Search

Monocular depth estimation from camera images is very important for surrounding scene evaluation in many technical fields from automotive to medicine. However, traditional triangulation methods using stereo cameras or multiple views with the assumption of a rigid environment are not applicable for endoscopic domains. Particularly in cystoscopies it is not possible to produce ground truth depth information to directly train machine learning algorithms for using a monocular image directly for depth prediction. This work considers first creating a synthetic cystoscopic environment for initial encoding of depth information from synthetically rendered images. Next, the task of predicting pixel-wise depth values for real images is constrained to a domain adaption between the synthetic and real image domains. This adaptation is done through added gated residual blocks in order to simplify the network task and maintain training stability during adversarial training. Training is done on an internally collected cystoscopy dataset from human patients. The results after training demonstrate the ability to predict reasonable depth estimations from actual cystoscopic videos and added stability from using gated residual blocks is shown to prevent mode collapse during adversarial training.

Item does not contain fulltext This Seminar presents the current best practice for the diagnosis and management of bladder cancer. The scope of this Seminar ranges from current challenges in pathology, such as the evolving histological and molecular classification of disease, to advances in personalised medicine and novel imaging approaches. We discuss the current role of radiotherapy, surgical management of non-muscle-invasive and muscle-invasive disease, highlight the challenges of treatment of metastatic bladder cancer, and discuss the latest developments in systemic therapy. This Seminar is intended to provide physicians with knowledge of current issues in bladder cancer.

The European Association of Urology (EAU) delved into the very latest in urological science at the 37th Annual Congress, EAU22, on 1st–4th July in Amsterdam, the Netherlands. The four days were filled to the brim with 2,580 presentations by more than 900 speakers, and 56 courses organised by the European School of Urology (ESU). This included six Game Changing sessions, a day of live surgery, eight plenary sessions, Section meetings, Urology Beyond Europe, poster presentations, prestigious awards, patient days, and industry sessions. EAU Nurses (EAUN) also held EAUN22 during this time. The congress attracted over 8,200 participants from 124 countries, with some also opting to participate via the virtual option. The opening night not only honoured pioneering and promising urologists, it also commemorated the beginning of the 50th celebratory year of the EAU. Secretary General Frans Debruyne was awarded a bronze bust to mark his extraordinary contributions to the EAU during its 50-year existence.

Background Receptor activator of NF kappa B (RANK) and its ligand have an essential role in T-cell regulation and the development of bone metastases. The role of RANK expression in muscle-invasive bladder cancer (MIBC) is unknown. Objective To assess the relevance of RANK expression in patients with MIBC. Design, setting, and participants Expression of RANK was assessed via immunohistochemistry of benign urothelium, MIBC tissue, and lymph node metastases from 153 patients undergoing radical cystectomy. Expression data from The Cancer Genome Atlas (TCGA) cohort were analyzed for potential associations with molecular subtypes and outcome. Outcome measurements and statistical analysis RANK expression was correlated with clinical and pathological parameters and to individual data for the clinical course of MIBC. Results and limitations Expression of RANK was significantly higher in both primary tumors (p = 0.02) and lymph node metastases (p = 0.01) compared to normal urothelium. In tumor tissue, RANK expression was significantly lower in patients with locally advanced disease and lymph node involvement compared to those with organ-confined disease (p = 0.0009) and node-negative MIBC (p = 0.0002). In univariable and multivariable analyses, high expression of RANK was associated with a longer time to recurrence (p = 0.0005 and 0.01) and better cancer-specific (p = 0.0004 and 0.007) and overall survival (p = 0.002 and 0.04). High expression of RANK was associated with better outcome for patients with luminal infiltrated tumors in the TCGA cohort. Conclusions RANK expression is increased in bladder cancer tissue compared to benign urothelium, with higher expression in organ-defined compared to locally advanced disease. High RANK expression indicates a favorable prognosis in MIBC. The prognostic role differs in tumors of different molecular subtypes. Patient summary Expression of a protein involved in bone turnover regulation (RANK) is higher in bladder cancer tissue than in benign bladder tissue. However, high levels of RANK on tumor cells indicate favorable prognosis for patients with bladder cancer that invades the muscle layer of the bladder.

