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Abstract Immune checkpoint inhibitors (ICI) improve overall survival in patients with metastatic urothelial cancer (mUC), but therapeutic success at the individual patient level varies significantly. Here we identify predictive markers of response, based on whole-genome DNA (n = 70) and RNA-sequencing (n = 41) of fresh metastatic biopsy samples, collected prior to treatment with pembrolizumab. We find that PD-L1 combined positivity score does not, whereas tumor mutational burden and APOBEC mutagenesis modestly predict response. In contrast, T cell-to-stroma enrichment (TSE) score, computed from gene expression signature data to capture the relative abundance of T cells and stromal cells, predicts response to immunotherapy with high accuracy. Patients with a positive and negative TSE score show progression free survival rates at 6 months of 67 and 0%, respectively. The abundance of T cells and stromal cells, as reflected by the TSE score is confirmed by immunofluorescence in tumor tissue, and its good performance in two independent ICI-treated cohorts of patients with mUC (IMvigor210) and muscle-invasive UC (ABACUS) validate the predictive power of the TSE score. In conclusion, the TSE score represents a clinically applicable metric that potentially supports the prospective selection of patients with mUC for ICI treatment.

AMG 256 is a bi-specific, heteroimmunoglobulin molecule with an anti-PD-1 antibody domain and a single IL-21 mutein domain on the C-terminus. Nonclinical studies in cynomolgus monkeys revealed that AMG 256 administration led to the development of immunogenicity-mediated responses and indicated that the IL-21 mutein domain of AMG 256 could enhance the anti-drug antibody response directed toward the monoclonal antibody domain. Anti-AMG 256 IgE were also observed in cynomolgus monkeys. A first-in-human (FIH) study in patients with advanced solid tumors was designed with these risks in mind. AMG 256 elicited ADA in 28 of 33 subjects (84.8%). However, ADA responses were only robust and exposure-impacting at the 2 lowest doses. At mid to high doses, ADA responses remained low magnitude and all subjects maintained exposure, despite most subjects developing ADA. Limited drug-specific IgE were also observed during the FIH study. ADA responses were not associated with any type of adverse event. The AMG 256 program represents a unique case where nonclinical studies informed on the risk of immunogenicity in humans, due to the IL-21-driven nature of the response.

Multiple prognostic biomarkers, including circulating tumour cell (CTC) counts, exist in metastatic castration‐resistant prostate cancer (mCRPC) patients, but none of them have been implemented into daily clinical care. The modified fast aneuploidy screening test‐sequencing system (mFast‐SeqS), which yields a genome‐wide aneuploidy score, is able to reflect the fraction of cell‐free tumour DNA (ctDNA) within cell‐free DNA (cfDNA) and may be a promising biomarker in mCRPC. In this study, we investigated the prognostic value of dichotomized aneuploidy scores (

Prostate cancer is known to have a variable response to androgen receptor signalling inhibitors. Here, the authors use machine learning to predict response to therapy from genomic, transcriptomic and clinical data.

Mutations in the estrogen receptor gene (ESR1), its transcriptional regulators, and the mitogen-activated protein kinase (MAPK) pathway are enriched in patients with endocrine-resistant metastatic breast cancer (MBC). Here, we integrated whole genome sequencing with RNA sequencing data from the same samples of 101 ER-positive/HER2-negative MBC patients who underwent a tumor biopsy prior to the start of a new line of treatment for MBC (CPCT-02 study, NCT01855477) to analyze the downstream effects of DNA alterations previously linked to endocrine resistance, thereby gaining a better understanding of the associated mechanisms. Hierarchical clustering was performed using expression of ESR1 target genes. Genomic alterations at the DNA level, gene expression levels, and last administered therapy were compared between the identified clusters. Hierarchical clustering revealed two distinct clusters, one of which was characterized by increased expression of ESR1 and its target genes. Samples in this cluster were significantly enriched for mutations in ESR1 and amplifications in FGFR1 and TSPYL. Patients in the other cluster showed relatively lower expression levels of ESR1 and its target genes, comparable to ER-negative samples, and more often received endocrine therapy as their last treatment before biopsy. Genes in the MAPK-pathway, including NF1, and ESR1 transcriptional regulators were evenly distributed. In conclusion, RNA sequencing identified a subgroup of patients with clear expression of ESR1 and its downstream targets, probably still benefiting from ER-targeting agents. The lower ER expression in the other subgroup might be partially explained by ER activity still being blocked by recently administered endocrine treatment, indicating that biopsy timing relative to endocrine treatment needs to be considered when interpreting transcriptomic data.

