The Risk of Thromboembolism in Patients with Muscle Invasive Bladder Cancer before and after Cystectomy Depending on Blood Group and Neoadjuvant Chemotherapy-A Multicentre Retrospective Cohort Study.

PURPOSE: Previous studies have indicated that patients with muscle-invasive bladder cancer with non-O blood types have an increased risk of experiencing thromboembolic events (TEEs). This is finding is in relation to neoadjuvant-chemotherapy (NAC)-naïve patients. AIM: to establish the risk of TEEs and any association with blood types among NAC patients as well as NAC-naïve patients. METHODS: Cystectomized patients at four centres treated from 2009 to 2018 (n = 244) were analysed. The quantities of patients corresponding to each blood group were as follows: A-108 (44%); O-99 (41%); B-30 (12%); and AB-7 (3%). NAC patients (n = 167) and NAC-naïve NAC-eligible patients (n = 77) were assessed. In total, 54 women (22%) and 190 men (78%), with a median age of 69 years, were included in the study. The occurrence of any type of TEE from six months pre-cystectomy to 12-24 months after was analysed using logistic regression adjusted for NAC and confounders. RESULTS: Sixty-six TEEs were detected in 21% of the patients (n = 52). Pulmonary embolus (n = 33) and deep venous thrombosis (n = 11) were the most common forms. No significant differences between blood types were found in the analysis, although B blood type had a nearly significant increased crude risk compared with O blood type, for which there was an OR of 2.48 (95% CI 0.98-6.36). Adjustment for NAC and covariates weakened the OR, which plummeted to 1.98 (95% CI 0.71-5.51). CONCLUSIONS: No significant associations were found between blood types and TEE occurrences in this cohort including both NAC and NAC-naïve NAC-eligible patients.

Infiltrating immune cells in prostate cancer tissue after androgen deprivation and radiotherapy.

OBJECTIVES: Androgen deprivation therapy (ADT) has long been a cornerstone in treatment of advanced prostate cancer (PCa), and is known to improve the results of radiotherapy (RT) for high-risk disease. The purpose of our study was to use a multiplexed immunohistochemical (mIHC) approach to investigate the infiltration of immune cells in PCa tissue after eight weeks of ADT and/or RT with 10 Gy. METHODS: From a cohort of 48 patients divided into two treatment arms, we obtained biopsies before and after treatment and used a mIHC method with multispectral imaging to analyze the infiltration of immune cells in tumor stroma and tumor epithelium, focusing on areas with high infiltration. RESULTS: Tumor stroma showed a significantly higher infiltration of immune cells compared to tumor epithelium. The most prominent immune cells were CD20+ B-lymphocytes, followed by CD68+ macrophages, CD8+ cytotoxic T-cells, FOXP3+ regulatory T-cells (Tregs), and T-bet+ Th1-cells. Neoadjuvant ADT followed by RT significantly increased the infiltration of all five immune cells. Numbers of Th1-cells and Tregs significantly increased after single treatment with ADT or RT. In addition, ADT alone increased the number of cytotoxic T-cells and RT increased the number of B-cells. CONCLUSIONS: Neoadjuvant ADT in combination with RT results in a higher inflammatory response compared to RT or ADT alone. The mIHC method may be a useful tool for investigating infiltrating immune cells in PCa biopsies to understand how immunotherapeutic approaches can be combined with current PCa therapies.

Immune-Activated B Cells Are Dominant in Prostate Cancer.

