC. In some embodiments, the APC is mPRPC. The drug effective at treating an HRD positive cancer may be any of the drugseffective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent.

[0121] Provided herein are methods that can be used to select treatment for an individual having cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual; further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if BRCA1 homozygous deletion and / or BRCA2 homozygous deletion are detected in the tissue biopsy sample from the individual, classifying the individual as a candidate to receive treatment with a drug effective in treating an HRD positive cancer. In some aspects, the cancer is APC. In some embodiments, the APC is mPRPC. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent.

[0122] Provided herein, are methods for selecting treatment for an individual having cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual; further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if a BRCA1 homozygous deletion and a BRCA2 homozygous deletion are not detected in the tissue biopsy sample, classifying the individual as not a candidate to receive treatment with a drug effective in treating an HRD positive cancer, or if a BRCA1 homozygous deletion or a BRCA2 homozygous deletion is detected in the tissue biopsy sample, classifying the individual as a candidate to receive treatment with a drug effective in treating an HRD positive cancer. In some aspects, if a BRCA1 homozygous deletion and a BRCA2 homozygous deletion are not detected in the tissue biopsy sample, the individual is classified as a candidate that will receive less of a benefit from treatment with a drug effective in treating an HRD positive cancer. In some embodiments, if a BRCA1 homozygous deletion and a BRCA2 homozygous deletion are not detected in the tissue biopsy sample, the individual may be classified as a candidate that will receive less of a benefit from treatment with a drug effective in treating an HRD positive cancer than an individual that is classifiedas a candidate to receive a benefit from the treatment. In some aspects, the cancer is APC. In some embodiments, the APC is mPRPC. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent.

[0123] Provided herein are methods that can be used to treat or delaying progression of advanced cancer in an individual said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual; further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual, and if a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion are detected in the tissue biopsy sample from the individual, administering to the individual an effective amount of a drug effective in treating an HRD positive cancer. In some aspects, the cancer is APC. In some embodiments, the APC is mPRPC. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent.

[0124] Provided herein are methods that can be used to treat or delaying progression of cancer in an individual said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual, further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if a BRCA1 homozygous deletion and a BRCA2 homozygous deletion are not detected in the tissue biopsy sample, administering to the individual an effective amount of a drug other than a drug effective in treating an HRD positive cancer, or if a BRCA1 homozygous deletion or a BRCA2 homozygous deletion is detected in the tissue biopsy sample, administering to the individual a drug effective in treating an HRD positive cancer. In some embodiments, if a BRCA1 homozygous deletion and a BRCA2 homozygous deletion are not detected in the tissue biopsy sample, an effective amount of a drug effective at treating an HRD positive cancer is administered, wherein the individual may not receive the most benefit from the treatment, the treatment may not be as effective for treating the individual or wherein the treatment may not be the most effective for treating the individual.In some aspects, the cancer is APC. In some embodiments, the APC is mPRPC. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent.

[0125] Provided herein are methods that can be used to predict survival of an individual having cancer treated with administration of a drug effective in treating an HRD positive cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual, further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion are detected in the tissue biopsy sample the individual is predicted to have longer survival after treatment with the administration of the drug effective in treating an HRD positive cancer, as compared to an individual whose cancer does not exhibit the BRCA1 homozygous deletion and / or BRCA2 homozygous deletion. In some aspects, survival may be measured using any survival measurement described herein, for example, but not limited to real- world overall survival (rwOS), time to next treatment (TTNT), or time to treatment discontinuation (TTD). In some aspects, the cancer is APC. In some embodiments, the APC is mPRPC. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent.

[0126] Provided herein are methods that can be used to predict survival of an individual having cancer treated with administration of a drug effective in treating an HRD positive cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual, further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if a BRCA1 homozygous deletion and a BRCA2 homozygous deletion are not detected in the tissue biopsy sample the individual is not predicted to have longer survival after treatment with the administration of the drug effective in treating an HRD positive cancer, as compared to an individual whose cancer does not exhibit the BRCA1 homozygous deletion and / orBRCA2 homozygous deletion, or if a BRCA1 homozygous deletion or a BRCA2 homozygous deletion is detected in the tissue biopsy sample, the individual is predicted to have longer survival after treatment with the administration of the drug effective in treating an HRD positive cancer, as compared to an individual whose cancer does not exhibit the BRCA1 homozygous deletion and / or BRCA2 homozygous deletion. In some aspects, survival may be measured using any survival measurement described herein, for example, but not limited to real-world overall survival (rwOS), time to next treatment (TTNT), or time to treatment discontinuation (TTD). In some aspects, the cancer is APC. In some embodiments, the APC is mPRPC. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to a PARPi or a platinum agent.

[0127] Provided herein are methods that can be used to generate a report an individual with cancer with a homozygous deletion in BRCA1 and / or BRCA2. The report may provide disclosure about the homozygous deletion of BRCA1 and / or BRCA2 identified in a tissue biopsy sample obtained from an individual with cancer. The report may explain that a tissue biopsy sample was used because the liquid biopsy sample had a TF of less than about 20% and a BRCA1 and a BRCA2 homozygous deletion were not identified in the liquid biopsy sample.

[0128] Generating the report may comprise determining the TF in a liquid biopsy sample from the individual and if the TF in the liquid biopsy sample is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detecting in the liquid biopsy sample from the individual, further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual. If a BRCA1 and / or BRCA2 homozygous deletion is detected in the tissue biopsy sample, the report may provide disclosure identifying the individual as an individual with cancer who is likely to respond to treatment with a drug effective at treating an HRD positive cancer. If a BRCA1 and BRCA2 homozygous deletion are not detected in the tissue biopsy sample, the report may provide disclosure identifying the individual as an individual with cancer who is less likely to respond to treatment with of a drug effective at treating an HRD positive cancer, is likely to response to treatment comprising a drug effective at treating an HRD positive cancer, or may not receive the most benefit from treatment with a drug effective at treating an HRD positive cancer. The drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but notlimited to a PARPi or a platinum agent. In some embodiments, the report may be generated and provided to a reporting party according to the methods described herein.Methods of detecting a homozygous deletion in BRCA1 and / or BRCA2

[0129] Provided herein are methods that can be used to detect a homozygous deletion (or loss) in BRCA1 and / or BRCA2. A homozygous deletion of BRCA1 and / or BRCA2 may be characterized as a deletion in both copies of BRCA1 and / or BRCA2. A heterozygous deletion of BRCA1 and / or BRCA2 may be characterized as a deletion in one copy of BRCA1 and / or BRCA2 respectively. Homozygous deletions likely result in a cell not being able to produce any functional BRCA1 and / or BRCA2 polypeptides. This is in comparison to a heterozygous deletion, wherein the cell may be able to produce a functional BRCA1 and / or BRCA2 polypeptide but at a level lower than if there was no deletion.

[0130] As described herein, a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion may be detected in a sample (e.g., a liquid biopsy sample or a tissue biopsy sample) by one or more of: a nucleic acid hybridization assay, an amplification-based assay, a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay, real-time PCR, a screening analysis, fluorescence in situ hybridization (FISH), spectral karyotyping, multicolor FISH (mFISH), comparative genomic hybridization, in situ hybridization, sequence- specific priming (SSP) PCR, high-performance liquid chromatography (HPLC), mass-spectrometric genotyping, or sequencing. In some embodiments, a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion may be detected in a sample by a nucleic acid hybridization assay. In some embodiments, a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion may be detected in a sample by an amplification-based assay. In some embodiments, a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion may be detected in a sample by PCR-RFLP. In some embodiments, a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion may be detected in a sample by real time PCR. In some embodiments, a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion may be detected in a sample by a screening analysis. In some embodiments, a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion may be detected in a sample by a FISH. In some embodiments, a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion may be detected in a sample by spectral karyotyping. In some embodiments, a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion may be detected in a sample by mFISH. In some embodiments, a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion may bedetected in a sample by comparative genomic hybridization. In some embodiments, a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion may be detected in a sample by in situ hybridization. In some embodiments, a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion may be detected in a sample by SSP PCR. In some embodiments, a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion may be detected in a sample by HPLC. In some embodiments, a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion may be detected in a sample by mass-spectrometric genotyping.

[0131] In some embodiments, the homozygous deletion of BRCA1 and / or BRCA2 may be detected among nucleic acid molecules isolated from the sample of the disclosure is detected using any suitable method known in the art, such as a nucleic acid hybridization assay, an amplification-based assay (e.g., polymerase chain reaction, PCR), a PCR-RFLP assay, realtime PCR, sequencing (e.g., Sanger sequencing or next-generation sequencing), a screening analysis (e.g., using karyotype methods), fluorescence in situ hybridization (FISH), break away FISH, spectral karyotyping, multiplex-FISH, comparative genomic hybridization, in situ hybridization, single specific primer-polymerase chain reaction (SSP-PCR), high performance liquid chromatography (HPLC), or mass-spectrometric genotyping. Methods of analyzing samples, e.g., to detect a nucleic acid molecule, are described in U.S. Patent No. 9,340,830 and in WO2012092426A1, which are hereby incorporated by reference in their entirety.

[0132] In some aspects, provided herein are reagents that can be used for detecting the presence or absence of BRCA1 and / or BRCA2 nucleic acid molecules of the disclosure or a fragment thereof, e.g., according to the methods of detection provided herein. In some embodiments, a detection reagent provided herein comprises a nucleic acid molecule, e.g., a DNA, RNA, or mixed DNA / RNA molecule, comprising a nucleotide sequence that is complementary to a nucleotide sequence on a target nucleic acid, e.g., a nucleic acid that comprises a deletion of BRCA1 and or BRCA2 described herein or a fragment or portion thereof.In Situ Hybridization

[0133] In some embodiments, the homozygous deletion of BRCA1 and / or BRCA2 among nucleic acid molecules isolated from the sample of the disclosure may be detected using an in situ hybridization method, such as a fluorescence in situ hybridization (FISH) method.

[0134] In some embodiments, FISH analysis is used to identify an RNA molecule comprising a BRCA1 and / or BRCA2 nucleic acid described herein. Methods for performing FISH areknown in the art and can be used in nearly any type of tissue. In FISH analysis, nucleic acid probes which are detectably labeled, e.g., fluorescently labeled, are allowed to bind to specific regions of DNA, e.g., a chromosome, or an RNA, e.g., an mRNA, and then examined, e.g., through a microscope. See, for example, U.S. Patent No. 5,776,688. DNA or RNA molecules are first fixed onto a slide, the labeled probe is then hybridized to the DNA or RNA molecules, and then visualization is achieved, e.g., using enzyme-linked label-based detection methods known in the art. Generally, the resolution of FISH analysis is on the order of detection of 60 to 100000 nucleotides, e.g., 60 base pairs (bp) up to 100 kilobase pairs of DNA. Nucleic acid probes used in FISH analysis comprise single stranded nucleic acids. Such probes are typically at least about 50 nucleotides in length. In some embodiments, probes comprise about 100 to about 500 nucleotides. Probes that hybridize with centromeric DNA and locus- specific DNA or RNA are available commercially, for example, from Vysis, Inc. (Downers Grove, Ill.), Molecular Probes, Inc. (Eugene, Oreg.) or from Cytocell (Oxfordshire, UK). Alternatively, probes can be made non-commercially from chromosomal or genomic DNA or other sources of nucleic acids through standard techniques. Examples of probes, labeling and hybridization methods are known in the art. In some embodiments, FISH probes designed to hybridize to BRCA1 and / or BRCA2. In some embodiments, the absence of FISH probes hybridizing to BRCA1 and / or BRCA2 may indicate a homozygous deletion of BRCA1 and / or BRCA2 respectively.

[0135] Several variations of FISH methods are known in the art and are suitable for use according to the methods of the disclosure, including single-molecule RNA FISH, Fiber FISH, Q-FISH, Flow-FISH, MA-FISH, break-away FISH, hybrid fusion-FISH, and multifluor FISH or mFISH.

[0136] Also provided herein are probes, e.g., nucleic acid molecules, suitable for the detection of a BRCA1 and / or BRCA2 nucleic acid molecule provided herein or suitable for the detection a deletion of BRCA1 and / or BRCA2. In some embodiments, a probe provided herein comprises a nucleic acid sequence configured to hybridize to a target nucleic acid molecule comprising a BRCA1 and / or BRCA2 nucleic acid molecule provided herein, or a fragment or portion thereof. In some embodiments, the probe comprises a nucleic acid sequence configured to hybridize to the BRCA1 and / or BRCA2 nucleic acid molecule, or the fragment or portion thereof, of the target nucleic acid molecule. In some embodiments, the probe comprises a nucleic acid sequence configured to hybridize to a fragment or portion of the BRCA1 and / or BRCA2 nucleic acid molecule of the target nucleic acid molecule. In some embodiments, the fragment or portion comprises between about 5 and about 25nucleotides, between about 5 and about 300 nucleotides, between about 100 and about 300 nucleotides, between about 130 and about 230 nucleotides, or between about 150 and about 200 nucleotides. In some instances, failure of the probe to detect BRCA1 and / or BRCA through hybridization indicates a deletion of BRCA1 and / or BRCA2.

[0137] In some embodiments, the probe comprises a nucleic acid molecule which is a DNA, RNA, or a DNA / RNA molecule. In some embodiments, the probe comprises a nucleic acid molecule comprising any of between about 10 and about 20 nucleotides, between about 12 and about 20 nucleotides, between about 10 and about 1000 nucleotides, between about 50 and about 500 nucleotides, between about 100 and about 500 nucleotides, between about 100 and about 300 nucleotides, between about 130 and about 230 nucleotides, or between about 150 and about 200 nucleotides. In some embodiments, the probe comprises a nucleic acid molecule comprising any of 10 nucleotides, 11 nucleotides, 12 nucleotides, 13 nucleotides, 14 nucleotides, 15 nucleotides, 16 nucleotides, 17 nucleotides, 18 nucleotides, 19 nucleotides, 20 nucleotides, 21 nucleotides, 22 nucleotides, 23 nucleotides, 24 nucleotides, 25 nucleotides, 26 nucleotides, 27 nucleotides, 28 nucleotides, 29 nucleotides, or 30 nucleotides. In some embodiments, the probe comprises a nucleic acid molecule comprising any of between about 40 nucleotides and about 50 nucleotides, about 50 nucleotides and about 100 nucleotides, about 100 nucleotides and about 150 nucleotides, about 150 nucleotides and about 200 nucleotides, about 200 nucleotides and about 250 nucleotides, about 250 nucleotides and about 300 nucleotides, about 300 nucleotides and about 350 nucleotides, about 350 nucleotides and about 400 nucleotides, about 400 nucleotides and about 450 nucleotides, about 450 nucleotides and about 500 nucleotides, about 500 nucleotides and about 550 nucleotides, about 550 nucleotides and about 600 nucleotides, about 600 nucleotides and about 650 nucleotides, about 650 nucleotides and about 700 nucleotides, about 700 nucleotides and about 750 nucleotides, about 750 nucleotides and about 800 nucleotides, about 800 nucleotides and about 850 nucleotides, about 850 nucleotides and about 900 nucleotides, about 900 nucleotides and about 950 nucleotides, or about 950 nucleotides and about 1000 nucleotides. In some embodiments, the probe comprises a nucleic acid molecule comprising between about 12 and about 20 nucleotides.

[0138] In some embodiments, a probe provided herein comprises a DNA, RNA, or a DNA / RNA molecule. In some embodiments, a probe provided herein includes a label or a tag. In some embodiments, the label or tag is a radiolabel (e.g., a radioisotope), a fluorescent label (e.g., a fluorescent compound), an enzymatic label, an enzyme co-factor, a sequence tag, biotin, or another ligand. In some embodiments, a probe provided herein includes adetection reagent such as a fluorescent marker. In some embodiments, a probe provided herein includes (e.g., is conjugated to) an affinity tag, e.g., that allows capture and isolation of a hybrid formed by a probe and a nucleic acid hybridized to the probe. In some embodiments, the affinity tag is an antibody, an antibody fragment, biotin, or any other suitable affinity tag or reagent known in the art. In some embodiments, a probe is suitable for solution phase hybridization.

[0139] In some embodiments, probes provided herein may be used according to the methods of detection of a deletion of BRCA1 and / or BRCA2 within nucleic acid molecules provided herein. For example, a probe provided herein may be used for detecting a BRCA1 and / or BRCA2 nucleic acid molecule provided herein in a sample, e.g., a sample obtained from an individual. In some embodiments, the probe may be used for identifying cells or tissues that express a BRCA1 and / or BRCA2 nucleic acid molecule provided herein, e.g., by determining levels of the BRCA1 and / or BRCA2 nucleic acid molecule. In some embodiments, the probe may be used for detecting levels of a BRCA1 and / or BRCA2 nucleic acid molecule, e.g., mRNA levels, in a sample of cells from an individual.

[0140] In some embodiments, probes provided herein may be used according to the methods of detection of a deletion of BRCA1 and / or BRCA2 within nucleic acid molecules provided herein. For example, a probe provided herein may be used for detecting a BRCA1 and / or BRCA2 nucleic acid molecule provided herein in a sample, e.g., a sample obtained from an individual, wherein detection of BRCA1 and / or BRCA2 may indicate lack of a homozygous deletion of BRCA1 and / or BRCA2. In some embodiments, the probe may be used for identifying cells or tissues that express a BRCA1 and / or BRCA2 nucleic acid molecule provided herein, e.g., by measuring levels of the BRCA1 and / or BRCA2 nucleic acid molecule, wherein detecting BRCA1 and / or BRCA2 expression may indicate a lack of a homozygous deletion of BRCA1 and / or BRCA2. In some embodiments, the probe may be used for detecting levels of a BRCA1 and / or BRCA2 nucleic acid molecule, e.g., mRNA levels, in a sample of cells from an individual, wherein detection of BRCA1 and / or BRCA2 mRNA may indicate lack of a homozygous deletion of BRCA1 and / or BRCA2. In some embodiments, a probe provided herein specifically hybridizes to a nucleic acid comprising a deletion of BRCA1 and / or BRCA2 nucleic acid molecules described herein.

[0141] In some embodiments, one or more probes provided herein are suitable for use in in situ hybridization methods, e.g., as described above, such as FISH.

