Prognostic Value of Low-Pass Whole Genome Sequencing of Circulating Tumor DNA in Metastatic Castration-Resistant Prostate Cancer.

BACKGROUND: Multiple treatments are available for metastatic castration-resistant prostate cancer (mCRPC), including androgen receptor signaling inhibitors (ARSI) enzalutamide and abiraterone, but therapy resistance remains a major clinical obstacle. We examined the clinical utility of low-pass whole-genome sequencing (LPWGS) of circulating tumor DNA (ctDNA) for prognostication in mCRPC. METHODS: A total of 200 plasma samples from 143 mCRPC patients collected at the start of first-line ARSI treatment (baseline) and at treatment termination (n = 57, matched) were analyzed by LPWGS (median: 0.50X) to access ctDNA% and copy number alteration (CNA) patterns. The best confirmed prostate specific antigen (PSA) response (≥50% decline [PSA50]), PSA progression-free survival (PFS), and overall survival (OS) were used as endpoints. For external validation, we used plasma LPWGS data from an independent cohort of 70 mCRPC patients receiving first-line ARSI. RESULTS: Baseline ctDNA% ranged from ≤3.0% to 73% (median: 6.6%) and CNA burden from 0% to 82% (median: 13.1%) in the discovery cohort. High ctDNA% and high CNA burden at baseline was associated with poor PSA50 response (P = 0.0123/0.0081), poor PFS (P < 0.0001), and poor OS (P < 0.0001). ctDNA% and CNA burden was higher at PSA progression than at baseline in 32.7% and 42.3% of the patients. High ctDNA% and high CNA burden at baseline was also associated with poor PFS and OS (P ≤ 0.0272) in the validation cohort. CONCLUSIONS: LPWGS of ctDNA provides clinically relevant information about the tumor genome in mCRPC patients. Using LPWGS data, we show that high ctDNA% and CNA burden at baseline is associated with short PFS and OS in 2 independent cohorts.

Low- versus standard- pneumoperitoneum in patients undergoing robot-assisted radical prostatectomy: a randomised, triple-blinded study.

OBJECTIVE: To investigate the effectiveness and impact of low-pressure pneumoperitoneum (Pnp) on postoperative quality of recovery (QoR) and surgical workspace (SWS) in patients with prostate cancer undergoing robot-assisted radical prostatectomy (RARP). PATIENTS AND METHODS: A randomised, triple-blinded trial was conducted in a single centre in Denmark from March 2021 to January 2022. A total of 98 patients with prostate cancer undergoing RARP were randomly assigned to either low-pressure Pnp (7 mmHg) or standard-pressure Pnp (12 mmHg). Co-primary outcomes were postoperative QoR measured via the QoR-15 questionnaire on postoperative Day 1 (POD1), POD3, POD14, and POD30, and SWS assessed intraoperatively by a blinded assessor (surgeon) via a validated SWS scale. Data analysis was performed according to the intention-to-treat principle. RESULTS: Patients who underwent RARP at low Pnp pressure demonstrated better postoperative QoR on POD1 (mean difference = 10, 95% confidence interval [CI] 4.4-15.5), but no significant differences were observed in the SWS (mean difference = 0.25, 95% CI -0.02 to 0.54). Patients allocated to low-pressure Pnp experienced statistically higher blood loss than those in the standard-pressure Pnp group (mean difference = 67 mL, P = 0.01). Domain analysis revealed significant improvements in pain (P = 0.001), physical comfort (P = 0.007), and emotional state (P = 0.006) for patients with low-pressure Pnp. This trial was registered at ClinicalTrials.gov, NCT04755452, on 16/02/2021. CONCLUSION: Performing RARP at low Pnp pressure is feasible without compromising the SWS and improves postoperative QoR, including pain, physical comfort, and emotional state, compared to the standard pressure.

Hemodynamic, renal and hormonal effects of lung protective ventilation during robot-assisted radical prostatectomy, analysis of secondary outcomes from a randomized controlled trial.