B cells are multifaceted immune cells responding robustly during immune surveillance against tumor antigens by presentation to T cells and switched immunoglobulin production. However, B cells are unstudied in prostate cancer (PCa). We used flow cytometry to analyze B-cell subpopulations in peripheral blood and lymph nodes from intermediate-high risk PCa patients. B-cell subpopulations were related to clinicopathological factors. B-cell-receptor single-cell sequencing and VDJ analysis identified clonal B-cell expansion in blood and lymph nodes. Pathological staging was pT2 in 16%, pT3a in 48%, and pT3b in 36%. Lymph node metastases occurred in 5/25 patients (20%). Compared to healthy donors, the peripheral blood CD19+ B-cell compartment was significantly decreased in PCa patients and dominated by naïve B cells. The nodal B-cell compartment had significantly increased fractions of CD19+ B cells and switched memory B cells. Plasmablasts were observed in tumor-draining sentinel lymph nodes (SNs). VDJ analysis revealed clonal expansion in lymph nodes. Thus, activated B cells are increased in SNs from PCa patients. The increased fraction of switched memory cells and plasmablasts together with the presence of clonally expanded B cells indicate tumor-specific T-cell-dependent responses from B cells, supporting an important role for B cells in the protection against tumors.

A Retrospective Analysis of the De Ritis Ratio in Muscle Invasive Bladder Cancer, with Focus on Tumor Response and Long-Term Survival in Patients Receiving Neoadjuvant Chemotherapy and in Chemo Naïve Cystectomy Patients-A Study of a Clinical Multicentre Database.

Background: A high pre-treatment De Ritis ratio, the aspartate transaminase/alanine aminotransferase ratio, has been suggested to be of prognostic value for mortality in muscle-invasive bladder cancer (MIBC). Our purpose was to evaluate if a high ratio was associated with mortality and downstaging. Methods: A total of 347 Swedish patients with clinically staged T2-T4aN0M0, with administered neoadjuvant chemotherapy (NAC) or eligible for NAC and undergoing radical cystectomy (RC) 2009−2021, were retrospectively evaluated with a low ratio < 1.3 vs. high ratio > 1.3, by Log Rank test, Cox regression and Mann−Whitney U-test (MWU), SPSS 27. Results: Patients with a high ratio had a decrease of up to 3 years in disease-free survival (DFS), cancer-specific survival (CSS) and overall survival (OS) (p = 0.009, p = 0.004 and p = 0.009) and 5 years in CSS and OS (p = 0.019 and p = 0.046). A high ratio was associated with increased risk of mortality, highest in DFS (HR, 1.909; 95% CI, 1.265−2.880; p = 0.002). No significant relationship between downstaging and a high ratio existed (p = 0.564 MWU). Conclusion: A high pre-treatment De Ritis ratio is on a population level, associated with increased mortality post-RC in endpoints DFS, CSS and OS. Associations decrease over time and require further investigations to determine how strong the associations are as meaningful prognostic markers for long-term mortality in MIBC. The ratio is not suitable for downstaging-prediction.

Adverse events during neoadjuvant chemotherapy for muscle invasive bladder cancer-a Swedish retrospective multicentre study of a clinical database.

Background: Adverse events (AEs) during neoadjuvant chemotherapy (NAC) for muscle invasive bladder cancer (MIBC) are known but insufficiently reported. Clinical implications include affected cardiac, pulmonary, urinary, vascular and haematological organ systems. The main purpose was to evaluate the incidence and severity of AEs for ascertaining possible clinical significance. Further investigating possible effects of AEs on downstaging outcomes-downstaging is considered a surrogate marker for overall survival (OS). Methods: A retrospective evaluation of AEs during ongoing NAC for MIBC patients analysing individual patient data in a clinical database. We identified 687 cystectomies between 2009-2020 at four Swedish urological centres. Inclusion criteria were cT2-4aN0M0 in 261 NAC patients undergoing radical cystectomy (RC). Medical files were reviewed and AEs were assessed and graded, including detailed measurements by the Common Terminology Criteria for Adverse Events (CTCAE) v.5. Data were retrospectively analysed in SPSS statistics 27.0 with Spearman rank-order correlation coefficient and Mann-Whitney U-test (MWU). Results: A total of 251/261 patients [95% confidence interval (CI), 93-98%] experienced AEs during NAC pre-RC (mean two AEs/patient). In total, 208 (80%) patients received methotrexate, vinblastine, adriamycin (doxorubicin) and cisplatin (MVAC). In the total cohort, 200 (76.6%) received all pre-planned NAC-cycles. Most common AEs were anaemia (88.9%), thrombocytopenia (44.8%) and acute kidney injury (40.6%). Patients with prematurely terminated cycles had higher AE-grades (P=0.042 MWU). A correlation between higher AE-grades and decrease in downstaging existed, in the entire cohort (-0.133; P=0.033) and in patients undergoing all pre-planned NAC-cycles (-0.148; P=0.038). Anaemia and acute kidney injury were individually associated with decreased downstaging (-0.360, P=0.025 and -0.183, P=0.010, respectively). Conclusions: NAC in MIBC poses a significant risk for AEs before RC with clinical implications. For instance, patients terminating chemotherapy prematurely, have higher AE-grades and decreased downstaging. Further, acute kidney injury and anaemia are individually associated with decreased downstaging. We propose that early detection and prevention of AEs may increase downstaging of the primary tumour.