[0142] In some embodiments, probes, such as probes for use in the FISH methods described herein, are labeled such that a chromosomal region or a region on an RNA to which theprobes hybridize can be detected. Probes typically are directly labeled with a fluorophore, allowing the probe to be visualized without a secondary detection molecule. Probes can also be labeled by nick translation, random primer labeling or PCR labeling. Labeling may be accomplished using fluorescent (direct)-or haptene (indirect)-labeled nucleotides. Representative, non-limiting examples of labels include: AMCA-6-dUTP, CascadeBlue-4- dUTP, Fluorescein- 12-dUTP, Rhodamine-6-dUTP, TexasRed-6-dUTP, Cy3-6-dUTP, Cy5- dUTP, Biotin(BIO)-l l-dUTP, Digoxygenin(DIG)-l l-dUTP and Dinitrophenyl (DNP)-l l- dUTP. Probes can also be indirectly labeled with biotin or digoxygenin or labeled with radioactive isotopes such as32P and3H, and secondary detection molecules are used, or further processing is performed, to visualize the probes. For example, a probe labeled with biotin can be detected by avidin conjugated to a detectable marker, e.g., avidin can be conjugated to an enzymatic marker such as alkaline phosphatase or horseradish peroxidase. Enzymatic markers can be detected in standard colorimetric reactions using a substrate and / or a catalyst for the enzyme. Catalysts for alkaline phosphatase include 5-bromo-4-chloro-3- indolylphosphate and nitro blue tetrazolium. Diaminobenzoate can be used as a catalyst for horseradish peroxidase. Probes can also be prepared such that a fluorescent or other label is added after hybridization of the probe to its target to detect that the probe hybridized to the target. For example, probes can be used that have antigenic molecules incorporated into the nucleotide sequence. After hybridization, these antigenic molecules are detected, for example, using specific antibodies reactive with the antigenic molecules. Such antibodies can, for example, themselves incorporate a fluorochrome, or can be detected using a second antibody with a bound fluorochrome. For fluorescent probes, e.g., used in FISH techniques, fluorescence can be viewed with a fluorescence microscope equipped with an appropriate filter for each fluorophore, or by using dual or triple band-pass filter sets to observe multiple fluorophores. Alternatively, techniques such as flow cytometry can be used to examine the hybridization pattern of the chromosomal probes.Array-Based Methods

[0143] In some embodiments, the homozygous deletion of BRCA1 and / or BRCA2 among nucleic acid molecules isolated from the sample of the disclosure may be detected using an array-based method, such as array-based comparative genomic hybridization (CGH) methods. In array-based CGH methods, a first sample of nucleic acids (e.g., from a sample, such as from a tumor) is labeled with a first label, while a second sample of nucleic acids (e.g., a control, such as from a healthy cell / tissue) is labeled with a second label. In someembodiments, equal quantities of the two samples are mixed and co-hybridized to a DNA microarray of several thousand evenly spaced cloned DNA fragments or oligonucleotides, which have been spotted in triplicate on the array. After hybridization, digital imaging systems are used to capture and quantify the relative fluorescence intensities of each of the hybridized fluorophores. The resulting ratio of the fluorescence intensities is proportional to the ratio of the copy numbers of DNA sequences in the two samples. In some embodiments, a copy number of zero may indicate a homozygous deletion of BRCA1 and / or BRC2. In some embodiments, Array-based CGH can also be performed with single-color labeling. In single color CGH, a control (e.g., control nucleic acid sample, such as from a healthy cell / tissue) is labeled and hybridized to one array and absolute signals are read, and a test sample (e.g., a nucleic acid sample obtained from an individual or from a tumor) is labeled and hybridized to a second array (with identical content) and absolute signals are read.

[0144] Provided herein are baits suitable for the detection of a BRCA1 or BRCA2 nucleic acid molecule of the disclosure. In some instances, the baits are suitable to detect a deletion if BRCA1 and / or BRCA2. In some embodiments, the bait comprises a capture nucleic acid molecule configured to hybridize to a target nucleic acid molecule comprising a BRCA1 and or BRCA2 nucleic acid molecule provided herein, or a fragment or portion thereof. In some embodiments, the bait comprises a capture nucleic acid molecule configured to hybridize to a target nucleic acid molecule comprising a deletion of BRCA1 and / or BRCA2 in a nucleic acid molecule provided herein, or a fragment or portion thereof. In some embodiments, the capture nucleic acid molecule is configured to hybridize to the BRCA1 and / or BRCA2 nucleic acid molecule of the target nucleic acid molecule. In some embodiments, the capture nucleic acid molecule is configured to hybridize to the genomic regions surrounding a BRCA1 and / or BRCA2 nucleic acid molecule of the target nucleic acid molecule.

[0145] In some embodiments, the capture nucleic acid molecule is configured to hybridize to a fragment of the BRCA1 and / or BRCA2 nucleic acid molecule. In some embodiments, the fragment comprises (or is) between about 5 and about 25 nucleotides, between about 5 and about 300 nucleotides, between about 100 and about 300 nucleotides, between about 130 and about 230 nucleotides, or between about 150 and about 200 nucleotides. In some embodiments, the capture nucleic acid molecule is between about 5 and about 25 nucleotides, between about 5 and about 300 nucleotides, between about 100 and about 300 nucleotides, between about 130 and about 230 nucleotides, or between about 150 and about 200 nucleotides. In some embodiments, the fragment comprises (or is) about 100 nucleotides, about 125 nucleotides, about 150 nucleotides, about 175 nucleotides, about 200 nucleotides,about 225 nucleotides, about 250 nucleotides, about 275 nucleotides, or about 300 nucleotides in length. In some embodiments, the capture nucleic acid molecule comprises (or is) about 100 nucleotides, about 125 nucleotides, about 150 nucleotides, about 175 nucleotides, about 200 nucleotides, about 225 nucleotides, about 250 nucleotides, about 275 nucleotides, or about 300 nucleotides in length.

[0146] In some embodiments, a bait provided herein comprises a DNA, RNA, or a DNA / RNA molecule. In some embodiments, a bait provided herein includes a label or a tag. In some embodiments, the label or tag is a radiolabel, a fluorescent label, an enzymatic label, a sequence tag, biotin, or another ligand. In some embodiments, a bait provided herein includes a detection reagent such as a fluorescent marker. In some embodiments, a bait provided herein includes (e.g., is conjugated to) an affinity tag, e.g., that allows capture and isolation of a hybrid formed by a bait and a nucleic acid hybridized to the bait. In some embodiments, the affinity tag is an antibody, an antibody fragment, biotin, or any other suitable affinity tag or reagent known in the art. In some embodiments, a bait is suitable for solution phase hybridization.

[0147] Baits can be produced and used according to methods known in the art, e.g., as described in WO2012092426 Al and / or or in Frampton et al (2013) Nat Biotechnol, 31:1023- 1031, incorporated herein by reference. For example, biotinylated baits (e.g., RNA baits) can be produced by obtaining a pool of synthetic long oligonucleotides, originally synthesized on a microarray, and amplifying the oligonucleotides to produce the bait sequences. In some embodiments, the baits are produced by adding an RNA polymerase promoter sequence at one end of the bait sequences, and synthesizing RNA sequences using RNA polymerase. In one embodiment, libraries of synthetic oligodeoxynucleotides can be obtained from commercial suppliers, such as Agilent Technologies, Inc., and amplified using known nucleic acid amplification methods.

[0148] In some embodiments, a bait provided herein is between about 100 nucleotides and about 300 nucleotides. In some embodiments, a bait provided herein is between about 130 nucleotides and about 230 nucleotides. In some embodiments, a bait provided herein is between about 150 nucleotides and about 200 nucleotides. In some embodiments, a bait provided herein comprises a target- specific bait sequence (e.g., a capture nucleic acid molecule described herein) and universal tails on each end. In some embodiments, the targetspecific sequence, e.g., a capture nucleic acid molecule described herein, is between about 40 nucleotides and about 300 nucleotides. In some embodiments, the target- specific sequence, e.g., a capture nucleic acid molecule described herein, is between about 100 nucleotides andabout 200 nucleotides. In some embodiments, the target- specific sequence, e.g., a capture nucleic acid molecule described herein, is between about 120 nucleotides and about 170 nucleotides. In some embodiments, the target- specific sequence, e.g., a capture nucleic acid molecule described herein, is about 150 nucleotides or about 170 nucleotides. In some embodiments, a bait provided herein comprises an oligonucleotide comprising about 200 nucleotides, of which about 150 nucleotides or about 170 nucleotides are target- specific (e.g., a capture nucleic acid molecule described herein), and the other 50 nucleotides or 30 nucleotides (e.g., 25 or 15 nucleotides on each end of the bait) are universal arbitrary tails, e.g., suitable for PCR amplification.Amplification-Based. Methods

[0149] In some embodiments, the homozygous deletion of BRCA1 and / or BRCA2 among nucleic acid molecules isolated from the sample of the disclosure may be detected use an amplification-based method, wherein post amplification of BRCA1 and / or BRCA2 nucleic acids, nucleic acids encoding BRCA1 and / or BRCA2 are ultimately not found. As is known in the art, in such amplification-based methods, a sample of nucleic acids, such as a sample obtained from an individual or from a tumor, is used as a template in an amplification reaction (e.g., Polymerase Chain Reaction (PCR)) using one or more oligonucleotides or primers, e.g., such as one or more oligonucleotides or primers provided herein. The presence of a homozygous deletion in BRCA1 and / or BRCA2 of the disclosure in the sample can be determined based on the presence or absence of an amplification product. Quantitative amplification methods are also known in the art and may be used according to the methods provided herein. Methods of measurement of DNA copy number at micro satellite loci using quantitative PCR analysis are known in the art. The known nucleotide sequence for genes is sufficient to enable one of skill in the art to routinely select primers to amplify any portion of the gene. Fluorogenic quantitative PCR can also be used. In Anorogenic quantitative PCR, quantitation is based on the amount of Auorescence signals, e.g., TaqMan and Sybr green.

[0150] Other amplification methods suitable for use according to the methods provided herein include, e.g., ligase chain reaction (LCR), transcription amplification, self-sustained sequence replication, dot PCR, and linker adapter PCR.

[0151] In some aspects, provided herein are oligonucleotides, e.g., useful as primers. In some embodiments, an oligonucleotide, e.g., a primer, provided herein comprises a nucleotide sequence configured to hybridize to a target nucleic acid molecule comprising a BRCA1 and / or BRCA2 nucleic acid molecule provided herein, or a fragment or portion thereof. Insome embodiments, the oligonucleotide comprises a nucleotide sequence configured to hybridize to the BRCA1 and / or BRCA2 nucleic acid molecule of the target nucleic acid molecule. In some embodiments, the oligonucleotide comprises a nucleotide sequence configured to hybridize to a fragment or portion of the BRCA1 and / or BRCA2 nucleic acid molecule of the target nucleic acid molecule. In some embodiments, the oligonucleotides may be used for targeted sequencing as described herein.

[0152] In some embodiments, the oligonucleotide comprises a nucleotide sequence corresponding to a BRCA1 and / or BRCA2 nucleic acid molecule provided herein. In some embodiments, the oligonucleotide comprises a nucleotide sequence corresponding to a fragment or a portion of a BRCA1 and / or BRCA2 nucleic acid molecule provided herein. In some embodiments, the fragment or portion comprises between about 10 and about 30 nucleotides, between about 12 and about 20 nucleotides, or between about 12 and about 17 nucleotides. In some embodiments, the oligonucleotide comprises a nucleotide sequence complementary to a BRCA1 and / or BRCA2 nucleic acid molecule provided herein. In some embodiments, the oligonucleotide comprises a nucleotide sequence complementary to a fragment or a portion of a BRCA1 and / or BRCA2 nucleic acid molecule provided herein. In some embodiments, the fragment or portion comprises between about 10 and about 30 nucleotides, between about 12 and about 20 nucleotides, or between about 12 and about 17 nucleotides.

[0153] In some embodiments, an oligonucleotide, e.g., a primer, provided herein comprises a nucleotide sequence that is sufficiently complementary to its target nucleotide sequence such that the oligonucleotide specifically hybridizes to a nucleic acid molecule comprising the target nucleotide sequence, e.g., under high stringency conditions. In some embodiments, an oligonucleotide, e.g., a primer, provided herein comprises a nucleotide sequence that is sufficiently complementary to its target nucleotide sequence such that the oligonucleotide specifically hybridizes to a nucleic acid molecule comprising the target nucleotide sequence under conditions that allow a polymerization reaction (e.g., PCR) to occur.

[0154] In some embodiments, an oligonucleotide, e.g., a primer, provided herein may be useful for initiating DNA synthesis via PCR (polymerase chain reaction) or a sequencing method. In some embodiments, the oligonucleotide may be used to amplify a nucleic acid molecule comprising a BRCA1 and / or BRCA2 nucleic acid molecule provided herein, or a fragment thereof, e.g., using PCR. In some embodiments, the oligonucleotide may be used to sequence a nucleic acid molecule comprising a BRCA1 and / or BRCA2 nucleic acid moleculeprovided herein, or a fragment thereof. In some embodiments, the oligonucleotide may be used to sequence a nucleic acid molecule comprising a BRCA1 and / or BRCA2 deletion.

[0155] In some embodiments, pairs of oligonucleotides, e.g., pairs of primers, are provided herein, which are configured to hybridize to a nucleic acid molecule comprising a BRCA1 and / or BRCA2 nucleic acid molecule provided herein, or a fragment thereof. In some embodiments, a pair of oligonucleotides of the disclosure may be used for directing amplification of the BRCA1 and / or BRCA2 nucleic acid molecule or fragment thereof, e.g., using a PCR reaction. In some embodiments, pairs of oligonucleotides, e.g., pairs of primers, are provided herein, which are configured to hybridize to a nucleic acid molecule comprising a BRCA1 and / or BRCA2 breakpoint provided herein, e.g., for use in directing amplification of the BRCA1 and / or BRCA2 nucleic acid molecule or fragment thereof, e.g., using a PCR reaction.

[0156] In some embodiments, an oligonucleotide, e.g., a primer, provided herein is a single stranded nucleic acid molecule, e.g., for use in sequencing or amplification methods described herein. In some embodiments, an oligonucleotide provided herein is a double stranded nucleic acid molecule. In some embodiments, a double stranded oligonucleotide is treated, e.g., denatured, to separate its two strands prior to use, e.g., in sequencing or amplification methods. Oligonucleotides provided herein comprise a nucleotide sequence of sufficient length to hybridize to their target, e.g., a BRCA1 and / or BRCA2 nucleic acid molecule provided herein, or a fragment thereof, and to prime the synthesis of extension products, e.g., during PCR or sequencing.

[0157] In some embodiments, an oligonucleotide, e.g., a primer, provided herein comprises 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55,56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80,81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, or more deoxyribonucleotides or ribonucleotides. In some embodiments, an oligonucleotide provided herein comprises at least about 8 deoxyribonucleotides or ribonucleotides. In some embodiments, an oligonucleotide provided herein comprises at least about 10 deoxyribonucleotides or ribonucleotides. In some embodiments, an oligonucleotide provided herein comprises at least about 12 deoxyribonucleotides or ribonucleotides. In some embodiments, an oligonucleotide provided herein comprises at least about 15 deoxyribonucleotides or ribonucleotides. In some embodiments, an oligonucleotide provided herein comprises at least about 20 deoxyribonucleotides or ribonucleotides. In someembodiments, an oligonucleotide provided herein comprises at least about 30 deoxyribonucleotides or ribonucleotides. In some embodiments, an oligonucleotide provided herein comprises between about 10 and about 30 deoxyribonucleotides or ribonucleotides. In some embodiments, an oligonucleotide provided herein comprises between about 10 and about 25 deoxyribonucleotides or ribonucleotides. In some embodiments, an oligonucleotide provided herein comprises between about 10 and about 20 deoxyribonucleotides or ribonucleotides. In some embodiments, an oligonucleotide provided herein comprises between about 10 and about 15 deoxyribonucleotides or ribonucleotides. In some embodiments, an oligonucleotide provided herein comprises between about 12 and about 20 deoxyribonucleotides or ribonucleotides. In some embodiments, an oligonucleotide provided herein comprises between about 17 and about 20 deoxyribonucleotides or ribonucleotides. In some embodiments, the length and nucleotide sequence of an oligonucleotide provided herein is determined according to methods known in the art, e.g., based on factors such as the specific application (e.g., PCR, sequencing library preparation, sequencing), reaction conditions (e.g., buffers, temperature), and the nucleotide composition of the nucleotide sequence of the oligonucleotide or of its target complementary sequence.

[0158] In some embodiments, an oligonucleotide, e.g., a primer, of the disclosure distinguishes a nucleic acid, e.g., a genomic or transcribed nucleic acid, e.g., a cDNA or RNA, having a BRCA1 and / or BRCA2 deletion as described herein, from a reference nucleotide sequence.

[0159] In some embodiments, the oligonucleotide, e.g., the primer, hybridizes to the BRCA1 and / or BRCA2, and a sequence on either side of the BRCA1 and / or BRCA2 (e.g., any of 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleotides on either side of the BRCA1 and / or BRCA2, or any of between 1 and about 5, about 5 and about 10, about 10 and about 15, about 15 and about 20, about 20 and about 25, about 25 and about 30, about 30 and about 35, about 35 and about 40, about 40 and about 45, about 45 and about 50, about 50 and about 55, about 55 and about 60, about 60 and about 65, about 70 and about 75, about 75 and about 80, about 80 and about 85, about 85 and about 90, about 90 and about 95, or about 95 and about 100, or more nucleotides on either side of the BRCA1 and / or BRCA2).Sequencing

[0160] In some embodiments, a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion may be detected in a sample by sequencing, as described herein, such as by RNA sequencing or DNA sequencing. In some embodiments, DNA sequencing can be used todetect a homozygous deletion in BRCA1 and / or BRCA2. In some embodiments, RNA sequencing can be used to detect a homozygous deletion in BRCA1 and / or BRCA2. In some embodiments, a homozygous deletion in BRCA1 and / or BRCA2 may appear in RNA sequencing as a lack of BRCA1 and / or BRCA2 mRNA transcripts. In some embodiments, the sequencing massively parallel sequencing (MPS) technique, whole genome sequencing (WGS), whole exome sequencing, targeted sequencing, direct sequencing, or a Sanger sequencing technique; an optionally wherein the massively parallel sequencing (MPS) technique comprises next- generation sequencing.

