Therapeutic biomarkers in metastatic castration-resistant prostate cancer: does the state matter?

The treatment of metastatic castration-resistant prostate cancer (mCRPC) has been fundamentally transformed by our greater understanding of its complex biological mechanisms and its entrance into the era of precision oncology. A broad aim is to use the extreme heterogeneity of mCRPC by matching already approved or new targeted therapies to the correct tumor genotype. To achieve this, tumor DNA must be obtained, sequenced, and correctly interpreted, with individual aberrations explored for their druggability, taking into account the hierarchy of driving molecular pathways. Although tumor tissue sequencing is the gold standard, tumor tissue can be challenging to obtain, and a biopsy from one metastatic site or primary tumor may not provide an accurate representation of the current genetic underpinning. Sequencing of circulating tumor DNA (ctDNA) might catalyze precision oncology in mCRPC, as it enables real-time observation of genomic changes in tumors and allows for monitoring of treatment response and identification of resistance mechanisms. Moreover, ctDNA can be used to identify mutations that may not be detected in solitary metastatic lesions and can provide a more in-depth understanding of inter- and intra-tumor heterogeneity. Finally, ctDNA abundance can serve as a prognostic biomarker in patients with mCRPC.The androgen receptor (AR)-axis is a well-established therapeutical target for prostate cancer, and through ctDNA sequencing, insights have been obtained in (temporal) resistance mechanisms that develop through castration resistance. New third-generation AR-axis inhibitors are being developed to overcome some of these resistance mechanisms. The druggability of defects in the DNA damage repair machinery has impacted the treatment landscape of mCRPC in recent years. For patients with deleterious gene aberrations in genes linked to homologous recombination, particularly BRCA1 or BRCA2, PARP inhibitors have shown efficacy compared to the standard of care armamentarium, but platinum-based chemotherapy may be equally effective. A hierarchy exists in genes associated with homologous recombination, where, besides the canonical genes in this pathway, not every other gene aberration predicts the same likelihood of response. Moreover, evidence is emerging on cross-resistance between therapies such as PARP inhibitors, platinum-based chemotherapy and even radioligand therapy that target this genotype. Mismatch repair-deficient patients can experience a beneficial response to immune checkpoint inhibitors. Activation of other cellular signaling pathways such as PI3K, cell cycle, and MAPK have shown limited success with monotherapy, but there is potential in co-targeting these pathways with combination therapy, either already witnessed or anticipated. This review outlines precision medicine in mCRPC, zooming in on the role of ctDNA, to identify genomic biomarkers that may be used to tailor molecularly targeted therapies. The most common druggable pathways and outcomes of therapies matched to these pathways are discussed.

Early On-treatment Circulating Tumor DNA Measurements and Response to Immune Checkpoint Inhibitors in Advanced Urothelial Cancer.

BACKGROUND: Immune checkpoint inhibitors (ICIs) can induce durable disease control in metastatic urothelial cancer (mUC), but only 20-25% of patients respond. Early identification of a nondurable response will improve management strategies. OBJECTIVE: To investigate whether on-treatment circulating tumor DNA (ctDNA) measurements can predict ICI responsiveness in mUC patients. DESIGN, SETTING, AND PARTICIPANTS: This study consists of a discovery cohort of 40 mUC patients and a prospective multicenter validation cohort of 16 mUC patients. Plasma cell-free DNA was collected at baseline and after 3 and 6 wk on ICIs. The ctDNA levels were calculated from targeted sequencing. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Outcome measurements were progression-free survival (PFS), overall survival (OS), and nondurable response (PFS ≤6 mo). Relationships with ctDNA were assessed using Cox regression. Changes in ctDNA level at 3 and 6 wk were categorized by an increase or decrease relative to baseline. RESULTS AND LIMITATIONS: In the discovery cohort, ctDNA was detected in 37/40 (93%) of patients at baseline. A ctDNA increase was observed in 12/15 (80%) and ten of 12 (83%) patients with a nondurable response at 3 and 6 wk, respectively. Of patients with a durable response (PFS >6 mo), 94% showed a decrease. A ctDNA increase at 3 wk was associated with shorter PFS (hazard ratio [HR] 7.8, 95% confidence interval [CI] 3.1-19.5) and OS (HR 8.0, 95% CI 3.0-21.0), independent of clinical prognostic variables. Similar results were observed at 6 wk. The 3-wk association with PFS was validated in a prospective cohort (HR 7.5, 95% CI 1.3-42.6). Limitations include the limited number of patients. CONCLUSIONS: Early changes in ctDNA levels are strongly linked to the duration of ICI benefit in mUC and may contribute to timely therapy modifications. PATIENT SUMMARY: Benefit from immunotherapy can be predicted after only 3 wk of treatment by investigating cancer DNA in blood. This could help in timely therapy changes for urothelial cancer patients with limited benefit from immunotherapy.