Central Venous Access and the Risk for Thromboembolic Events in Patients Undergoing Neoadjuvant Chemotherapy and Radical Cystectomy for Muscle-Invasive Bladder Cancer.

Thromboembolic events (TEE) are high-risk complications in patients undergoing neoadjuvant chemotherapy (NAC) and radical cystectomy (RC) for urothelial muscle-invasive bladder cancer (MIBC). The purpose of the study was to investigate any differences in TEE-incidence, comparing peripherally inserted central catheter (PICC) versus a totally implanted port (PORT) as CVA (central venous access) during NAC. We identified 947 cystectomized MIBC-patients from four Swedish medical centers in 2009-2021. Inclusion criteria were cT2-T4aN0M0 and 375 patients were finally eligible and evaluated, divided into: NAC-administered (n = 283) resp. NAC-naïve-NAC-eligible (n = 92), the latter as tentative control group. Data on TEEs and types of CVA were retrospectively collected and individually validated, from final transurethral resection of the bladder tumor (TUR-B) to 30 days post-RC. Adjusted logistic regression and log rank test were used for statistical analyses. Amongst NAC-administered, 83% (n = 235) received PICCs and 15% (n = 42) PORTs. Preoperative TEEs occurred in 38 PICC-patients (16.2%) and in one PORT-patient (2.4%), with 47 individual events registered. We found a significantly increased odds ratio of TEE in NAC-administered PICC-patients compared to in PORT-patients (OR: 8.140, p-value: 0.042, 95% CI 1.078-61.455). Our findings indicate a greater risk for pre-RC TEEs with PICCs than with PORTs, suggesting favoring the usage of PORTs for MIBC-NAC-patients.

Proteomic Profiling of Tissue Exosomes Indicates Continuous Release of Malignant Exosomes in Urinary Bladder Cancer Patients, Even with Pathologically Undetectable Tumour.

Invasive urothelial bladder cancer (UBC) has high recurrence rates even after radical cystectomy (RC). Exosomes are membrane-bound nanovesicles, which have been shown to contribute to carcinogenesis and metastasis. We previously showed that urinary exosomes display a malignant profile in UBC patients despite the absence of detectable tumour. Here, we investigated exosomes from sampling sites close to or distant from the former tumour, aiming to understand the effect of the tumour on the local milieu. Ten patients scheduled for cystectomy after transurethral bladder resection (TUR-B), without remaining detectable tumour, were included. Exosomes were isolated from tissue explants of both the previous tumour site and distant bladder tissue. Proteins were quantified by mass spectrometry in seven patients. Exosomes from the previous tumour site were enriched in inflammatory but not cancer-related pathways compared to distant tissue. However, the 69 most abundant proteins in tissue-derived exosomes regardless of site, 20 of which were also found in urinary exosomes from our previous study, were enriched for cancer-related metabolic pathways and associated with poor prognosis in an external mRNA dataset. The enrichment of cancer-related pathways in the most abundant proteins, regardless of sampling site, confirms our hypothesis that despite the absence of detectable tumour, the entire bladder releases exosomes that contribute to metastasis and highlights the need for early RC.

IL-16 processing in sentinel node regulatory T cells is a factor in bladder cancer immunity.