[0161] In some embodiments, a homozygous deletion of BRCA1 and or BRCA2 of the disclosure is detected using a sequencing method. Any method of sequencing known in the art can be used to detect a homozygous deletion of BRCA1 and / or BRCA2. Exemplary sequencing methods that may be used to detect a homozygous deletion of BRCA1 and / or BRCA2 include those based on techniques developed by Maxam and Gilbert or Sanger. Automated sequencing procedures may also be used, e.g., including sequencing by mass spectrometry.

[0162] In some embodiments, the sequencing may comprise the sequencing a massively parallel sequencing (MPS) technique, whole genome sequencing (WGS), whole exome sequencing, targeted sequencing, direct sequencing, or a Sanger sequencing technique; an optionally wherein the massively parallel sequencing (MPS) technique comprises nextgeneration sequencing. In some embodiments, the sequencing may be whole genome sequencing, whole exome sequencing, or targeted sequencing. In some embodiments, targeted sequencing may be used to sequence HRR genes, such as BRCA1 and / or BRCA2.

[0163] In some embodiments, a homozygous deletion of BRCA1 and / or BRCA nucleic acid molecules of the disclosure is detected using hybrid capture -based sequencing (hybrid capture -based NGS), e.g., using adaptor ligation-based libraries. See, e.g., Frampton, G.M. et al. (2013) Nat. Biotech. 31:1023-1031. In some embodiments, a homozygous deletion of BRCA1 and / or BRCA2 of the disclosure is detected using next-generation sequencing (NGS). Next-generation sequencing includes any sequencing method that determines the nucleotide sequence of either individual nucleic acid molecules or clonally expanded proxies for individual nucleic acid molecules in a highly parallel fashion (e.g., greater than or equal to 105molecules may be sequenced simultaneously). Next generation sequencing methods suitable for use according to the methods provided herein are known in the art and include, without limitation, massively parallel short-read sequencing, template-based sequencing, pyrosequencing, real-time sequencing comprising imaging the continuous incorporation ofdye-labeling nucleotides during DNA synthesis, nanopore sequencing, sequencing by hybridization, nano-transistor array based sequencing, polony sequencing, scanning tunneling microscopy (STM)-based sequencing, or nanowire-molecule sensor based sequencing. See, e.g., Metzker, M. (2010) Nature Biotechnology Reviews 11:31-46, which is hereby incorporated by reference. Exemplary NGS methods and platforms that may be used to detect a [biomarker] nucleic acid molecule provided herein include, without limitation, the HeliScope Gene Sequencing system from Helicos BioSciences (Cambridge, MA., USA), the PacBio RS system from Pacific Biosciences (Menlo Park, CA, USA), massively parallel short-read sequencing such as the Solexa sequencer and other methods and platforms from Illumina Inc. (San Diego, CA, USA), 454 sequencing from 454 LifeSciences (Branford, CT, USA), Ion Torrent sequencing from ThermoFisher (Waltham, MA, USA), or the SOLiD sequencer from Applied Biosystems (Foster City, CA, USA). Additional exemplary methods and platforms that may be used to detect a [biomarker] nucleic acid molecule provided herein include, without limitation, the Genome Sequencer (GS) FEX System from Roche (Basel, CHE), the G.007 polonator system, the Solexa Genome Analyzer, HiSeq 2500, HiSeq3000, HiSeq 4000, and NovaSeq 6000 platforms from Illumina Inc. (San Diego, CA, USA).Protein Detection Based. Methods

[0164] In some embodiments, the homozygous deletion of BRCA1 and / or BRCA2 from the sample of the disclosure may be detected using methods for detecting a BRCA1 and / or BRCA2 polypeptide. In some embodiments, absence of a BRCA1 and / or BRCA2 polypeptide may indicate a homozygous deletion of BRCA1 and / or BRCA2 respectively.

[0165] In some embodiments, methods of detecting a BRCA1 and / or BRCA2 polypeptide comprise using any method known in the art, such as using antibodies (e.g., an antibody described herein), mass spectrometry (e.g., tandem mass spectrometry), a reporter assay (e.g., a fluorescence-based assay), immunoblots such as a Western blot, immunoassays such as enzyme-linked immunosorbent assays (EEISA), immunohistochemistry, other immunological assays (e.g., fluid or gel precipitin reactions, immunodiffusion, immunoelectrophoresis, radioimmunoassay (RIA), immunofluorescent assays), and analytic biochemical methods (e.g., electrophoresis, capillary electrophoresis, high performance liquid chromatography (HPLC), thin layer chromatography (TLC), hyperdiffusion chromatography).

[0166] In some embodiments, antibodies or antibody fragments that specifically bind to a BRCA1 and / or a BRCA2 polypeptide may be used to detect a homozygous deletion in BRCA1 and / or BRCA2 respectively. The antibody may be of any suitable type of antibody orantibody fragment, including, but not limited to, a monoclonal antibody, a polyclonal antibody, a multi- specific antibody (e.g., a bispecific antibody), or an antibody fragment, so long as the antibody or antibody fragment exhibits a specific antigen binding activity (e.g., binding to a BRCA1 and / or BRCA2 polypeptide of the disclosure, or a portion thereof). In some embodiments, the antibody may be a chimeric or humanized antibody. In certain embodiments, an antibody provided herein is a human antibody.Methods of identifying individuals with cancer with a homozygous deletion in BRCA1 and / or BRCA2

[0167] In some embodiments, the methods disclosed herein may be used to identify individuals with cancer having a homozygous deletion in BRCA1 and / or BRCA2. In some embodiments, the methods may be used to identify an individual having cancer that may benefit from a treatment with a drug effective in treating an HRD positive cancer to the individual, comprising detecting a BRCA1 homozygous deletion in a sample from the individual, wherein the presence of a BRCA1 homozygous deletion in the sample identifies the individual as one who may benefit from the treatment. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0168] In some aspects, provided herein are methods that can be used to diagnose or characterize a homozygous deletion of BRCA1 and / or BRCA2 in cancer, in an individual with cancer. In some embodiments, the methods comprise acquiring knowledge of the presence or absence of a homozygous deletion in BRCA1 and / or BRCA2 in a sample obtained from the individual. In some embodiments, the methods comprise detecting a BRCA1 and / or BRCA2 nucleic acid molecule provided herein in a sample obtained from the individual. In some embodiments, the BRCA1 and / or BRCA2 nucleic acid molecule is detected in a sample obtained from the individual using any method known in the art, such as one or more of the methods of detection of BRCA1 and / or BRCA2 nucleic acid molecules or polypeptides described herein. In some embodiments, the methods further comprise providing a diagnosis or an assessment of the homozygous deletion of BRCA1 and / or BRCA2. In some embodiments, the diagnosis or assessment identifies the presence or absence of the homozygous deletion in BRCA1 and / or BRCA2 in the sample. In some embodiments, the diagnosis or assessment identifies the cancer, as likely to respond to a treatment with a drug effective at treating an HRD positive cancer. In some embodiments, the presence of the homozygous deletion of BRCA1 and / or BRCA2 in the sample identifies the cancer as likely to respond to a treatment with a drug effective at treating an HRD positivecancer. In some embodiments, the sample is a sample described herein. In some embodiments, the sample comprises cells from the cancer or is obtained from cells from the cancer. In some embodiments, the individual has cancer, is suspected of having cancer, is being tested for cancer, is being treated for cancer, or is being tested for a susceptibility to cancer. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0169] In some aspects, provided herein are methods that can be used to screen an individual having cancer, suspected of having cancer, being tested for cancer, being treated for cancer, or being tested for a susceptibility to cancer. In some embodiments, the individual is being treated with a drug effective at treating an HRD positive cancer. In some embodiments, the methods comprise acquiring knowledge of a homozygous deletion of BRCA1 and / or BRCA2 in a sample from the individual. In some embodiments, the methods comprise detecting a homozygous deletion of BRCA1 and / or BRCA2 a sample from the individual. In some embodiments, responsive to acquiring knowledge of a homozygous deletion of BRCA2 in the sample, the individual is predicted to have increased risk of cancer recurrence, aggressive cancer, anti-cancer therapy resistance, or poor prognosis, for example, as compared to an individual whose cancer does not exhibit the homozygous deletion of BRCA1 and / or BRCA2. In some embodiments, responsive to detecting a homozygous deletion of BRCA1 and / or BRCA2 in the sample, the individual is predicted to have increased risk of cancer recurrence, aggressive cancer, anti-cancer therapy resistance, or poor prognosis, for example, as compared to an individual whose cancer does not exhibit the homozygous deletion of BRCA1 and / or BRCA2. In some embodiments, the methods further comprise providing a diagnosis or an assessment. In some embodiments, the diagnosis or assessment identifies the presence or absence of the homozygous deletion of BRCA1 and / or BRCA2 in the sample. In some embodiments, the diagnosis or assessment identifies the individual as being predicted to have increased risk of cancer recurrence, aggressive cancer, anti-cancer therapy resistance, or poor prognosis, for example, as compared to an individual whose cancer does not exhibit the homozygous deletion of BRCA2. In some embodiments, the sample is a sample described herein. In some embodiments, the sample comprises cells from the cancer or is obtained from cells from the cancer, such as ctDNA. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).Methods of predicting survival for an individual with cancer with a homozygous deletion in BRCA1 and / or BRCA2

[0170] Provided herein are methods of predicting survival for individuals with cancer with a homozygous deletion in BRCA1 and / or BRCA1. In some embodiments, survival may be measured as overall survival, such as real-world overall survival (rwOS), time to next treatment (TTNT), or time to treatment discontinuation (TTD). In some embodiments, predicting survival may comprise predicting survival wherein the individual is predicted to have longer survival after treatment with a drug effective at treating an HRD positive cancer when the cancer has a homozygous deletion in BRCA1 and / or BRCA2. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0171] In some embodiments, the method of predicting survival of an individual having cancer treated with a drug effective in treating an HRD positive cancer, comprises acquiring knowledge of a BRCA1 homozygous deletion in a sample from the individual, wherein responsive to the acquisition of said knowledge, the individual is predicted to have longer survival after treatment with the drug effective in treating an HRD positive cancer, as compared to an individual whose cancer does not exhibit the BRCA1 homozygous deletion. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0172] In some aspects, provided herein are methods that can be used to predict survival of an individual having cancer. In some instances, survival may be measured as overall survival, a progression-free survival, an objective response rate, such as objective response rate is a change in prostate specific antigen (PSA) from baseline, a time to tumor progression, a time to treatment failure, a durable complete response, a time to death or a time to next treatment. In some embodiments, the individual is being treated a drug effective at treating an HRD positive cancer, such as drug effective at treating an HRD positive cancer described herein. In some embodiments, the methods comprise acquiring knowledge of a homozygous deletion of BRCA1 and / or BRCA2 in a sample from the individual. In some embodiments, the methods comprise detecting a homozygous deletion of BRCA1 and / or BRCA2 in a sample from the individual. In some embodiments, responsive to acquiring knowledge of a homozygous deletion of BRCA1 and / or BRCA2, the individual is predicted to have longer survival after treatment with a drug effective at treating an HRD positive cancer, such as drug effective at treating an HRD positive cancer provided herein, for example, as compared to an individual whose cancer does not exhibit the homozygous deletion of BRCA1 and / or BRCA2. In someembodiments, responsive to detecting a homozygous deletion of BRCA1 and / or BRCA2 in the sample, the individual is predicted to have longer survival after treatment with a drug effective at treating an HRD positive cancer, such as drug effective at treating an HRD positive cancer provided herein, for example, as compared to an individual whose cancer does not exhibit the homozygous deletion of BRCA1 and / or BRCA2. In some embodiments, the methods further comprise providing a diagnosis or an assessment. In some embodiments, the diagnosis or assessment identifies the presence or absence of the homozygous deletion of BRCA1 and / or BRCA2 in the sample. In some embodiments, the diagnosis or assessment identifies the individual as being predicted to have longer survival after treatment with a drug effective at treating an HRD positive cancer, such as drug effective at treating an HRD positive cancer provided herein, for example, as compared to an individual whose cancer does not exhibit the homozygous deletion of BRCA1 and / or BRCA2. In some embodiments, the sample is a sample described herein. In some embodiments, the sample comprises cells from the cancer or is obtained from cells from the cancer, such as ctDNA. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0173] In some aspects, provided herein are methods that can be used to predict progression- free survival of an individual having cancer. In some aspects, provided herein are methods that can be used to predict an objective response rate, such as objective response rate is a change in prostate specific antigen (PSA) from baseline, of cancer to the treatments described herein for an individual having cancer. In some aspects, provided herein are methods that can be used to predict a time to treatment failure of an individual having cancer. In some aspects, provided herein are methods that can be used to predict a time to tumor progression of cancer in an individual having a cancer. In some aspects, provided herein are methods that can be used to predict a durable complete response rate to a treatment described herein for an individual having cancer. In some aspects, provided herein are methods that can be used to predict a time to death of an individual having cancer. In some aspects, provided herein are methods that can be used to predict a time to next treatment, using a treatment described herein, of an individual having a cancer. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).Methods of selecting treatment for individuals with cancer with a homozygous deletion in BRCA1 and / or BRCA2

[0174] In some embodiments, the methods disclosed herein can be used to select a treatment for individuals with cancer with homozygous deletions in BRCA1 and / or BRCA2. The treatment may be selected in response to characterizing a cancer in an individual and containing a homozygous deletion of BRCA1 and / or BRCA2, as described herein. The method may comprise characterizing the cancer as having a homozygous deletion in BRCA1 and / or BRCA2 and classifying the individual as a candidate to receive a treatment comprising the administration of a drug effective at treating an HRD positive cancer. The individual may be classified as a candidate for receiving a treatment comprising the administration of a drug effective at treating an HRD positive cancer because the drug is likely to be effective at treating the cancer. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC). In some embodiments, the drug effective at treating an HRD positive cancer is a PARPi.

[0175] In some embodiments, the method of selecting treatment for an individual having cancer, comprises acquiring knowledge of a BRCA1 homozygous deletion in a sample from an individual having cancer, wherein responsive to the acquisition of said knowledge: (i) the individual is classified as a candidate to receive treatment with administration of a drug effective in treating an HRD positive cancer and / or (ii) the individual is identified as likely to respond to a treatment that comprises administration of the drug effective in treating an HRD positive cancer. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC). In some embodiments, the drug effective at treating an HRD positive cancer is a PARPi.

[0176] In some aspects, provided herein are methods that can be used to identify an individual having cancer, who may benefit from a treatment comprising administration of a drug effective at treating an HRD positive cancer. In some embodiments, the methods comprise detecting a homozygous deletion of BRCA1 and / or BRCA2 in a sample obtained from the individual. In some embodiments, the methods comprise acquiring knowledge of the presence of a homozygous deletion of BRCA1 and / or BRCA2 in a sample obtained from the individual. In some embodiments, the presence of the homozygous deletion of BRCA1 and / or BRCA2 in the sample identifies the individual as one who may benefit from a treatment with a drug effective at treating an HRD positive cancer. In some embodiments, detection of the homozygous deletion of BRCA1 and / or BRCA2 in the sample identifies the individual as one who may benefit from a treatment with a drug effective at treating an HRD positive cancer.In some embodiments, responsive to knowledge of the homozygous deletion of BRCA1 and / or BRCA2 in the sample, the individual is identified as one who may benefit from a treatment with a drug effective at treating an HRD positive cancer. In some embodiments, the homozygous deletion of BRCA1 and / or BRCA2 is detected using any suitable method known in the art or described herein. In some embodiments, the sample is a sample described herein. In some embodiments, the sample comprises cells from the cancer or is obtained from cells from the cancer, such as ctDNA. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0177] In some embodiments, the methods described herein may be used to identify one or more treatment options for an individual having cancer. In some embodiments, the method may comprise identifying one or more treatment option for an individual having cancer may comprise detecting a BRCA1 homozygous deletion in a sample from the individual; and generating a report comprising one or more treatment options identified for the individual based at least in part on the presence of the BRCA1 homozygous deletion in the sample, wherein the one or more treatment options comprise a drug effective in treating an HRD positive cancer, such as any of the drugs effective at treating an HRD positive cancer described herein.

[0178] Also provided herein are methods that can be used to identify or select a treatment, a therapy, or one or more treatment options for an individual having cancer. In some embodiments, the cancer comprises a homozygous deletion of BRCA1 and / or BRCA2. In some embodiments, the methods comprise detecting the homozygous deletion of BRCA1 and / or BRCA2 in a sample obtained from an individual having cancer. In some embodiments, the homozygous deletion of BRCA1 and / or BRCA2 is detected using any suitable method known in the art or described herein. In some embodiments, detection of a homozygous deletion of BRCA1 and / or BRCA2 in a sample obtained from an individual having cancer described herein, identifies the individual as one who may benefit from a treatment with a drug effective at treating an HRD positive cancer. In some embodiments, the presence of a homozygous deletion of BRCA1 and / or BRCA2 in a sample obtained from an individual having cancer, identifies the individual as one who may benefit from a treatment with a drug effective at treating an HRD positive cancer. In some embodiments, responsive to detection of homozygous deletion of BRCA1 and / or BRCA2 in a sample obtained from an individual having cancer, the individual is classified as a candidate to receive treatment with a drug effective at treating an HRD positive cancer. In some embodiments, responsive to detection of a homozygous deletion of BRCA1 and / or BRCA2 in a sample obtained from anindividual having cancer, the individual is classified or identified as likely to respond to treatment with a drug effective at treating an HRD positive cancer. In some embodiments, the sample is a sample described herein. In some embodiments, the sample comprises cells from the cancer or is obtained from cells from the cancer, such as ctDNA. In some embodiments, the methods further comprise generating a report, e.g., as described herein. In some embodiments, the report comprises a treatment, a therapy, or one or more treatment options identified or selected for the individual, e.g., based at least in part on detection of a homozygous deletion of BRCA1 and / or BRCA2 in the sample. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).In some embodiments, the methods disclosed herein can be used to select a treatment for individuals with cancer with homozygous deletions in BRCA1 and / or BRCA2. In some aspects, the treatment may comprise administration of a drug effective at treating an HRD positive cancer to the individual. In some embodiments, treatment of the disclosure includes one or more therapeutic agents, e.g., for treating cancer associated with a homozygous deletion of BRCA1 and / or BRCA2 as described herein. In some instances, the treatments comprise treatment by administration of a drug effective in treating an HRD positive cancer. In some instances, the drug effective in treating an HRD positive cancer is a drug that targets double strand break repair. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0179] In some embodiments, the drug effective in treating an HRD positive cancer is a PARP inhibitor (e.g., talazoparib, niraparib, rucaparib, olaparib), a RAD51 inhibitor (e.g., RI- 1), or an inhibitor of a DNA damage response kinase, e.g., CHCK1 (e.g., AZD7762), ATM (e.g., KU-55933, KU-60019, NU7026, or VE-821), and ATR (e.g., NU7026).

