Circulating Tumor Cell Conversion Research Articles

A phase 2 study of immunogenic cell death inducer PT-112 in patients with metastatic castration-resistant prostate cancer.

TPS292 Background: PT-112 is a novel small I-O molecule currently in development for several cancers. Preclinical studies have shown the robust induction of immunogenic cell death by PT-112, leading to an anti-cancer adaptive immune response. Biodistribution experiments in mice showed PT-112’s partial bone affinity. In two prior phase I studies, late-line metastatic castration-resistant prostate cancer patients (mCRPC pts) were treated with evidence of anti-cancer activity, including tumor volume reductions, improvements in PET and bone scans, PSA and alkaline phosphatase (ALP) reductions, and anecdotal cases of improvement in disease-related pain. This, taken together with the immunogenic and osteotropic properties observed in preclinical work, provided a strong rationale to further explore PT-112 in mCRPC, an immunologically “cold” disease with a high prevalence of bone metastases. Moreover, in an analysis of >6,000 mCRPC pts, circulating tumor cell (CTC) declines (CTC0 and CTC conversions) were correlated with improvements in survival to a greater extent than PSA declines (PSA50) (Heller et al., JCO, 36:6 2017), prompting incorporation of CTC changes as a secondary endpoint. Methods: The primary objective of the study is to define the dose regimen of PT-112 for pivotal study. Pts are randomized to one of three arms, receiving IV PT-112 on 28-day cycles via one hour infusions on (1) Days 1 and 15 at 250 mg/m2, (2) Days 1 and 15 at 360 mg/m2, or (3) Days 1 and 15 of cycle 1 at 360 mg/m2 followed by 250 mg/m2 on Day 15 of subsequent cycles. Key eligibility criteria include radiographically progressive disease at study entry; ≥3 life-prolonging therapies for metastatic disease including 1-2 taxanes, ≥1 new generation anti-androgen therapies, and other drugs FDA approved on the basis of survival; and allowing bone-only metastatic disease. Efficacy assessments comprise disease control rate at 4 months (DCR4) using RECIST and PCWG3 criteria, objective response rate, CTC0 and CTC conversion, PSA50 and ALP reductions. Additionally, T cell receptor sequencing is conducted to characterize treatment-induced changes in T cell fraction and clonal expansions, as a means of generating meaningful supportive data on the immune mechanism of PT-112 monotherapy. A Fleming two-stage design will be used to assess each arm, with a 20% DCR4 as the null hypotheses, requiring ≥6 of 25 pts responding in stage one and ≥14 of 45 in total to reject the null. In addition, other efficacy measures, such as CTC responses, as well as exposure/response and exposure/safety analyses will be applied to characterize the risk/benefit ratio and select the optimal dose regimen for PT-112. As of October 11th, 2022, 38 pts have been enrolled out of a planned maximum of 135. Clinical trial information: NCT02266745 .

Differential efficacy of PARP inhibitors in metastatic castration-resistant prostate cancer with DNA repair defects: A systematic review and meta-analysis.

134 Background: Recent trials have suggested poly ADP-ribose polymerase inhibitors (PARPi) sensitivity in patients with treatment-refractory metastatic castration-resistant prostate cancer (mCRPC) harboring mutations in DNA damage and response repair genes (DDR). However, the efficacy profile might differ with different genes and is still being explored in multiple studies. Methods: We systematically searched several electronic databases (MEDLINE, EMBASE), and conference proceedings to include abstracts and full-text publications of phase II/III trials evaluating PARPi in treatment-refractory mCRPC with DDR gene alterations. Outcomes of interest included objective response rate (ORR), complete response (CR), prostate specific antigen (PSA) 50% response, circulating tumor cells conversion (CTCc) from ≥ 5 to < 5 cells, composite response (CoR) and risk of progression. Data was pooled using Inverse-variance approach. A DerSimonian and Laird random-effect meta-analysis was conducted to estimate pooled proportion (PP) of events using the Freeman-Tukey transformation in each gene. Clopper-Pearson method was used to estimate the associated 95% confidence intervals (CI). Results: This systematic review and meta-analysis included five unique trials with 648 patients and four PARPi (niraparib, olaparib, rucaparib, talazoparib). Objective response to PARPi was observed in 44.7% of patients with mutations in either BRCA1 or 2 or both (95% CI: 36.0%-53.6%; I2: 0%), 23.5% with ATM (95% CI: 3.91%-51.0%; I2: 65%), 25.0% with CDK12 (95% CI: 8.66%-49.1%), and 45.3% with PALB2 alterations (95% CI: 14.8%-77.5%; I2: 0%). Consistent results were observed for CR outcome. PSA50% response to PARPi was observed in 60.3% of patients with BRCA1/2 mutations (95% CI: 50.3%-70.0%; I2: 57%), 5.54% (95% CI: 0.01-16.6; I2: 0%) with ATM, and 70.1% (95% CI: 35.6%-96.6%; I2: 0%) with PALB2 mutations while no PSA 50% response was observed in patients with CDK12 mutations (PP: 0.00%; 95% CI: 0%-16.84%). Moreover, CTCc was observed in 68.3% of patients with BRCA1/2 mutations (95% CI: 45.0%-87.8%; I2: 77), 34.2% (95% CI: 11.2-61.3%; I2: 45%) with ATM, 41.7% (95% CI: 15.2%-72.3%) with CDK12, and 48.5% (95% CI: 6.39%-92.9%; I2: 49%) with PALB2. CoR was observed in 76.5% patients with BRCA1/2 mutations (95% CI: 65.4%-86.1%; I2: 22%), 30.4% (95% CI: 16.0%-46.9%) with ATM, 25% (95% CI: 8.66%-49.1%) with CDK12, and 63.9% (95% CI: 31.0%-91.8%; I2: 0%) with PALB2 mutations. Similarly, the risk of progression was 50.9% in patients with BRCA1/2 mutations (95% CI: 37.7%-63.9%), 70.9% (95% CI: 60.0%-80.40%; I2: 0%) with ATM, and 75% (95% CI: 19.4-99.3%) with PALB2. Conclusions: Current evidence is sparse regarding efficacy by specific genes. Limited data favors the use of PARPi in mCRPC patients harboring mutations in BRCA1/2 and PALB2 genes.