BACKGROUND: Lung protective ventilation with low tidal volume (TV) and increased positive end-expiratory pressure (PEEP) can have unfavorable effects on the cardiovascular system. We aimed to investigate whether lung protective ventilation has adverse impact on hemodynamic, renal and hormonal variables. METHODS: In this randomized, single-blinded, placebo-controlled study, 24 patients scheduled for robot-assisted radical prostatectomy were included. Patients were equally randomized to receive either ventilation with a TV of 6 ml/IBW and PEEP of 10 cm H2O (LTV-h.PEEP) or ventilation with a TV of 10 ml/IBW and PEEP of 4 cm H2O (HTV-l.PEEP). Before, during and after surgery, hemodynamic variables were measured, and blood and urine samples were collected. Blood samples were analyzed for plasma concentrations of electrolytes and vasoactive hormones. Urine samples were analyzed for excretions of electrolytes and markers of nephrotoxicity. RESULTS: Comparable variables were found among the two groups, except for significantly higher postoperative levels of plasma brain natriuretic peptide (p = 0.033), albumin excretion (p = 0.012) and excretion of epithelial sodium channel (p = 0.045) in the LTV-h.PEEP ventilation group compared to the HTV-l.PEEP ventilation group. In the combined cohort, we found a significant decrease in creatinine clearance (112.0 [83.4;126.7] ml/min at baseline vs. 45.1 [25.4;84.3] ml/min during surgery) and a significant increase in plasma concentrations of renin, angiotensin II, and aldosterone. CONCLUSION: Lung protective ventilation was associated with minor adverse hemodynamic and renal effects postoperatively. All patients showed a substantial but transient reduction in renal function accompanied by activation of the renin-angiotensin-aldosterone system. TRIAL REGISTRATION: ClinicalTrials, NCT02551341 . Registered 13 September 2015.

Urinary tract infections and risk of squamous cell carcinoma bladder cancer: A Danish nationwide case-control study.

Schistosoma haematobium infection can lead to squamous cell carcinomas (SCC) of the bladder. Whether this also applies to more common urinary tract infections (UTIs) is unclear. We therefore aimed to investigate the association between UTIs, reflected by the use of specific antibiotics and risk of SCC of the bladder. We conducted a Danish nationwide case-control study and identified histologically verified bladder cancer cases (2000-2015; n = 12,271) and age- and sex-matched cancer-free controls. We computed odds ratios (ORs) with 95% confidence intervals (CI) associating the use of UTI-specific antibiotics with SCC bladder cancer, using conditional logistic regression. We applied a 2-year lag-time to minimize reverse causation. To aid interpretation, similar analyses were performed for other bladder cancer types and other antibiotics. We identified 333 SCC cases (2.7% of all bladder cancers). Compared to no use (0-1 prescription), high-use (≥10 prescriptions) of UTI-specific antibiotics was associated with SCC with an OR of 11.4 (CI 7.6-17.2) and a clear dose-response pattern (ptrend < 0.001). Use of phenoxymethylpenicillin, an antibiotic not used against UTIs, was not associated with SCC after adjustment for use of UTI-specific antibiotics (OR 0.5). Furthermore, UTI-specific antibiotic use was not associated with urothelial carcinomas (n = 11,029; OR 1.13; CI 0.97-1.32). Excluding patients with known urogenital disease did not influence the SCC estimates (overall OR 10.8; CI 6.2-18.9). Data on smoking were lacking, however, a quantitative bias analysis suggested this to be of limited importance. In conclusion, common UTIs are strong, dose-dependent and specifically associated with risk of SCC of the bladder.

Implementing a Multimodal Prehabilitation Program to Radical Cystectomy in a Comprehensive Cancer Center: A Pilot Study to Assess Feasibility and Outcomes.

BACKGROUND: Radical cystectomy(RC) often leads to postoperative morbidity and complications. We conducted a pilot study on the effectiveness of multimodal prehabilitation, a preoperative conditioning method shown to be effective for colorectal surgery, in bladder cancer patients soon to undergo RC. We assessed patients' adherence to the prehabilitation regimen and changes in their physical condition. METHODS: Thirty-two bladder cancer patients at Memorial Sloan Kettering from February to August 2015 scheduled for RC were included in a standardized prehabilitation program. The 2-week program consisted of general physical exercises for the major muscle groups used for everyday activities, and sufficient protein intake. Patients received a program journal to document physical and nutritional achievements. Patients were physically tested using handgrip strength and bio-impedance at 2 weeks pre-surgery, day of surgery, and 6 weeks post-surgery. Additionally, a six-minute walk test (6MWT) 2 weeks before and 6 weeks after surgery were measured. RESULTS: Adherence to the exercises and nutritional recommendations respectively, was 62% (95% confidence interval [CI] 42-78%) for the exercise component and 81% (95% CI 62-93) for the nutritional component. The 6MWT results, showing physical capacity, significantly improved from baseline to 6-week follow-up, with an increase of 9.2% (95% CI 0.3-20.99; p=0.03). The handgrip strength, a proxy for nutritional status, improved 6.8% (95% CI 1.4-14.4; p=0.001) from baseline to admission, and maintained until 6-week follow-up (p=0.7). CONCLUSION: In a United States comprehensive cancer center, implementing a multimodal prehabilitation program is feasible in clinical practice and maintained. or even improved, physical functioning post-surgery compared to baseline.