[0180] In some embodiments, the drug effective at treating an HRD positive cancer is a platinum-based chemotherapeutic agent. In some embodiments, the methods provided herein comprise administering to the individual a platinum-based chemotherapeutic agent, e.g., in combination with another anti-cancer therapy. Platinum-based chemotherapeutic agents are chemotherapeutic agents that comprise an organic compound containing platinum as an integral part of the molecule. In some embodiments, a chemotherapeutic agent is a platinum agent. In some such embodiments, the platinum agent is selected from cisplatin, carboplatin, oxaliplatin, nedaplatin, triplatin tetranitrate, phenanthriplatin, picoplatin, or satraplatin.

[0181] In some aspects, the methods disclosed herein can be used to select a treatment for an individual having cancer. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC). In some embodiments, the treatmentcomprises one or more anti-cancer therapy. In some embodiments, the anti-cancer therapy comprises a radiosensitizer. In some embodiments, the one or more anti-cancer therapies comprise administering to the individual a radiosensitizer, e.g., in combination with another anti-cancer therapy. Exemplary radiosensitizers include hypoxia radiosensitizers such as misonidazole, metronidazole, and trans-sodium crocetinate, a compound that helps to increase the diffusion of oxygen into hypoxic tumor tissue. The radiosensitizer can also be a DNA damage response inhibitor interfering with base excision repair (BER), nucleotide excision repair (NER), mismatch repair (MMR), recombinational repair comprising homologous recombination (HR) and non-homologous end-joining (NHEJ), and direct repair mechanisms. Single strand break (SSB) repair mechanisms include BER, NER, or MMR pathways, while double stranded break (DSB) repair mechanisms consist of HR and NHEJ pathways. Radiation causes DNA breaks that, if not repaired, are lethal. SSBs are repaired through a combination of BER, NER and MMR mechanisms using the intact DNA strand as a template. The predominant pathway of SSB repair is BER, utilizing a family of related enzymes termed poly-(ADP-ribose) polymerases (PARP). Thus, the radiosensitizer can include DNA damage response inhibitors such as PARP inhibitors.

[0182] In some embodiments, the one or more anti-cancer therapies comprises a hormonal therapy. In some instances, treatment comprises administration of a hormonal therapy. In some instances, the hormonal therapy is an androgen receptor blocker. In some instances, the hormonal therapy comprises one or more of abiraterone, enzalutamide, apalutamide, and darolutamide, or any combination thereof. In some embodiments, the one or more anti-cancer therapies comprises a second-generation hormonal therapy

[0183] In some instances, the individual with cancer has received a prior anti-cancer treatment or is being treated with an anti-cancer treatment. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC). In some instances, the prior anti-cancer treatment comprises one or more of a small molecule inhibitor, a chemotherapeutic agent, a cancer immunotherapy, an antibody, a cellular therapy, a nucleic acid, a surgery, a radiotherapy, an anti- angiogenic therapy, an anti-DNA repair therapy, an anti-inflammatory therapy, an anti-neoplastic agent, a growth inhibitory agent, a cytotoxic agent, a vaccine, a small molecule agonist, a virus-based therapy, an antibody-drug conjugate, a recombinant protein, a fusion protein, a natural compound, a peptide, a PROteolysis-Targeting Chimera (PROTAC), or any combination thereof.

[0184] In some instances, the individual with cancer has not received a prior anti-cancer treatment. In some instances, the individual has not received prior anti-cancer treatment suchas one or more of a small molecule inhibitor, a chemotherapeutic agent, a cancer immunotherapy, an antibody, a cellular therapy, a nucleic acid, a surgery, a radiotherapy, an anti-angiogenic therapy, an anti-DNA repair therapy, an anti-inflammatory therapy, an anti- neoplastic agent, a growth inhibitory agent, a cytotoxic agent, a vaccine, a small molecule agonist, a virus-based therapy, an antibody-drug conjugate, a recombinant protein, a fusion protein, a natural compound, a peptide, a PROteolysis-Targeting Chimera (PROTAC), or any combination thereof. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0185] In some embodiments, the methods disclosed herein can be used to select a treatment for individuals with cancer with homozygous deletions in BRCA1 and / or BRCA2. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC). In some instances, the treatment may comprise administration of a therapeutic formulation comprising an anti-cancer therapy provided herein and a pharmaceutically acceptable carrier, excipient, or stabilizer. A formulation provided herein may contain more than one active compound, e.g., an anti-cancer therapy provided herein and one or more additional agents (e.g., anti-cancer agents). Acceptable carriers, excipients, or stabilizers are non-toxic to recipients at the dosages and concentrations employed, and include, for example, one or more of: buffers such as phosphate, citrate, and other organic acids; antioxidants, including ascorbic acid and methionine; preservatives such as octadecyldimethylbenzyl ammonium chloride, hexamethonium chloride, benzalkonium chloride, benzethonium chloride, phenol, butyl or benzyl alcohol, alkyl parabens such as methyl or propyl paraben, catechol, resorcinol, cyclohexanol, 3-pentanol, or m-cresol; low molecular weight polypeptides (e.g., less than about 10 residues); proteins such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates including glucose, mannose, or dextrins; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose or sorbitol; salt-forming counter-ions such as sodium; metal complexes (e.g., Zn-protein complexes); surfactants such as non-ionic surfactants; or polymers such as polyethylene glycol (PEG).

[0186] The active ingredients may be entrapped in microcapsules. Such microcapsules may be prepared, for example, by coacervation techniques or by interfacial polymerization, for example, hydroxy methylcellulose or gelatin-microcapsules and poly-(methylmethacylate) microcapsules, respectively; in colloidal drug delivery systems (for example, liposomes,albumin microspheres, microemulsions, nano-particles and nano-capsules); or in macroemulsions. Such techniques are known in the art.

[0187] Sustained-release compositions may be prepared. Suitable examples of sustained- release compositions include semi-permeable matrices of solid hydrophobic polymers containing an anti-cancer therapy of the disclosure. Such matrices may be in the form of shaped articles, e.g., films, or microcapsules. Examples of sustained-release matrices include polyesters, hydrogels (for example, poly(2-hydroxyethyl-methacrylate), or poly(vinylalcohol)), polylactides, copolymers of L-glutamic acid and y ethyl-L-glutamate, non-degradable ethylene- vinyl acetate, degradable lactic acid-glycolic acid copolymers such as the LUPRON DEPOT™ (injectable microspheres composed of lactic acid-glycolic acid copolymer and leuprolide acetate), and poly-D-(-)-3-hydroxybutyric acid.

[0188] Formulations to be used for in vivo administration are sterile. This is readily accomplished by filtration through sterile filtration membranes or other methods known in the art.

[0189] In some embodiments, the anti-cancer therapy is administered as a monotherapy. In some embodiments, the anti-cancer therapy is administered in combination with one or more additional anti-cancer therapies or treatments. In some embodiments, the one or more additional anti-cancer therapies or treatments include one or more anti-cancer therapies described herein. In some embodiments, the additional anti-cancer therapy comprises one or more of surgery, radiotherapy, chemotherapy, anti-angiogenic therapy, anti-DNA repair therapy, and anti-inflammatory therapy. In some embodiments, the additional anti-cancer therapy comprises an anti-neoplastic agent, a chemotherapeutic agent, a growth inhibitory agent, an anti- angiogenic agent, a radiation therapy, a cytotoxic agent, or combinations thereof. In some embodiments, an anti-cancer therapy may be administered in conjunction with a chemotherapy or chemotherapeutic agent. In some embodiments, the chemotherapy or chemotherapeutic agent is a platinum-based agent (including, without limitation cisplatin, carboplatin, oxaliplatin, and staraplatin). In some embodiments, an anti-cancer therapy may be administered in conjunction with a radiation therapy.

[0190] In some embodiments, discloser herein are drugs for use in any of the methods described herein. In some instances, the drugs for use in any of the methods described herein may comprise administration of a drug effective in treating an HRD positive cancer such as PARPi and a second-generation hormonal therapy such as an androgen receptor blocker.

[0191] Any of the anti-cancer therapies (optionally as monotherapies or in combination with another therapy or treatment) may find use in any of the methods described herein.

[0192] Randomized clinical trials have shown efficacy for inhibition of PARP in men with HRR-deficient mCRPC (de Bono et al., Olaparib for Metastatic Castration-Resistant Prostate Cancer, New England Journal of Medicine; 382:2091-102. (2020)) On the basis of these trials, olaparib was the first PARPi approved by the US Food and Drug Administration (FDA) for men with mCRPC and germline or somatic HRR mutations. Rucaparib was subsequently approved as a single agent for men with mCRPC and BRCA1 / 2 mutations (BRCAalt) previously treated with next generation androgen-receptor directed therapy and prior taxane therapy (Abida et al., Rucaparib in Men With Metastatic Castration-Resistant Prostate Cancer Harboring a BRCA1 or BRCA2 Gene Alteration, Journal of Clinical Oncology;38:3763-72.(2020)) More recently, talazoparib plus enzalutamide and niraparib plus abiraterone have been approved for mCRPC patients with any HRR mutation (talazoparib) or BRCA1 / 2 mutations only (niraparib) based on phase 3 data from TAEAPRO- 2 and MAGNITUDE, respectively (Agarwal et al., Talazoparib plus enzalutamide in men with first-line metastatic castration-resistant prostate cancer (TALAPRO-2): a randomised, placebo-controlled, phase 3 trial, The Lancet 3;402:291-303 (2023) , Chi et al., Detection of BRCA1, BRCA2, and ATM Alterations in Matched Tumor Tissue and Circulating Tumor DNA in Patients with Prostate Cancer Screened in PROfound, Journal of Clinical Oncology 41:3339-51. (2023)).Methods of treatment for individuals with cancer with a homozygous deletion of BRCA1 and / or BRCA2

[0193] Provided herein are methods that can be used to treat an individual with cancer with a homozygous deletion of BRCA1 and / or BRCA2. The treatment may comprise administration of an effective amount of a drug effective at treating an HRD positive cancers, such as any of the drugs effective at treating an HRD positive cancer disclosed here. In some embodiments, the method may be used to delay progression of the cancer. In some embodiments, treating the cancer comprises delaying progression of the tumor in the individual. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0194] In some embodiments, the method of treating or delaying progression of thecancer may comprise, responsive to knowledge of a BRCA1 homozygous deletion in a sample from an individual, administering to the individual an effective amount of a drug effective in treating an HRD positive cancer, such as any of the drugs effective at treating an HRDpositive cancer herein. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0195] Also provided herein are methods that can be used to treat or delay progression of a cancer in an individual. In some embodiments, the individual has cancer comprising a homozygous deletion of BRCA1 and / or BRCA2. In some embodiments, the methods of treating or delaying progression of a cancer of the disclosure in an individual, e.g., comprising a homozygous deletion of BRCA1 and / or BRCA2, comprise administering to the individual a therapeutically effective amount of a drug effective at treating a HRD positive cancer. In some embodiments, the methods of treating or delaying progression of cancer of the disclosure in an individual, e.g., comprising a homozygous deletion of BRCA1 and / or BRCA2, comprise administering to the individual an effective amount a drug effective at treating a HRD positive cancer, responsive to knowledge of the presence of the homozygous deletion of BRCA1 and / or BRCA2 in a sample obtained from the individual. In some embodiments, the sample is a sample described herein. In some embodiments, the sample comprises cells from the cancer or is obtained from cells from the cancer, such as ctDNA. The methods of treatment disclosed herein may include administration of a drug effective at treating an HRD positive cancer such as any of the drugs effective at treating an HRD positive cancer described herein. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0196] Also provided herein are methods that can be used to treat or delay progression of a cancer in an individual. In some embodiments, the individual has a cancer comprising a homozygous deletion of BRCA1 and / or BRCA2. In some embodiments, the methods of treating or delaying progression of a cancer of the disclosure in an individual, e.g., comprising a homozygous deletion of BRCA1 and / or BRCA2, comprise administering to the individual a therapeutically effective amount of a drug effective at treating an HRD positive cancer, such as PARPi. In some embodiments, the methods of treating or delaying progression of a cancer of the disclosure in an individual, e.g., comprising a homozygous deletion of BRCA1 and / or BRCA2, comprise administering to the individual an effective amount of a drug effective at treating an HRD positive cancer, such as PARPi, responsive to knowledge of the presence of the homozygous deletion of BRCA1 and / or BRCA2 in a sample obtained from the individual. In some embodiments, the sample is a sample described herein. In some embodiments, the sample comprises cells from the cancer or is obtained from cells from the cancer, such as ctDNA. The methods of treatment disclosed herein may include administration of a drug effective at treating an HRD positive cancer, such as any of thedrugs effective at treating an HRD positive cancer described herein. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0197] In some aspects, the methods of treatment comprise treatment with a drug effective at treating an HRD positive cancer. In some embodiments, the drug effective at treating an HRD positive cancer may be any of the drugs effective at treating an HRD positive cancer described herein, such as but not limited to PARPi and a platinum agent. In some instances, the treatment may comprise one or more anti-cancer therapies described herein. In some instances, the individual with cancer has received a prior anti-cancer treatment or therapy. In some instances, the individual with cancer has not received a prior anti-cancer treatment or therapy. In some instances, the treatment may comprise administration of a therapeutic formulation comprising an anti-cancer therapy provided herein and a pharmaceutically acceptable carrier, excipient, or stabilizer. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).Methods of generating a report about a cancer with a homozygous deletion in BRCA1 and / or BRCA2

[0198] Comprised herein are methods that can be used to generate a report about a cancer with a homozygous deletion in BRCA1 and / or BRCA2. In some embodiments, the methods provided herein may comprise generating a report and / or provided a report to a party. In some embodiments, the report may comprise the results of characterizing a cancer with a homozygous deletion in BRCA1 and / or BRCA2 in an individual. In some embodiments, the report may comprise one or more treatment options for the individual having the cancer. In some embodiments, the report may comprise a predicted survival for an individual having cancer. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0199] In some embodiments, a report according to the present disclosure comprises information about one or more of: a homozygous deletion of BRCA1 and / or BRCA2 of the disclosure; a cancer of the disclosure, e.g., comprising a homozygous deletion of BRCA1 and / or BRCA2 the disclosure; or a treatment, a therapy, or one or more treatment options for an individual having cancer, such as a cancer of the disclosure (e.g., comprising a homozygous deletion of BRCA1 and / or BRCA2 as described herein). In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0200] In some embodiments, a report according to the present disclosure comprises information about the presence or absence of a homozygous deletion of BRCA1 and / or BRCA2 in a sample obtained from an individual, such as an individual having cancer. In one embodiment, a report according to the present disclosure indicates that a homozygous deletion of BRCA1 and / or BRCA2 is present in a sample obtained from the individual. In one embodiment, a report according to the present disclosure indicates that a homozygous deletion of BRCA1 and / or BRCA2 is not present in a sample obtained from the individual. In one embodiment, a report according to the present disclosure indicates that a homozygous deletion of BRCA1 and / or BRCA2 has been detected in a sample obtained from the individual. In one embodiment, a report according to the present disclosure indicates that a homozygous deletion of BRCA1 and / or BRCA2 has not been detected in a sample obtained from the individual. In some embodiments, the report comprises an identifier for the individual from which the sample was obtained. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0201] In some embodiments, the report includes information on the role of a homozygous deletion in BRCA1 and / or BRCA2, or its wild type counterparts, in disease, such as in cancer. Such information can include one or more of: information on prognosis of a cancer, such as a cancer provided herein, e.g., comprising a homozygous deletion of BRCA1 and / or BRCA2; information on resistance of cancer, such as a cancer provided herein, e.g., comprising a homozygous deletion of BRCA1 and / or BRCA2, to one or more treatments; information on potential or suggested therapeutic options (e.g., such as a treatment with a drug effective at treating an HRD positive cancer provided herein, or a treatment selected or identified according to the methods provided herein); or information on therapeutic options that should be avoided. In some embodiments, the report includes information on the likely effectiveness, acceptability, and / or advisability of applying a therapeutic option (e.g., such as an a treatment with a drug effective at treating an HRD positive cancer provided herein, or a treatment selected or identified according to the methods provided herein) to an individual having a cancer, such as a canvecer provided herein, e.g., comprising a homozygous deletion of BRCA1 and / or BRCA2 and identified in the report. In some embodiments, the report includes information or a recommendation on the administration of a treatment (e.g., such as a treatment with a drug effective at treating an HRD positive cancer provided herein, or a treatment selected or identified according to the methods provided herein). In some embodiments, the information or recommendation includes the dosage of the treatment and / or a treatment regimen (e.g., in combination with other treatments, such as a second therapeuticagent). In some embodiments, the report comprises information or a recommendation for at least one, at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, at least ten, or more treatments. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0202] Also provided herein are methods that can be used to generate a report according to the present disclosure. In some embodiments, a report according to the present disclosure is generated by a method comprising one or more of the following steps: obtaining a sample, such as a sample described herein, from an individual, e.g., an individual having cancer, such as a cancer described herein; detecting a homozygous deletion of BRCA1 and / or BRCA2 in the sample, or acquiring knowledge of the presence of a homozygous deletion of BRCA1 and / or BRCA2 in the sample; and generating a report. In some embodiments, a report generated according to the methods provided herein comprises one or more of: information about the presence or absence of a homozygous deletion of BRCA1 and / or BRCA2 in the sample; an identifier for the individual from which the sample was obtained; information on the role of a homozygous deletion of BRCA1 and / or BRCA2, or its wild type counterparts, in disease (e.g., such as in cancer); information on prognosis, resistance, or potential or suggested therapeutic options (such as an a treatment with a drug effective at treating an HRD positive cancer provided herein, or a treatment selected or identified according to the methods provided herein); information on the likely effectiveness, acceptability, or the advisability of applying a therapeutic option (such as an a treatment with a drug effective at treating an HRD positive cancer provided herein, or a treatment selected or identified according to the methods provided herein) to the individual; a recommendation or information on the administration of a treatment (such as an a treatment with a drug effective at treating an HRD positive cancer provided herein, or a treatment selected or identified according to the methods provided herein); or a recommendation or information on the dosage or treatment regimen of a treatment (such as an anti-cancer therapy provided herein, or a treatment selected or identified according to the methods provided herein), e.g., in combination with other treatments (e.g., a second therapeutic agent). In some embodiments, the report generated is a personalized cancer report. In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).