Value of Early Circulating Tumor Cells Dynamics to Estimate Docetaxel Benefit in Metastatic Castration-Resistant Prostate Cancer (mCRPC) Patients.

Simple SummaryThe prognostic role of CTC enumeration in mCRPC patients has been established in several studies, demonstrating a higher prognostic performance than post-treatment changes in PSA levels in patients treated with AR signaling inhibitors, but not taxanes. We carried out a pooled analysis of two prospective studies in mCRPC patients treated with docetaxel. The results of this study showed a greater ability of early changes in circulating tumor cells (CTCs) compared to PSA response endpoints to predict overall survival in metastatic castration-resistant prostate cancer (mCRPC) patients treated with docetaxel. These results encourage the clinical usefulness of CTC enumeration to determine the outcome of mCRPC patients.Circulating tumor cell (CTC) enumeration and changes following treatment have been demonstrated to be superior to PSA response in determining mCRPC outcome in patients receiving AR signaling inhibitors but not taxanes. We carried out a pooled analysis of two prospective studies in mCRPC patients treated with docetaxel. CTCs were measured at baseline and 3–6 weeks post treatment initiation. Cox regression models were constructed to compare 6-month radiographical progression-free survival (rPFS), CTCs and PSA changes predicting outcome. Among the subjects, 80 and 52 patients had evaluable baseline and post-treatment CTC counts, respectively. A significant association of higher baseline CTC count with worse overall survival (OS), PFS and time to PSA progression (TTPP) was observed. While CTC response at 3–6 weeks (CTC conversion (from ≥5 to <5 CTCs), CTC30 (≥30% decline in CTC) or CTC0 (decline to 0 CTC)) and 6-month rPFS were significantly associated with OS (all p < 0.005), the association was not significant for PSA30 or PSA50 response. CTC and PSA response were discordant in over 50% of cases, with outcome driven by CTC response in these patients. The c-index values for OS were superior for early CTC changes compared to PSA response endpoints, and similar to 6-month rPFS. Early CTC declines were good predictors of improved outcomes in mCRPC patients treated with docetaxel in this small study, offering a superior and/or earlier estimation of docetaxel benefit in comparison to PSA or rPFS that merits further confirmation in larger studies.

Phase I study of AMG 160, a half-life extended bispecific T-cell engager (HLE BiTE) immune therapy targeting prostate-specific membrane antigen (PSMA), in patients with metastatic castration-resistant prostate cancer (mCRPC).

TPS261 Background: AMG 160 is a novel HLE BiTE immune therapy that redirects T cells to kill tumor cells by binding to PSMA on tumor cells and CD3 on T cells. Methods: Primary objectives of this open-label, ascending, multiple-dose, phase 1 study (NCT03792841) are to evaluate safety and tolerability and determine the maximum tolerated dose (MTD) or recommended phase 2 dose (RP2D) of AMG 160 in men with mCRPC; secondary objectives are to characterize pharmacokinetics (PK) and evaluate preliminary efficacy. The dose exploration will estimate the MTD or RP2D by Bayesian logistic regression modeling. The dose expansion will assess safety, efficacy, PK, and pharmacodynamics (PD) of the selected dose and provide further safety and efficacy data. PD biomarkers and potential patient selection biomarkers will be explored. Preliminary antitumor activity will be assessed by objective response per RECIST 1.1 with PCWG3 modifications, PSA response, duration of response, time to progression, PFS (radiographic and PSA)/OS, and circulating tumor cell (CTC) response (CTC0 and CTC conversion). Imaging will include CT/MRI, bone scan, 68Ga-PSMA-11 PET/CT, and 18F-FDG PET/CT. In cycle 1, patients will be pretreated with dexamethasone before short-term IV infusion of AMG 160 and will be hospitalized for 72 h after each AMG 160 dose. Key inclusion criteria: age ≥18 y; histologically/cytologically confirmed mCRPC that progressed after novel hormone therapy; failure of 1–2 taxane-based regimens or have refused a taxane regimen; bilateral orchiectomy or continuous androgen-deprivation therapy; evidence of progressive disease; total serum testosterone ≤50 ng/dL. Key exclusion criteria: active autoimmune disease or diseases requiring immunosuppressive therapy (low-dose prednisone permitted); CNS metastases, leptomeningeal disease, or spinal cord compression; prior PSMA-targeted therapy (177Lu-PSMA-617 may be allowed). The study will enroll 30–50 patients in the dose exploration and 50 patients in the dose expansion globally. The study opened in January 2019; dose exploration is ongoing. Clinical trial information: NCT03792841.

Niraparib in patients (pts) with metastatic castration-resistant prostate cancer (mCRPC) and biallelic DNA-repair gene defects (DRD): Correlative measures of tumor response in phase II GALAHAD study.

118 Background: Niraparib, a highly potent and selective poly (ADP-ribose) polymerase inhibitor (PARPi) received breakthrough designation by US FDA for treatment of pts with BRCA1,2 mutant mCRPC who progressed on taxane and androgen receptor-targeted therapy. Circulating tumor cells (CTC) detection associates with poor outcomes, with declining counts consistent with improved survival [1,2]. Methods: GALAHAD study assessed niraparib (300 mg daily) in pts with mCRPC+DRD (NCT02854436). Patients with non-measurable soft tissue disease by RECIST 1.1 were required to have a baseline CTC count ≥1 cell/7.5 mL blood. CTC response was defined as CTC conversion to &lt;5 for pts with baseline CTC≥5 and CTC drop to 0 post-baseline for pts with ≥1 baseline CTC. Alkaline phosphatase (ALP) was collected at each monthly cycle. Results: For primary efficacy population of pts with BRCA1/2 mutations, the objective response rate (ORR) by RECIST 1.1 criteria was 41.4%. CTC response rates for this population were as high as ORR regardless of measurability (Table). Time to CTC response for each CTC responder will be shown. Radiographic progression-free survival (rPFS) durations were similar for patients with a measurable disease response and patients with CTC conversion. Median duration of treatment for responders of any type was 6.7mo (range: 2–27). DRD status, both BRCA and non- BRCA, for each responder will also be discussed. Trends in disease burden and markers of bone metabolism will also be quantitatively explored including 24% pts who were on treatment for at least one cycle who had ≥25% decreased unfractionated ALP from baseline. Conclusions: Niraparib showed clinical activity with CTC response and decline in ALP levels in mCRPC pts having biallelic BRCA mutations, which further supports its recent breakthrough designation. Clinical trial information: NCT02854436. [Table: see text]