In the effort of developing new immunotherapies, the sentinel node (SN) has proven a promising source from which to harness an effective antitumour T cell response. However, tumour immune escape, a process in which regulatory T cells (Tregs) play a central role, remains a major limiting factor. Therefore, there is a clear need to increase the knowledge of Treg function and signalling in sentinel nodes. Here, we set out to explore whether the proteome in SN-resident T cells is altered by the tumour and to identify key proteins in SN T cell signalling, focusing on Tregs. Five patients with muscle-invasive urothelial bladder cancer were prospectively included. Mass spectrometry was performed on two patients, with validation and functional studies being performed on three additional patients and four healthy donors. At cystectomy, SN, non-SN lymph nodes and peripheral blood samples were collected from the patients and T cell subsets isolated through flow cytometry before downstream experiments. Proteomic analysis indicated that growth and immune signalling pathways are upregulated in SN-resident Tregs. Furthermore, centrality analysis identified the cytokine IL-16 to be central in the SN-Treg signalling network. We show that tumour-released factors, through activating caspase-3, increase Treg IL-16 processing into bioactive forms, reinforcing Treg suppressive capacity. In conclusion, we provide evidence that Tregs exposed to secreted factors from bladder tumours show increased immune and growth signalling and altered IL-16 processing which translates to enhanced Treg suppressive function, indicating altered IL-16 signalling as a novel tumour immune escape mechanism.

Urinary Exosomes from Bladder Cancer Patients Show a Residual Cancer Phenotype despite Complete Pathological Downstaging.

Invasive urinary bladder cancer shows high recurrence rates after cystectomy even with apparent complete downstaging at cystectomy. Exosomes are nano-sized vesicles important in cell-cell communication, which have been hypothesized to contribute to cancer dissemination and recurrence. The aim of this study was to investigate if pro-carcinogenic exosomes could be detected in urine from histologically downstaged bladder cancer patients. 13 Patients were included in this study. Paired ureter and urine samples from nine patients underwent mass spectrometry, while samples from the remaining patients were used for exosome characterization. At cystectomy, exosomes were isolated from bladder and ureter urine, whereafter quantitative proteome profiling was performed. Urinary exosomes clustered based on whether they came from the bladder, with tumour contact, or the ureters, without tumour contact, even though all came from completely downstaged patients. Proteins overexpressed in exosomes derived from bladder urine contained several oncogenes and were mainly associated with tumour metabolism pathways. Although patients were histologically tumour-free at cystectomy, the bladder urine contained exosomes with a carcinogenic metabolic profile. This suggests a continuous release of exosomes from the bladder, which may promote recurrence at distant sites through metabolic rewiring, even after apparent complete downstaging. These exosomes, coming from either undetected cancer cells or partly transformed cells, are likely to increase the risk of metastasis and encourages cystectomy even in completely downstaged patients.

Blood transfusions during neoadjuvant chemotherapy for muscle-invasive urinary bladder cancer may have a negative impact on overall survival.

Objective: To evaluate the extent and plausible effects of blood transfusions given during cisplatin-based neoadjuvant chemotherapy (NAC) on overall survival in patients with muscle-invasive urothelial bladder cancer (MIBC) undergoing NAC and radical cystectomy (RC).Background: Several studies have demonstrated a decreased survival for MIBC patients receiving allogenic peri- and postoperative blood transfusions in conjunction with RC. No studies have previously investigated the effects of blood transfusions during NAC.Materials and methods: 120 patients with MIBC (cT2-T4aN0M0) undergoing NAC and RC between 2008 and 2014 at four Swedish cystectomy centers were retrospectively evaluated. Clinicopathological data were obtained, including data of allogenic blood administration. Survival data was analyzed by Kaplan-Meier plotting and Cox regression.Results: One third of the cohort received blood transfusions during the period of NAC. In univariate analysis, blood transfusions during NAC, nodal stage and advanced tumor stage (pT >2) were negative prognostic factors for survival. In multivariate analysis, only pNx and pT >2 remained significant negative prognostic factors. In a subgroup analysis consisting of patients with localized tumors without dissemination (n = 96), patients that received transfusions during NAC showed an 18.5% absolute risk increase of death at five years of observation, although without statistical significance (p = .197).Conclusions: This is the first time that the extent and plausible effects of allogenic blood transfusions during NAC is examined in MIBC. Data suggest that there may be an association between blood transfusion and poor pathological and oncological outcome. Firm conclusions are difficult to draw due to few study participants and the retrospective nature of the study.