[0203] A report according to the present disclosure may be in an electronic, web-based, or paper form. The report may be provided to an individual or a patient (e.g., an individual or a patient with cancer, such as a cancer provided herein, e.g., comprising a homozygous deletion of BRCA2), or to an individual or entity other than the individual or patient (e.g., other thanthe individual or patient with the cancer), such as one or more of a caregiver, a physician, an oncologist, a hospital, a clinic, a third party payor, an insurance company, or a government entity. In some embodiments, the report is provided or delivered to the individual or entity within any of about 1 day or more, about 7 days or more, about 14 days or more, about 21 days or more, about 30 days or more, about 45 days or more, or about 60 days or more from obtaining a sample from an individual (e.g., an individual having a cancer). In some embodiments, the report is provided or delivered to an individual or entity within any of about 1 day or more, about 7 days or more, about 14 days or more, about 21 days or more, about 30 days or more, about 45 days or more, or about 60 days or more from detecting a homozygous deletion of BRCA1 and / or BRCA2 in a sample obtained from an individual (e.g., an individual having cancer). In some embodiments, the report is provided or delivered to an individual or entity within any of about 1 day or more, about 7 days or more, about 14 days or more, about 21 days or more, about 30 days or more, about 45 days or more, or about 60 days or more from acquiring knowledge of the presence of a homozygous deletion of BRCA1 and / or BRCA2 in a sample obtained from an individual (e.g., an individual having cancer). In some respects, the cancer is APC. In some aspects, the APC is metastatic castration resistant prostate cancer (mCRPC).The method steps of the methods described herein are intended to include any suitable method of causing one or more other parties or entities to perform the steps unless a different meaning is expressly provided or otherwise clear from the context. Such parties or entities need not be under the direction or control of any other party or entity and need not be located within a particular jurisdiction. Thus, for example, a description or recitation of "adding a first number to a second number" includes causing one or more parties or entities to add the two numbers together. For example, if person X engages in an arm's length transaction with person Y to add the two numbers, and person Y indeed adds the two numbers, then both persons X and Y perform the step as recited: person Y by virtue of the fact that he actually added the numbers, and person X by virtue of the fact that he caused person Y to add the numbers. Furthermore, if person X is located within the United States and person Y is located outside the United States, then the method is performed in the United States by virtue of person X's participation in causing the step to be performed.Methods of determining a tumor fraction in a liquid biopsy

[0204] Certain liquid biopsy assays have been validated to detect loss of BRCA, but even when these tools are used, the lack of detection of loss BRCA should be interpreted as“uninformative” rather than “negative” in the absence of very high ctDNA fraction, whereas properly validated tissue biopsy assays can definitively rule out presence of loss of BRCA. For example, in a post hoc analysis of a mCRPC study, Chi et al., found that while concordance of liquid CGP for detection of nonsense, splice, and frameshift mutations found on matched tissue CGP were high (each >85%), concordance of homozygous loss was only 27% (Chi et al. Clinical Cancer Research 2023;29:81-91 (2023). Liquid biopsy assays as described herein have much higher sensitivity (requiring less ctDNA tumor fraction) to detect mutations, rearrangements, and deletions.

[0205] Accordingly, certain aspects of the present disclosure comprise determining a tumor fraction (TF) in a liquid biopsy collected from an individual with cancer (e.g., APC). In some aspects determining a TF provides an assessment of tumor DNA content in relation to the overall cell-free DNA content in a liquid biopsy sample. Determining a TF is important because at a low TF a liquid biopsy sample may be classified as “uninformative” rather than “negative” when a BRCA1 and / or BRCA2 homozygous deletion is not detected. In such cases, a tissue biopsy sample may be used to properly classify a cancer as positive or negative for a BRCA1 and / or BRCA2 homozygous deletion.

[0206] Methods of determining a tumor fraction may comprise approaches based on: (i) a derived physical relationship between the allele frequency of variants and tumor content that’s dictated by the copy number state of the genomic location(s) of the variant(s) and a tumor average ploidy, and (ii) access to a database collection of individual genomic profile data. In some embodiments, the individual genomic profile data may comprise information germline ploidy for the individual. Exemplary methods for determining a tumor fraction using such approach are described in PCT patent application PCT / US2023 / 070229 and in PCT publication WO2024 / 015973, which are hereby incorporated by reference in their entirety.

[0207] Additional exemplary methods for determining a tumor fraction are described in US patent application publication US2022-0243279-A1 and in PCT publication WO2020 / 236941, which are hereby incorporated by reference in their entirety. Additional exemplary methods for determining a tumor fraction are described in PCT application PCT / US2021 / 038547, PCT publication WO2020 / 271159, and US patent application US 18 / 570,600, which are hereby incorporated by reference in their entirety.Cancers

[0208] In some embodiments, a cancer of the disclosure, is acute lymphoblastic leukemia (“ALL”), acute myeloid leukemia (“AML”), adenocarcinoma, adenocarcinoma of the lung, adrenocortical cancer, adrenocortical carcinoma, anal cancer, appendiceal cancer, B-cell derived leukemia, B-cell derived lymphoma, B-cell lymphoma, bladder cancer, brain cancer, breast cancer (e.g., triple negative breast cancer (TNBC) or non-triple negative breast cancer), cancer of the fallopian tube(s), cancer of the testes, carcinoma, cerebral cancer, cervical cancer, cholangiocarcinoma, choriocarcinoma, chronic myelogenous leukemia, central nervous system (CNS) tumor, CNS cancer, colon cancer, colorectal cancer (e.g., colon adenocarcinoma), diffuse intrinsic pontine glioma (DIPG), diffuse large B cell lymphoma (“DLBCL”), embryonal rhabdomyosarcoma (ERMS), endometrial cancer, epithelial cancer, epithelial neoplasm, thymoma, esophageal cancer, Ewing’s sarcoma, eye cancer (e.g., uveal melanoma), eyelid cancer, follicular lymphoma (“FL”), gall bladder cancer, gastric cancer, gastrointestinal cancer, glioblastoma, polycythemia vera, glioblastoma multiforme, glioma (e.g., lower grade glioma), gullet cancer, head and neck cancer, a hematological cancer, hepatocellular cancer, hepatocellular carcinoma, Hodgkin’s lymphoma (HL), a heavy chain disease, intestinum rectum cancer, renal cancer, kidney cancer (e.g., kidney clear cell cancer, kidney chromophobe cancer, kidney clear cell cancer, kidney papillary cancer), large B-cell lymphoma, large intestine cancer, laryngeal cancer, leucosis, leukemia, liver cancer, lung cancer (e.g., lung adenocarcinoma, or non-small cell lung cancer), lymphoma, mammary gland cancer, melanoma (e.g., metastatic malignant melanoma), Hodgkin’s disease, Waldenstrom’s macroglobulinemia, Merkel cell carcinoma, mesothelioma, monocytic leukemia, multiple myeloma, myeloma, myogenic sarcoma, nasopharyngeal cancer, neuroblastic-derived CNS tumor (e.g., neuroblastoma (NB)), neuroma, astrocytoma, pilocytic astrocytoma, anaplastic astrocytoma, medulloblastoma, craniopharyngioma, ependymoma, pinealoma, hemangioblastoma, acoustic neuroma, oligodendroglioma, meningioma, vestibular schwannoma, adenoma, metastatic brain tumor, spinal tumor, non-Hodgkin’s lymphoma (NHL), oral cancer, oral cavity cancer, osteosarcoma, ovarian cancer, ovarian carcinoma, pancreatic adenocarcinoma, pancreatic cancer, peritoneal cancer, pheochromocytoma, primary mediastinal B-cell lymphoma, primary peritoneal cancer, prostate cancer (e.g., hormone refractory prostate adenocarcinoma), rectal cancer (rectum carcinoma), relapsed or refractory classic Hodgkin’s Lymphoma (cHL), salivary gland cancer (e.g., salivary gland tumor), skin cancer, small cell lung cancer, small intestine cancer, soft tissue sarcoma, squamous carcinoma, squamous cellcarcinoma (e.g., squamous cell carcinoma of the anogenital region, squamous cell carcinoma of the anus, squamous cell carcinoma of the cervix, squamous cell carcinoma of the esophagus, squamous cell carcinoma of the head and neck (SCHNC), squamous cell carcinoma of the lung, squamous cell carcinoma of the penis, squamous cell carcinoma of the vagina, or squamous cell carcinoma of the vulva), stomach cancer, T-cell derived leukemia, T-cell lymphoma, testicular cancer, testicular tumor, thymic cancer, thyroid cancer (thyroid carcinoma), tongue cancer, tunica conjunctiva cancer, urinary bladder cancer, urothelial cell carcinoma, uterine cancer (e.g., uterine endometrial cancer or uterine sarcoma such as uterine carcinosarcoma), uterine endometrial cancer, uterus cancer, vaginal cancer, vulvar cancer, or Wilms’ tumor.

[0209] In some embodiments, a cancer of the disclosure is a hematologic cancer (e.g., a hematologic malignancy), such as diffuse large B cell lymphoma (“DLBCL”), Hodgkin’s lymphoma (“HL”), Non-Hodgkin’s lymphoma (“NHL”), follicular lymphoma (“FL”), acute myeloid leukemia (“AML”), acute lymphoblastic leukemia (“ALL”), multiple myeloma (“MM”), acute lymphoblastic B-cell leukemia, acute lymphoblastic T-cell leukemia, acute myeloblastic leukemia , acute promyelocytic leukemia (“APL”), acute monoblastic leukemia, acute erythroleukemic leukemia, acute megakaryoblastic leukemia, acute myelomonocytic leukemia, acute nonlymphocyctic leukemia, acute undifferentiated leukemia, chronic myelocytic leukemia (“CML”), chronic lymphocytic leukemia (“CLL”), or hairy cell leukemia. In some embodiments, a hematologic cancer of the disclosure, is an acute or a chronic leukemia, such as a lymphoblastic, myelogenous, lymphocytic, or myelocytic leukemia. In some embodiments, a hematologic cancer of the disclosure is a lymphoma (e.g., Hodgkin’s lymphoma, such as relapsed or refractory classic Hodgkin’s Lymphoma (cHL), a non-Hodgkin’s lymphoma, a diffuse large B-cell lymphoma, or a precursor T-lymphoblastic lymphoma), a lymphoepithelial carcinoma, or a malignant histiocytosis.

[0210] In some embodiments, a cancer of the disclosure, is a solid tumor (e.g., a solid malignancy), such as fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, lymphangioendotheliosarcoma, synovioma, mesothelioma, Ewing’s tumor, leiomyosarcoma, rhabdomyosarcoma, osteosarcoma, colon cancer, colorectal cancer, kidney cancer, pancreatic cancer, bone cancer, breast cancer, ovarian cancer, prostate cancer, esophageal cancer, stomach cancer, oral cancer, nasal cancer, throat cancer, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinoma, cystadenocarcinoma, medullary carcinoma,bronchogenic carcinoma, renal cell carcinoma, hepatoma, bile duct carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilms tumor, cervical cancer, uterine cancer, testicular cancer, non-small cell lung cancer (NSCLC), small cell lung carcinoma, bladder carcinoma, lung cancer, epithelial carcinoma, skin cancer, melanoma, neuroblastoma (NB), or retinoblastoma.

[0211] In certain embodiments, a cancer of the disclosure, is a cancer of the adrenal glands (such as neuroblastoma), bladder cancer (such as urothelial (transitional cell) carcinoma), brain cancer (such as anaplastic astrocytoma or glioblastoma), bone cancer (such as osteosarcoma), bone marrow cancer (such as B-cell acute leukemia (B-ALL) or multiple myeloma), breast cancer (such as invasive ductal carcinoma), head and neck cancer (such as adenocarcinoma, mucoepidermoid carcinoma, squamous cell carcinoma), lymph node cancer, lung cancer (e.g., mucoepidermoid carcinoma, sarcoma, small cell undifferentiated carcinoma, adenocarcinoma, adenosquamous carcinoma, large cell carcinoma, large cell neuroendocrine carcinoma, non-small cell lung carcinoma, non-small cell lung carcinoma not otherwise specified, or squamous cell carcinoma), female reproductive cancer (e.g., cancer of the fallopian tubes such as fallopian tube serous carcinoma; ovarian cancer, such as epithelial carcinoma, epithelial carcinoma not otherwise specified, high grade serous carcinoma, low grade serous carcinoma, serous carcinoma; and uterine cancer, such as carcinosarcoma, endometrial adenocarcinoma, endometrial adenocarcinoma not otherwise specified, papillary serous endometrial adenocarcinoma, leiomyosarcoma, sarcoma, sarcoma not otherwise specified, or smooth muscle tumor of uncertain malignant potential (STUMP)), gallbladder cancer (such as adenocarcinoma), cancer of the gastroesophageal junction (such as adenocarcinoma), lymph node cancer (such as anaplastic large cell lymphoma, B-cell lymphoma, B-cell lymphoma not otherwise specified, diffuse large B cell lymphoma, nonHodgkin’s lymphoma, non-Hodgkin’s lymphoma not otherwise specified), colon cancer (such as adenocarcinoma), colorectal cancer, skin cancer (such as melanoma or squamous cell carcinoma), small intestine cancer (adenocarcinoma), soft tissue cancer (such as Ewing sarcoma, fibrosarcoma, histiocytosis, histiocytosis not otherwise specified, juvenile xanthogranuloma or non-Langerhans cell histiocytosis, inflammatory myofibroblastic tumor, leiomyosarcoma, neurofibroma, neuroblastoma, sarcoma not otherwise specified, sarcoma, undifferentiated sarcoma, or an undifferentiated soft tissue cancer), pancreatic cancer (such as carcinoma, carcinoma not otherwise specified, ductal adenocarcinoma, or mucinous cystadenocarcinoma), prostate cancer (such as acinar adenocarcinoma), pericardium cancer (such as mesothelioma), peritoneum cancer (such as mesothelioma), salivary gland cancer(such as carcinoma or carcinoma not otherwise specified), stomach cancer (such as adenocarcinoma, adenocarcinoma not otherwise specified, or diffuse type cancer), kidney cancer (such as renal cell carcinoma or renal cell carcinoma not otherwise specified), thyroid cancer (such as carcinoma, carcinoma not otherwise specified, or papillary carcinoma), or a cancer of unknown primary origin (such as adenocarcinoma, carcinoma, carcinoma not otherwise specified, leiomyosarcoma, malignant neoplasm, malignant neoplasm not otherwise specified, melanoma, myoepithelial carcinoma, squamous cell carcinoma (SCC), or undifferentiated neuroendocrine carcinoma).Advanced prostate cancer (APC)

[0212] Certain aspects of the present disclosure relate to APC. APC is an advanced form of prostate cancer. APC may be advanced and metastatic. APC originates in the cells of the prostate and then has the potential to spread to other parts of the body. Several variants of APC exist, including but not limited to metastatic castration-sensitive prostate cancer (mCSPC), metastatic hormone- sensitive prostate cancer, or metastatic castration-resistant prostate cancer (mCRPC).

[0213] Metastatic castration resistant prostate cancer (mCRPC) is a genetically heterogenous disease and 20-30% of men with mCRPC harbor somatic or germline loss of function mutations in genes involved in DNA damage repair (Pritchard et al., Inherited DNA-Repair Gene Mutations in Men with Metastatic Prostate Cancer, New England Journal of Medicine 375:443-53. (2016), Robinson et al. Integrative Clinical Genomics of Advanced Prostate Cancer. Cell;161:1215-28 (2015)). Deleterious mutations in homologous recombination repair (HRR) genes are often associated with distant metastases and worse overall survival (Castro et al., Germline BRCA Mutations Are Associated With Higher Risk of Nodal Involvement, Distant Metastasis, and Poor Survival Outcomes in Prostate Cancer. Journal of Clinical Oncology, 2013;31:1748-57).Samples

[0214] The disclosed methods and systems may comprise detecting the presence of a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in a sample. In some instances, the sample may be any of a variety of samples (also referred to herein as specimens) comprising nucleic acids (e.g., DNA or RNA) that are collected from a subject (e.g., a patient). Examples of a sample include, but are not limited to, a tumor sample, a tissue sample, a biopsy sample (e.g., a tissue biopsy, a liquid biopsy, or both), a blood sample (e.g.,a peripheral whole blood sample), a blood plasma sample, a blood serum sample, a lymph sample, a saliva sample, a sputum sample, a urine sample, a gynecological fluid sample, a circulating tumor cell (CTC) sample, a cerebral spinal fluid (CSF) sample, a pericardial fluid sample, a pleural fluid sample, an ascites (peritoneal fluid) sample, a feces (or stool) sample, or other body fluid, secretion, and / or excretion sample (or cell sample derived therefrom). In certain instances, the sample may be frozen sample or a formalin-fixed paraffin-embedded (FFPE) sample.

[0215] In some instances, the sample may be collected by tissue resection (e.g., surgical resection), needle biopsy, bone marrow biopsy, bone marrow aspiration, skin biopsy, endoscopic biopsy, fine needle aspiration, oral swab, nasal swab, vaginal swab or a cytology smear, scrapings, washings or lavages (such as a ductal lavage or bronchoalveolar lavage), etc.

[0216] In some instances, the sample is a liquid biopsy sample, and may comprise, e.g., whole blood, blood plasma, blood serum, urine, stool, sputum, saliva, or cerebrospinal fluid. In some instances, the sample may be a liquid biopsy sample and may comprise circulating tumor cells (CTCs). In some instances, the sample may be a liquid biopsy sample and may comprise cell-free DNA (cfDNA), circulating tumor DNA (ctDNA), or any combination thereof.