Background: Radiotherapy constitutes an important therapeutic option for prostate cancer. However, prostate cancer cells often acquire resistance during cancer progression, limiting the cytotoxic effects of radiotherapy. Among factors regulating sensitivity to radiotherapy are members of the Bcl-2 protein family, known to regulate apoptosis at the mitochondrial level. Here, we analyzed the role of anti-apoptotic Mcl-1 and USP9x, a deubiquitinase stabilizing Mcl-1 protein levels, in prostate cancer progression and response to radiotherapy. Methods: Changes in Mcl-1 and USP9x levels during prostate cancer progression were determined by immunohistochemistry. Neutralization of Mcl-1 and USP9x was achieved by siRNA-mediated knockdown. We analyzed Mcl-1 stability after translational inhibition by cycloheximide. Cell death was determined by flow cytometry using an exclusion assay of mitochondrial membrane potential-sensitive dye. Changes in the clonogenic potential were examined by colony formation assay. Results: Protein levels of Mcl-1 and USP9x increased during prostate cancer progression, and high protein levels correlated with advanced prostate cancer stages. The stability of Mcl-1 reflected Mcl-1 protein levels in LNCaP and PC3 prostate cancer cells. Moreover, radiotherapy itself affected Mcl-1 protein turnover in prostate cancer cells. Particularly in LNCaP cells, the knockdown of USP9x expression reduced Mcl-1 protein levels and increased sensitivity to radiotherapy. Conclusion: Posttranslational regulation of protein stability was often responsible for high protein levels of Mcl-1. Moreover, we demonstrated that deubiquitinase USP9x as a factor regulating Mcl-1 levels in prostate cancer cells, thus limiting cytotoxic response to radiotherapy.

Supplementary Tables/Figures: 7

Drugs targeting the programmed cell death protein 1 (PD-1) pathway are approved as therapies for an increasing number of cancer entities, including renal cell carcinoma. Despite a significant increase in overall survival, most treated patients do not show durable clinical responses. A combination of checkpoint inhibitors could provide a promising improvement. The aim of the study was to determine the most promising checkpoint blockade combination for renal cell carcinoma patients. Tumor-infiltrating lymphocytes (TIL) and autologous peripheral blood mononuclear cells (PBMC) were isolated from patients undergoing surgery for primary tumors. Cells were stained for multicolor flow cytometry to determine the (co)expression of five inhibitory receptors (iR), PD-1, LAG-3, Tim-3, BTLA, and CTLA-4, on T-cell populations. The function of these TILs was assessed by intracellular cytokine staining after in vitro stimulation in the presence or absence of PD-1 ± LAG-3 or Tim-3–specific antibodies. Although the percentage of iR+ T cells was low in PBMCs, both CD4+ and CD8+ T cells showed increased frequencies of PD-1+, LAG-3+, and Tim-3+ cells on TILs. The most frequent iR combination was PD-1 and LAG-3 on both CD4+ and CD8+ TILs. Blockade of PD-1 resulted in significant LAG-3, but not Tim-3, upregulation. The dual blockade of PD-1 and LAG-3, but not PD-1 and Tim-3, led to increased IFNγ release upon in vitro stimulation. Together, these data suggest that dual blockade of PD-1 and LAG-3 is a promising checkpoint blockade combination for renal cell carcinoma.

The significance of lactate transporters has been recognized in various cancer types, but their role in urothelial carcinoma remains mostly unknown. The aim of this study was to investigate the functional importance of the monocarboxylate transporter (MCT) 4 in preclinical models of urothelial carcinoma and to assess its relevance in patient tumors. The association of MCT4 expression with molecular subtypes and outcome was determined in The Cancer Genome Atlas (TCGA) cohort and two independent cohorts of patients with urothelial carcinoma. Silencing of MCT4 was performed using siRNAs in urothelial carcinoma cell lines. Effects of MCT4 inhibition on cell growth, apoptosis, and production of reactive oxygen species (ROS) were assessed. Moreover, effects on lactate efflux were determined. The in vivo effects of MCT4 silencing were assessed in an orthotopic xenograft model. MCT4 expression was higher in the basal subtype. Decreased MCT4 methylation and increased RNA and protein expression were associated with worse overall survival (OS). Inhibition of MCT4 led to a reduction in cell growth, induction of apoptosis, and an increased synthesis of ROS. MCT4 inhibition resulted in intracellular accumulation of lactate. In vivo, stable knockdown of MCT4 reduced tumor growth. The expression of MCT4 in urothelial carcinoma is associated with features of aggressive tumor biology and portends a poor prognosis. Inhibition of MCT4 results in decreased tumor growth in vitro and in vivo. Targeting lactate metabolism via MCT4 therefore provides a promising therapeutic approach for invasive urothelial carcinoma, especially in the basal subtype.