Phase Ib study of niraparib plus androgen receptor-targeted therapy (ART) in patients (pts) with metastatic castration-resistant prostate cancer (mCRPC).

122 Background: Poly ADP-ribose polymerase (PARP) inhibitor and abiraterone acetate + prednisone (AA+P) has activity in pts with mCRPC regardless of DNA repair gene defects (DRD). Methods: This phase 1b study (Bedivere; NCT02924766) began with a 3+3 dose escalation of niraparib (Part 1), followed by expansion (Part 2) of the recommended phase 2 dose (RP2D) of nira + AA+P. Pts with mCRPC received nira 200 or 300 mg/day with AA+P. Dose-limiting toxicities (DLTs) were evaluated the first 28 days. Serial PK for nira were collected on cycle 1 day 1 (C1D1) over 24 h; PK for nira and abiraterone were collected on C2D1 over 10 h. Results: Nira 200 mg + 1000 mg AA+P was selected as the RP2D dose. 4 pts in 200 mg cohort and 8 pts in 300 mg cohort were enrolled in Part 1. During Part 1, 1 patient in the 300 mg cohort had two DLTs: Gr 3 fatigue and Gr 4 increased gamma glutamyltransferase. Therefore, the 200 mg dose was evaluated in 15 pts in Part 2. In 19 total pts treated with 200 mg niraparib, 12 (63%) pts had Gr 3-4 AEs, 5 (26%) pts had an AE with outcome of drug discontinuation and 1 (5%) patient had an AE with outcome of death (general physical health deterioration; unrelated). AEs of special interest were: 6 pts (32%) thrombocytopenia, 6 pts (32%) hypertension, 5 pts (26%) anemia and 3 pts (16%) neutropenia of any grade. 21% pts in 200 mg cohort had treatment emergent serious AE (TESAE) while 50% treated with 300 mg had TESAE. The exposure (AUC24h) on Day 1 of Cycle 2 for 200 mg niraparib with 1000 mg of AA were 17745±9380 ng.h/mL and 712±140 ng.h/mL, respectively. These exposures are consistent with single agent PK. Summaries of all responses will be provided, including a patient with primarily bone disease and carrying an ATM mutation who had a circulating tumor cell conversion and was on study treatment for 22.1 mo. Conclusions: The safety and PK findings support the choice of nira 200 mg in combination with AA+P in pts with mCRPC. The efficacy of nira + AA+P is being evaluated in an ongoing phase 3 MAGNITUDE study (NCT03748641). Clinical trial information: NCT02924766.

Hydrodynamic shear stress promotes epithelial-mesenchymal transition by downregulating ERK and GSK3\u03b2 activities

BackgroundEpithelial-mesenchymal transition (EMT) occurs in the tumor microenvironment and presents an important mechanism of tumor cell intravasation, stemness acquisition, and metastasis. During metastasis, tumor cells enter the circulation to gain access to distant tissues, but how this fluid microenvironment influences cancer cell biology is poorly understood.Methods and resultsHere, we present both in vivo and in vitro evidence that EMT-like transition also occurs in circulating tumor cells (CTCs) as a result of hydrodynamic shear stress (+SS), which promotes conversion of CD24middle/CD44high/CD133middle/CXCR4low/ALDH1low primary patient epithelial tumor cells into specific high sphere-forming CD24low/CD44low/CD133high/CXCR4high/ALDH1high cancer stem-like cells (CSLCs) or tumor-initiating cells (TICs) with elevated tumor progression and metastasis capacity in vitro and in vivo. We demonstrate that conversion of CSLCs/TICs from epithelial tumor cells via +SS is dependent on reactive oxygen species (ROS)/nitric oxide (NO) generation, and suppression of extracellular signal-related kinase (ERK)/glycogen synthase kinase (GSK)3β, a mechanism similar to that operating in embryonic stem cells to prevent their differentiation while promoting self-renewal.ConclusionFluid shear stress experienced during systemic circulation of human breast tumor cells can lead to specific acquisition of mesenchymal stem cell (MSC)-like potential that promotes EMT, mesenchymal-epithelial transition, and metastasis to distant organs. Our data revealed that biomechanical forces appeared to be important microenvironmental factors that not only drive hematopoietic development but also lead to acquisition of CSLCs/TIC potential in cancer metastasis. Our data highlight that +SS is a critical factor that promotes the conversion of CTCs into distinct TICs in blood circulation by endowing plasticity to these cells and by maintaining their self-renewal signaling pathways.

The Promise of Circulating Tumor Cells in Metastatic CRPC

The Promise of Circulating Tumor Cells in Metastatic CRPC

Genomic and phenotypic evidence for prostate cancer osteomimicry in circulating tumor cells from men with metastatic castration resistant prostate cancer (mCRPC) treated with radium-223.