[0217] In some instances, the sample may comprise one or more premalignant or malignant cells. Premalignant, as used herein, refers to a cell or tissue that is not yet malignant but is poised to become malignant. In certain instances, the sample may be acquired from a solid tumor, a soft tissue tumor, or a metastatic lesion. In certain instances, the sample may be acquired from a hematologic malignancy or pre-malignancy. In other instances, the sample may comprise a tissue or cells from a surgical margin. In certain instances, the sample may comprise tumor-infiltrating lymphocytes. In some instances, the sample may comprise one or more non-malignant cells. In some instances, the sample may be, or is part of, a primary tumor or a metastasis (e.g., a metastasis biopsy sample). In some instances, the sample may be obtained from a site (e.g., a tumor site) with the highest percentage of tumor (e.g., tumor cells) as compared to adjacent sites (e.g., sites adjacent to the tumor). In some instances, the sample may be obtained from a site (e.g., a tumor site) with the largest tumor focus (e.g., the largest number of tumor cells as visualized under a microscope) as compared to adjacent sites (e.g., sites adjacent to the tumor).

[0218] The disclosed methods and systems may be applied to the analysis of homozygous deletions in BRCA1 and / or BRCA2 in nucleic acids extracted from any of variety of tissuesamples (or disease states thereof), e.g., solid tissue samples, soft tissue samples, metastatic lesions, or liquid biopsy samples. Examples of tissues include, but are not limited to, connective tissue, muscle tissue, nervous tissue, epithelial tissue, and blood. Tissue samples may be collected from any of the organs within an animal or human body. Examples of human organs include, but are not limited to, the brain, heart, lungs, liver, kidneys, pancreas, spleen, thyroid, mammary glands, uterus, prostate, large intestine, small intestine, bladder, bone, skin, etc.

[0219] In some instances, the nucleic acids extracted from the sample may comprise deoxyribonucleic acid (DNA) molecules. Examples of DNA that may be suitable for analysis by the disclosed methods include, but are not limited to, genomic DNA or fragments thereof, mitochondrial DNA or fragments thereof, cell-free DNA (cfDNA), and circulating tumor DNA (ctDNA). Cell-free DNA (cfDNA) is comprised of fragments of DNA that are released from normal and / or cancerous cells during apoptosis and necrosis and circulate in the blood stream and / or accumulate in other bodily fluids. Circulating tumor DNA (ctDNA) is comprised of fragments of DNA that are released from cancerous cells and tumors that circulate in the blood stream and / or accumulate in other bodily fluids.

[0220] In some instances, the methods disclosed herein may be used to detect a homozygous deletion of BRCA1 and / or BRCA2 from a liquid biopsy sample. In some instances, the methods comprise determining a TF of the liquid biopsy sample according to the methods described herein. In some instances, the sample is a liquid biopsy with a tumor fraction (TF) of less than about 40% and greater than or equal to about 1%. In some instances, the liquid biopsy has a TF of less than about 35% and greater than or equal to about 1%, less than about 30% and greater than or equal to about 1%, less than about 25% and greater than or equal to about 1%, less than about 20% and greater than or equal to about 1%, less than about 15% and greater than or equal to about 1%, or less than about 10% and greater than or equal to about 1%. In some instances, the liquid biopsy has a TF of about 40%, 30%, 25%, 20%, 15%, 10%, 5% or 1%.

[0221] In some instances, the methods disclosed herein may be used to detect a homozygous deletion of BRCA1 and / or BRCA2 from a liquid biopsy sample. In some instances, the methods comprise determining a TF of the liquid biopsy sample according to the methods described herein. In some instances, the sample is a liquid biopsy with a tumor fraction (TF) of greater than or equal to about 20%. In some instances, the liquid biopsy has a TF greater than or equal to about 25%, greater than or equal to about 30%, greater than or equal to about 35%, greater than or equal to about 40%, greater than or equal to about 45%, or and greaterthan or equal to about 50%. In some instances, the liquid biopsy has a TF of about 25%, 30%, 35%, 40%, 45%, 50%, 65% or 70%.

[0222] In some instances, DNA is extracted from nucleated cells from the sample. In some instances, a sample may have a low nucleated cellularity, e.g., when the sample is comprised mainly of erythrocytes, lesional cells that contain excessive cytoplasm, or tissue with fibrosis. In some instances, a sample with low nucleated cellularity may require more, e.g., greater, tissue volume for DNA extraction.

[0223] In some instances, the nucleic acids extracted from the sample may comprise ribonucleic acid (RNA) molecules. Examples of RNA that may be suitable for analysis by the disclosed methods include, but are not limited to, total cellular RNA, total cellular RNA after depletion of certain abundant RNA sequences (e.g., ribosomal RNAs), cell-free RNA (cfRNA), messenger RNA (mRNA) or fragments thereof, the poly(A)-tailed mRNA fraction of the total RNA, ribosomal RNA (rRNA) or fragments thereof, transfer RNA (tRNA) or fragments thereof, and mitochondrial RNA or fragments thereof. In some instances, RNA may be extracted from the sample and converted to complementary DNA (cDNA) using, e.g., a reverse transcription reaction. In some instances, the cDNA is produced by random-primed cDNA synthesis methods. In other instances, the cDNA synthesis is initiated at the poly(A) tail of mature mRNAs by priming with oligo(dT)-containing oligonucleotides. Methods for depletion, poly(A) enrichment, and cDNA synthesis are well known to those of skill in the art.

[0224] In some instances, the sample may comprise a tumor content (e.g., comprising tumor cells or tumor cell nuclei), or a non-tumor content (e.g., immune cells, fibroblasts, and other non-tumor cells). In some instances, the tumor content of the sample may constitute a sample metric. In some instances, the sample may comprise a tumor content of at least 5-50%, 10- 40%, 15-25%, or 20-30% tumor cell nuclei. In some instances, the sample may comprise a tumor content of at least 5%, at least 10%, at least 20%, at least 30%, at least 40%, or at least 50% tumor cell nuclei. In some instances, the percent tumor cell nuclei (e.g., sample fraction) is determined (e.g., calculated) by dividing the number of tumor cells in the sample by the total number of all cells within the sample that have nuclei.

[0225] In some instances, as noted above, the sample comprises nucleic acid (e.g., DNA, RNA (or a cDNA derived from the RNA), or both), e.g., from a tumor or from normal tissue. In certain instances, the sample may further comprise a non-nucleic acid component, e.g., cells, protein, carbohydrate, or lipid, e.g., from the tumor or normal tissue.Kits

[0226] Also provided herein are kits that may be used to characterized a cancer with a homozygous deletion of BRCA1 and / or BRCA2 as described herein. In some embodiments, characterizing a cancer may comprise detecting a homozygous deletion of BRCA1 and / or BRCA2 of the disclosure in a sample from an individual with a cancer. In some embodiments, the kit may comprise reagents for processing a sample, such as a tissue biopsy or a liquid biopsy from an individual with a cancer. In some aspects, the kit may comprise reagents for processing a liquid biopsy with a low tumor fraction. In some aspects, the kit may comprise instructions for selecting a treatment for an individual with a cancer with a homozygous deletion in BRCA1 and / or BRCA2. In some embodiments, the treatment may comprise administration of a drug effective at treating an HRD positive cancer. In some aspects, the kit may comprise instructions for predicting survival of an individual with a cancer treated with a drug effecting at treating an HRD positive cancer as described herein. In some embodiments, the kit may comprise instructions for generating a report about a cancer with a homozygous deletion in BRCA1 and / or BRCA2. In some embodiments the cancer is APC. In some embodiments, the APC is metastatic castration resistant prostate cancer (mPRPC). In some embodiments, the APC is hormone sensitive prostate cancer. In some embodiments, the APC is de novo metastatic prostate cancer.

[0227] In some embodiments, a kit provided herein comprises a reagent for detecting a homozygous deletion of BRCA1 and / BRCA2 provided herein. In some embodiments, the kit comprises a reagent for detecting a wild-type counterpart of a BRCA1 and or BRCA2 nucleic acid molecule provided herein. In some embodiments, the reagent comprises one or more oligonucleotides, primers, probes, or baits of the present disclosure capable of hybridizing to a nucleic acid molecule with a deletion in BRCA1 and / or BRCA2 provided herein, or to a wild-type counterpart of a BRCA1 and / or BRCA2 nucleic acid molecule provided herein. In some embodiments, the reagent comprises one or more oligonucleotides, primers, probes, or baits of the present disclosure capable of distinguishing a nucleic acid molecule with a deletion in BRCA1 and / or BRCA2 provided herein from a wild-type counterpart of the BRCA1 and / or BRCA2 nucleic acid molecule provided herein. In some embodiments, the kit is for use according to any method of detecting deletions in BRCA1 and or BRCA2 nucleic acid molecules known in the art or described herein, such as sequencing, PCR, in situ hybridization methods, a nucleic acid hybridization assay, an amplification-based assay, a PCR-RFLP assay, real-time PCR, sequencing, next-generation sequencing, a screening analysis, FISH, spectral karyotyping, MFISH, comparative genomic hybridization, in situhybridization, sequence-specific priming (SSP) PCR, HPLC, and mass-spectrometric genotyping. In some embodiments, a kit provided herein further comprises instructions for detecting a homozygous deletion of BRCA1 and / or BRCA2 of the disclosure, e.g., using one or more oligonucleotides, primers, probes, or baits of the present disclosure.EXEMPLARY EMBODIMENTS

[0228] The following exemplary embodiments are representative of some aspects of the invention:

[0229] Embodiment 1. A method of identifying an individual having advanced prostate cancer (APC) that may benefit from a treatment of a drug effective in treating an HRD positive cancer to the individual, said method comprising detecting a BRCA1 homozygous deletion in a sample from the individual, wherein the presence of a one BRCA1 homozygous deletion in the sample identifies the individual as one who may benefit from the treatment.

[0230] Embodiment 2. A method of selecting treatment for an individual having advanced prostate cancer (APC), comprising acquiring knowledge of a BRCA1 homozygous deletion in a sample from an individual having APC, wherein responsive to the acquisition of said knowledge: (i) the individual is classified as a candidate to receive treatment with a drug effective in treating an HRD positive cancer and / or (ii) the individual is identified as likely to respond to a treatment with the drug effective in treating an HRD positive cancer.

[0231] Embodiment 3. A method of treating or delaying progression of advanced prostate cancer (APC) comprising, responsive to knowledge of a BRCA1 homozygous deletion in a sample from an individual, administering to the individual an effective amount of a drug effective in treating an HRD positive cancer.

[0232] Embodiment 4. A method of identifying one or more treatment options for an individual having advanced prostate cancer (APC), the method comprising:(a) detecting a BRCA1 homozygous deletion in a sample from the individual; and(b) generating a report comprising one or more treatment options identified for the individual based at least in part on the presence of the BRCA1 homozygous deletion in the sample, wherein the one or more treatment options comprise a drug effective in treating an HRD positive cancer.

[0233] Embodiment 5. A method of predicting survival of an individual having advanced prostate cancer (APC) treated with a drug effective in treating an HRD positive cancer, comprising acquiring knowledge of a BRCA1 homozygous deletion in a sample from the individual, wherein responsive to the acquisition of said knowledge, the individual ispredicted to have longer survival after treatment with the drug effective in treating an HRD positive cancer, as compared to an individual whose APC does not exhibit the BRCA1 homozygous deletion.

[0234] Embodiment 6. A method of diagnosing / assessing a BRCA1 homozygous deletion, the method comprising:(a) detecting in a sample from an individual with advanced prostate cancer (APC); and(b) providing a diagnosis of the BRCA1 homozygous deletion.

[0235] Embodiment 7. The method of embodiment 6, further comprising recommending treatment options including administration of a HRR pathway drug.

[0236] Embodiment 8. The method of any one of embodiments 1-7, wherein the BRCA1 homozygous deletion comprises a homozygous deletion of the full BRCA1 gene.

[0237] Embodiment 9. The method of any of embodiments 1-8, wherein the sample comprises a tissue biopsy sample or a liquid biopsy.

[0238] Embodiment 10. The method of embodiment 9, wherein the sample is a tissue biopsy and compromises a tumor biopsy.

[0239] Embodiment 11. The method of embodiment 9, wherein the sample is a liquid biopsy sample and comprises blood, serum, plasma, cerebrospinal fluid, sputum, stool, urine, or saliva.

[0240] Embodiment 12. The method of any of embodiments 1-11, wherein the sample comprises cells and / or nucleic acids from the APC.

[0241] Embodiment 13. The method of embodiment 12, wherein the sample comprises mRNA, DNA, circulating tumor DNA (ctDNA), cell-free DNA, cell-free RNA from the cancer, or any combination thereof.

[0242] Embodiment 14. The method of embodiment 12, wherein the sample is a liquid biopsy sample and comprises circulating tumor cells (CTCs).

[0243] Embodiment 15. The method of embodiment 12, wherein the sample is a liquid biopsy sample and comprises cell-free DNA (cfDNA), circulating tumor DNA (ctDNA), or any combination thereof.

[0244] Embodiment 16. The method of embodiments 2 or 5, wherein acquiring knowledge of the BRCA1 homozygous deletion comprises detecting the BRCA1 homozygous deletion.

[0245] Embodiment 17. The method of any one of embodiments 1-16 , wherein the BRCA1 homozygous deletion is detected in the sample by one or more of: a nucleic acid hybridization assay, an amplification-based assay, a polymerase chain reaction-restrictionfragment length polymorphism (PCR-RFLP) assay, real-time PCR, a screening analysis, fluorescence in situ hybridization (FISH), spectral karyotyping, multicolor FISH (mFISH), comparative genomic hybridization, in situ hybridization, sequence-specific priming (SSP) PCR, high-performance liquid chromatography (HPLC), mass-spectrometric genotyping, or sequencing.

[0246] Embodiment 18. The method of any one of embodiments 1-17, wherein the advanced prostate cancer (APC) is metastatic castration resistant prostate cancer (mPRPC).

[0247] Embodiment 19. The method of any one of embodiments 1-17, wherein the advanced prostate cancer (APC) is hormone sensitive prostate cancer.

[0248] Embodiment 20. The method of any one of embodiments 1-17, wherein the advanced prostate cancer (APC) is de novo metastatic prostate cancer.

[0249] Embodiment 21. A method of identifying if an individual having advanced prostate cancer (APC) that may benefit from a treatment with a drug effective in treating an HRD positive cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to about 20% or less than about 20% and a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion is detected in the liquid biopsy sample, identifying the individual as one who may benefit from the treatment of a drug effective in treating an HRD positive cancer.

[0250] Embodiment 22. A method of identifying if an individual having advanced prostate cancer (APC) that may benefit from a treatment with a drug effective in treating an HRD positive cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to about 20% and a BRCA1 homozygous deletion and a BRCA2 homozygous deletion is not detected in the liquid biopsy sample, identifying the individual as one who may not receive the most benefit from the treatment with a drug effective in treating an HRD positive cancer.

[0251] Embodiment 23. A method of selecting treatment for an individual having advanced prostate cancer (APC), said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual;detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to 20% or less than about 20% and a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion is detected in the liquid biopsy sample from the individual, classifying the individual as a candidate to receive treatment with a drug effective in treating an HRD positive cancer.

[0252] Embodiment 24. A method of selecting treatment for an individual having advanced prostate cancer (APC), said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to 20% and BRCA1 homozygous deletion and BRCA2 homozygous deletion is not detected in the liquid biopsy sample, classifying the individual as one who may not receive the most benefit from treatment with a drug effective in treating an HRD positive cancer.

[0253] Embodiment 25. A method of treating or delaying progression of advanced prostate cancer (APC) in an individual said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to 20% or less than about 20%, and a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion is detected in the liquid biopsy sample from the individual, administering to the individual an effective amount of a drug effective in treating an HRD positive cancer.

[0254] Embodiment 26. A method of treating or delaying progression of advanced prostate cancer (APC) in an individual said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to 20% and a BRCA1 homozygous deletion and a BRCA2 homozygous deletion is not detected in the liquid biopsy sample from the individual,administering to the individual an effective amount of a drug other than a drug effective in treating an HRD positive cancer.

[0255] Embodiment 27. A method of predicting survival of an individual having advanced prostate cancer (APC) treated with a drug effective in treating an HRD positive cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to about 20% or less than about 20%, and a BRCA1 homologous deletion and / or a BRCA2 homozygous deletion is detected in the liquid biopsy sample from the individual, the individual is predicted to have longer survival after treatment with a drug effective in treating an HRD positive cancer, as compared to an individual whose APC does not exhibit the BRCA1 homozygous deletion and / or BRCA2 homozygous deletion.

[0256] Embodiment 28. A method of predicting survival of an individual having advanced prostate cancer (APC) treated with administration of a drug effective in treating an HRD positive cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual, the individual is predicted to have shorter survival after treatment with the drug effective in treating an HRD positive cancer, as compared to an individual whose APC does exhibit the BRCA1 homozygous deletion and / or BRCA2 homozygous deletion.

[0257] Embodiment 29. A method of identifying if an individual having cancer that may benefit from a treatment with a drug effective in treating an HRD positive cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample;if the TF in the liquid biopsy sample is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detecting in the liquid biopsy sample from the individual, further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if BRCA1 homozygous deletion and / or BRCA2 homozygous deletion are detected in the tissue biopsy sample from the individual, identifying the individual as one who may benefit from the treatment with a drug effective in treating an HRD positive cancer.

[0258] Embodiment 30. A method of identifying if an individual having cancer that may benefit from a treatment with a drug effective in treating an HRD positive cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample from the individual, if the TF in the liquid biopsy sample is less than about 20% and BRCA1 homozygous deletion and BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual; further detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if a BRCA1 homozygous deletion and a BRCA2 homozygous deletion are not detected in the tissue biopsy sample, identifying the individual as one who may not benefit from the treatment with a drug effective in treating an HRD positive cancer; or if a BRCA1 homozygous deletion or a BRCA2 homozygous deletion is detected in the tissue biopsy sample, identifying the individual as one who may benefit from the treatment with a drug effective in treating an HRD positive cancer.

[0259] Embodiment 31. A method of selecting treatment for an individual having cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual; further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; andif BRCA1 homozygous deletion and / or BRCA2 homozygous deletion are detected in the tissue biopsy sample from the individual, classifying the individual as a candidate to receive treatment with a drug effective in treating an HRD positive cancer.

[0260] Embodiment 32. A method of selecting treatment for an individual having cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual; further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if a BRCA1 homozygous deletion and a BRCA2 homozygous deletion are not detected in the tissue biopsy sample, classifying the individual as not a candidate to receive treatment with a drug effective in treating an HRD positive cancer, or if a BRCA1 homozygous deletion or a BRCA2 homozygous deletion is detected in the tissue biopsy sample, classifying the individual as a candidate to receive treatment with a drug effective in treating an HRD positive cancer.