Metastatic and locally-advanced neuroendocrine neoplasms (aNEN) display heterogeneous clinical and genetic characteristics. Here, the authors investigate the mutational landscape of 85 aNEN by whole genome sequencing and identify distinct subpopulations, tumour mutational burden patterns, drivers and actionable somatic alterations.

Abstract Here, we describe a novel approach for rapid discovery of a set of tumor-specific genomic structural variants (SVs), based on a combination of low coverage cancer genome sequencing using Oxford Nanopore with an SV calling and filtering pipeline. We applied the method to tumor samples of high-grade ovarian and prostate cancer patients and validated on average ten somatic SVs per patient with breakpoint-spanning PCR mini-amplicons. These SVs could be quantified in ctDNA samples of patients with metastatic prostate cancer using a digital PCR assay. The results suggest that SV dynamics correlate with and may improve existing treatment-response biomarkers such as PSA. https://github.com/UMCUGenetics/SHARC .

Prostate cancer is the most dominant male malignancy worldwide. The clinical presentation of prostate cancer ranges from localized indolent to rapidly progressing lethal metastatic disease. Despite a decline in death rate over the past years, with the advent of early diagnosis and new treatment options, challenges remain towards the management of metastatic prostate cancer, particularly metastatic castration sensitive prostate cancer (mCSPC) and castration resistant prostate cancer (mCRPC). Current treatments involve a combination of chemotherapy with androgen deprivation therapy and/or androgen receptor signalling inhibitors. However, treatment outcomes are heterogeneous due to significant tumor heterogeneity indicating a need for better prognostic biomarkers to identify patients with poor outcomes. Liquid biopsy has opened a plethora of opportunities from early diagnosis to (personalized) therapeutic disease interventions. In this review, we first provide recent insights about (metastatic) prostate cancer and its current treatment landscape. We highlight recent studies involving various circulating biomarkers such as circulating tumor cells, genetic markers, circulating nucleic acids, extracellular vesicles, tumor-educated platelets, and the secretome from (circulating) tumor cells and tumor microenvironment in metastatic prostate cancer. The comprehensive array of biomarkers can provide a powerful approach to understanding the spectrum of prostate cancer disease and guide in developing improved and personalized treatments for patients.

Molecular landscapes of metastatic colorectal cancers (mCRC) have often been restricted to coding regions or low numbers of patients. Here the authors present a whole-genome landscape of 429 mCRC patients, revealing the mutational impact of prior therapies and potential actionable targets.

Treatment-induced AR alterations, including AR alternative splice variants (AR-Vs), have been extensively linked to harboring roles in primary and acquired resistance to conventional and next-generation hormonal therapies in prostate cancer and therefore have gained momentum. Our aim was to uniformly determine recurrent AR-Vs in metastatic castration-resistant prostate cancer (mCRPC) using whole transcriptome sequencing in order to assess which AR-Vs might hold potential diagnostic or prognostic relevance in future research. This study reports that in addition to the promising AR-V7 as a biomarker, AR45 and AR-V3 were also seen as recurrent AR-Vs and that the presence of any AR-V could be associated with higher AR expression. With future research, these AR-Vs may therefore harbor similar or complementary roles to AR-V7 as predictive and prognostic biomarkers in mCRPC or as proxies for abundant AR expression.

Some cancer patients first present with metastases where the location of the primary is unidentified; these are difficult to treat. In this study, using machine learning, the authors develop a method to determine the tissue of origin of a cancer based on whole sequencing data.