Plasma ctDNA as a treatment response biomarker in metastatic cancers: evaluation by the RECIST working group.

Early indicators of metastatic cancer response to therapy are important for evaluating new drugs and stopping ineffective treatment. The Response Evaluation Criteria in Solid Tumors (RECIST) based on repeat cancer imaging are widely adopted in clinical trials, are used to identify active regimens that may change practice, and contribute to regulatory approvals. However, these criteria do not provide insight before 6 - 12 weeks of treatment and typically require that patients have measurable disease. Recent data suggests that measuring on-treatment changes in the amount or proportion of circulating tumor DNA (ctDNA) in peripheral blood plasma may accurately identify responding and non-responding cancers at earlier time points. Over the past year, the RECIST working group has evaluated current evidence for plasma ctDNA kinetics as a treatment response biomarker in metastatic cancers and early endpoint in clinical trials, to identify areas of focus for future research and validation. Here, we outline the requirement for large standardized trial datasets, greater scrutiny of optimal ctDNA collection time points and assay thresholds, and consideration of regulatory body guidelines and patient opinions. In particular, clinically-meaningful changes in plasma ctDNA abundance are likely to differ by cancer type and therapy class, and must be assessed before ctDNA can be considered as a potential pan-cancer response evaluation biomarker. Despite the need for additional data, minimally-invasive on-treatment ctDNA measurements hold promise to build upon existing response assessments such as RECIST, and offer opportunities for developing novel early endpoints for modern clinical trials.

Prediction of plasma ctDNA fraction and prognostic implications of liquid biopsy in advanced prostate cancer.

No consensus strategies exist for prognosticating metastatic castration-resistant prostate cancer (mCRPC). Circulating tumor DNA fraction (ctDNA%) is increasingly reported by commercial and laboratory tests but its utility for risk stratification is unclear. Here, we intersect ctDNA%, treatment outcomes, and clinical characteristics across 738 plasma samples from 491 male mCRPC patients from two randomized multicentre phase II trials and a prospective province-wide blood biobanking program. ctDNA% correlates with serum and radiographic metrics of disease burden and is highest in patients with liver metastases. ctDNA% strongly predicts overall survival, progression-free survival, and treatment response independent of therapeutic context and outperformed established prognostic clinical factors. Recognizing that ctDNA-based biomarker genotyping is limited by low ctDNA% in some patients, we leverage the relationship between clinical prognostic factors and ctDNA% to develop a clinically-interpretable machine-learning tool that predicts whether a patient has sufficient ctDNA% for informative ctDNA genotyping (available online: https://www.ctDNA.org ). Our results affirm ctDNA% as an actionable tool for patient risk stratification and provide a practical framework for optimized biomarker testing.

Early On-treatment Changes in Circulating Tumor DNA Fraction and Response to Enzalutamide or Abiraterone in Metastatic Castration-Resistant Prostate Cancer.

PURPOSE: Androgen receptor pathway inhibitors (ARPI) are standard of care for treatment-naïve metastatic castration-resistant prostate cancer (mCRPC), but rapid resistance is common. Early identification of resistance will improve management strategies. We investigated whether changes in circulating tumor DNA (ctDNA) fraction during ARPI treatment are linked with mCRPC clinical outcomes. EXPERIMENTAL DESIGN: Plasma cell-free DNA was collected from 81 patients with mCRPC at baseline and after 4 weeks of first-line ARPI treatment during two prospective multicenter observational studies (NCT02426333; NCT02471469). ctDNA fraction was calculated from somatic mutations in targeted sequencing and genome copy-number profiles. Samples were classified into detected versus undetected ctDNA. Outcome measurements were progression-free survival (PFS) and overall survival (OS). Nondurable treatment response was defined as PFS ≤6 months. RESULTS: ctDNA was detected in 48/81 (59%) baseline and 29/81 (36%) 4-week samples. ctDNA fraction for samples with detected ctDNA was lower at 4 weeks versus baseline (median 5.0% versus 14.5%, P = 0.017). PFS and OS were shortest for patients with persistent ctDNA at 4 weeks (univariate HR, 4.79; 95% CI, 2.62-8.77 and univariate HR, 5.49; 95% CI, 2.76-10.91, respectively), independent of clinical prognostic factors. For patients exhibiting change from detected to undetected ctDNA by 4 weeks, there was no significant PFS difference versus patients with baseline undetected ctDNA. ctDNA change had a positive predictive value of 88% and negative predictive value of 92% for identifying nondurable responses. CONCLUSIONS: Early changes in ctDNA fraction are strongly linked to duration of first-line ARPI treatment benefit and survival in mCRPC and may inform early therapy switches or treatment intensification. See related commentary by Sartor, p. 2745.