Suppl. Table 1: Characteristics of the TCGA cohort; Suppl. Table 2: Characteristics of cohort A. CIS= carcinoma in situ; Suppl. Table 3: Characteristics of cohort B Suppl. Fig. 1. Correlation of degree of DNA methylation at CpG site cg10183885 in the SLC16A3 gene region (see Suppl. Table 4) and MCT4 mRNA expression; Suppl. Fig. 2. Correlation of gene expression and protein expression in cohort B. Suppl. Fig. 3. Representative H&E (upper panel) and immunohistochemical images in matched bladder cancer samples (each column is one patient). CD8 was used to stain T cells (middle panel) in comparison with MCT4 expression (lower panel). Suppl. Fig. 4. Protein expression of MCT4 in benign urothelium, primary tumor tissue and lymph node metastases (cohort A). Suppl. Fig. 5. Protein expression in primary tumors according to tumor stage (left) and lymph node involvement (right) (cohort A).; Suppl. Fig. 6. MCT4 mRNA expression correlates with molecular subtypes in the TCGA cohort using the updated TCGA nomenclature (n=408); Suppl. Fig. 7. MCT4 gene expression does not correlate with overall survival in a cohort of patients undergoing neoadjuvant chemotherapy; Suppl. Fig. 8. Expression of MCT4 in urothelial carcinoma cell lines detected by Western Blot (A) and qPCR (B); Suppl. Fig. 9. Effect of MCT4 silencing on cell growth in hypoxic conditions determined by MTT assay. Suppl. Fig. 10. Effect of inhibition of MCT4 using siRNA on cell growth of the MCT4 negative cell line RT4 determined by MTT assay; Suppl. Fig 11. Effect of MCT4 silencing on cell growth in glucose free conditions determined by MTT assay; Suppl.Fig. 12. Effect of different concentrations of MCT4 specific antisense olignucleotide (ASO) on cell growth determined by MTT assay. Suppl. Fig. 13. Oxygen consumption rates at baseline (A), following glucose injection (B) and following oligomycin injection (C) in cell lines treated with control siRNA or MCT4 specific siRNAs.

PDF file, 405K, Supplementary methods, including detailed description of patient cohort, MCT4 immunohistochemical staining and TMA analysis, MCT4 knockdown experiments, Western Blot analysis, RNA isolation and quantification, DNA isolation and promoter methylation analyses, promoter reporter constructs and in vitro methylation, transient transfection and reporter assays as well as statistical analyses. Supplementary Table S1: Comparison of the three independent patient cohorts. Supplementary Table S2: Correlation of MCT4 protein expression in ccRCC tissue with clinicopathological parameters in the TuCP1-cohort. Supplementary Table S3: CpG sites analyzed by the Infinium HumanMethylation450 BeadChip and by MALDI-TOF MS. Supplementary Table S4: Correlation of MCT4 protein expression in ccRCC tissue with clinicopathological parameters in the TuCP2-cohort. Supplementary Table S5: Univariate Cox regression analysis of prognostic factors for survival in the TuCP1-, TuCP2- and the TCGA-cohort. Supplementary Table S6: Results from ROC analysis for survival in the TuCP2- and TCGA-cohort. Supplementary Figure S1: Validation of MCT4 antibody by MCT4 knockdown and representative pictures of stained non-tumor and ccRCC tissue for the evaluation of MCT4 protein expression by the composite IHC score. Supplementary Figure S2: MCT4 protein and mRNA expression and correlation with specific CpG sites in the TuCP2-cohort. Supplementary Figure S3: Kaplan-Meier curve of cancer-specific survival based on percentage of MCT4 protein expression in ccRCC tissue of the TuCP2-cohort. Supplementary Figure S4: DNA methylation at the SLC16A3 promoter in the TCGA-cohort (Infinium Human Methylation 450 Bead Chip) depending on VHL mutations.

