In recent years, there has been increasing interest in developing drugs that increase the precision targeting of molecular alterations in prostate cancer cells. Traditionally, tissue samples are used to screen for and study such alterations, but liquid biopsies emerge as very practical and potentially superior methodologies to longitudinally characterize cancer cell biology and evolution. The expression of androgen receptor variants correlates with the development of castrate resistance prostate cancer (CRPC), and the most studied variant 7 (AR-V7) is associated with CRPC, disease progression, and resistance to certain AR-pathway targeted treatments and is readily detectable in circulating tumor cells (CTCs). Here, we describe a method of AR-V7 detection in CTCs using highly sensitive droplet digital PCR (ddPCR).

Precision medicine prostate cancer drug development, which targets patient-specific molecular alterations in prostate cancer cells, is rapidly progressing. Traditionally, tissue samples are used to screen for and study molecular alterations, but liquid biopsies are emerging as practical and potentially superior methodologies to longitudinally characterize cancer cell biology and evolution. The expression of androgen receptor variants correlates with the development of castrate resistance prostate cancer (CRPC), and the most studied variant 7 (AR-V7) is associated with CRPC, disease progression, and resistance to certain AR-pathway targeted treatments and is readily detectable in circulating tumor cells (CTCs). While mainly detected at the transcript level, subcellular localization of the protein defines its activity as a transcription factor, and emerging evidence suggests that detection of nuclear AR-V7 in CTCs is central to its utility as a poor-prognosis biomarker. Thus, transcript and protein detection of the AR-V7 biomarker in CTCs may complement each other to achieve the highest sensitivity and specificity. Here, we describe a method of AR-V7 protein detection in CTCs using immunocytostaining.

Background: Melanoma patients with pre-existing autoimmune diseases (AIDs) have been excluded from clinical trials of immune checkpoint inhibitors (ICIs), due to a risk of flare and immune-related adverse effects (irAEs). Materials and methods: A comprehensive literature search of Medline, Embase, CINAHL and Scopus was carried out. Studies of melanoma patients with pre-existing AIDs were included. Two reviewers used the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines. Outcomes assessed included all-grade and grade 3 or 4 de novo irAEs, AID flare, treatment discontinuation owing to adverse effect(s), treatment-related mortality and objective response rate (ORR). Results: Thirty-two percent of patients with AIDs developed a flare, with 80% requiring immunosuppression. In patients with AIDs, meta-analysis revealed a higher risk of all-grade irAEs [risk ratio (RR) 1.16, 95% confidence interval (CI) 1.01-1.33, P = 0.0106] but not grade 3 or 4 (RR 1.21, 95% CI 0.99-1.47, P = 0.0663). Treatment discontinuation, treatment-related mortality and ORR were not different among patients with AIDs. Conclusions: Melanoma patients with pre-existing AIDs are at significant risk of flare with ICI use. These patients had an increased risk of irAEs of any grade. Although there was a trend towards increased risk of severe irAEs, this was not statistically significant. irAEs did not result in treatment-related deaths, and there were no differences in treatment response. Risks should be discussed, coupled with close monitoring.

PURPOSE:Adherence to active surveillance in patients with stage 1 testicular cancers may be influenced by factors affecting capacity and motivation to attend appointments. The aims of this study were to assess adherence to active surveillance and analyze factors which may impact adherence. PATIENTS AND METHODS:A retrospective cohort study was conducted in patients diagnosed with stage 1 testicular cancer between 2005 and 2020, and managed with active surveillance at 3 institutions in South Western Sydney, Australia. Adherence with active surveillance was followed to 2023 and patients were subsequently classified into 3 groups: "Optimal," "Adequate" or "Loss to follow-up" (LTFU). Factors for adherence were analyzed using multivariable logistic regression. Disease recurrence was analyzed using multivariable Cox regression. RESULTS:In 125 patients, adherence with active surveillance was assessed as "Optimal" in 64 (51%), "Adequate" in 14 (11%), and LTFU in 47 (38%). Multivariable analysis demonstrated that patients had higher odds of being in the "Optimal" or "Adequate" categories if they were from a culturally and linguistically diverse background (OR 4.86, P = .026), nonsmokers (OR 7.63, P = .0002), not employed (OR 4.93, P = .0085), had a partner (OR 2.74, P = .0326), or were diagnosed after June 2016 (OR 5.22, P = .0016). Recurrence occurred in 21 patients (17%). The risk of recurrence increased with the presence of multiple pathological risk factors (HR 5.77, P = .0032), if patients were unemployed (HR 2.57, P = .032), or if they had "Optimal" or "Adequate" adherence (HR 12.74, P = .0136). CONCLUSION:Adherence with active surveillance was poorer in this cohort of stage 1 testicular cancer patients. Patients from culturally and linguistically diverse backgrounds and those who were nonsmokers, unemployed, with a partner, and later date of diagnosis, were more likely to be adherent with active surveillance.