Prospective comparison of 68Ga-PSMA PET/CT, 18F-sodium fluoride PET/CT and diffusion weighted-MRI at for the detection of bone metastases in biochemically recurrent prostate cancer.

PURPOSE: To prospectively compare diagnostic accuracies for detection of bone metastases by 68Ga-PSMA PET/CT, 18F-NaF PET/CT and diffusion-weighted MRI (DW600-MRI) in prostate cancer (PCa) patients with biochemical recurrence (BCR). METHODS: Sixty-eight PCa patients with BCR participated in this prospective study. The patients underwent 68Ga-PSMA PET/CT, a 18F-NaF PET/CT and a DW600-MRI (performed in accordance with European Society of Urogenital Radiology guidelines, with b values of 0 and 600 s/mm2). Bone lesions were categorized using a three-point scale (benign, malignant or equivocal for metastases) and a dichotomous scale (benign or metastatic) for each imaging modality by at least two experienced observers. A best valuable comparator was defined for each patient based on study-specific imaging, at least 12 months of clinical follow-up and any imaging prior to the study and during follow-up. Diagnostic performance was assessed using a sensitivity analysis where equivocal lesions were handled as non-metastatic and then as metastatic. RESULTS: Ten of the 68 patients were diagnosed with bone metastases. On a patient level, sensitivity, specificity and the area under the curve (AUC) by receiver operating characteristic analysis were, respectively, 0.80, 0.98-1.00 and 0.89-0.90 for 68Ga-PSMA PET/CT (n = 68 patients); 0.90, 0.90-0.98 and 0.90-0.94 for 18NaF PET/CT (n = 67 patients); and 0.25-0.38, 0.87-0.92 and 0.59-0.62 for DW600-MRI (n = 60 patients). The diagnostic performance of DW600-MRI was significantly lower than that of 68Ga-PSMA PET/CT and 18NaF PET/CT for diagnosing bone metastases (p < 0.01), and no significant difference in the AUC was seen between 68Ga-PSMA PET/CT and 18NaF PET/CT (p = 0.65). CONCLUSION: 68Ga-PSMA PET/CT and 18F-NaF PET/CT showed comparable and high diagnostic accuracies for detecting bone metastases in PCa patients with BCR. Both methods performed significantly better than DW600-MRI, which was inadequate for diagnosing bone metastases when conducted in accordance with European Society of Urogenital Radiology guidelines.

Prehabilitation for major abdominal urologic oncology surgery.

PURPOSE OF REVIEW: Whether prehabilitation in radical cystectomy adds to the effort of reducing postoperative morbidity and impairments in the survivorship phase has until recently received limited attention. This narrative review aims to summarize the current evidence base on prehabilitaion interventions focusing on the efficacy of procedure-specific interventions and the influence on postoperative outcomes. RECENT FINDINGS: Given the oncological risk, there is a relative short window to intervene and proactively optimize the patient before radical cystectomy. Preliminary results are however promising and a single-center randomized controlled trial (RCT) has shown that home-based short-term physical prehabilitation is feasible and effective and significantly improves early mobilization, time to perform activities of daily living and health-related quality of life (HRQoL). No significant impact on length of stay or complications was found. Limited evidence support preoperative nutritional interventions in cancer surgery, although evidence suggests improved outcome if malnourished individuals are adequately fed 7-10 days before surgery. No RCTs have evaluated the effect of smoking or alcohol cessation interventions on complications or HRQoL in radical cystectomy. Patient education interventions focusing on stoma care improve significantly self-efficacy in regards to independently change of stoma-appliance up to 1 year postoperatively. Currently, there is no evidence of early intervention considering psychological well being, sexual health or shared decision-making. SUMMARY: Published data indicate that a group of preoperative multiprofessional interventions including physical exercises, supportive nutritional care and stoma education can postoperatively improve early mobilization, self-efficacy and HRQoL. No evidence for further reduction of length of stay or complications was found.

FGFR3, TERT and OTX1 as a Urinary Biomarker Combination for Surveillance of Patients with Bladder Cancer in a Large Prospective Multicenter Study.