Fewer tumour draining sentinel nodes in patients with progressing muscle invasive bladder cancer, after neoadjuvant chemotherapy and radical cystectomy.

PURPOSE: To examine the relationship between the number of tumour draining sentinel nodes (SNs) and pathoanatomical outcomes, in muscle-invasive bladder cancer (MIBC), in patients undergoing neoadjuvant chemotherapy (NAC) and radical cystectomy (RC). MATERIALS AND METHODS: In an ongoing prospective multicenter study, we included 230 patients with suspected urothelial MIBC from ten Swedish urological centers. All underwent TURb and clinical staging. From the cohort, 116 patients with urothelial MIBC; cT2-cT4aN0M0, underwent radical cystectomy (RC) and lymphadenectomy with SN-detection (SNd). 83 patients received cisplatin-based NAC and 33 were NAC-naïve. The number and locations of detected SNs and non-SNs were recorded for each patient. The NAC treated patients were categorized by pathoanatomical outcomes post-RC into three groups: complete responders (CR), stable disease (SD) and progressive disease (PD). Selected covariates with possible impact on SN-yield were tested in uni -and multivariate analyses for NAC-treated patients only. RESULTS: In NAC treated patients, the mean number of SNs was significantly higher in CR patients (3.3) and SD patients (3.6) compared with PD patients (1.4) (p = 0.034). In a linear multivariate regression model, the number of harvested nodes was the only independent variable that affected the number of SNs (p = 0.0004). CONCLUSIONS: The number of tumor-draining SNs in NAC-treated patients was significantly lower in patients with progressive disease.

Increased CD4+ T cell lineage commitment determined by CpG methylation correlates with better prognosis in urinary bladder cancer patients.

BACKGROUND: Urinary bladder cancer is a common malignancy worldwide. Environmental factors and chronic inflammation are correlated with the disease risk. Diagnosis is performed by transurethral resection of the bladder, and patients with muscle invasive disease preferably proceed to radical cystectomy, with or without neoadjuvant chemotherapy. The anti-tumour immune responses, known to be initiated in the tumour and draining lymph nodes, may play a major role in future treatment strategies. Thus, increasing the knowledge of tumour-associated immunological processes is important. Activated CD4+ T cells differentiate into four main separate lineages: Th1, Th2, Th17 and Treg, and they are recognized by their effector molecules IFN-γ, IL-13, IL-17A, and the transcription factor Foxp3, respectively. We have previously demonstrated signature CpG sites predictive for lineage commitment of these four major CD4+ T cell lineages. Here, we investigate the lineage commitment specifically in tumour, lymph nodes and blood and relate them to the disease stage and response to neoadjuvant chemotherapy. RESULTS: Blood, tumour and regional lymph nodes were obtained from patients at time of transurethral resection of the bladder and at radical cystectomy. Tumour-infiltrating CD4+ lymphocytes were significantly hypomethylated in all four investigated lineage loci compared to CD4+ lymphocytes in lymph nodes and blood (lymph nodes vs tumour-infiltrating lymphocytes: IFNG -4229 bp p < 0.0001, IL13 -11 bp p < 0.05, IL17A -122 bp p < 0.01 and FOXP3 -77 bp p > 0.05). Examination of individual lymph nodes displayed different methylation signatures, suggesting possible correlation with future survival. More advanced post-cystectomy tumour stages correlated significantly with increased methylation at the IFNG -4229 bp locus. Patients with complete response to neoadjuvant chemotherapy displayed significant hypomethylation in CD4+ T cells for all four investigated loci, most prominently in IFNG p < 0.0001. Neoadjuvant chemotherapy seemed to result in a relocation of Th1-committed CD4+ T cells from blood, presumably to the tumour, indicated by shifts in the methylation patterns, whereas no such shifts were seen for lineages corresponding to IL13, IL17A and FOXP3. CONCLUSION: Increased lineage commitment in CD4+ T cells, as determined by demethylation in predictive CpG sites, is associated with lower post-cystectomy tumour stage, complete response to neoadjuvant chemotherapy and overall better outcome, suggesting epigenetic profiling of CD4+ T cell lineages as a useful readout for clinical staging.