Supplementary-Figure 1 Tumor cell infiltration of other organs after PTH/ZA treatment (A) The combined treatment with PTH/ZA does not affect the homing of MDA tumor cells to other organs; n=14/15 in 2 experiments. Evaluation of blood vessel numbers and binding of cancer cells to vascular cells after PTH/ZA treatment (B) PTH/ZA did not affect blood vessel numbers in the treated mice. Proximal tibiae from 4 mice/group were evaluated and the number of blood vessels adjusted to the evaluated area. (C) Binding of cancer cells to immune cells (CD45+) or vascular cells (CD31 or PLVAP) was not affected by treatment with PTH/ZA, n=13/11. Bone marrow from control and treated mice that received cancer cells 24 hours prior to bone marrow isolation was subjected to addition of magnetic beads precoated with CD45 antibody and carefully removed. Addition of either CD31 or PLVAP antibody coated beads followed and beads were used for DNA isolation and evaluation of cancer cell presence. The amount of cancer cell specific DNA was corrected to murine -actin. PTH/ZA treatment does not affect colony forming units of hematopoietic populations (D) There were no changes in various progenitor colonies when bone marrow from treated mice was cultured in vitro for 2 weeks in specialized media, stained with crystal violet and counted; n=6/6/6/6 in 3 experiments. CFU-GEMM is shown in Fig. 1C. Examples of the various colonies are shown below the graphs (light microscopy, 40x).

Supplementary-Figure 3 Correlations between MSCs, CFU-f and homing (A) The percentage of detected MSCs in vivo in control mice and after PTH/ZA treatment correlates negatively with the number of detected tumor cells in the bone marrow, n=40, Pearson''s correlation r=-0.59. (B) The percentage of detected MSCs in vivo in control mice and after PTH/ZA treatment correlates positively with the number of CFU-fs in vitro. (n=27; Pearson correlation r=0.57). (C) In the absence of tumor cells, the combination of PTH/ZA diminishes the MSC subpopulation. Please compare to Fig. 1G obtained in the presence of cancer cells, n=9/12/9/9 in three experiments. Apoptosis after PTH/ZA treatment (D) Apoptosis in CD45- Ter119- CD31+ cells increased after treatment with PTH/ZA and ZA alone, whereas apoptosis in CD45- Ter119- CD146+ cells only increased in the PTH/ZA-treated group, n=10/12/7/7 in 3 experiments.

Supplementary-Table 1A. The markers for murine mesenchymal stromal cells were selected based on literature. Supplementary-Table 1B. Model summary of stepwise multiple-regression analysis of the relation of the selected murine MSC markers and homing of tumor cells; *p

Supplementary-Figure 5 Flow cytometry of MSC markers in human bone marrow biopsies Bone marrow aspirates from patients with prostate cancer (A) and breast cancer (B) who had an evaluation of their bone marrows for the presence of cytokeratin-stained cells to detect disseminated cancer cells (DTCs) were stained for the various MSC markers and analyzed by flow cytometry. The expression of the following markers was not different between cytokeratin negative or positive biopsies: MSCA1, Stro1, CD146, FAP, CD44, CD144, CD73, CD349, CD56 and CD140b in prostate cancer biopsies, n=15/21 (A) and MSCA1, Stro1, CD146, FAP, CD44, CD73 and CD349 in breast cancer biopsies, n=30/30 (B). (C) To exclude any effect of neoadjuvant chemotherapy on the MSC population, the breast cancer patient samples were divided into samples from patients with or without neoadjuvant chemotherapy. Both groups showed an increase in the MSC subpopulation in samples without evidence of cytokeratin-stained cells in bone marrow, n= 10/11/20/19.

Excel file with all five supplemental tables, each contained on a separate tab. Table S1 describes the patient cohort and clinical characteristics. Table S2 describes the targets of our custom sequencing approach. Table S3 relates cfDNA yields and subsequent ctDNA fractions from all samples. Table S4 describes the somatic mutations identified. Table S5 describes the chromosomal rearrangements identified.

Purpose: To elicit a long-lasting antitumor immune response, CD8+ and CD4+ T cells should be activated. We attempted to isolate HLA-DR–presented peptides directly from dissected solid tumors, in particular from renal cell carcinoma, to identify MHC class II ligands from tumor-associated antigens (TAA) for their use in peptide-based immunotherapy.Experimental Design: Tumor specimens were analyzed by immunohistochemical staining for their HLA class II expression. HLA class II peptides were subsequently isolated and identified by mass spectrometry. Gene expression analysis was done to detect genes overexpressed in tumor tissue. Peptides from identified TAAs were used to induce peptide-specific CD4+ T-cell responses in healthy donors and in tumor patients.Results: In the absence of inflammation, expression of MHC class II molecules is mainly restricted to cells of the immune system. To our surprise, we were able to isolate and characterize hundreds of class II peptides directly from primary dissected solid tumors, especially from renal cell carcinomas, and from colorectal carcinomas and transitional cell carcinomas. Infiltrating leukocytes expressed MHC class II molecules and tumor cells, very likely under the influence of IFNγ. Our list of identified peptides contains ligands from several TAAs, including insulin-like growth factor binding protein 3 and matrix metalloproteinase 7. The latter bound promiscuously to HLA-DR molecules and were able to elicit CD4+ T-cell responses.Conclusions: Thus, our direct approach will rapidly expand the limited number of T-helper epitopes from TAAs for their use in clinical vaccination protocols.