160 Background: Radium-223 is a targeted alpha therapy that improves survival in men with mCRPC. The biologic basis for radium-223 efficacy is not completely understood. We hypothesized that PC osteomimicry, a form of epithelial plasticity leading to an osteoblastic phenotype, may contribute to the intralesional deposition of radium-223 and subsequent irradiation of the tumor microenvironment. Methods: We conducted a pharmacodynamic study of radium-223 in men with bone predominant mCRPC to investigate genomic and phenotypic alterations in circulating tumor cells (CTCs), ctDNA, and metastases. Prior to radium and 3 and 6 months after radium, liquid and metastatic biopsies were collected, including CTCs for phenotypic characterization and CTC/ctDNA genomic analysis. The primary objective was to describe the prevalence of CTC bone alkaline phosphatase (BAP) over time. We measured radium-223 decay products in tumor and surrounding normal bone during treatment. Results: We enrolled 20 men with heavily pre-treated symptomatic bone predominant mCRPC and treated with radium-223 over a median of 6 doses. 55% had elevated serum BAP. PFS was 5.5 mo; OS was 13.3 mo; 10% had unfavorable (≥5) to favorable ( &lt; 5) Cellsearch CTC conversion; 5% had ≥30% PSA decline. We found evidence of persistent CTC BAP expression in both EpCAM+ and EpCAM- CTCs in the majority of men over time during radium treatment. We identified genomic gain of key osteomimicry regions in CTC DNA, including loci for BAP, osteopontin, and OB-cadherin. Radium-223 uptake was observed in tumor to a greater degree than surrounding normal bone. CTC DNA and matched ctDNA and CTC cultures suggested persistence of CTCs with aggressive genomic alterations such as AR, FOXA1, and MYC gain and PTEN and RB1 loss. We established multiple CTC cultures and one CTC PDX; cells exhibited evidence of epithelial plasticity with BAP expression. Conclusions: Osteomimicry may contribute to the uptake of Radium-223 within bone metastases and may thereby enhance the therapeutic benefit of radium-223. We found genomic and phenotypic evidence of osteomimicry in CTCs and CTC cultures from men with mCRPC. Clinical trial information: NCT02204943.

Association of CTC detection by AdnaTest with outcome on enzalutamide in chemotherapy-naïve castration-resistant prostate cancer: Exploratory results from PREMIERE—A SOGUG trial.

5052 Background: Circulating tumor cells (CTCs) enumeration using CellSearch correlates with clinical outcome in prostate cancer, but is limited for gene expression analysis. AdnaTest ProstateCancer is a commercially available CTC immune-enrichment and PCR-related detection method that allows gene expression studies (Antonarakis E, NEJM 2014). It has demonstrated incremental detection of CTCs in patients with no CTCs identified by CellSearch (Samoila A, ASCO 2013) but needs to be clinically qualified. There is a strong need for studies to assess the association with the clinical outcome in CRPC. Methods: Between February and November 2015, 98 asymptomatic or oligo-symptomatic chemotherapy-naïve mCRPC pts were recruited in 16 institutions. Although initially designed to study the predictive value of TMPRSS2-ETS, data emerging after the trial was initiated led the group to prioritize alternative predefined exploratory biomarkers, including plasma AR (Grande E, ASC0 2017 #) and CTC characterization (Grande E, ESMO 2016). Outcome measures included PSA-PFS (sPFS), radiographic PFS (rPFS) and OS. Cox regression was used for survival analyses and Fisher’s exact test for PSA response. Results: Ninety-eight patients had CTC blood samples available. CTCs were detected at baseline, 12 weeks and progression in 36% (35/98), 27% (26/95) and 78% (32/41), respectively. The CTC conversion rate (positive to negative after 12 w) was 43% (15/35). All CTC conversions had ≥50% decline in PSA (15/15) whereas only 35% (7/20) of pts with persistent CTCs. At first interim analysis, with a median follow-up of 10.6 months, detection of CTCs at baseline was associated with worse sPFS (median, 7.59 m versus NR, HR, 3.67; 95% CI 1.90-7.10; P &lt; 0.001), rPFS (median, 12.9 m versus NR; HR, 7.61; 95% CI, 2.80-20.64; P &lt; 0.001) and OS (medians NR, HR, 9.51; 95% CI, 1.11-81.52; P = 0.0398). CTC positive pts were less likely to have a ≥90% decline in PSA (OR, 2.88; 95% CI, 1.13-7.72; P = 0.02). Conclusions: CTC detection using AdnaTest is associated with an adverse outcome in chemotherapy-naïve asymptomatic or oligo-symptomatic mCRPC pts. Clinical trial information: NCT02288936.

Circulating tumor cell (CTC) number as a response endpoint in metastatic castration resistant (mCRPC) compared with PSA across five randomized phase 3 trials.

5007 Background: Radiographic progression and overall survival (OS) are the traditional clinical benefit measures for mCRPC trials. Reliable indicators of response that occur early are a critical unmet need in practice and clinical research. We explored a week 13 CTC and prostate-specific antigen (PSA) endpoint relative to baseline in 5 prospective randomized phase 3 registration trials that enrolled 5912 pts. OS was the primary endpoint. Methods: CTC number (CellSearch) and PSA values in patients who survived at least 13 weeks were evaluated as response endpoints in COU-AA-301, AFFIRM, ELM-PC-5, ELM-PC-4 and COMET-1. Pts with missing values at week 13 were considered non-responders. The endpoints considered are shown in Table 1. Results: The discriminatory strength of the response endpoints with respect to OS was estimated using the weighted c-index. To summarize discrimination results for each measure, the mean and the standard deviation based on the weighted c-indices from the 5 studies were computed. Conclusions: CTC0 and CTC conversion endpoints had the highest discriminatory power for OS relative to the % decline in CTC or PSA endpoints. The percent of pts eligible and evaluable for the CTC0 endpoint was significantly higher than the conversion endpoint, 75% vs. 51%, respectively. These two absolute measures of CTC can be considered meaningful response indicators in mCRPC clinical trials. [Table: see text] [Table: see text]

Association of changes in circulating cell-free plasma DNA (cfDNA) and circulating tumor cells (CTC) during treatment with clinical outcome from olaparib in castration-resistant prostate cancer (CRPC): Exploratory analyses from the TOPARP-A trial.