[0261] Embodiment 33. A method of treating or delaying progression of advanced cancer in an individual said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual; further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual, and if a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion are detected in the tissue biopsy sample from the individual, administering to the individual an effective amount of a drug effective in treating an HRD positive cancer.

[0262] Embodiment 34. A method of treating or delaying progression of cancer in an individual said method comprising:determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual, further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if a BRCA1 homozygous deletion and a BRCA2 homozygous deletion are not detected in the tissue biopsy sample, administering to the individual an effective amount of a drug other than a drug effective in treating an HRD positive cancer, or if a BRCA1 homozygous deletion or a BRCA2 homozygous deletion is detected in the tissue biopsy sample, administering to the individual a drug effective in treating an HRD positive cancer.

[0263] Embodiment 35. A method of predicting survival of an individual having cancer treated with administration of a drug effective in treating an HRD positive cancer, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual, further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion are detected in the tissue biopsy sample the individual is predicted to have longer survival after treatment with the administration of the drug effective in treating an HRD positive cancer, as compared to an individual whose cancer does not exhibit the BRCA1 homozygous deletion and / or BRCA2 homozygous deletion.

[0264] Embodiment 36. A method of predicting survival of an individual having cancer treated with administration of a drug effective in treating an HRD positive cancer, said method comprising:determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual, further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual; and if a BRCA1 homozygous deletion and a BRCA2 homozygous deletion are not detected in the tissue biopsy sample the individual is not predicted to have longer survival after treatment with the administration of the drug effective in treating an HRD positive cancer, as compared to an individual whose cancer does not exhibit the BRCA1 homozygous deletion and / or BRCA2 homozygous deletion, or if a BRCA1 homozygous deletion or a BRCA2 homozygous deletion is detected in the tissue biopsy sample, the individual is predicted to have longer survival after treatment with the administration of the drug effective in treating an HRD positive cancer, as compared to an individual whose cancer does not exhibit the BRCA1 homozygous deletion and / or BRCA2 homozygous deletion.

[0265] Embodiment 37. The method of any of embodiments 18-34, wherein the liquid biopsy sample comprises blood, serum, plasma, cerebrospinal fluid, sputum, stool, urine, or saliva.

[0266] Embodiment 38. The method of any one of embodiments 29-37, wherein the tissue sample is a tissue biopsy and comprises a tumor biopsy.

[0267] Embodiment 39. The method of any one of embodiments 21-38, wherein the liquid biopsy sample comprises mRNA, DNA, circulating tumor DNA (ctDNA), cell-free DNA, cell-free RNA from the cancer, or any combination thereof.

[0268] Embodiment 40. The method of embodiment 39, wherein the liquid biopsy sample comprises circulating tumor cells (CTCs).

[0269] Embodiment 41. The method of embodiment 39, wherein the sample comprises cell- free DNA (cfDNA), circulating tumor DNA (ctDNA), or any combination thereof.

[0270] Embodiment 42. The method of any one of embodiments 21-41, wherein the BRCA1 homozygous deletion and / or the or BRCA2 homozygous deletion is detected in the liquid biopsy sample by one or more of: a nucleic acid hybridization assay, an amplification-based assay, a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay, real-time PCR, a screening analysis, fluorescence in situ hybridization (FISH), spectralkaryotyping, multicolor FISH (mFISH), comparative genomic hybridization, in situ hybridization, sequence- specific priming (SSP) PCR, high-performance liquid chromatography (HPLC), mass-spectrometric genotyping, or sequencing.

[0271] Embodiment 43. The method of any one of embodiments 29-41, wherein the BRCA1 homozygous deletion and / or the or BRCA2 homozygous deletion is detected in the tissue biopsy sample by one or more of: a nucleic acid hybridization assay, an amplification-based assay, a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay, real-time PCR, a screening analysis, fluorescence in situ hybridization (FISH), spectral karyotyping, multicolor FISH (mFISH), comparative genomic hybridization, in situ hybridization, sequence- specific priming (SSP) PCR, high-performance liquid chromatography (HPLC), mass-spectrometric genotyping, or sequencing.

[0272] Embodiment 44. The method of any one of embodiments 29-43, wherein the cancer is advanced prostate cancer (APC).

[0273] Embodiment 45. The method of any one of embodiments 21-28 and 44, wherein the advanced prostate cancer (APC) is metastatic castration resistant prostate cancer (mPRPC).

[0274] Embodiment 46. The method of any one of embodiments 21-28 and 44, wherein the advanced prostate cancer (APC) is hormone sensitive prostate cancer.

[0275] Embodiment 47. The method of any one of embodiments 21-28 and 44, wherein the advanced prostate cancer (APC) is de novo metastatic prostate cancer.

[0276] Embodiment 48. The method of any one of embodiments 1-5 and 7-47, wherein the drug effective in treating an HRD positive cancer is a Poly ADP-ribose polymerase inhibitor (PARPi).

[0277] Embodiment 49. The method of embodiment 44, wherein the PARPi is selected from a group consisting of niraparib, olaparib, talazoparib, or rucaparib.

[0278] Embodiment 50. The method of any one of embodiments 1-5 and 7-47, wherein the drug effective in treating an HRD positive cancer is a platinum agent.

[0279] Embodiment 51. The method of any one of embodiments 1-5 and 7-47, wherein the drug effective in treating an HRD positive cancer is a drug that targets double strand break repair.

[0280] Embodiment 52. The method of any one of embodiments 1-5 and 7-51 wherein the treatment comprises administration of a hormonal therapy.

[0281] Embodiment 53. The method of embodiment 52, wherein the hormonal therapy is an androgen receptor blocker.

[0282] Embodiment 54. The method of any one of embodiments 52-53, wherein the hormonal therapy comprises one or more of abiraterone, enzalutamide, apalutamide, and darolutamide, or any combination thereof.

[0283] Embodiment 55. The method of any of embodiments 1-54, wherein the individual has received a prior anti-cancer treatment or is being treated with an anti-cancer treatment.

[0284] Embodiment 56. The method of clause 55, wherein the prior anti-cancer treatment comprises one or more of a small molecule inhibitor, a chemotherapeutic agent, a cancer immunotherapy, an antibody, a cellular therapy, a nucleic acid, a surgery, a radiotherapy, an anti-angiogenic therapy, an anti-DNA repair therapy, an anti-inflammatory therapy, an anti- neoplastic agent, a growth inhibitory agent, a cytotoxic agent, a vaccine, a small molecule agonist, a virus-based therapy, an antibody-drug conjugate, a recombinant protein, a fusion protein, a natural compound, a peptide, a PROteolysis-Targeting Chimera (PROTAC), or any combination thereof.

[0285] Embodiment 57. The method of any of embodiments 1-54, wherein the individual has not received a prior anti-cancer treatment.

[0286] Embodiment 58. The method of any one of embodiments 5-17, 27, 28, and 35-57, wherein the survival is an overall survival, a progression-free survival, a disease-free survival, an objective response rate, a time to tumor progression, a time to treatment failure, a durable complete response, a time to death, or a time to next treatment.

[0287] Embodiment 59. The method of embodiment 58, wherein the objective response rate is a change in prostate specific antigen (PSA) from baseline.

[0288] Embodiment 60. The method of any one of embodiments 1-20, further comprising obtaining the sample from the individual.

[0289] Embodiment 61. The method of any one of embodiments 21-59, further comprising obtaining the liquid biopsy sample from the individual.

[0290] Embodiment 62. The method of any one of embodiments 29-59, further comprising obtaining the tissue biopsy sample from the individual.

[0291] Embodiment 63. The method of any one of embodiment 17 or embodiment 42 or embodiment 43, wherein the sequencing comprises a massively parallel sequencing (MPS) technique, whole genome sequencing (WGS), whole exome sequencing, targeted sequencing, direct sequencing, or a Sanger sequencing technique; an optionally wherein the massively parallel sequencing (MPS) technique comprises next- generation sequencing.

[0292] Embodiment 64. The method of embodiment 17 or embodiment 63 wherein the sequencing comprises:(a) providing a plurality of nucleic acid molecules obtained from the sample, wherein the plurality of nucleic acid molecules comprises a mixture of tumor nucleic acid molecules and non-tumor nucleic acid molecules;(b) optionally, ligating one or more adapters onto one or more nucleic acid molecules from the plurality of nucleic acid molecules;(c) amplifying nucleic acid molecules from the plurality of nucleic acid molecules;(d) optionally, capturing nucleic acid molecules from the amplified nucleic acid molecules, wherein the captured nucleic acid molecules are captured from the amplified nucleic acid molecules by hybridization to one or more bait molecules; and(e) sequencing, by a sequencer, the captured nucleic acid molecules to obtain a plurality of sequence reads corresponding to one or more genomic loci within a subgenomic interval in the sample.

[0293] Embodiment 65. The method of embodiment 42 or embodiment 63 wherein the sequencing comprises:(a) providing a plurality of nucleic acid molecules obtained from the liquid biopsy sample, wherein the plurality of nucleic acid molecules comprises a mixture of tumor nucleic acid molecules and non-tumor nucleic acid molecules;(b) optionally, ligating one or more adapters onto one or more nucleic acid molecules from the plurality of nucleic acid molecules;(c) amplifying nucleic acid molecules from the plurality of nucleic acid molecules;(d) optionally, capturing nucleic acid molecules from the amplified nucleic acid molecules, wherein the captured nucleic acid molecules are captured from the amplified nucleic acid molecules by hybridization to one or more bait molecules; and(e) sequencing, by a sequencer, the captured nucleic acid molecules to obtain a plurality of sequence reads corresponding to one or more genomic loci within a subgenomic interval in the liquid biopsy sample.

[0294] Embodiment 66. The method of embodiment 43 or embodiment 63 wherein the sequencing comprises:(a) providing a plurality of nucleic acid molecules obtained from the tissue biopsy sample, wherein the plurality of nucleic acid molecules comprises a mixture of tumor nucleic acid molecules and non-tumor nucleic acid molecules;(b) optionally, ligating one or more adapters onto one or more nucleic acid molecules from the plurality of nucleic acid molecules;(c) amplifying nucleic acid molecules from the plurality of nucleic acid molecules;(d) optionally, capturing nucleic acid molecules from the amplified nucleic acid molecules, wherein the captured nucleic acid molecules are captured from the amplified nucleic acid molecules by hybridization to one or more bait molecules; and(e) sequencing, by a sequencer, the captured nucleic acid molecules to obtain a plurality of sequence reads corresponding to one or more genomic loci within a subgenomic interval in the tissue biopsy sample.

[0295] Embodiment 67. The method of any one of embodiments 64-66, wherein the adapters comprise one or more of amplification primer sequences, flow cell adapter hybridization sequences, unique molecular identifier sequences, substrate adapter sequences, or sample index sequences.

[0296] Embodiment 68. The method of any one of embodiments 64-67, wherein amplifying nucleic acid molecules comprises performing a polymerase chain reaction (PCR) technique, a non-PCR amplification technique, or an isothermal amplification technique.

[0297] Embodiment 69. The method of any one of embodiments 64-68, wherein the one or more bait molecules comprise one or more nucleic acid molecules, each comprising a region that is complementary to a region of a captured nucleic acid molecule.

[0298] Embodiment 70. The method of embodiment 69, wherein the one or more bait molecules each comprise a capture moiety.

[0299] Embodiment 71. The method of embodiment 70, wherein the capture moiety is biotin.

[0300] Embodiment 72. The method of any of embodiments 1-71, wherein the individual is a patient.

[0301] Embodiment 73. The method of any of embodiments 1- 72, wherein the individual is a human.

[0302] Embodiment 74. A drug effective in treating an HRD positive cancer for use in a method of treating or delaying progression of advanced prostate cancer (APC), wherein the method comprises administering the drug effective in treating an HRD positive cancer to an individual, wherein a BRCA1 homozygous deletion is detected in a sample obtained from the individual.

[0303] Embodiment 75. A drug effective in treating an HRD positive cancer for use in the manufacture of a medicament for treating or delaying progression of advanced prostate cancer (APC), wherein the medicament is to be administered to an individual, wherein a BRCA1 homozygous deletion has been detected in a sample obtained from the individual.

[0304] Embodiment 76. A drug effective in treating an HRD positive cancer for use in a method of treating or delaying progression of clinically advanced prostate cancer (APC),wherein the method comprises administering the drug effective in treating an HRD positive cancer to an individual, wherein a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion is detected in a liquid biopsy sample obtained from the individual; and wherein the same is a liquid biopsy has a TF of greater than or equal to about 20%.

[0305] Embodiment 77. A drug effective in treating an HRD positive cancer for use in a method of treating or delaying progression of clinically advanced prostate cancer (APC), wherein the method comprises administering the drug effective in treating an HRD positive cancer to an individual, wherein a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion is detected in a tissue biopsy sample obtained from the individual; wherein a liquid biopsy sample has been collected from the individual and the liquid biopsy sample has a TF of less than about 20%.

[0306] Embodiment 78. A drug effective in treating an HRD positive cancer for use in the manufacture of a medicament for treating or delaying progression of advanced prostate cancer (APC), wherein the medicament is to be administered to an individual, wherein a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion has been detected in a sample obtained from the individual; and wherein the same is a liquid biopsy has a TF of greater than or equal to about 20%.

[0307] Embodiment 79. A drug effective in treating an HRD positive cancer for use in the manufacture of a medicament for treating or delaying progression of advanced prostate cancer (APC), wherein the medicament is to be administered to an individual, wherein a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion is detected in a tissue biopsy sample obtained from the individual; wherein a liquid biopsy sample has been collected from the individual and the liquid biopsy sample has a TF of less than about 20%.EXAMPLES

[0308] The invention will be more fully understood by reference to the following examples. They should not, however, be construed as limiting the scope of the invention. It is understood that the examples and embodiments described herein are for illustrative purposes only and that various modifications or changes in light thereof will be suggested to persons skilled in the art and are to be included within the spirit and purview of this application and scope of the appended claims.Example 1 - Durable Benefit from PARP Inhibitors for Patients with Metastatic Prostate Cancer in Routine Practice: Biomarker Associations and. Implications for Optimal Clinical NGS Testing

[0309] This example provides a non-limiting example of the use real world data to show poly ADP-ribose polymerase inhibitors (PARPi) in patients with metastatic castration resistant prostate cancer (mCRPC) and BRCA1 or BRCA2 alterations (BRCAalt).

[0310] Study Design and Patient Selection. Patient-level de-identified clinical data from a deidentified nationwide (U.S. -based) cancer clinico-genomic database between January 2011 and September 2023 were extracted. The de-identified clinical data originated from approximately 280 US cancer clinics (-800 sites of care) was obtained. Retrospective longitudinal clinical data were derived from electronic health records (EHR), comprising patient-level structured and unstructured data, curated via technology-enabled abstraction of clinical notes and radiology / pathology reports, which were linked to genomic data derived from FMI testing by de-identified, deterministic matching (Singal et al., Association of Patient Characteristics and Tumor Genomics With Clinical Outcomes Among Patients With Non-Small Cell Lung Cancer Using a Clinicogenomic Database, JAMA;321:1391-9 (2019)). Clinical data included demographics, clinical and laboratory features, timing of treatment exposure, treatment progression, and survival.

[0311] Patients had received a first line single-agent PARPi (olaparib, rucaparib, or niraparib) in the mCRPC setting and genomic profiling by tissue or liquid biopsy. Patients were grouped by genes with deleterious alterations detected: BRCA1 / 2 (BRCAalt), ATM, other HRR (ATR, ATRX, BAP1, BARD1, BRIP1, CHEK1, CHEK2, CDK12, FANCA, FANCL, MRE11, RAD51B, RAD51C, RAD51D, RAD54E, PAEB2), or no HRR (if negative by tissue or liquid profiling). Patients BRCA status were further subclassified into BRCA homozygous loss and all other BRCA mutations.

[0312] Comprehensive Genomic Profiling. Hybrid capture-based next-generation sequencing (NGS) assays were performed on patient tumor or blood specimens in a Clinical Eaboratory Improvement Amendments (CEIA)-certified, College of American Pathologists (CAP)- accredited laboratory (FMI, Cambridge, MA). FoundationOne® or FoundationOne®CDx assays interrogated all exons from a minimum of 324 cancer related genes, plus select introns from minimum of 28 genes for rearrangement detection. Samples were evaluated for alterations as previously described (Frampton et al., Development and validation of a clinical cancer genomic profiling test based on massively parallel DNA sequencing, Nat Biotechnol 31:1023-31 (2013)). FoundationOne®Eiquid CDx is a validated, FDA-approved next-generation sequencing panel assay interrogating 324 cancer-related genes, and reporting single-nucleotide variants, insertions / deletions, genomic rearrangements, copy number amplifications, as well as losses.

[0313] Cell-free DNA was extracted from whole blood and CGP performed using hybridization-captured, adaptor ligation-based libraries (Woodhouse et al., Clinical and analytical validation ofFoundationOne Liquid CDx, a novel 324-Gene cfDNA-based comprehensive genomic profiling assay for cancers of solid tumor origin, PLoS One;15:e0237802- (2020)).

[0314] HRDsig is a machine learning algorithm developed to predict genomic scarring consistent with homologous recombination deficiency (Moore et al., Pan-Cancer Analysis of Copy-Number Features Identifies Recurrent Signatures and a Homologous Recombination Deficiency Biomarker to Predict Poly (ADP-Ribose) Polymerase Inhibitor Response, JCO Precis Oncol :e2300093 (2023).). HRDsig scores range from 0 to 1 and a cutoff of 0.7 was pre-specified based on prior analyses demonstrating 90% sensitivity of detecting biallelic BRCA alterations in BRCA-driven tumors.

[0315] Predominant genetic ancestry was assigned by training a random forest classifier to distinguish the 5 ancestral superpopulations of the 1000 Genomes Project (Carrot-Zhang et al., Comprehensive Analysis of Genetic Ancestry and its Molecular Correlated in Cancer, Cancer Cell, 37(5):639-54 e 6 (2020); Auton et al. A global reference for human genetic variation, Nature 526:68-74 (2015)), as previously described (Mata et al., Disparities According to Genetic Ancestry in the Use of Precision Oncology Assays, New England Journal of Medicine; 388:281-3 (2023)). Then the closest match for each specimen was determined.