Supplementary-Figure 2 Characterization of the affected stromal population after PTH/ZA treatment (A-B) Using flow cytometry, potential markers of mesenchymal stromal cells were evaluated in bone marrow of PTH/ZA-treated mice. Gating was applied after the exclusion of hematopoietic and erythroid cells (CD45- Ter119-). (B) PTH/ZA diminished the size of populations expressing the following surface markers: Sca1+, CD29+, CD31+, CD44-, αSMA+, leptin receptor+, and CD146+; n=15/15 in 3 experiments.

Purpose: The monocarboxylate transporter 4 (MCT4) is a metabolic target in tumor biology because it mediates lactate transport across membranes resulting in antiapoptotic effects. Cell experiments support the importance of MCT4 in clear cell renal cell carcinoma (ccRCC). In this study, we assessed the prognostic potential of MCT4 expression in ccRCC and its epigenetic regulation by DNA methylation as novel predictive marker for patient outcome using independent ccRCC cohorts.Experimental Design: MCT4 protein expression was quantified in 207 ccRCC and corresponding nontumor tissues. Data of an independent ccRCC cohort from The Cancer Genome Atlas (TCGA) were analyzed on MCT4 mRNA (n = 482) and DNA methylation (n = 283) level. The findings on MCT4 expression and DNA methylation in the SLC16A3 promoter were validated in a third cohort (n = 64). Promoter activity assays were conducted in four RCC cell lines.Results: MCT4 protein expression was upregulated (P < 0.0001) in ccRCC and showed significant association with cancer-related death. Upregulation of MCT4 mRNA expression (P < 0.00001) was confirmed in the TCGA cohort. Single CpG sites correlated inversely with mRNA expression and were associated with overall survival in Kaplan–Meier analyses [HR = 0.39; 95% confidence interval (CI), 0.24–0.64; P[log-rank] = 1.23e−04]. Promoter activity studies confirmed MCT4 regulation by DNA methylation. The significant correlation between MCT4 protein and gene expression or DNA methylation at single CpG sites was validated in a third cohort. Again, higher methylation at individual CpG sites was associated with prolonged survival [HR = 0.05; 95% CI, 0.01–0.40; P[log-rank] = 6.91e−05].Conclusion: We identified SLC16A3 promoter DNA methylation as a novel epigenetic mechanism for MCT4 regulation in ccRCC with first evidence of a biological rationale for prognosis and clinical outcome. Clin Cancer Res; 19(18); 5170–81. ©2013 AACR.

Purpose: Targeted agents and immunotherapies promise to transform the treatment of metastatic bladder cancer, but therapy selection will depend on practical tumor molecular stratification. Circulating tumor DNA (ctDNA) is established in several solid malignancies as a minimally invasive tool to profile the tumor genome in real-time, but is critically underexplored in bladder cancer.Experimental Design: We applied a combination of whole-exome sequencing and targeted sequencing across 50 bladder cancer driver genes to plasma cell-free DNA (cfDNA) from 51 patients with aggressive bladder cancer, including 37 with metastatic disease.Results: The majority of patients with metastasis, but only 14% of patients with localized disease, had ctDNA proportions above 2% of total cfDNA (median 16.5%, range 3.9%–72.6%). Twelve percent of estimable samples had evidence of genome hypermutation. We reveal an aggressive mutational landscape in metastatic bladder cancer with 95% of patients harboring deleterious alterations to TP53, RB1, or MDM2, and 70% harboring a mutation or disrupting rearrangement affecting chromatin modifiers such as ARID1A. Targetable alterations in MAPK/ERK or PI3K/AKT/mTOR pathways were robustly detected, including amplification of ERBB2 (20% of patients) and activating hotspot mutations in PIK3CA (20%), with the latter mutually exclusive to truncating mutations in TSC1. A novel FGFR3 gene fusion was identified in consecutive samples from one patient.Conclusions: Our study demonstrates that ctDNA provides a practical and cost-effective snapshot of driver gene status in metastatic bladder cancer. The identification of a wide spectrum of clinically informative somatic alterations nominates ctDNA as a tool to dissect disease pathogenesis and guide therapy selection in patients with metastatic bladder cancer. Clin Cancer Res; 23(21); 6487–97. ©2017 AACR.