141 Background: Response biomarkers are needed to optimize treatment switch decisions in CRPC patients. CTC and cfDNA may have clinical utility as response biomarkers; we studied them during olaparib treatment in a phase II trial in CRCP (Mateo et al NEJM 2015). Methods: CTC were enumerated using CellSearch (Jannsen Diagnostics) and cfDNA was extracted with the QIASymphony circulating DNA kit (Qiagen) from blood samples taken at baseline, 4- and 8-weeks (wk) of therapy. Radiological progression-free survival (rPFS) was defined as time from starting treatment to progression by RECIST 1.1, bone scan (PCWG2) or death. Overall survival (OS) was defined as time from starting treatment to death. CTC changes were categorized based on conversion from ≥ 5 to &lt; 5 CTC/7.5ml blood and on ≥ 30% decline (Lorente et al Eur Urol 2016). cfDNA changes were evaluated as percentage change from baseline (continuous and binary). The prognostic value of CTC and cfDNA changes were assessed by Landmark analysis and Cox models with time-varying covariates; p-value &lt; 0.01 were considered significant to account for multiple tests. Results: Overall, 13/47 (28%) and 16/42 (38%) evaluable patients had a CTC conversion at 4- and 8-wk respectively. A CTC conversion after 4-wk of olaparib associated with longer rPFS (median 8.9 vs 2.7 months [m], p = 0.001); a similar association was found at 8-wk. A 30% CTC decline at 4-wk also associated with longer rPFS (median 4.4 vs 2.6 m, p = 0.004). CTC conversion as a time-varying covariate associated with longer OS (HR 0.26, 95%CI 0.14-0.50, p &lt; 0.001). Median baseline cfDNA was 31.6 ng/ml (IQR 19.4-57.1); 46 and 42 patients were evaluable for cfDNA changes at 4- and 8-wk. Percentage changes in cfDNA at 4- and 8- wk associated with rPFS (HR 1.01 and 1.005; p = 0.005 and 0.002 respectively) but association with OS was not significant. cfDNA declines ≥ 50% at 8- wk associated with longer rPFS (median 8.9 vs 2.7 m, p = 0.007) and OS (17.0 vs 10.1 m, p = 0.004). Conclusions: Decreases in CTC counts and cfDNA concentration associate with improved outcome from olaparib (rPFS, OS) in the TOPARP-A trial. Clinical trial information: NCT01682772.

A phase I dose-escalation study of enzalutamide in combination with the AKT inhibitor AZD5363 in patients with mCRPC.

135 Background: Activation of the PI3K/AKT/mTOR pathway may contribute to resistance to androgen receptor targeted therapies in metastatic CRPC (mCRPC). The phase I/II RE-AKT trial (NCT02525068) investigates the safety and activity of enzalutamide (enza) in combination with the AKT inhibitor, AZD5363, in patients (pts) with mCRPC. Results of the phase I run-in are reported. Methods: mCRPC pts progressing after 1-2 lines of taxane chemotherapy and at least 12 weeks (wks) of abiraterone or enza were treated with enza (160mg od) and AZD5363 bid 4-days on, 3-days off, in a 3+3 dose escalation design. Co-primary endpoints were to assess toxicity (CTCAE), and identify the recommended phase II dose (RP2D) of AZD5363; antitumour activity and pharmacokinetics (PK) were secondary endpoints. Response was assessed by PSA, RECIST v1.1 and circulating tumor cell (CTC) conversion. Pts were considered evaluable for response if they completed 12 wks of treatment. Results: 16 pts were enrolled between 12/2014 &amp; 04/2016 with 15 receiving treatment. At the AZD5363 320mg dose 3 pts were treated with no dose limiting toxicity (DLT). At the AZD5363 480mg dose, 5 pts were treated with 2 DLTs of grade (G) 3 maculopapular rash (MPR) related to AZD5363. An intermediate dose level of AZD5363 400mg was selected with 7 pts treated. 1 pt withdrew consent prior to completing the DLT window. 1 DLT of G3 MPR occurred and this dose was selected as the RP2D. Non-DLT treatment related (TR) G3/4 adverse events (AEs) were hyperglycaemia (n = 4, 26.7%), neutropenia (n = 1, 6.7%) &amp; diarrhea (n = 1, 6.7%). All other TR AEs were G1 or G2, with diarrhea (n = 9, 60%), anorexia (n = 8, 53.3%) &amp; nausea (n = 7, 46.7%) being most common. Of the 10 pts who completed 12 wks of treatment, 3 met at least one of the criteria for response. 1 pt (AZD5363 320mg) who had previously progressed on enza exhibited RECIST v1.1 partial response, &gt; 50% PSA response, and CTC conversion by wk 13. Enza decreased AZD5363 PK exposure; robust modulation of pS6, pGSK3b and pPRAS40 was demonstrated. Conclusions: AZD5363 at the RP2D of 400mg bid 4 days on, 3 days off combined with enza 160mg od is safe and tolerable. Antitumor activity is reported, suggesting that AZD5363 may be able to overcome resistance to enza. Clinical trial information: NCT02525068.

The End of the Beginning: Circulating Tumor Cells As a Biomarker in Castration-Resistant Prostate Cancer