Abstract Background Patients with advanced cancer for whom standard systemic treatment is no longer available may be offered participation in early phase clinical trials. In the decision making process, both medical-technical information and patient values and preferences are important. Since patients report decisional conflict after deciding on participation in these trials, improving the decision making process is essential. We aim to develop and evaluate an Online Value Clarification Tool (OnVaCT) to assist patients in clarifying their values around this end-of-life decision. This improved sharing of values is hypothesized to support medical oncologists in tailoring their information to individual patients’ needs and, consequently, to support patients in taking decisions in line with their values and reduce decisional conflict. Methods In the first part, patients’ values and preferences and medical oncologists’ views hereupon will be explored in interviews and focus groups to build a first prototype OnVaCT using digital communication (serious gaming). Next, we will test feasibility during think aloud sessions, to deliver a ready-to-implement OnVaCT. In the second part, the OnVaCT, with accompanied training module, will be evaluated in a pre-test (12–18 months before implementation) post-test (12–18 months after implementation) study in three major Dutch cancer centres. We will include 276 patients (> 18 years) with advanced cancer for whom standard systemic therapy is no longer available, and who are referred for participation in early phase clinical trials. The first consultation will be recorded to analyse patient-physician communication regarding the discussion of patients’ values and the decision making process. Three weeks afterwards, decisional conflict will be measured. Discussion This project aims to support the discussion of patient values when considering participation in early phase clinical trials. By including patients before their first appointment with the medical oncologist and recording that consultation, we are able to link decisional conflict to the decision making process, e.g. the communication during consultation. The study faces challenges such as timely including patients within the short period between referral and first consultation. Furthermore, with new treatments being developed rapidly, molecular stratification may affect the patient populations included in the pre-test and post-test periods. Trial registration Netherlands Trial Registry number: NTR7551 (prospective; July 17, 2018).

Detecting genomic abnormalities in metastatic castration-resistant prostate cancer (mCRPC) may impact clinical treatment. Here, the authors present whole-genome sequencing of metastatic biopsies from 197 mCRPC patients, highlighting the landscape of microsatellite stability, homologous repair deficiency, and other genomic subgroups.

Background: Biomarkers predicting treatment response may be used to stratify pancreatic ductal adenocarcinoma (PDAC) patients for therapy. The aim of this study was to identify circulating tumor DNA (ctDNA) mutations that associate with tumor progression during FOLFIRINOX chemotherapy, and overall survival (OS). Methods: Circulating cell-free DNA was analyzed with a 57 gene next-generation sequencing panel using plasma samples of 48 PDAC patients of all disease stages. Patients received FOLFIRINOX as initial treatment. Chemotherapy response was determined on CT scans as disease control ( n = 30) or progressive disease ( n = 18) within eight cycles of FOLFIRINOX, based on RECIST 1.1 criteria. Results: Detection of a TP53 ctDNA mutation before start of FOLFIRINOX [odds ratio (OR) 10.51, 95% confidence interval (CI) 1.40–79.14] and the presence of a homozygous TP53 Pro72Arg germline variant (OR 6.98, 95% CI 1.31–37.30) were predictors of early tumor progression during FOLFIRINOX in multivariable analysis. Five patients presented with the combination of a TP53 ctDNA mutation before start of FOLFIRINOX and the homozygous Pro72Arg variant. All five patients showed progression during FOLFIRINOX. The combination of the TP53 mutation and TP53 germline variant was associated with shorter survival (median OS 4.4 months, 95% CI 2.6–6.2 months) compared with patients without any TP53 alterations (median OS 13.0 months, 95% CI 8.6–17.4 months). Conclusion: The combination of a TP53 ctDNA mutation before start of FOLFIRINOX and a homozygous TP53 Pro72Arg variant is a promising biomarker, associated with early tumor progression during FOLFIRINOX and poor OS. The results of this exploratory study need to be validated in an independent cohort.

Circulating tumor cell (CTC)- and/or tumor-derived extracellular vesicle (tdEV) loads in the blood of metastatic castration-resistant prostate cancer (CRPC) patients are associated with worse overall survival and can be used as predictive markers of treatment response. In this study, we investigated the quantity/quality of CTCs and tdEVs in metastatic castration-naive prostate cancer (CNPC) and CRPC patients, and whether androgen deprivation therapy (ADT) affects CTCs and tdEVs. We included 104 CNPC patients before ADT initiation and 66 CRPC patients. Blood samples from 31/104 CNPC patients were obtained 6 months after ADT. CTCs and tdEVs were identified using ACCEPT software. Based on the morphology, CTCs of metastatic CNPC and CRPC patients were subdivided by manual reviewing into six subclasses. The numbers of CTCs and tdEVs were correlated in both CNPC and CRPC patients, and both CTCs (p = 0.013) and tdEVs (p = 0.005) were significantly lower in CNPC compared to CRPC patients. Qualitative differences in CTCs were observed: CTC clusters (p = 0.006) and heterogeneously CK expressing CTCs (p = 0.041) were significantly lower in CNPC patients. CTC/tdEV numbers declined 6 months after ADT. Our study showed that next to CTC-load, qualitative CTC analysis and tdEV-load may be useful in CNPC patients.