Urothelial bladder cancer may suppress perforin expression in CD8+ T cells by an ICAM-1/TGFß2 mediated pathway.

The immune system plays a significant role in urothelial bladder cancer (UBC) progression, with CD8+ T cells being capable to directly kill tumor cells using perforin and granzymes. However, tumors avoid immune recognition by escape mechanisms. In this study, we aim to demonstrate tumor immune escape mechanisms that suppress CD8+ T cells cytotoxicity. 42 patients diagnosed with UBC were recruited. CD8+ T cells from peripheral blood (PB), sentinel nodes (SN), and tumor were analyzed in steady state and in vitro-stimulated conditions by flow cytometry, RT-qPCR, and ELISA. Mass spectrometry (MS) was used for identification of proteins from UBC cell line culture supernatants. Perforin was surprisingly found to be low in CD8+ T cells from SN, marked by 1.8-fold decrease of PRF1 expression, with maintained expression of granzyme B. The majority of perforin-deficient CD8+ T cells are effector memory T (TEM) cells with exhausted Tc2 cell phenotype, judged by the presence of PD-1 and GATA-3. Consequently, perforin-deficient CD8+ T cells from SN are low in T-bet expression. Supernatant from muscle invasive UBC induces perforin deficiency, a mechanism identified by MS where ICAM-1 and TGFß2 signaling were causatively validated to decrease perforin expression in vitro. Thus, we demonstrate a novel tumor escape suppressing perforin expression in CD8+ T cells mediated by ICAM-1 and TGFß2, which can be targeted in combination for cancer immunotherapy.

Neoadjuvant Chemotherapy Reinforces Antitumour T cell Response in Urothelial Urinary Bladder Cancer.

Evidence indicates that neoadjuvant chemotherapy (NAC) may promote antitumour immune responses by activating T cells. The tumour-draining sentinel node (SN) is a key site to study tumour-specific T cell activation, being the primary immunological barrier against the tumour. In this prospective study, we set out to elucidate the effects of NAC on T cell subsets in the SNs of patients with muscle-invasive urothelial bladder cancer. We found that CD8+ effector T (Teff) cell exhaustion was reduced after NAC treatment, while cytotoxicity was increased. Additionally, in complete responders (CR patients), these cells were functionally committed effectors, as displayed by epigenetic analysis. In CD4+ Teffs, NAC treatment was associated with increased clonal expansion of tumour-specific SN-derived cells, as demonstrated by a specific cell reactivity assay. In contrast, we observed an attenuating effect of NAC on regulatory T cells (Tregs) with a dose-dependent decrease in Treg frequency and reduced effector molecule expression in the remaining Tregs. In addition, multicolour flow cytometry analysis revealed that CR patients had higher Teff to activated Treg ratio, promoting antitumoural T cell activation. These results suggest that NAC reinforces the antitumour immune response by activating the effector arm of the T cell compartment and diminishing the influence of suppressive Tregs. PATIENT SUMMARY: In this report, we analysed the effect of chemotherapy on immune cell subsets of 40 patients with advanced bladder cancer. We found that chemotherapy has a positive effect on immune effector T cells, whereas an opposite, diminishing effect was observed for immune-suppressive regulatory T cells. We conclude that chemotherapy reinforces the antitumour immune response in bladder cancer patients.

Urinary Bladder Cancer Tregs Suppress MMP2 and Potentially Regulate Invasiveness.