PURPOSE: Patients with nonmuscle invasive bladder cancer are followed with frequent cystoscopies. In this study FGFR3, TERT and OTX1 were investigated as a diagnostic urinary marker combination during followup of patients with primary nonmuscle invasive bladder cancer. MATERIALS AND METHODS: In this international, multicenter, prospective study 977 patients with nonmuscle invasive bladder cancer were included. A total of 2,496 urine samples were collected prior to cystoscopy during regular visits. Sensitivity was estimated to detect concomitant recurrences. Kaplan-Meier curves were used to estimate the development of future recurrences after urinalysis and a negative cystoscopy. RESULTS: Sensitivity of the assay combination for recurrence detection was 57% in patients with primary low grade, nonmuscle invasive bladder cancer. However, sensitivity was 83% for recurrences that were pT1 or muscle invasive bladder cancer. Of the cases 2% progressed to muscle invasive bladder cancer. Sensitivity for recurrence detection in patients with primary high grade disease was 72% and 7% of them had progression to muscle invasive bladder cancer. When no concomitant tumor was found by cystoscopy, positive urine samples were more frequently followed by a recurrence over time compared to a negative urine sample (58% vs 36%, p <0.001). High stage recurrences were identified within 1 year after a positive urine test and a negative cystoscopy. CONCLUSIONS: Recurrences in patients with primary nonmuscle invasive bladder cancer can be detected by a combination of urine assays. This study supports the value of urinalysis as an alternative diagnostic tool in patients presenting with low grade tumors and as a means to identify high stage tumors earlier.

DaPeCa-3: promising results of sentinel node biopsy combined with (18) F-fluorodeoxyglucose positron emission tomography/computed tomography in clinically lymph node-negative patients with penile cancer - a national study from Denmark.

OBJECTIVES: To estimate the diagnostic accuracy of sentinel node biopsy (SNB) combined with preoperative (18) F-fluorodeoxyglucose positron emission tomography/computed tomography (FDG PET/CT) for inguinal lymph node (LN) evaluation in patients with invasive penile squamous cell carcinoma (PSCC) with no clinical evidence of inguinal metastases (cN0) at two tertiary centres with complete clinical follow-up. PATIENTS AND METHODS: From April 2010 in Centre one and from January 2013 in Centre two, we prospectively enrolled patients diagnosed with invasive PSCC and scheduled for SNB at the only two university centres treating penile cancer in Denmark. All patients had FDG PET/CT before SNB. The sentinel LNs were preoperatively located by planar lymphoscintigraphy in 134 groins (68 patients) and by single-photon emission CT/CT in 120 groins (61 patients). The primary endpoints were the sensitivity, specificity, and false-negative rate of SNB combined with FDG PET/CT. The secondary endpoint was SNB-related morbidity. RESULTS: We examined 254 groins in 129 patients by SNB combined with FDG PET/CT. The median (interquartile range, IQR) follow-up of survivors was 23 (14-35) months. Of 201 LN-negative groins, two were false negatives, and despite radio-chemotherapy treatment, both patients died from penile cancer. Four of 23 radiotracer-silent groins, had a FDG PET/CT-positive LNs and were surgically explored. In one of four of the explored groins, a positive LN was found. Combined FDG PET/CT-SNB sensitivity was 94.4% (95% confidence interval [CI] 81-99%) per groin. The false-negative rate was 5.6% (95% CI 1-19%) per groin. In 15 patients (11.6%) there were 25 SNB-related complications of Clavien-Dindo grades I-IIIa. The only Clavien-Dindo IIIa complication was an inguinal lymphocele treated by aspiration. CONCLUSION: In this study, we present a favourable SNB false-negative rate of 5.6% in a national cohort of clinically LN-negative patients with invasive PSCC with a pre-SNB FDG PET/CT scan. The combination of FDG PET/CT and SNB seems to be a promising diagnostic approach. Even so, a false-negative SNB was fatal in two of two cases and we are determined to continue the development of our SNB technique. The SNB-related morbidity was limited.

DaPeCa-1: diagnostic accuracy of sentinel lymph node biopsy in 222 patients with penile cancer at four tertiary referral centres - a national study from Denmark.