BACKGROUND Continued androgen receptor (AR) signaling constitutes a key target for treatment in metastatic castration-resistant prostate cancer (CRPC). Studies have identified 11-ketotestosterone (11KT) as a potent AR agonist, but it is unknown if 11KT is present at physiologically relevant concentrations in patients with CRPC to drive AR activation. The goal of this study was to investigate the circulating steroid metabolome including all active androgens in patients with CRPC.METHODS Patients with metastatic CRPC (n = 29) starting a new line of systemic therapy were included. Sequential plasma samples were obtained for measurement of circulating steroid concentrations by multisteroid profiling employing liquid chromatography–tandem mass spectrometry. Metastatic tumor biopsy samples were obtained at baseline and subjected to RNA sequencing.RESULTS 11KT was the most abundant circulating active androgen in 97% of patients with CRPC (median 0.39 nmol/L, range: 0.03–2.39 nmol/L), constituting 60% (IQR 43%–79%) of the total active androgen (TA) pool. Treatment with glucocorticoids reduced 11KT by 84% (49%–89%) and testosterone by 68% (38%–79%). Circulating TA concentrations at baseline were associated with a distinct intratumor gene expression signature comprising AR-regulated genes.CONCLUSION The potent AR agonist 11KT is the predominant circulating active androgen in patients with CRPC and, therefore, one of the potential drivers of AR activation in CRPC. Assessment of androgen status should be extended to include 11KT, as current clinical approaches likely underestimate androgen abundance in patients with CRPC.TRIAL REGISTRATION Netherlands Trial Register: NL5625 (NTR5732).FUNDING Daniel den Hoed Foundation and Wellcome Trust (Investigator Award WT209492/Z/17/Z).

The emerging interest in circulating tumor DNA (ctDNA) analyses for clinical trials has necessitated the development of a high‐throughput method for fast, reproducible, and efficient isolation of ctDNA. Currently, the majority of ctDNA studies use the manual QIAamp (QA) platform to isolate DNA from blood. The purpose of this study was to compare two competing automated DNA isolation platforms [Maxwell (MX) and QIAsymphony (QS)] to the current ‘gold standard’ QA to facilitate high‐throughput processing of samples in prospective trials. We obtained blood samples from healthy blood donors and metastatic cancer patients for plasma isolation. Total cell‐free DNA (cfDNA) quantity was assessed by TERT quantitative PCR. Recovery efficiency was investigated by quantitative PCR analysis of spiked‐in synthetic plant DNA. In addition, a β‐actin fragmentation assay was performed to determine the amount of contamination by genomic DNA from lysed leukocytes. ctDNA quality was assessed by digital PCR for somatic variant detection. cfDNA quantity and recovery efficiency were lowest using the MX platform, whereas QA and QS showed a comparable performance. All platforms preferentially isolated small (136 bp) DNA fragments over large (420 and 2000 bp) DNA fragments. Detection of the number variant and wild‐type molecules was most comparable between QA and QS. However, there was no significant difference in variant allele frequency comparing QS and MX to QA. In summary, we show that the QS platform has comparable performance to QA, the ‘gold standard’, and outperformed the MX platform depending on the readout used. We conclude that the QS can replace the more laborious QA platform, especially when high‐throughput cfDNA isolation is needed.

When standard treatment options are not available anymore, patients with advanced cancer may participate in early phase clinical trials. Improving this complex decision-making process may improve their quality of life. Therefore, this prospective multicenter study with questionnaires untangles several contributing factors to decisional conflict (which reflects the quality of decision-making) in patients with advanced cancer who recently decided upon early phase clinical trial participation (phase I or I/II). We hypothesized that health-related quality of life, health literacy, sense of hope, satisfaction with the consultation, timing of the decision, and the decision explain decisional conflict. Mean decisional conflict in 116 patients was 30.0 (SD = 16.9). Multivariate regression analysis showed that less decisional conflict was reported by patients with better global health status (β = −0.185, p = 0.018), higher satisfaction (β = −0.246, p = 0.002), and who made the decision before (β = −0.543, p < 0.001) or within a week after the consultation (β = −0.427, p < 0.001). These variables explained 37% of the variance in decisional conflict. Healthcare professionals should realize that patients with lower global health status and who need more time to decide may require additional support. Although altering such patient intrinsic characteristics is difficult, oncologists can impact the satisfaction with the consultation. Future research should verify whether effective patient-centered communication could prevent decisional conflict.