Although circulating tumor cells (CTCs) were first observed from patients with metastatic malignancies more than 100 years ago, only recently has the clinical and research potential of CTCs become widely recognized. Because CTCs are thought to represent cells that are shed by primary or metastatic tumors, significant interest has focused on examining CTCs as prognostic or predictive biomarkers in several malignancies, including breast and prostate cancer. One could hypothesize that the number of CTCs could be prognostically important by potentially estimating the total-body burden of disease and/or a tumor’s invasiveness. Alternatively, changes in CTC counts might indicate sensitivity or resistance to an anticancer therapy. CTCs could also represent research tools to personalize treatment by serving as a so-called liquid biopsy of an individual’s disease. Quantitation and characterization of CTCs may therefore represent a means to directly inform clinical care but also serve as an investigational platform to elucidate basic mechanisms of tumor biology. The most extensively studied clinical role for CTCs is as a prognostic biomarker; that is, counts at a specific time point are associated with a later discrete clinical end point. In this capacity, CTC counts that are enumerated using the CellSearch platform (Veridex, Raritan, NJ) are the most studied. CellSearch enriches for CTCs with antibodies directed against the epithelial cell adhesion molecule (EpCAM), which is expressed on epithelially derived cells. CTCs are then identified and quantitated on the basis of nuclear staining with 4,6diamidino-2-phenylindole (DAPI), and anticytokeratin and antiCD45 antibodies, which distinguishes CD45 epithelial cells from CD45 leukocytes. In clinical trials, CellSearch CTC counts are prognostic for overall survival (OS) in metastatic breast, colon, and prostate cancers. In prostate cancer, specifically, the Circulating Tumor Cells and the Prediction of Overall Survival in Patients With Androgen Independent Prostate Cancer Entering Onto Chemotherapy (IMMC38) study demonstrated the prognostic value of CellSearch CTCs in 238 men with metastatic castration-resistant disease (mCRPC) starting a new first, second, or third line of chemotherapy. In this study, a pretreatment CTC threshold of 5 CTCs/7.5 mL of blood distinguished patients into favorable and unfavorable groups, the latter having shorter median OS (11.5 v 21.7 months; hazard ratio, 3.3; P .001). In their modeling, the best predictor of survival incorporated baseline high-serum lactate dehydrogenase (LDH) levels with baseline CTC counts; the addition of prostate-specific antigen (PSA) did not improve prediction of survival. In patients with unfavorable baseline counts, conversion to a favorable count after the initiation of treatment was also associated with improved survival. Conversely, conversion to unfavorable counts after treatment initiation was associated with worse survival. Perhaps most importantly, CTC counts outperformed PSA in predicting survival at all time points. More recently, the prognostic value of 5 CTCs/7.5 mL of blood (with the CellSearch assay) was confirmed in a randomized phase III study of abiraterone acetate, a novel hormonal therapy, in men with mCRPC who were previously treated with docetaxel. In this study, CTCs were enumerated at baseline and during the first three cycles. CTC conversion from 5 CTCs to 5 CTCs, along with changes in serum LDH, was strongly predictive of OS. Similar results were recently observed in a phase III study of docetaxel with or without lenalidomide. Building on these findings, in the article that accompanies this editorial, Goldkorn et al present an analysis of the impact of baseline and post-treatment CTC counts on OS and disease response in a subset of men with mCRPC who were treated as part of the Southwest Oncology Group (SWOG) trial 0421, a phase III study comparing the effectiveness of docetaxel plus atrasentan, an endothelin receptor antagonist, versus docetaxel alone. The study itself failed to meet its coprimary end points of improved OS and progression-free survival with the addition of atrasentan and was halted early for futility. Nevertheless, as part of the study, CTCs were enumerated at baseline and 21 days after the first dose of the study treatment. In contrast to the IMMC-38 study, this study had a uniform cohort of men with chemotherapy-naive CRPC and a longer duration of median follow-up (24 v 16 months). In their analysis, the authors showed that baseline CTC counts were correlated with recognized prognostic markers, including PSA, alkaline phosphatase, bone pain, liver disease, and hemoglobin, although relationship with LDH, a widely recognized prognostic marker, was not assessed. Baseline CTC count was associated with subsequent PSA declines and with objective responses. Importantly, median survival for patients with 5 CTCs/7.5 mL of blood at baseline was far better than for those with 5 CTCs (26 v 13 months), and after adjustment for known prognostic factors, the baseline CTC JOURNAL OF CLINICAL ONCOLOGY E D I T O R I A L VOLUME 32 NUMBER 11 APRIL 1

A phase II trial of prostate-specific membrane antigen antibody drug conjugate (PSMA ADC) in taxane-refractory metastatic castration-resistant prostate cancer (mCRPC).

83 Background: The abundant expression of prostate-specific membrane antigen (PSMA) on prostate cancer cells provides a rationale for antibody therapy. PSMA antibody drug conjugate (ADC) is a fully human antibody to PSMA linked to the microtubule disrupting agent monomethyl auristatin E (MMAE). It binds PSMA and is internalized and cleaved by lysosomal enzymes releasing free MMAE causing cell cycle arrest and apoptosis. We enrolled 70 patients (pts) in a phase II trial of PSMA ADC in taxane-refractory metastatic castration-resistant prostate cancer (mCRPC). Methods: Pts with progressive mCRPC following taxane and ECOG PS 0 or 1 were eligible. PSMA ADC was administered Q3 week IV for up to eight cycles. Safety, tumor response by prostate-specific antigen (PSA), circulating tumor cells (CTC), imaging, biomarkers and clinical progression were assessed. Dosing was initiated at 2.5 mg/kg and adjustment for tolerability was allowed. Results: Thirty five pts began treatment at 2.5 mg/kg. Due to neutropenia, the remaining 35 pts began at 2.3 mg/kg. All pts received prior docetaxel and abiraterone and/or enzalutamide. Forty one percent also received cabazitaxel. Adverse events (AEs) were consistent with what was seen in phase I; most common significant AEs were neutropenia (grade 4, 6.7% and 11.4% at 2.3 and 2.5 mg/kg, respectively) and peripheral neuropathy (grade 3 or higher, 6.7% (2.3) and 5.7% (2.5)). Two pts at 2.5 mg/kg died of sepsis. 43% of pts at 2.3 and 37% of pts at 2.5 had declines in CTC from 5 or more to less than 5 cells/7.5 ml blood and 57.1% (2.3) and 74.1% (2.5) had 50% or more CTC declines; 26.1% (2.3) and 16.1% (2.5) had PSA declines of 30% or more thus far. PSA and CTC responses were associated with higher PSMA expression on CTC and lower neuroendocrine (NE) markers. The CTC conversion rate (5 or more to less than 5) was approximately 80% in pts with low NE markers. Prior cabazitaxel or abiraterone and/or enzalutamide did not appear to affect response. Centralized assessments of images by RECIST of all pts are currently planned and will be presented. Conclusions: PSMA ADC at 2.3 mg/kg was generally well tolerated in pts with progressive mCRPC previously treated with taxane. Anti-tumor activity, CTC and PSA reductions were observed at 2.3 and 2.5 mg/kg. Updated safety, tumor response and radiographic assessments from the full cohorts of 2.3 and 2.5 mg/kg will be presented. Testing in taxane naïve pts is also ongoing. Clinical trial information: NCT01695044.