Background Lutetium-177 [Lu-177]Lu-prostate-specific membrane antigen (PSMA)-617 improves survival and quality of life in patients with metastatic castration-resistant prostate cancer, but whether it confers a benefit in hormone-sensitive disease is unknown. We aimed to evaluate [Lu-177]Lu-PSMA-617 before docetaxel treatment in patients with de-novo high-volume metastatic hormone-sensitive prostate cancer. Methods UpFrontPSMA was an investigator-initiated, multicentre, open-label, randomised, phase 2 trial done at 11 Australian hospitals. Eligible patients had prostate adenocarcinoma without clinically significant neuroendocrine differentiation or small-cell histology, were aged 18 years or older, had less than 4 weeks on androgen deprivation therapy, had an Eastern Cooperative Oncology Group performance status of 0-2, and had high-volume PSMA-avid disease on [Ga-68]Ga-PSMA-11 PET-CT with no major discordance on 2-[F-18] fluorodeoxyglucose-PET-CT. Patients were randomly assigned (1:1) to the experimental treatment ([Lu-177]Lu-PSMA-617 followed 6 weeks later by docetaxel) or standard-of-care treatment (docetaxel alone) using computer-based block randomisation with random block sizes, stratified by disease volume by conventional imaging and duration of androgen deprivation therapy at the time of registration. Neither patients nor investigators were masked to treatment assignment. Patients in the experimental group received two cycles of [Lu-177]Lu-PSMA-617 75 GBq every 6 weeks intravenously, followed 6 weeks later by six cycles of docetaxel 75 mg/m(2) every 3 weeks intravenously, whereas patients in the standard-of-care treatment group received six cycles of docetaxel 75 mg/m(2) every 3 weeks intravenously. All patients received continuous androgen deprivation therapy. The primary endpoint was undetectable prostate-specific antigen (<= 02 ng/mL) at 48 weeks, assessed using a modified intention-to-treat analysis. The trial is registered with ClinicalTrials.gov, NCT04343885. Findings Between May 5, 2020, and April 18, 2023, 130 patients were randomly assigned, 63 (48%) to [Lu-177]Lu-PSMA-617 plus docetaxel and 67 (52%) to docetaxel alone. All patients were male and no race or ethnicity data were collected. Median follow-up was 25 years (IQR 18-30). Four patients in the docetaxel alone group withdrew consent after randomisation and no data beyond screening were collected. An additional four patients were not evaluable for the primary endpoint at 48 weeks (two in each group). 25 (41%) of 61 patients (95% CI 30-54) in the [Lu-177]Lu-PSMA-617 plus docetaxel group had undetectable PSA at 48 weeks compared with ten (16%) of 61 patients (9-28) in the docetaxel alone group (OR 388, 95% CI 161-938; p=00020). The most common grade 3 or 4 treatment-related adverse events were febrile neutropenia (seven [11%] of 63 patients in the [Lu-177]Lu-PSMA-617 plus docetaxel group vs six [10%] of 63 patients in the docetaxel alone group) and diarrhoea (four [6%] of 63 patients vs none). Serious adverse events occurred in 16 (25%) patients in the [Lu-177]Lu-PSMA-617 plus docetaxel group (none were definitely related to [Lu-177]Lu-PSMA-617) and 16 (25%) patients in the docetaxel alone group. No treatment-related deaths occurred. Interpretation [Lu-177]Lu-PSMA-617 followed by docetaxel improved antitumour activity in patients with de-novo high-volume metastatic hormone-sensitive prostate cancer compared with docetaxel alone, without increased toxic effects. Our data potentially support a role for [Lu-177]Lu-PSMA-617 in metastatic hormone-sensitive prostate cancer.

CYP2D6 genotype and phenotype for each subject

Endoxifen level for patients on moderate or strong CYP2D6 inhibitors

Immune checkpoint inhibitors (ICIs) have transformed the landscape of solid cancer management. These drugs carry a risk of novel side effects, which have become known as immune-related adverse events (irAEs).Traditionally, irAEs have been managed empirically with corticosteroids. A subset of these may be steroid refractory and as more evidence emerges about their distinct pathogeneses, a more tailored approach is required. Here, we report the use of a Janus kinase (JAK) inhibitor, baricitinib, in a patient with chronic inflammatory demyelinating polyneuropathy secondary to ICI use. We also review the current literature with regards to the use of these inhibitors in the management of irAEs.Modulation of the JAK pathway warrants further investigation in the targeted management of irAEs.