Methods for blood processing and DNA isolation.

File containing all 13 supplementary figures, complete with figure legends.

Breast and prostate cancer cells home to the bone marrow, where they presumably hijack the hematopoietic stem cell niche. We characterize here the elusive premetastatic niche by examining the role of mesenchymal stromal cells (MSC) in cancer cell homing. Decreasing the number of MSC pharmacologically enhanced cancer cell homing to the bone marrow in mice. In contrast, increasing the number of these MSCs by various interventions including G-CSF administration diminished cancer cell homing. The MSC subpopulation that correlated best with cancer cells expressed stem, endothelial, and pericytic cell markers, suggesting these cells represent an undifferentiated component of the niche with vascular commitment. In humans, a MSC subpopulation carrying markers for endothelial and pericytic cells was lower in the presence of cytokeratin+ cells in bone marrow. Taken together, our data show that a subpopulation of MSC with both endothelial and pericytic cell surface markers suppresses the homing of cancer cells to the bone marrow. Similar to the presence of cytokeratin+ cells in the bone marrow, this MSC subpopulation could prove useful in determining the risk of metastatic disease, and its manipulation might offer a new possibility for diminishing bone metastasis formation.Significance: These findings establish an inverse relationship between a subpopulation of mesenchymal stromal cells and cancer cells in the bone marrow. Cancer Res; 78(1); 129–42. ©2017 AACR.

Supplementary-Figure 1. Tumor cell infiltration of other organs after PTH/ZA treatment. Evaluation of blood vessel numbers and binding of cancer cells to vascular cells after PTH/ZA treatment. PTH/ZA treatment does not affect colony forming units of hematopoietic populations. Supplementary-Figure 2. Characterization of the affected stromal population after PTH/ZA treatment. Supplementary-Figure 3. Correlations between MSCs, CFU-f and homing. Apoptosis after PTH/ZA treatment. Supplementary-Figure 4. Absolute numbers of HSPCs and MSCs after G-CSF and G-CSF PTH/ZA treatment and in Mx 1fl/fl knockout mice. Expression of MSC markers on low-passage human MSCs. Supplementary-Figure 5. Flow cytometry of MSC markers in human bone marrow biopsies.

Methods, Tables 1-3, Figures 1-4

Supplementary Figure Legend from Simultaneous Infiltration of Polyfunctional Effector and Suppressor T Cells into Renal Cell Carcinomas

Renal cell carcinoma is frequently infiltrated by cells of the immune system. This makes it important to understand interactions between cancer cells and immune cells so they can be manipulated to bring clinical benefit. Here, we analyze subsets and functions of T lymphocytes infiltrating renal cell tumors directly ex vivo following mechanical disaggregation and without any culture step. Subpopulations of memory and effector CD4+ Th1, Th2, and Th17 and CD8+ Tc1 cells were identified based on surface phenotype, activation potential, and multicytokine production. Compared with the same patient's peripheral blood, T lymphocytes present inside tumors were found to be enriched in functional CD4+ cells of the Th1 lineage and in effector memory CD8+ cells. Additionally, several populations of CD4+ and CD8+ regulatory T cells were identified that may synergize to locally dampen antitumor T-cell responses. [Cancer Res 2009;69(21):8412–9]

Supplementary Figure 1 from Simultaneous Infiltration of Polyfunctional Effector and Suppressor T Cells into Renal Cell Carcinomas

OBJECTIVE: The insulin/insulin-like growth factor 1 (IGF1) pathway is emerging as a crucial component of prostate cancer progression. Therefore, we investigated the role of the novel insulin/IGF1 signaling modulator inceptor in prostate cancer. METHODS: We analyzed the expression of inceptor in human samples of benign prostate epithelium and prostate cancer. Further, we performed signaling and functional assays using prostate cancer cell lines. RESULTS: We found that inceptor was expressed in human benign and malignant prostate tissue and its expression positively correlated with various genes of interest, including genes involved in androgen signaling. In vitro, total levels of inceptor were increased upon androgen deprivation and correlated with high levels of androgen receptor in the nucleus. Inceptor overexpression was associated with increased cell migration, altered IGF1R trafficking and higher IGF1R activation. CONCLUSIONS: Our in vitro results showed that inceptor expression was associated with androgen status, increased migration, and IGF1R signaling. In human samples, inceptor expression was significantly correlated with markers of prostate cancer progression. Taken together, these data provide a basis for investigation of inceptor in the context of prostate cancer.