Regulatory T cells (Treg) have long been considered one-sided suppressors of antitumor immune responses and hence associated with poor patient outcome in cancer. However, evidence is mounting of a paradoxical positive prognostic effect of Tregs on certain malignancies, including urinary bladder cancer (UBC). This discrepancy has partly been attributed to the shear misidentification of Tregs, but also to the inflammatory profile of the tumor. Our aim was to determine whether tumor-infiltrating Forkhead box P3+ (FOXP3+) cells confer a stable Treg phenotype and to investigate putative beneficial Treg functions, focusing on tumor-promoting inflammatory pathways in UBC. Patients (n = 52) with suspected UBC were prospectively included. We show, by using a broad range of analytical approaches, that tumor-infiltrating CD4+FOXP3+ T cells in UBC phenotypically, functionally, and epigenetically represent a true Treg population. At the invasive front of UBC tumors, we found an inverse relationship between Treg frequency and expression of matrix metalloproteinase 2 (MMP2), a key proinvasive factor induced by tumor-promoting inflammation. Correspondingly, a significant, dose-dependent Treg-mediated downregulation of MMP2 protein and mRNA expression was observed in both macrophages and UBC cells. Also, we found that Treg frequency specifically at the invasive front positively correlated with survival. Thus, we identify Treg-mediated suppression of MMP2 in the tumor microenvironment as a mechanism explaining the paradoxical positive prognostic impact of tumor-infiltrating Tregs in UBC. Cancer Immunol Res; 6(5); 528-38. ©2018 AACR.

Sentinel node detection in muscle-invasive urothelial bladder cancer is feasible after neoadjuvant chemotherapy in all pT stages, a prospective multicenter report.

PURPOSE: To determine whether sentinel node detection (SNd) in muscle-invasive urothelial bladder cancer (MIBC) can be performed in patients undergoing neoadjuvant chemotherapy (NAC) and determine whether SNd is feasible in all pT stages, including pT0. BACKGROUND: Previous published series of SNd in MIBC have not included patients undergoing NAC, and systematic reports of pT0 patients w/wo NAC were absent. Translational immunological tumor research on MIBC focusing on SNd, in the era of NAC, requires technical feasibility. Additionally, SNd in MIBC requests further evaluations as a method for nodal staging. MATERIALS AND METHODS: Ninety-nine patients with suspected urothelial MIBC were prospectively selected from six urological centers. After TUR-B and primary staging, 65 MIBC patients qualified for radical cystectomy. Precystectomy staging was cT2a-T4aN0M0, including 47 NAC patients and 18 chemo-naïve patients. All 65 patients underwent intraoperative SNd by peritumoral injection of 80 Mbq Technetium and Geiger probe detection. Postcystectomy staging was pT0-T4aN0-N2M0. SNs were defined by two calculations, SNdef1 and SNdef2. RESULTS: Totally 1063 lymph nodes were removed (total SNs; 222-227). NAC patients with pT0 (n = 24) displayed a true positive detection in 91.7 % by either SNdef, with a median of 3.0 SNs. NACpT >0 patients had a true positive detection in 87 % (SNdef1) and 91.3 % (SNdef2). In a univariate analysis, patient group neither NAC nor tumor downstaging influenced detection rates, regardless of SN definition. In total eight patients, 4/22 metastatic nodes were SNs while 18/22 were non-SNs. CONCLUSIONS: Sentinel node detection in MIBC is feasible also in NAC patients, regardless of pT stage. SNd played no role in nodal staging.

Detection of micrometastases by flow cytometry in sentinel lymph nodes from patients with renal tumours.