Circulating Tumor DNA-Based Disease Monitoring of Patients with Locally Advanced Esophageal Cancer.

Patients diagnosed with locally advanced esophageal cancer are often treated with neoadjuvant chemoradiotherapy followed by surgery. This study explored whether detection of circulating tumor DNA (ctDNA) in plasma can be used to predict residual disease during treatment. Diagnostic tissue biopsies from patients with esophageal cancer receiving neoadjuvant chemoradiotherapy and surgery were analyzed for tumor-specific mutations. These tumor-informed mutations were used to measure the presence of ctDNA in serially collected plasma samples using hybrid capture-based sequencing. Plasma samples were obtained before chemoradiotherapy, and prior to surgery. The association between ctDNA detection and progression-free and overall survival was measured. Before chemoradiotherapy, ctDNA was detected in 56% (44/78) of patients and detection was associated with tumor stage and volume (p = 0.05, Fisher exact and p = 0.02, Mann-Whitney, respectively). After chemoradiotherapy, ctDNA was detected in 10% (8/78) of patients. This preoperative detection of ctDNA was independently associated with recurrent disease (hazard ratio 2.8, 95% confidence interval 1.1-6.8, p = 0.03, multivariable Cox-regression) and worse overall survival (hazard ratio 2.9, 95% confidence interval 1.2-7.1, p = 0.02, multivariable Cox-regression).Ultradeep sequencing-based detection of ctDNA in preoperative plasma of patients with locally advanced esophageal cancer may help to assess which patients have a high risk of recurrence after neoadjuvant chemoradiotherapy and surgery.

Homologous Recombination Repair Deficiency and Implications for Tumor Immunogenicity.

Homologous recombination repair deficiency (HRD) can be observed in virtually all cancer types. Although HRD sensitizes tumors to DNA-damaging chemotherapy and poly(ADP-ribose) polymerase (PARP) inhibitors, all patients ultimately develop resistance to these therapies. Therefore, it is necessary to identify therapeutic regimens with a more durable efficacy. HRD tumors have been suggested to be more immunogenic and, therefore, more susceptible to treatment with checkpoint inhibitors. In this review, we describe how HRD might mechanistically affect antitumor immunity and summarize the available translational evidence for an association between HRD and antitumor immunity across multiple tumor types. In addition, we give an overview of all available clinical data on the efficacy of checkpoint inhibitors in HRD tumors and describe the evidence for using treatment strategies that combine checkpoint inhibitors with PARP inhibitors.

Plasma BRAF Mutation Detection for the Diagnostic and Monitoring Trajectory of Patients with LDH-High Stage IV Melanoma.

For patients with newly diagnosed metastatic melanoma, rapid BRAF mutation (mBRAF) assessment is vital to promptly initiate systemic therapy. Additionally, blood-based biomarkers are desired to monitor and predict treatment response. Circulating tumor DNA (ctDNA) has shown great promise for minimally invasive mBRAF assessment and treatment monitoring, but validation studies are needed. This prospective study utilized longitudinal plasma samples at regular timepoints (0, 6, 12, 18 weeks) to address the clinical validity of ctDNA measurements in stage IV melanoma patients with elevated serum lactate dehydrogenase (LDH > 250U/L) starting first-line systemic treatment. Using droplet digital PCR, the plasma mBRAF abundance was assessed in 53 patients with a BRAFV600 tissue mutation. Plasma mBRAF was detected in 50/51 patients at baseline (98% sensitivity; median fraction abundance of 19.5%) and 0/17 controls (100% specificity). Patients in whom plasma mBRAF became undetectable during the first 12-18 weeks of treatment had a longer progression-free survival (30.2 vs. 4.0 months; p < 0.001) and cancer-specific survival (not reached vs. 10.2 months; p < 0.001) compared to patients with detectable mBRAF. The ctDNA dynamics outperformed LDH and S100 dynamics. These results confirm the clinical validity of ctDNA measurements as a minimally invasive biomarker for the diagnostic and monitoring trajectory of patients with LDH-high stage IV melanoma.