C-X-C Motif Chemokine Receptor 4 (CXCR4) is part of the human chemokine system and involved in progression and metastasis in renal cell carcinoma (RCC). However, the role of CXCR4 protein expression in RCC remains controversial. In particular, data regarding the subcellular distribution of CXCR4 in RCC and RCC metastasis as well as CXCR4 expression in renal tumors of variant histology are limited. The aim of the present study was the evaluation of the differential CXCR4 expression in RCC primary tumor and metastatic tissue as well as in variant renal histologies. In addition, the prognostic capacity of CXCR4 expression in organ-confined clear cell RCC (ccRCC) was evaluated. Three independent renal tumor cohorts (primary ccRCC cohort n1 = 64; cohort of various histological entities n2 = 146; metastatic RCC tissue cohort n3 = 92) were evaluated using tissue microarrays (TMA). After immunohistochemical staining for CXCR4, nuclear and cytoplasmic expression patterns were evaluated. CXCR4 expression was correlated with validated pathologic prognosticators, clinical data, and overall and cancer-specific survival. Positive cytoplasmic staining was observed in 98% of the benign and 38.9% of the malignant samples. Nuclear staining was positive for 94.1% of the benign samples and 83% of the malignant samples. The median cytoplasmic expression score was found to be higher in benign tissue than in ccRCC (130.00 vs. 0.00); median nuclear expression score analysis indicated the opposite (56.0 vs. 71.0). Within malignant subtypes, the highest expression score was seen in papillary renal cell carcinomas (cytoplasmic: 117.50, nuclear: 41.50). Within benign renal tumors, high cytoplasmic and nuclear CXCR4 expression scores were seen for oncocytomas (cytoplasmic: 100.00, nuclear: 31.00). Expression scores in RCC metastasis ranked between benign renal tissue and ccRCC in cytoplasmic and nuclear expression. Cytoplasmic CXCR4 expression was identified as a prognostic factor for OS and CSS (p = 0.042; p = 0.019). Multivariate analysis including clinicopathological parameters did not reveal an independent prognostic character of CXCR4 expression. CXCR4 expression differs significantly within benign lesions and renal neoplasms. Cytoplasmic and nuclear expression of CXCR4 could be detected across all RCC subtypes. The prognostic value of CXCR4 in ccRCC was confirmed in univariate analysis.

Purpose Complete metastasectomy of renal cell carcinoma (RCC) is receding into the past due to the progress of immuno-oncology-based combinations (IO) in systemic therapy. The prognostic impact of curative intended complete metastasectomy vs. immediate IO-based therapy or tyrosine kinase inhibition (TKI) on progression-free survival (PFS) and cancer-specific survival (CSS) was investigated in the first-line setting. Methods 205 patients with synchronous or metachronous metastasis received complete metastasectomy (n = 80) or systemic therapy (n = 125, TKI: 87, TKI–IO: 13, IO–IO: 25) as first-line therapy. The prognostic impact of these therapies was assessed using Cox regression and Kaplan–Meier analyses. Results First-line complete metastasectomy significantly improved CSS compared to both TKI monotherapy (6.1 vs. 2.6 years, HR 0.45, p p = 0.007). Repetitive complete metastasectomy without ever receiving systemic therapy vs. systemic therapy in first-line significantly prolonged CSS (11.3 vs. 3.1 years, HR 0.34, p = 0.002). First-line complete metastasectomy and subsequent systemic therapy at tumor progression was associated with a significant CSS benefit vs. systemic therapy (5.8 vs. 3.1 years, HR 0.53, p = 0.003), also compared to IO-based combinations (5.8 vs. 3.5 years, HR 0.30, p = 0.017). Median PFS was improved by IO-based therapy compared to TKI monotherapy in the first-line setting (HR 0.61, p = 0.05), with maximal benefit of the TKI–IO combination vs. TKI monotherapy (HR 0.27, p = 0.01), as well as compared to PFS of complete metastasectomy (HR 0.34, p = 0.035). Conclusion Despite the progress of IO-based combination therapies in first line, complete metastasectomy remains an integral part of the multimodality treatment of metastatic RCC.