OBJECTIVES: To estimate the diagnostic accuracy of sentinel lymph node biopsy (SNB) in patients with penile cancer and assess SNB complications in a national multicentre setting. PATIENTS AND METHODS: Retrospectively data were collected from records in four university centres by one medical doctor covering all SNBs performed in Denmark between 1 January 2000 and 31 December 2010. Patients had either impalpable lymph nodes (LNs) in one or both groins, or had a palpable inguinal mass from which aspiration cytology failed to reveal malignancy. Patients were injected with nanocolloid technetium and had a scintigram recorded before the SNB. The primary endpoint was LN recurrence on follow-up. The secondary endpoint was complications after SNB. Diagnostic accuracy was computed. RESULTS: In all, 409 groins in 222 patients were examined by SNB. The median (interquartile range) follow-up of patients who survived was 6.6 (5-10) years. Of 343 negative groins, eight were false negatives. The sensitivity was 89.2% (95% confidence interval 79.8-95.2%) per groin. Interestingly, four of 67 T1G1 patients had a positive SNB. In all, 28 of 222 (13%) patients had complications of Clavien-Dindo grade I-IIIa. CONCLUSION: Penile cancer SNB with a close follow-up stages LN involvement reliably and has few complications in a national multicentre setting. Inguinal LN dissection was avoided in 76% of patients.

Impact of low pneumoperitoneum on renal function and acute kidney injury biomarkers during robot-assisted radical prostatectomy (RARP): a randomised clinical trial.

The objective of this study was to evaluate the effect of low pneumoperitoneum pressure (Pnp) on renal function and renal injury biomarkers during robot-assisted radical prostatectomy (RARP). A single-centre, triple-blinded, randomised clinical trial was conducted with 98 patients undergoing RARP, who were assigned to either standard Pnp of 12 mmHg or low Pnp of 7 mmHg. The primary outcome was urinary neutrophil gelatinase-associated lipocalin (u-NGAL), and several other kidney injury biomarkers were assessed as secondary outcomes. Acute kidney injury (AKI) was evaluated using the Kidney Disease Improving Global Outcomes (KDIGO) criteria, the gold standard method for defining AKI. The trial was registered on ClinicalTrials.gov (NCT04755452). Patients in the low Pnp group had significantly lower levels of u-NGAL (mean difference - 39.9, 95% CI - 73.7 to - 6.1, p = 0.02) compared to the standard Pnp group. No significant differences were observed for other urinary biomarkers. Interestingly, there was a significant difference in intraoperative urine production between the groups (low Pnp median: 200 mL, IQR: 100-325 vs. standard Pnp median: 100 mL, IQR: 50-200, p = 0.01). Similarly, total postoperative urine production also varied significantly (low Pnp median: 1325 mL, IQR: 1025-1800 vs. standard Pnp median: 1000 mL, IQR: 850-1287, p = 0.001). The occurrence of AKI, as defined by the KDIGO criteria, did not differ significantly between the groups. Low Pnp during RARP resulted in lower u-NGAL levels, suggesting a potential benefit in terms of reduced renal injury. However, the lack of a notable difference in AKI as defined by the KDIGO criteria indicates that the clinical significance of this finding may be limited. Further research is needed to validate and expand on these results, ultimately defining the optimal Pnp strategy for RARP and improving patient outcomes.

Validation of a surgical workspace scale during robot-assisted surgery.

BACKGROUND: A sufficient surgical workspace is crucial to avoid complications. Within classic laparoscopy, many subjective surgical rating scales (SRSs) have previously been used to evaluate the surgical workspace. This study aimed to validate a modified version of the 5-point SRS during robot-assisted radical nephrectomy (RARN). METHODS: Thirty-two intra-operative videos of intraperitoneal spaces were recorded from eight patients who underwent RARN. To attain the visualisation of different types of workspaces, we recorded 20 s panoramic videos of different pneumoperitoneum, namely 3, 5, 7 and 12 mmHg. The videos were randomised and presented two times to eight experienced robotic surgeons to evaluate the workspace using our modified 5-point SRS. Both inter-and intra-rater reliabilities were tested. RESULTS: The results of the validation study showed moderate inter-rater and good to excellent intra-rater reliability. CONCLUSION: This is a valid tool that can be confidently used by future researchers in the field of robot-assisted surgery.

DaBlaCa-13 Study: Oncological Outcome of Short-Term, Intensive Chemoresection With Mitomycin in Nonmuscle Invasive Bladder Cancer: Primary Outcome of a Randomized Controlled Trial.