Frequency of CYP2D6 alleles in study subjects compared to published frequencies for selected race/ ethnic groups

Change in tamoxifen and metabolite levels in response to tamoxifen dose increase

PARPi have become new treatments in mCRPC. Toxicities of treatment are less well established. Our review encompassess all published studies of PARPi in mCRPC outlining both hematological and non-hematological toxicities. Overall PARPi appear to be have consistent hematological toxicities across malignancies with an overall low toxicity burden. There is emerging evidence for the use of poly (ADP-ribose) polymerase inhibitors (PARPi) in patients with mCRPC with patients harboring germline or somatic mutations deriving clinical benefit. However, the toxicity profile of PARPi in mCRPC is not well established. In March 2022 a literature search was conducted across 4 databases - Medline, PubMed, Cochrane Library and Embase. In total, 14 relevant studies were identified cumulating in 2066 patients that were treated with PARPi. The overall ORR to PARPi alone or in combination with other therapy was 37% (246/666). In 5trials that investigated PARPi alone, the ORR was 39% (141/361). Treatment emergent adverse events (TEAEs) of any grade were reported in 96% (1034/1080) in PARPi treatment arms. TEAEs of grade > = 3 were reported in 57% (611/1080). 45% (457/1006) experienced treatment interruption whilst 31% (310/989) required dose reductions. 11% (114/1006) of patients had their treatment discontinued directly as the result of toxicity associated with the trial medications. The most common hematological toxicity was anemia, reported in 490/1160 (42%) patients. and lowered white blood cell count were the next 2most common toxicities, reported in 186/655 (28%) and 133/729 (18%) respectively. The 3most common non-hematological toxicities reported were nausea, fatigue and anorexia reported in 440/1013 (43%), 340/1013 (34%) and 274/1013 (27%) patients respectively. Overall, TRAEs associated with individual PARPi are still emerging with hematological toxicities being most apparent. Further toxicities will be informed from future clinical trials to allow improved treatment selection, education and management of toxicities in prostate cancer.

Androgen receptor variant 7 (AR-V7) is an important biomarker to guide treatment options for castration-resistant prostate cancer (CRPC) patients. Its detectability in circulating tumour cells (CTCs) opens non-invasive diagnostic avenues. While detectable at the transcript level, AR-V7 protein detection in CTCs may add additional information and clinical relevance. The aim of this study was to compare commercially available anti-AR-V7 antibodies and establish reliable AR-V7 immunocytostaining applicable to CTCs from prostate cancer (PCa) patients. We compared seven AR-V7 antibodies by western blotting and immmunocytostaining using a set of PCa cell lines with known AR/AR-V7 status. The emerging best antibody was validated for detection of CRPC patient CTCs enriched by negative depletion of leucocytes. The anti-AR-V7 antibody, clone E308L emerged as the best antibody in regard to signal to noise ratio with a specific nuclear signal. Moreover, this antibody detects CRPC CTCs more efficiently compared to an antibody previously shown to detect AR-V7 CTCs. We have determined the best antibody for AR-V7 detection of CTCs, which will open future studies to correlate AR-V7 subcellular localization and potential co-localization with other proteins and cellular structures to patient outcomes.

Androgen Receptor (AR) alterations (amplification, point mutations, and splice variants) are master players in metastatic castration resistant prostate cancer (CRPC) progression and central therapeutic targets for patient management. Here, we have developed two multiplexed droplet digital PCR (ddPCR) assays to detect AR copy number (CN) and the key point mutation T877A. Overcoming challenges of determining gene amplification from liquid biopsies, these assays cross-validate each other to produce reliable AR amplification and mutation data from plasma cell free DNA (cfDNA) of advanced prostate cancer (PC) patients. Analyzing a mixed PC patient cohort consisting of CRPC and hormone sensitive prostate cancer (HSPC) patients showed that 19% (9/47) patients had AR CN amplification. As expected, only CRPC patients were positive for AR amplification, while interestingly the T877A mutation was identified in two patients still considered HSPC at the time. The ddPCR based analysis of AR alterations in cfDNA is highly economic, feasible, and informative to provide biomarker detection that may help to decide on the best follow-up therapy for CRPC patients.