BACKGROUND: Stage is an important prognostic factor in renal tumours and dissemination to regional lymph nodes is associated with poor outcomes. Lymph nodes are routinely assessed by immunohistochemistry and microscopic evaluation, a time-consuming process where micrometastases might go undiagnosed. We evaluate an alternative method for detecting metastatic cells in sentinel nodes (SNs) by flow cytometry. METHODS: A total of 15 nodes from 5 patients diagnosed with renal tumours were analysed by flow cytometry. Staining for the intracellular marker cytokeratin 18 (CK18) with the surface markers carbonic anhydrase IX (CA9) and Cadherin 6 were used in flow cytometry analysis. Peripheral blood mononuclear cells (PBMCs) with the addition of known concentrations of cancer cell lines were analysed to investigate the sensitivity of micrometastasis detection. RESULTS: Stability of the assay was marked by low intra-assay variability (coefficient of variance ⩽16%) and low inter-assay variability (R2=0.9996-1). Eight nodes in four patients were positive for metastasis; six of them were considered being micrometastatic. These metastases were undetected by routine pathology and the patients were restaged from pN0 to pN1. CONCLUSIONS: Flow cytometry is able to detect micrometastases in lymph nodes of renal tumour patients that were undetected under H&E examination.

Angiogenesis and other markers for prediction of survival in metastatic renal cell carcinoma.

OBJECTIVE: The treatment of metastatic renal cell carcinoma (RCC) remains a clinical challenge. Factors predicting any benefit of different therapies would therefore be useful. Angiogenesis is important in tumor progression and the development of metastases. The aim of this study in patients with distant metastases at diagnosis was to evaluate possible outcome information obtained with a number of soluble angiogenic variables in serum. MATERIAL AND METHODS: Serum samples were taken at diagnosis from 120 consecutive patients with metastatic RCC who were operated on with radical nephrectomy. Different clinicopathological variables and vascular endothelial growth factor (VEGF), vascular endothelial growth factor receptor-1 (VEGFR-1), basic fibroblast growth factor (bFGF) and erythropoietin levels in serum were compared with the clinical course. RESULTS: The median survival time for all patients was 9 months. Six patients (5%) died during the postoperative period, all of whom had a performance status (PS) of 2 or 3. None of the angiogenic factors (VEGF, VEGFR-1, bFGF, erythropoietin) gave any prognostic information, except that VEGF was associated with survival (p = 0.0234) in patients with a good PS. A number of other variables gave prognostic information in univariate analysis but, after multivariate analysis, only PS (p = 0.002), the number of metastatic sites (p = 0.003) and capsule invasion (p = 0.017) remained as independent predictive factors. CONCLUSIONS: Among predictive factors, only PS, the number of metastatic sites and capsule invasion independently predicted survival in patients with metastatic RCC, while soluble angiogenic factors in serum gave no prognostic information. Nephrectomy in patients with metastatic RCC remains controversial but long-term survival can be achieved in selected patients, especially those with a good PS.

Adrenal metastasis in renal cell carcinoma: a recommendation for adjustment of the TNM staging system.

OBJECTIVES: To study the incidence of adrenal metastasis in patients with renal cell carcinoma (RCC) of all stages, its correlation with clinicopathological variables and its impact on survival. Furthermore, the need for adrenalectomy as part of the routine radical nephrectomy was assessed. MATERIAL AND METHODS: The medical records of 321 patients with RCC of all stages who were operated on with the aim of performing adrenalectomy combined with radical nephrectomy between 1982 and 2000 were reviewed. The accuracy of the available preoperative radiological examinations was evaluated and any adrenal involvement was compared with other clinical and histopathological findings. RESULTS: Ipsilateral adrenal tumour involvement was detected in 17/321 patients (5.3%). In four of these patients, the adrenal gland was the only preoperatively found metastatic site. Factors predicting the presence of ipsilateral adrenal metastases were male gender, tumour size, vein invasion, renal capsule and perirenal fat invasion. Tumour location within the kidney and tumour side had no predictive value for the presence of adrenal metastasis. The presence of ipsilateral adrenal involvement was a significant adverse prognostic variable, indicating a short survival time (p<0.001). CONCLUSIONS: Ipsilateral adrenal metastasis is a highly adverse prognostic factor. In the TNM staging system, adrenal gland involvement should be staged as M1a. Ipsilateral adrenalectomy in conjunction with radical nephrectomy should be performed if an adrenal lesion cannot be cleared of suspicion after preoperative radiological imaging, as in locally advanced tumours. The adrenal gland can be left in situ if the ipsilateral adrenal gland is assessed as normal at the preoperative investigation and perioperatively by the surgeon.