Sequential monitoring and characterization of circulating tumor cells (CTCs) using the epic sciences platform in metastatic castration-resistant prostate cancer (mCRPC) patients (pts) treated with recently approved therapeutics.

78 Background: The enumeration and molecular characterization of circulating tumor cells (CTCs) may allow sequential evaluation of drug-induced changes and mechanisms of drug resistance that cannot be practically performed by tissue biopsies. Methods: We utilized the Epic Sciences platform to identify and characterize traditional CTCs (CK-CD45+) and other CTC subpopulations (CK-CD45- CTCs and small CTCs) in sequential samples obtained from metastatic castration-resistant prostate cancer (mCRPC) patients (pts) receiving abiraterone acetate (AA), enzalutamide (E), or cabazitaxel (C). Peripheral blood was collected pre-treatment, during treatment (at 1 to 4 weeks, to evaluate pharmacodynamics effects) and at progression. Nucleated cells were plated onto glass slides and subjected to immunofluorescence staining for CK, CD45, and AR, and CTC identification by fluorescent scanners. CTCs from a subset of patients were tested for PTEN loss and ERG rearrangements by FISH. CTCs from a second blood sample collected at the same time were captured and enumerated using CellSearch (Veridex). Fresh CRPC biopsies were available for 9 pts. Results: Initial results are available from 24 mCRPC pts (16 pts treated with AA; four with E; four with C; 17 out of 24 previously treated with docetaxel). Pre-treatment, the median CK+CD45- CTC count was 3.5 (range 0 to 98) cells/mL blood. Eleven pts had more than or equal to 5 CK+CD45- CTCs/mL pre-treatment and 5 out of 11 pts had an Epic CTC conversion to less than 5 CK+CD45- CTCs/mL with treatment. 8 out of 11 pts had a decline in CK+CD45- CTC count greater than 30%. Changes in AR expression and AR subcellular localization were observed during treatment as well as changes in count of CTC subpopulations. CellSearch count was available for 22 out of 24 pts. Twelve out of 22 had more than or equal to five CTCs/7.5mls captured using CellSearch, including 7 out of 12 with more than or equal to five CTCs/ml using Epic platform. Conclusions: Enumeration and molecular characterization of CTCs on the Epic Sciences platform during treatment is feasible and demonstrates drug-induced changes associated with changes in CTC count, AR expression and AR subcellular localization. These assays warrant further studies to validate their role as clinically useful biomarkers in mCRPC.

An exploratory analysis of bone scan lesion area (BSLA), circulating tumor cell (CTC) change, pain reduction, and overall survival (OS) in patients (pts) with castration-resistant prostate cancer (CRPC) treated with cabozantinib (cabo): Updated results of a phase II nonrandomized expansion (NRE) cohort.

5026 Background: The results of 144 pts with metastatic CRPC treated in a phase II NRE cohort with daily cabo 100 mg and 40 mg starting doses have been previously reported. Substantial rates of bone scan improvement, reductions in CTC counts and pain relief were observed. To better understand the implication of these effects, the association with OS was explored. Methods: Relevant baseline variables (LDH, BSLA, visceral disease, pain, hemoglobin, CTCs) and post-treatment changes at week 6: ≥30% reduction in BSLA using computer-aided assessment, CTC conversion (&gt;5 vs. 4 or less/7.5 ml of blood), and pain intensity (7 day averaged worst pain score; BPI scale; using an IVR system) were associated with OS in 144 CRPC pts with bone metastasis who progressed within 6 months of docetaxel (D) treatment (≥225 mg/m2) in either bone or soft tissue. Median OS was compared between responders and non-responders for each of the above outcomes categories using a Cox proportional hazard model. The findings were examined further after adjusting for significant baseline covariates selected from a stepwise Cox regression model. Results: See Table. Conclusions: Recognizing the limitations of associating response with survival, this retrospective analysis of decreases in BSLA, CTC conversions and reductions in pain intensity support further study in ongoing phase III trials. Clinical trial information: NCT00940225. [Table: see text]

TERRAIN: A randomized, double-blind, phase II study comparing MDV3100 with bicalutamide (Bic) in men with metastatic castrate-resistant prostate cancer (CRPC).

TPS4698^ Background: MDV3100 is a novel androgen receptor (AR) signaling inhibitor (ARSI) in clinical development for treatment of prostate cancer (PCa). Compared with Bic in nonclinical CRPC models, MDV3100 showed higher affinity AR binding, inhibited nuclear translocation and AR-DNA binding, showed no evidence of AR agonism, and caused tumor regression in Bic-resistant xenografts. MDV3100 has shown antitumor activity in men with advanced PCa. We present the study design of a trial comparing MDV3100 and Bic in men with progressive metastatic PCa. Methods: Multinational study (Table) with planned enrollment of 370 patients (randomized 1:1 to MDV3100 160 mg/d or Bic 50 mg/d). Inclusion criteria include metastatic (≥2 bone lesions or soft tissue disease at screening) progressive CRPC (≥3 rising prostate-specific antigen [PSA] levels or new bone/soft tissue disease), ongoing stable gonadotropin-releasing hormone (GnRH) analog therapy or surgical castration (serum testosterone &lt;50 ng/dL), Eastern Cooperative Oncology Group performance status 0–1, and life expectancy ≥1 year. Exclusion criteria include previous chemotherapy, current/prior antiandrogens (except if administered for &lt;12 wk and discontinued no less than 6 mo before study). Patients will be stratified by time of bilateral orchiectomy or GnRH analog initiation (before/after diagnosis of metastases) and will receive treatment until occurrence of radiographic progression/skeletal-related event, start of new antineoplastic therapy, or development of an adverse event requiring discontinuation. Results: The primary endpoint is progression-free survival; secondary endpoints are safety, PSA response, and time to PSA progression. Exploratory endpoints include circulating tumor cell conversion rate and quality of life. Patients are currently enrolling. Conclusions: This ongoing, head-to-head, phase II trial is the first to prospectively assess whether MDV3100 can provide improved antitumor effects vs Bic in men with metastatic CRPC. [Table: see text]

Evaluation of circulating tumor cells (CTCs) enumeration and 18F-Choline positron emission tomography/computed tomography (FCH PET/CT) as early efficacy response biomarkers in metastatic castration-resistant prostate cancer (CRPC) patients (pts) treated with abiraterone acetate.