PURPOSE: This study aimed to assess long-term follow-up after chemoresection with mitomycin (MMC), a nonsurgical treatment modality for recurrent nonmuscle invasive bladder cancer (NMIBC). At the time of recurrence, chemoresection has previously been shown to reduce the number of patients requiring a procedure (transurethral resection of bladder tumors [TURBT] or office biopsy) by more than 50%. This study investigated the number of patients requiring a procedure during initial treatment and 2-year follow-up in patients treated with short-term, intensive chemoresection with MMC compared with patients undergoing standard surgical treatment of recurrent NMIBC. METHODS: A randomized, controlled trial was conducted in two urological departments in Denmark from January 2018 to August 2021. In total, 120 patients with a history of Ta low- or high-grade NMIBC were included upon recurrence. The intervention group received intravesical MMC (40 mg/40 mL) three times a week for 2 weeks and TURBT or office biopsy only if the response was incomplete. The control group received TURBT or office biopsy and 6 weekly adjuvant instillations. The primary outcome was the number of patients undergoing a procedure within 2 years from inclusion, which was compared between groups using the chi-squared test. Recurrence-free survival was analyzed using the Kaplan-Meier method. RESULTS: Significantly fewer patients were in need of a procedure in the intervention group than in the control group: 71% (95% CI, 57 to 81) and 100% (95% CI, 94 to 100), P < .001. The 12-month recurrence-free survival was 36% (95% CI, 24 to 50) and 43% (95% CI, 30 to 56) in the intervention and control groups, respectively (P = .5). CONCLUSION: Short-term intensive chemoresection is an effective treatment strategy for recurrent NMIBC that leads to a reduced number of required procedures without compromising long-term oncological safety.

Cell-Free Urine and Plasma DNA Mutational Analysis Predicts Neoadjuvant Chemotherapy Response and Outcome in Patients with Muscle-Invasive Bladder Cancer.

PURPOSE: To investigate the use of plasma and urine DNA mutation analysis for predicting neoadjuvant chemotherapy (NAC) response and oncological outcome in patients with muscle-invasive bladder cancer. EXPERIMENTAL DESIGN: Whole-exome sequencing of tumor and germline DNA was performed for 92 patients treated with NAC followed by radical cystectomy (RC). A custom NGS-panel capturing approximately 50 mutations per patient was designed and used to track mutated tumor DNA in plasma and urine. A total of 447 plasma samples, 281 urine supernatants, and 123 urine pellets collected before, during, and after treatment were analyzed. Patients were enrolled from 2013 to 2019, with a median follow-up time of 41.3 months after RC. RESULTS: We identified tumor DNA before NAC in 89% of urine supernatants, 85% of urine pellets, and 43% of plasma samples. Tumor DNA levels were higher in urine supernatants and urine pellets compared with plasma samples (P < 0.001). In plasma, detection of circulating tumor DNA (ctDNA) before NAC was associated with a lower NAC response rate (P < 0.001). Detection of tumor DNA after NAC was associated with lower response rates in plasma, urine supernatant, and urine pellet (P < 0.001, P = 0.03, P = 0.002). Tumor DNA dynamics during NAC was predictive of NAC response and outcome in urine supernatant and plasma (P = 0.006 and P = 0.002). A combined measure from plasma and urine supernatant tumor DNA dynamics stratified patients by outcome (P = 0.003). CONCLUSIONS: Analysis of tumor DNA in plasma and urine samples both separately and combined has a potential to predict treatment response and outcome.

Impact of germline DNA repair gene variants on prognosis and treatment of men with advanced prostate cancer.

The clinical importance of germline variants in DNA repair genes (DRGs) is becoming increasingly recognized, but their impact on advanced prostate cancer prognosis remains unclear. A cohort of 221 newly diagnosed metastatic castration-resistant prostate cancer (mCRPC) patients were screened for pathogenic germline variants in 114 DRGs. The primary endpoint was progression-free survival (PFS) on first-line androgen signaling inhibitor (ARSI) treatment for mCRPC. Secondary endpoints were time to mCRPC progression on initial androgen deprivation therapy (ADT) and overall survival (OS). Twenty-seven patients (12.2%) carried a germline DRG variant. DRG carrier status was independently associated with shorter PFS on first-line ARSI [HR 1.72 (1.06-2.81), P = 0.029]. At initiation of ADT, DRG carrier status was independently associated with shorter progression time to mCRPC [HR 1.56, (1.02-2.39), P = 0.04] and shorter OS [HR 1.99, (1.12-3.52), P = 0.02]. Investigating the contributions of individual germline DRG variants on PFS and OS revealed CHEK2 variants to have little effect. Furthermore, prior taxane treatment was associated with worse PFS on first-line ARSI for DRG carriers excluding CHEK2 (P = 0.0001), but not for noncarriers. In conclusion, germline DRG carrier status holds independent prognostic value for predicting advanced prostate cancer patient outcomes and may potentially inform on optimal treatment sequencing already at the hormone-sensitive stage.

Circulating Tumor DNA Analysis in Advanced Urothelial Carcinoma: Insights from Biological Analysis and Extended Clinical Follow-up.