Prostate adenocarcinoma with testicular metastasis is rare, present in up to 4% of autopsy diagnoses, and presents symptomatically in less than 0.5% of cases. We report an unusual case of a 55-year-old male who developed a symptomatic testicular metastasis from primary prostate cancer 4 years after initial diagnosis of metastatic castrate-sensitive prostate cancer with nodal and bone-only involvement. The patient had orchidectomy, histologically confirming the metastasis and revealing sparing of the spermatic cord. Prior treatment for his metastatic castrate-sensitive prostate cancer had included androgen deprivation therapy and upfront docetaxel chemotherapy. He had received palliative radiotherapy for symptomatic bone metastasis and managed on enzalutamide for castrate-resistant disease for the preceding 22 months with ongoing PSA response at the time of diagnosis of new testicular metastasis, with a further significant PSA response following his “testicular metastasectomy.” At the time of diagnosis of testicular metastasis, he did not have any evidence of other visceral metastases, and his metastatic disease otherwise remained radiologically stable. We describe his disease course, treatment and outline the rare nature of his case of testicular metastasis from prostate cancer.

Molecular testing of tumor biopsies allows for the identification of the key mutations driving a patient’s cancer. However, this is limited to singular nodes and may not accurately reflect cancer heterogeneity. Circulating tumor cell (CTC) analyses offer a noninvasive method of interrogating the genomic profile of patient-derived tumor material. To date, molecular analysis of CTCs has relied on the characterization of bulk or pooled CTC lysates, limiting the detection of minor tumorigenic CTC subclones. Here, we show a workflow enabling BRAFV600E/NRASQ61R mutation detection from single cultured melanoma cells by combining micromanipulation and genomic material amplification methods. This workflow can be directly integrated into circulating tumor cell analysis applications.

Personalised medicine targeted to specific biomarkers such as BRAF and c-Kit has radically improved the success of melanoma therapy. More recently, further advances have been made using therapies targeting the immune response. In particular, therapies targeting the PD-1/PD-L1 or CTLA-4 axes alone or in combination have shown more sustained responses in 30-60% of patients. However, these therapies are associated with considerable toxicities and useful biomarkers to predict responders and non-responders are slow to emerge. Here we developed a reliable melanoma circulating tumor cell (CTC) detection method with PD-L1 evaluation on CTCs. A set of melanoma cell surface markers was tested as candidates for targeted melanoma CTC isolation and a melanoma specific immunostaining-based CTC identification protocol combined with PD-L1 detection was established. In vitro testing of the effect of exposure to blood cells on melanoma cell PD-L1 expression was undertaken. Immunomagnetic targeting isolated melanoma CTCs in up to 87.5% of stage IV melanoma patient blood samples and 3 8.6% of these had some PD-L1 expressing CTCs. Our in vitro data demonstrate PD-L1 induction on melanoma cells in the blood.This study established a robust, reliable method to isolate melanoma CTCs and detect expression of PD-L1 on these cells.

AIM:To evaluate disease presentation, treatment practices, and outcomes of patients with germ cell tumor (GCT) treated in a high-volume cancer center in Australia.METHODS:This is a retrospective analysis of 609 patients diagnosed with GCT in the Sydney West Cancer Network between 1990 and 2013. Cause and date of death, and second malignancy information was sourced from The Centre for Health Record Linkage.RESULTS:The median age was 33 years (range, 14-85). Primary site was testis in 590 (96.9%), mediastinum in nine (1.5%), and retroperitoneum in nine (1.5%). History of undescended testis was present in 48 (7.9%). Pure seminoma was seen in 334 (54.8%), with 274 (82.0%) being stage I. There was a decline in use of adjuvant radiotherapy from 83% in 1990-1997 to 29% in 2006-2013. Nonseminoma GCT (NSGCT) was diagnosed in 275 (45.2%), with 162 (58.9%) being stage 1. Active surveillance has increased as the initial treatment, from 58% between 1990 and 1997 to 89% between 2006 and 2013. Metastatic disease at presentation was seen in 162 (26.6%): 55 (34.0%) seminoma and 107 (66.0%) NSGCT. With median of 15-year follow-up, 18 (3.0%) have died from GCT and 70 (11.5%) from all causes. Ten-year overall survival was 93% and GCT-specific survival was 97%. Forty patients developed a secondary malignancy, with 38 receiving chemotherapy, radiotherapy, or both.CONCLUSIONS:This large Australian series illustrates a changing pattern of care and outcomes and compares them favorably with other series. This serves as a basis for future comparison of outcomes for this malignancy.