63 Background: Circulating tumor cells (CTCs) and FCH PET/CT are both promising tools for treatment monitoring in CRPC pts. In this study, we assessed prostate specific antigen (PSA), CTC and FCH PET/CT for monitoring treatment response in metastatic CRPC pts receiving recently approved abiraterone acetate therapy after chemotherapy. Methods: Twenty nine metastatic CRPC pts progressing after docetaxel chemotherapy received abiraterone acetate 1,000 mg daily with prednisone 5 mg twice daily in continuous 28-day cycles. Pts were evaluated monthly for efficacy and safety. CTCs counts (using the CellSearch assay) and FCH PET/CT were performed at baseline and at 4 weeks (PERCIST criteria and SUV Peak). Results: After one month of treatment, 12 pts (41%) had a PSA rise and 17 pts (59%) had a PSA decline. At baseline, 18 pts had detectable CTCs (at least 5 CTCs), and 6 of them (33%) had a decline from at least 5 to less than 5 CTCs after 4 weeks of treatment. 10 of 29 pts (34%) had a &gt; or = 30% decline in CTCs. Amongst 22 pts who were evaluable by FCH PET/CT, response assessment was as follows: response (n=8), stable disease (n=12), progression (n=1). PSA decline and CTCs levels correlated in 16 of 29 pts (55%), and were discordant in 13 of 29 pts (45%). In all 12/29 pts (41%) with a 50% or more decline in PSA both a CTCs response and a FCH PET/CT response was observed. In patients with a PSA rise while on abiraterone acetate, 8 pts did not have a CTCs increase or a CTC conversion from unfavourable to favorable, and 7 of 8 pts had a PSA decline at 3 months or a FCH PET/CT response. No flare phenomenon on FCH PET/CT was observed. Conclusions: Monitoring pts with CRPC who receive abiraterone using both CTCs and metabolic imaging indicate discrepancy in 45% with response assessment using PSA. Association between CTC drop, early FCH PET/CT response and clinical benefit is currently under study.

Abstract C9: Treatment response to abiraterone acetate (AA) in patients with castration-resistant prostate cancer (CRPC) based on circulating tumor cells (CTCs), prostate-specific antigen (PSA), and imaging

Abstract Background: AA, a potent oral irreversible inhibitor of CYP17A1, has been recently approved for patients with CRPC progressing after docetaxel. CTC count ≥ 5/7.5 ml correlates with worse survival in CRPC. In this analysis we evaluated PSA responses, radiological responses and CTC conversion rates in relation to patient outcome. Methods: 141 patients (ECOG Performance Status 0-2; Median Age: 69.7; 85 post-docetaxel, 56 pre-docetaxel) treated with AA at the Royal Marsden NHS Foundation Trusts between December 2005 and March 2011 were included in this retrospective analyses. CTCs were enumerated using CellSearch at screening, baseline, at multiple time points during the study and at treatment discontinuation. PSA and imaging (Computer Tomography and Bone Scans) were obtained at the same time points as CTC collection. The Kaplan-Meier method was utilized for these analyses. Results: Baseline CTCs of ≥ 5 cells/7.5 ml were detectable in 17/53 (32.1%) pre-docetaxel patients (3 inevaluable) and 58/77 (75.3%) post-docetaxel patients (8 inevaluable at baseline). CTC conversion (≥ 5 to &amp;lt; 5 cells/7.5 ml) was observed in 10/17 (58.8%) pre-docetaxel patients and in 12/52 (23.1%) (6 inevaluable) post-docetaxel patients. PSA responses by PCWG II were observed in 16/22 (72.7%) of patients who attained a CTC conversion. PSA responses were seen in 8/47 (17.0%) patients in the absence of a corresponding CTC conversion. The median duration of radiological SD was 12.13 months (n=55) and 5.4 months (n=75) in patients with a baseline CTCs count of ≤ 5 cells/7.5mls and ≥ 5 cells/7.5 mls respectively. Overall, radiological stable disease was observed for a median of 6 months (95% CI: 4.2, 8.4; range 0.3, 35.6) beyond PSA progression in the 105 patients who experienced PSA progression. Radiological and clinical disease stability during treatment of ≥ 1 year, ≥ 2 years and ≥ 3 years was observed in 43/141 (30.5%), 21/141 (14.9%) and 12/141 (8.5%) respectively. Conclusion: Radiological and clinical disease stabilization beyond PSA progression is seen in a significant proportion of patients treated with AA. CTCs conversion correlates with PSA response and radiological disease stability. Citation Format: Diletta Bianchini, Alison Reid, Gerhardt Attard, Johann De Bono, Shahneen Sandhu, Amy Cassidy Mulick, Deborah Mukherji, Carmel Pezaro, Andrea Zivi, Aurelius Omlin, Ajit Sarvadikar, Emilda Thompson. Treatment response to abiraterone acetate (AA) in patients with castration-resistant prostate cancer (CRPC) based on circulating tumor cells (CTCs), prostate-specific antigen (PSA), and imaging [abstract]. In: Proceedings of the AACR Special Conference on Advances in Prostate Cancer Research; 2012 Feb 6-9; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(4 Suppl):Abstract nr C9.