PURPOSE: To investigate whether circulating tumor DNA (ctDNA) assessment in patients with muscle-invasive bladder cancer predicts treatment response and provides early detection of metastatic disease. EXPERIMENTAL DESIGN: We present full follow-up results (median follow-up: 68 months) from a previously described cohort of 68 neoadjuvant chemotherapy (NAC)-treated patients who underwent longitudinal ctDNA testing (712 plasma samples). In addition, we performed ctDNA evaluation of 153 plasma samples collected before and after radical cystectomy (RC) in a separate cohort of 102 NAC-naïve patients (median follow-up: 72 months). Total RNA sequencing of tumors was performed to investigate biological characteristics of ctDNA shedding tumors. RESULTS: Assessment of ctDNA after RC identified metastatic relapse with a sensitivity of 94% and specificity of 98% using the expanded follow-up data for the NAC-treated patients. ctDNA dynamics during NAC was independently associated with patient outcomes when adjusted for pathologic downstaging (HR = 4.7; P = 0.029). For the NAC-naïve patients, ctDNA was a prognostic predictor before (HR = 3.4; P = 0.0005) and after RC (HR = 17.8; P = 0.0002). No statistically significant difference in recurrence-free survival for patients without detectable ctDNA at diagnosis was observed between the cohorts. Baseline ctDNA positivity was associated with the Basal/Squamous (Ba/Sq) subtype and enrichment of epithelial-to-mesenchymal transition and cell cycle-associated gene sets. CONCLUSIONS: ctDNA is prognostic in NAC-treated and NAC-naïve patients with more than 5 years follow-up and outperforms pathologic downstaging in predicting treatment efficacy. Patients without detectable ctDNA at diagnosis may benefit significantly less from NAC, but additional studies are needed.

Evaluation of different lymph node (LN) variables as prognostic markers in patients undergoing radical cystectomy and extended LN dissection to the level of the inferior mesenteric artery.

OBJECTIVE: To evaluate the prognostic impact of lymph node (LN) variables in patients undergoing radical cystectomy (RC) and extended LN dissection. PATIENTS AND METHODS: From January 2004 to January 2009, 167 patients with bladder cancer underwent RC and extended LN dissection to the level of the inferior mesenteric artery in a surgery-only series with no neoadjuvant or adjuvant chemotherapy. Correlation to prognosis of different LN variables according to presence of LN metastasis, number, localization, extracapsular extension (ECE), size, volume, LN density and N-stage according to two different Tumour-Node-Metastasis (TNM) classifications were analysed. RESULTS: In all, 43 patients (26%) had LN metastases. In univariate analysis, gender, T-stage and several different LN variables stratified by presence of LN metastasis, number of positive LNs, anatomical localisation, ECE, LN density, size and volume of positive LNs, were significant prognostic predictors. Female gender, advanced T-stage, presence of LN metastasis, non-regional LN metastases (M-positive) and number of positive LNs (1 vs >1) were significant adverse prognostic predictors in multivariate analysis, whereas the other LN variables were not. Inclusion of the common iliac LNs in the regional LNs as suggested in the seventh edition of the TNM classification was relevant regarding prognosis. However, subclassification based on location was not correlated to prognosis. The new N3 category therefore seems superfluous. CONCLUSIONS: LN-positive patients have a poor prognosis, especially if >1 positive LN is present. Despite several different suggestions of new LN-dependent prognostic factors, none of the tested variables were independently significant in the present series.

Expression of TIP60 (tat-interactive protein) and MRE11 (meiotic recombination 11 homolog) predict treatment-specific outcome of localised invasive bladder cancer.