A Systematic Review and Meta-Analysis on the Predictive Value of Cell-Free DNA-Based Androgen Receptor Copy Number Gain in Patients With Castration-Resistant Prostate Cancer.

PURPOSE: It has been suggested that androgen receptor copy number gain (AR gain) detected in cell-free DNA (cfDNA) can predict treatment response to androgen receptor signaling inhibitors (ARSIs) in patients with castration-resistant prostate cancer (CRPC). But it is unclear whether cfDNA-based AR gain is a true resistance mechanism to ARSIs or mainly a reflection of the tumor burden. In this systematic review, we aim to summarize current literature and comment on the potential of cfDNA-based AR gain as a predictive biomarker to guide therapy choices. METHODS: A literature search was conducted in PubMed/Medline, Cochrane, Embase, and Web of Science databases. Sixteen articles published before November 2019 were selected for the meta-analysis, representing more than 1,000 patients. By using a random effects model, the progression-free survival (PFS) and overall survival (OS) were compared between patients with and without cfDNA-based AR gain who had been treated with ARSIs or with taxane chemotherapy. RESULTS: Upon treatment with ARSIs, the PFS (hazard ratio [HR], 2.33; 95% CI, 2.00 to 2.72; P < .0001) and the OS (HR, 3.83; 95% CI, 3.11 to 4.70; P < .0001) were worse for patients with cfDNA-based AR gain, independent of the line and type of ARSIs. The OS and PFS in patients treated with first-line docetaxel or second-line or third-line cabazitaxel seemed to be unaffected by AR gain, despite a higher disease burden in patients with AR gain. AR gain was associated with reduced response with later lines of docetaxel. CONCLUSION: In patients with CRPC, cfDNA-based AR gain is associated with a worse response to ARSIs. The effect on patients who are receiving taxane chemotherapy seems to be dependent on the type and line, although data are limited. Future prospective studies are essential to assess the true potential of cfDNA-based AR gain as a minimally invasive biomarker to guide therapy choice.

Metabolic stress in cancer cells induces immune escape through a PI3K-dependent blockade of IFNγ receptor signaling.

BACKGROUND: T-cell mediated immunotherapy brought clinical success for many cancer patients. Nonetheless, downregulation of MHC class I antigen presentation, frequently occurring in solid cancers, limits the efficacy of these therapies. Unraveling the mechanisms underlying this type of immune escape is therefore of great importance. We here investigated the immunological effects of metabolic stress in cancer cells as a result of nutrient deprivation. METHODS: TC1 and B16F10 tumor cell lines were cultured under oxygen- and glucose-deprivation conditions that mimicked the tumor microenvironment of solid tumors. Presentation of peptide antigens by MHC class I molecules was measured by flow cytometry and via activation of tumor-specific CD8 T cell clones. The proficiency of the IFNy-STAT1 pathway was investigated by Western blots on phosphorylated proteins, transfection of constitutive active STAT1 constructs and qPCR of downstream targets. Kinase inhibitors for PI3K were used to examine its role in IFNy receptor signal transduction. RESULTS: Combination of oxygen- and glucose-deprivation resulted in decreased presentation of MHC class I antigens on cancer cells, even in the presence of the stimulatory cytokine IFNy. This unresponsiveness to IFNy was the result of failure to phosphorylate the signal transducer STAT1. Forced expression of constitutive active STAT1 fully rescued the MHC class I presentation. Furthermore, oxygen- and glucose-deprivation increased PI3K activity in tumor cells. Pharmacological inhibition of this pathway not only restored signal transduction through IFNy-STAT1 but also improved MHC class I presentation. Importantly, PI3K inhibitors also rendered tumor cells sensitive for recognition by CD8 T cells in culture conditions of metabolic stress. CONCLUSIONS: These data revealed a strong impact of metabolic stress on the presentation of tumor antigens by MHC class I and suggest that this type of tumor escape takes place at hypoxic areas even during times of active T cell immunity and IFNy release.