Docetaxel entrapped nanoparticle CPC634 is associated with dose-related skin toxicity that resembles conventional docetaxel (Cd)-related skin toxicity. This study compared the cutaneous pharmacokinetics and pharmacodynamics of docetaxel and CPC634. In this randomised cross-over study, patients with solid tumours received one cycle of CPC634 and Cd (both at 75 mg/m2). Skin biopsies were taken at baseline and at day 8 of both cycles. Released and total docetaxel (released docetaxel plus entrapped docetaxel) concentrations and histopathological changes in the skin biopsies were evaluated. Twenty patients underwent paired skin biopsies for pharmacokinetic analysis and 10 patients had biopsies available for histopathological assessment. The total skin docetaxel concentration was 369% (95%CI: 229% to 569%, p < 0.001) higher after CPC634 administration compared to Cd while the released docetaxel concentrations were not statistically different (95%CI: −9% to 63%, p = 0.169). The CPC634 released docetaxel concentration in the skin was positively correlated with plasma concentrations (Pearson’s correlation 0.48, p = 0.03). Histopathological examination revealed increased apoptosis, mitotic cells with nuclear atypia, and micronucleation with an enhanced Ki-67 index for both compounds. In conclusion, both CPC634 and Cd treatment result in docetaxel exposure in the skin causing cutaneous anti-mitotic effects such as micronucleation, which could induce an inflammatory reaction leading to skin toxicity.

(1) Background: Markers identifying which patients with metastatic, castration-resistant prostate cancer (mCRPC) will benefit from cabazitaxel therapy are currently lacking. Therefore, the aim of this study was to identify markers associated with outcome to cabazitaxel therapy based on counts and gene expression profiles of circulating tumor cells (CTCs). (2) Methods: From 120 mCRPC patients, CellSearch enriched CTCs were obtained at baseline and after 6 weeks of cabazitaxel therapy. Furthermore, 91 genes associated with prostate cancer were measured in mRNA of these CTCs. (3) Results: In 114 mCRPC patients with an evaluable CTC count, the CTC count was independently associated with poor progression-free survival (PFS) and overall survival (OS) in multivariable analysis with other commonly used variables associated with outcome in mCRPC (age, prostate specific antigen (PSA), alkaline phosphatase, lactate dehydrogenase (LDH), albumin, hemoglobin), together with alkaline phosphatase and hemoglobin. A five-gene expression profile was generated to predict for outcome to cabazitaxel therapy. However, even though this signature was associated with OS in univariate analysis, this was not the case in the multivariate analysis for OS nor for PFS. (4) Conclusion: The established five-gene expression profile in CTCs was not independently associated with PFS nor OS. However, along with alkaline phosphatase and hemoglobin, CTC-count is independently associated with PFS and OS in mCRPC patients who are treated with cabazitaxel.

We and others have previously demonstrated that the acute release of progenitor cells in response to chemotherapy actually reduces the efficacy of the chemotherapy. Here, we take these data further and investigate the clinical relevance of circulating endothelial (progenitor) cells (CE(P)Cs) and modulatory cytokines in patients after chemotherapy with relation to progression-free and overall survival (PFS/OS). Patients treated with various chemotherapeutics were included. Blood sampling was performed at baseline, 4 hours, and 7 and 21 days after chemotherapy. The mononuclear cell fraction was analyzed for CE(P)C by FACS analysis. Plasma was analyzed for cytokines by ELISA or Luminex technique. CE(P)Cs were correlated with response and PFS/OS using Cox proportional hazard regression analysis. We measured CE(P)Cs and cytokines in 71 patients. Only patients treated with paclitaxel showed an immediate increase in endothelial progenitor cell 4 hours after start of treatment. These immediate changes did not correlate with response or survival. After 7 and 21 days of chemotherapy, a large and consistent increase in CE(P)C was found (P < .01), independent of the type of chemotherapy. Changes in CE(P)C levels at day 7 correlated with an increase in tumor volume after three cycles of chemotherapy and predicted PFS/OS, regardless of the tumor type or chemotherapy. These findings indicate that the late release of CE(P)C is a common phenomenon after chemotherapeutic treatment. The correlation with a clinical response and survival provides further support for the biologic relevance of these cells in patients' prognosis and stresses their possible use as a therapeutic target.