[0316] Time to Event Outcomes. Time to Next Treatment (TTNT) was calculated from treatment start date until the start of next treatment line (due to any cause) or death. Patients not yet reaching next treatment line or death were censored at date of last clinical visit or structured activity. Time to PARPi discontinuation (TTD) was calculated from treatment start date until the cessation of PARPi use (due to any cause), or death. Patients still on PARPi were censored at date of last clinical visit or structured activity. Real- world overall survival (rwOS) was calculated from start of 1stline treatment to death from any cause, and patients with no record of mortality were right censored at the date of last clinic visit. Because patients cannot enter the database until a CGP report is delivered, rwOS risk intervals were left truncated to the date of CGP report to account for immortal time. Truncation independence with censoring was evaluated with Kendall’s tau, with p < 0.05 consideredacceptable. Flatiron Health database mortality information is a composite derived from 3 sources: documents within the EHR, Social Security Death Index, and a commercial death dataset mining data from obituaries and funeral homes, with validations reported in comparison to the National Death Index (Zhang et al., Validation analysis of a composite real-world mortality endpoint for patients with cancer in the United States, Health Serv Res ;56:1281— 7 (2021)).

[0317] Prostate Specific Antigen (PSA) Response. A line of PARPi therapy was eligible for PSA response assessment if a PSA result was available within 60 days prior to PARPi initiation and a separate PSA result was available 1-180 days after. If multiple results were available, respective values most proximal to treatment initiation and 12 weeks on treatment were used. PSA response calculated as: (on treatment PSA - baseline PSA) / (baseline PSA + 0.01).

[0318] Statistical Analysis. Differences in time-to-event outcomes were assessed with the log-rank test and Cox proportional hazard (PH) models. Chi-square tests and Wilcoxon rank sum tests were used to assess differences between groups of categorical and continuous variables, respectively. Multiple comparison adjustments were not performed; p-values are reported to quantify the strength of association for biomarker and each outcome, not for null hypothesis significance testing, and interpretations adopted broadly considering consistency of multiple outcome measures in concert (TTNT, TTD, PSA Response, rwOS) with no outcome measure standing on its own.

[0319] Missing values were handled by simple imputation with expected values determined using random forests with the R package ‘missForest.’ In subsequent analyses, imputed values were treated identically to measured values. R software was used for all statistical analyses.

[0320] Characteristics of Analysis Cohort. 4,559 patients were identified with mCRPC and CGP data, of which 445 unique patients were treated with single agent PARPi and met inclusion criteria (FIG. 1). There were 224 patients who had tumor tissue CGP and 231 who had liquid biopsy CGP. Of these patients, 170 (38.2%) had BRCAalt, 110 (24.7%) had ATM mutations, 109 (24.5%) had other HRR mutations, and 56 (12.6%) had no known HRR alteration (Table 1).Table 1 : Participant demographics and clinical criteria

[0321] BRCAalt was more highly represented in the tissue CGP cohorts compared to liquid CGP (46.7% vs. 30.3% respectively, p<0.001). Timing of PARPi therapy was variable: it was administered as a first-line treatment in 10.1% (45 patients), second-line in 28.3% (126 patients), third-line in 23.1% (103 patients), and fourth-line in 38.4% (171 patients). This was similar between men with tissue or liquid biopsy genomic profiling. The vast majority of patients (n=390; 87.6%) had undergone prior novel hormonal therapy (NHT), 225 (50.6%) received prior taxane therapy, and 29 (6.5%) received prior platinum therapy. Most patients were treated with Olaparib (95.5%), and the remainder received rucaparib (4.0%) and niraparib (0.5%). Differences between molecular subgroups are listed in Table 2.Table 2: Patient differences between molecular subgroups

[0322] Outcomes of PARPi Stratified by Molecular Subgroup. TTNT, TTD, and rwOS, stratified by molecular subgrouping, was then assessed. A granular analysis of gene alterations associated with outcomes of individual patients is shown with swimmer plots for TTNT for patients with tissue biopsy CGP (FIGS. 2A-2C) and liquid biopsy CGP (FIGS. 2D-2F). In the tissue cohort, several patients with the longest TTNT (>12 months) were patients with BRCA1 or BRCA2 homozygous loss, which were less commonly detected in the liquid cohort. Several patients with non-BRCA alterations (FANCA, RAD51C, PALB2,CHEK2, and BRIP1) had greater than 12 months TTNT. A small group of patients were administered PARPi without detection of any HRR subgroup.

[0323] Using the no HRR group as a reference, multivariable analyses for TTNT, TTD, and rwOS in the tissue and liquid CGP cohorts was performed, adjusting for clinical factors that might confound time-to-event analyses (e.g., pre-tx PSA, ECOG performance status, etc.). In the tissue CGP cohort, BRCAalt was associated with more favorable TTNT compared to men with no HRR alterations (n=100, HR: 0.31, 95%CI: 0.19 - 0.51, p<0.001). ATM mutations (n=35, HR: 0.99, 95%, CI: 0.55 - 1.76, p=0.968) and other HRR mutations (n=51, HR: 0.77, 95%CI: 0.45 - 1.30, p=0.324) were not associated with more favorable TTNT compared to the no HRR cohort (FIG. 3A). rwOS was also more favorable in the BRCAalt group relative to no HRR (n=100, HR: 0.47, 95% CI: 0.27 - 0.81, p=0.006). ATM (n=35, HR: 0.75, 95%CI: 0.39 - 1.44, p=0.386) and other HRR (n=51, HR: 1.11, 95%CI: 0.62 - 2.00, p=0.728) cohorts were not statistically different than no HRR (FIG. 3B).

[0324] In the liquid CGP cohort, adjusted TTNT was more favorable in BRCAalt subgroup (n=70, HR: 0.46, 95%CI: 0.26 - 0.82, p=0.009) and similarly in the other HRR subgroup (n=58, HR: 0.51, 95% CI: 0.29 - 0.88, p=0.017) compared to no HRR. ATM was not statistically different than no HRR (FIG. 3C). Additionally, rwOS was similarly favorable in BRCAalt (n=70, HR: 0.50, 95%CI: 0.25 - 1.01, p=0.053) and other HRR subgroups (n=58, HR: 0.53, 95%CI: 0.27 - 1.04, p=0.064) compared to no HRR. No difference was observed in the ATM subgroup (n=75, HR: 0.95, 95%CI: 0.51 - 1.78, p=0.877) (FIG. 3D).

[0325] Multivariable models for TTD for both tissue and liquid cohorts was very similar to TTNT and is shown in FIGS. 4A-4B. Unadjusted, univariable associations between HRR groups can be found in FIGS. 5A-5F.

[0326] HRD Signature. Evaluation of the association of genomic scars consistent with homologous repair deficiency with outcomes on PARPi were completed. Compared to HRDsig(-), HRDsig(+) was associated with more favorable TTNT (aHR 0.44, 95%CI: 0.32 - 0.62, p<0.0001), TTD (aHR 0.48, 95%CI: 0.35 - 0.67, p<0.0001), and rwOS (aHR 0.49, 95%CI: 0.34-071, p<0.001) (FIGS. 6A- 61).

[0327] PSA Response. Baseline and 12-week PSA kinetics were evaluable in 198 patients (n=95 with tissue biopsy and n=103 with liquid biopsy). In the tissue CGP cohort, 16 (34.7%) of patients with BRCAalt had a >50% decline in PSA at 12-weeks after PARPi initiation. In contrast, 0 patients in the ATM alteration subgroup, 1 patient in the other-HRR subgroup (4.2%), and 0 patients in the no HRR subgroup had a PSA decline of 50% or greater (FIG. 7A). Several of the deepest responders were notably had BRCAloss. In the liquid CGPcohort, 7 patients (26.9%) with BRCAalt had a 12-week 50% PSA response (FIG. 7B). In this cohort 0 patients with ATM alterations, 3 patients (12.5%) with other HRR alterations, and 2 patients (14.0%) with no HRR alterations had a 50% PSA response. Patient characteristics were broadly similar between those with an evaluable PSA response (n=198) and those unevaluable for PSA response (n=247) (Table 3)).Table 3: Patient characteristics comparing between patients with an evaluable and. unevaluable PSA response.

[0328] Outcomes stratified by BRCA alteration type. The association between homozygous BRCA loss detected on tissue CGP with more favorable outcomes to single agent PARPi compared to other BRCAalt was assessed. For men with homozygous BRCA loss, there was more favorable TTNT (aHR 0.38, 95%CI: 0.2 - 0.72, p=0.003), TTD (aHR 0.43, 95%CI: 0.23 - 0.8, p=0.007), and rwOS (aHR 0.42, 95%CI: 0.2-0.84, p=0.015) compared to other BRCAalt (FIGS. 8A-11B). To further evaluate that BRCA homozygous loss are challenging to detect in liquid biopsy in the absence of high levels of ctDNA tumor fraction, the prevalence of detection of BRCA homozygous loss in FDA-approved CGP assays for tissue and liquid biopsy, regardless of the treatment received was explored. 2,667 tissue specimens profiled with FoundationOne® CDx were identified. Among these, 3.1% have BRCA homozygous loss. Among 5,064 liquid biopsy specimens profiled with FoundationOne®Liquid CDx, 1.0% have homozygous BRCA loss. Considering only the specimens with a minimum of 1% TF and 20% TF, the prevalence is 2% and 3.5%, respectively (FIG. 8C). A visualization of the TF associated with BRCA alteration types in liquid biopsy is also shown (FIG. 8D). Time to treatment discontinuation associations were similar to TTNT (FIG. 9) and multivariable assessments adjusting for standard baseline prognostic factors estimate similar magnitude of effect as seen in univariable assessments (FIGS. 10A-10C).

[0329] Data showed that men with mCRPC treated with single agent PARPi were most likely to have favorable outcomes and durable benefit in the setting of homozygous BRCA1 or BRCA2 loss compared to those in other biomarker defined subgroups (other BRCAalterations, ATM alterations, other HRR alterations, or no HRR alterations). These findings were robust across multiple endpoints — PSA response, TTNT, TTD and rwOS — and are in line with reports showing PARPi benefit is largely restricted to men with deleterious BRCA1 / 2 alterations, and smaller cohorts reporting extreme benefit in the presence of BRCA loss (Mateo et al., Olaparib for the Treatment of Patients With Metastatic Castration- Resistant Prostate Cancer and Alterations in BRCA1 and / or BRCA2 in the PROfound Trial, Journal of Clinical Oncology; 42:571-83 (2023).) These data provide support that complete loss of BRCA1 or BRCA2 confer inability to evolve resistance for PARPi through reversion mutations.

[0330] The possibility that a subset of liquid CGP patients had clonal hematopoiesis (CH) interference, which may also have impacted the interpretation of PARPi outcomes was considered. Liquid biopsy CGP assay sensitivity was dependent upon the amount of circulating tumor DNA (ctDNA) shed and measured by TF on liquid biopsy assays.(Woodhouse et al. Clinical and analytical validation of FoundationOne Liquid CDx, a novel 324-Gene cfDNA-based comprehensive genomic profiling assay for cancers of solid tumor origin. PLoS One ;15:e0237802- (2020); Reichert et al., Prognostic value of plasma circulating tumor DNA fraction across four common cancer types: a real-world outcomes study, Annals of Oncology ;34: 111-20. (2023)) It was observed that the prevalence of BRCA loss detection was similar between TBx and LBx when ctDNA TF on was very high (20% or greater) (FIGS 8A-8D). This was consistent with the differences observed between the cohort of outcomes associated with TBx and LBx detection of HRR alterations (FIGS. 2A- 2G). At face value, without consideration of confounders, in men who underwent liquid biopsy CGP, the BRCAalt group did not have more favorable outcomes compared to the other HRR alterations. Without being bound to this theory, this may be due to BRCA homozygous losses only being detectable in liquid biopsy with a level of ctDNA TF that most patients in routine practice will not have, that potential extreme responders were under- represented in the liquid biopsy cohort presented here.

[0331] The data support the use of tumor tissue for CGP for discovery of actionable HRR alterations in mCRPC. However, biopsy is not always feasible, and biopsies of bone metastases in particular have a high rate of failure of CGP (Abida et al., Prospective Genomic Profiling of Prostate Cancer Across Disease States Reveals Germline and Somatic Alterations That May Affect Clinical Decision Making, JCO Precis Oncol 2017:1-16.(2017)). The possibility that HRR alterations are largely truncal and would be present inarchival tissue biopsy was assessed. It was observed that similar prevalence of BRCaloss in specimens originating from the prostate and from other tissues (FIG. 8C).

[0332] PARPi therapy has not shown a consistent clinical benefit for patients with ATM mutations (Marshall et al., Differential Response to Olaparib Treatment Among Men with Metastatic Castration-resistant Prostate Cancer Harboring BRCA1 or BRCA2 Versus ATM Mutations, Eur Urol 2019;76:452-8(2019)), and results demonstrated herein are consistent with prior reports. It was observed that the small fraction of men with non-BRCA HRR and durable TTNT and TTD had alterations in other HRR genes (RAD51B, FANCA, BRIP1) as well.

[0333] A scar-based HRD signature assessed using CGP of the tissue cohort was evaluated. Scar-based copy number signatures are approved in ovarian cancer (Ray-Coquard et al., Olaparib plus Bevacizumab as First-Line Maintenance in Ovarian Cancer, New England Journal of Medicine; 381 :2416-28 (2019).) and are beginning to be used for prospective PARPi trial enrollment in other tumor types (NCT06065059), with the potential advantage of identifying both alterations in HRR genes leading to an HRD phenotype as well as non- genomic mechanisms of HRD such as BRCA promoter methylation. In this study, HRDsig positivity was associated with improved PARPi outcomes in the overall prostate cancer cohort. In sub-analysis, favorable trends for HRDsig were observed; however, the potential added benefit of HRDsig in BRCAalt and BRCAwt cohorts was less clear. The BRCAwt analysis was underpowered (n = 9 HRDsig+) and evaluation of additional cohorts is of interest for future study.

[0334] Observational and / or retrospective analyses are more prone to false discovery than prospective randomized trials, due to multiple hypothesis testing and potential imbalances between groups. Rigorous adjustment of prognostic factors was performed to reduce potential imbalances between groups that might confound survival comparisons. However, these adjustments do not account for all potential imbalances, and there are some biases that are known that must be carefully considered in order to generalize results, such as BRCA loss being easier to detect with tissue biopsy. Clinical annotation and reporting can vary between laboratories. Therefore, the results from the study may or may not be generalizable to biomarker performance of all NGS platforms.

[0335] Conclusions. In this analysis of real- world effectiveness of PARPi, patients with BRCA alterations identified on tissue biopsy CGP had more favorable outcomes across all endpoints relative to other patient subgroups in the setting of PARPi monotherapy. Patients with homozygous BRCA loss had significantly more favorable outcomes relative to all otherBRCA alterations. While BRCAloss can be detected with validated liquid biopsy assays, most patients with mCRPC do not have high enough ctDNA TF to rule out presence of BRCAloss using liquid biopsy. Our results suggest that tissue CGP should be prioritized when clinically feasible for the detection of alterations in HRR related genes, especially for those associated with durable benefit from PARPi.

Claims

CLAIMSWhat is claimed is:

1. A method of treating or delaying progression of advanced prostate cancer (APC) comprising, responsive to knowledge of a BRCA1 homozygous deletion in a sample from an individual, administering to the individual an effective amount of a drug effective in treating an HRD positive cancer.

2. The method of claim 1, wherein the BRCA1 homozygous deletion comprises a homozygous deletion of the full BRCA1 gene.

3. The method of claim 1, wherein the advanced prostate cancer (APC) is metastatic castration resistant prostate cancer (mPRPC), hormone sensitive prostate cancer, or de novo metastatic prostate cancer.

4. The method of claim 1, wherein the drug effective in treating an HRD positive cancer is a Poly ADP-ribose polymerase inhibitor (PARPi).

5. The method of claim 1, wherein the drug effective in treating an HRD positive cancer is a platinum agent.

6. A method of treating or delaying progression of advanced prostate cancer (APC) in an individual said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is greater than or equal to 20% or less than about 20%, and a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion is detected in the liquid biopsy sample from the individual, administering to the individual an effective amount of a drug effective in treating an HRD positive cancer.

7. The method of claim 6, wherein the BRCA1 homozygous deletion comprises a homozygous deletion of the full BRCA1 gene.

8. The method of claim 6, wherein the BRCA2 homozygous deletion comprises a homozygous deletion of the full BRCA2 gene.

9. The method of claim 6, wherein the advanced prostate cancer (APC) is metastatic castration resistant prostate cancer (mPRPC), hormone sensitive prostate cancer, or de novo metastatic prostate cancer.

10. The method of claim 6, wherein the drug effective in treating an HRD positive cancer is a Poly ADP-ribose polymerase inhibitor (PARPi).

11. The method of claim 6, wherein the drug effective in treating an HRD positive cancer is a platinum agent.

12. A method of treating or delaying progression of cancer in an individual, said method comprising: determining a tumor fraction (TF) in a liquid biopsy sample collected from the individual; detecting a BRCA1 homozygous deletion and / or BRCA2 homozygous deletion in the liquid biopsy sample; and if the TF is less than about 20% and a BRCA1 homologous deletion and a BRCA2 homozygous deletion are not detected in the liquid biopsy sample from the individual; further detecting a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion in a tissue biopsy sample from the individual, and if a BRCA1 homozygous deletion and / or a BRCA2 homozygous deletion are detected in the tissue biopsy sample from the individual, administering to the individual an effective amount of a drug effective in treating an HRD positive cancer.

13. The method of claim 12, wherein the BRCA1 homozygous deletion comprises a homozygous deletion of the full BRCA1 gene.

14. The method of claim 12, wherein the BRCA2 homozygous deletion comprises a homozygous deletion of the full BRCA2 gene.

15. The method of claim 12, wherein the cancer is advanced prostate cancer (APC).

16. The method of claim 15, wherein the advanced prostate cancer (APC) is metastatic castration resistant prostate cancer (mPRPC), hormone sensitive prostate cancer, or de novo metastatic prostate cancer.

17. The method of claim 12, wherein the drug effective in treating an HRD positive cancer is a Poly ADP-ribose polymerase inhibitor (PARPi).

18. The method of claim 12, wherein the drug effective in treating an HRD positive cancer is a platinum agent.

19. The method of claim 12, wherein the treatment comprises administration of a hormonal therapy.

20. The method of claim 12, further comprising obtaining the liquid biopsy sample from the individual.

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