UNLABELLED: What's known on the subject? and What does the study add? Several studies have shown that defects in DNA-damage response are associated with good survival after chemotherapy and radiotherapy. Furthermore, loss of cell cycle regulators may be prognostic indicators of poor survival after cystectomy. However, the potential clinical impact of previous findings is hampered by insufficient validation of significant results in suitable cystectomy and radiotherapy cohorts. Here we use a large cohort of patients receiving radiotherapy to successfully validate the importance of MRE11 as a predictive marker of disease-specific survival (DSS). Furthermore, using two independent patient cohorts we show for the first time that TIP60 is a predictive marker of DSS after cystectomy. We show that combined use of TIP60 and MRE11 may hold the potential to guide treatment decisions. OBJECTIVE: • To determine the association between the proteins: tat-interactive protein 60 kDa (TIP60), p16, meiotic recombination 11 homolog (MRE11), phosphorylated ataxia telangiectasia mutated (ATM), retinoblastoma protein (Rb), Ki67, and p53 and clinical outcome in invasive lymph node-negative bladder cancer. PATIENTS AND METHODS: • Protein expression was measured by immunohistochemistry in cancer specimens from two independent cohorts of patients with bladder cancer treated with cystectomy (162 patients and 273) and one cohort of patients receiving radiotherapy (148). • Disease-specific survival (DSS) was used as the outcome measure, and patients with no disease-specific death were followed for a minimum of 36 months. RESULTS: • TIP60 was significantly correlated with DSS in both cystectomy cohorts (hazard ratio [HR] 0.42, 95% confidence interval [CI] 0.26-0.68, P < 0.001 and HR 0.45, 95% CI 0.28-0.72, P = 0.001). • MRE11 was significantly correlated with DSS in the cohort receiving radiotherapy (HR 0.64, 95% CI 0.47-0.86, P = 0.005). • P16 was significantly correlated with DSS in all three cohorts (HR 0.46, 95% CI 0.30-0.75, P = 0.032; HR 0.60, 95% CI 0.37-0.97, P = 0.032; HR 0.52, 95% CI 0.28-0.96, P = 0.001). • Rb was significantly correlated with DSS in one cystectomy cohort (HR 1.71, 95% CI 1.13-2.75, P = 0.017). • Ki67, p53, and pATM were not significantly correlated with DSS in any of the cohorts. CONCLUSIONS: • TIP60 protein expression was a predictive marker for DSS after cystectomy in two independent cohorts. This novel marker was the strongest predictive factor in multivariate analysis in patients receiving cystectomy. • MRE11 was shown to be a predictive marker for DSS after radiotherapy. • We have shown that TIP60 and MRE11 hold the potential to guide patients with invasive bladder cancer to either cystectomy or radiotherapy. This study was based on retrospective material and consequently we suggest that these markers should be validated in a prospective study.

Differential RNA aptamer affinity profiling on plasma as a potential diagnostic tool for bladder cancer.

The molecular composition of blood is a signature of human health, reflected in the thousands of blood biomarkers known for human diseases. However, establishing robust disease markers is challenging due to the diversity of individual samples. New sequencing methods have simplified biomarker discovery for circulating DNA and RNA while protein profiling is still laborious and costly. To harness the power of high-throughput sequencing to profile the protein content of a biological sample, we developed a method termed APTASHAPE that uses oligonucleotide aptamers to recognize proteins in complex biofluids. We selected a large pool of 2'Fluoro protected RNA sequences to recognize proteins in human plasma and identified a set of 33 cancer-specific aptamers. Differential enrichment of these aptamers after selection against 1 µl of plasma from individual patients allowed us to differentiate between healthy controls and bladder cancer-diagnosed patients (91% accuracy) and between early non-invasive tumors and late stage tumors (83% accuracy). Affinity purification and mass spectrometry of proteins bound to the predictive aptamers showed the main target proteins to be C4b-binding protein, Complement C3, Fibrinogen, Complement factor H and IgG. The APTASHAPE method thus provides a general, automated and highly sensitive platform for discovering potential new disease biomarkers.

PAI-1 is a potential transcriptional silencer that supports bladder cancer cell activity.

The extracellular activity of Plasminogen activator inhibitor-1 (PAI-1) is well described, acting as an inhibitor of tissue plasminogen activator and urokinase-type plasminogen activator, impacting fibrinolysis. Recent studies have revealed a pro-tumorigenic role of PAI-1 in human cancers, via the regulation of angiogenesis and tumor cell survival. In this study, immunohistochemical staining of 939 human bladder cancer specimens showed that PAI-1 expression levels correlated with tumor grade, tumor stage and overall survival. The typical subcellular localization of PAI-1 is cytoplasmic, but in approximately a quarter of the cases, PAI-1 was observed to be localized to both the tumor cell cytoplasm and the nucleus. To investigate the potential function of nuclear PAI-1 in tumor biology we applied chromatin immunoprecipitation (ChIP)-sequencing, gene expression profiling, and rapid immunoprecipitation mass spectrometry to a pair of bladder cancer cell lines. ChIP-sequencing revealed that PAI-1 can bind DNA at distal intergenic regions, suggesting a role as a transcriptional coregulator. The downregulation of PAI-1 in bladder cancer cell lines caused the upregulation of numerous genes, and the integration of ChIP-sequence and RNA-sequence data identified 57 candidate genes subject to PAI-1 regulation. Taken together, the data suggest that nuclear PAI-1 can influence gene expression programs and support malignancy.