Bladder outlet obstruction (BOO) is a type of chronic disease that is mainly caused by benign prostatic hyperplasia. Previous studies discovered the involvements of both serum/glucocorticoid-regulated kinase 1 (SGK1) and activated T cell nuclear factor transcription factor 2 (NFAT2) in the proliferation of smooth muscle cells after BOO. However, the relationship between these two molecules is yet to be explored. Thus, this study explored the specific mechanism of the SGK1-NFAT2 signaling pathway in mouse BOO-mediated bladder smooth muscle cell proliferation in vivo and in vitro. In vivo experiments were performed by suturing 1/2 of the external urethra of female BALB/C mice to cause BOO for 2 weeks. In vitro, mouse bladder smooth muscle cells (MBSMCs) were treated with dexamethasone (Dex) or dexamethasone + SB705498 for 12 h and were transfected with SGK1 siRNA for 48 h. The expression and distribution of SGK1, transient receptor potential oxalate subtype 1 (TRPV1), NFAT2, and proliferating cell nuclear antigen (PCNA) were measured by Western blotting, polymerase chain reaction, and immunohistochemistry. The relationship between SGK1 and TRPV1 was analyzed by coimmunoprecipitation. The proliferation of MBSMCs was examined by 5-ethynyl-2'-deoxyuridine and cell counting kit 8 assays. Bladder weight, smooth muscle thickness, and collagen deposition in mice after 2 weeks of BOO were examined. Bladder weight, smooth muscle thickness, the collagen deposition ratio, and the expression of SGK1, TRPV1, NFAT2, and PCNA were significantly increased in mice after 2 weeks of BOO. Compared with the control, 10 μM Dex promoted the expression of these four molecules and the proliferation of MBSMCs. After inhibiting TRPV1, only the expression of SGK1 was not affected, and the proliferation of MBSMCs was inhibited. After silencing SGK1, the expression of these four molecules and the proliferation of MBSMCs decreased. Coimmunoprecipitation suggested that SGK1 acted directly on TRPV1. In this study, SGK1 targeted TRPV1 to regulate the proliferation of MBSMCs mediated by BOO in mice through NFAT2 and then affected the process of bladder remodeling after BOO. This finding may provide a strategy for BOO drug target screening.

Objective To describe the use of artificial intelligence (AI) in medical literature and trial data extraction, and its applications in uro-oncology. This bridging review, which consolidates information from the diverse applications of AI, highlights how AI users can investigate more sophisticated queries than with traditional methods, leading to synthesis of raw data and complex outputs into more actionable and personalized results, particularly in the field of uro-oncology. Methods Literature and clinical trial searches were performed in PubMed, Dimensions, Embase and Google (1999-2020). The searches focused on the use of AI and its various forms to facilitate literature searches, clinical guidelines development, and clinical trial data extraction in uro-oncology. To illustrate how AI can be applied toaddress questions about optimizing therapeutic decision making and individualizing treatment regimens, the Dimensions-linked information platform was searched for "prostate cancer" keywords (76 publications were identified; 48 were included). Results AI offers the promise of transforming raw data and complex outputs into actionable insights. Literature and clinical trial searches can be automated, enabling clinicians to develop and analyze publications expeditiously on complex issues such as therapeutic sequencing and to obtain updates on documents that evolve at the pace and scope of the landscape. An AI-based platform inclusive of 12 trial databases and >100 scientific literature sources enabled the creation of an interactive visualization. Conclusion As the literature and clinical trial landscape continues to grow in complexity and with increasing speed, the ability to pull the right information at the right time from different search engines and resources while excluding social media bias becomes more challenging. This review demonstrates that by applying natural language processing and machine learning algorithms, validated and optimized AI leads to a speedier, more personalized, efficient and focused search compared with traditional methods.

Background This study aims at describing the imaging features of the metastatic presentation of clear cell renal cell carcinoma (ccRCC) in arterial (AP) and portal venous phase (PVP) of contrast-enhanced-computed-tomography (CECT) during clinical follow-up (FU) and to evaluate the necessity of a dual phase approach for metastasis detection. Methods We identified a total of 584 patients that were diagnosed with ccRCC between January 2016 and April 2020. Inclusion criteria were histologically proven ccRCC with metastatic spread, proven by histology or interim follow-up of at least 2 years and follow-up CT examination with AP and PVP CECT including thorax/abdomen and pelvis. Exclusion criteria were defined by missing or incomplete CT-scans or lack of sufficient follow-up. CT studies of 43 patients with histologically proven ccRCCs were analyzed in retrospect. AP and PVP images were analyzed by two radiologists for metastases, two additional independent radiologists analyzed PVP images only. A 5-point Likert scale was used to evaluate the likelihood off the presence of metastasis. Imaging patterns of the metastases were analyzed visually. Results 43 patients (16 female; mean age: 67±10 years) with recurrent ccRCC and metastatic disease were included. Three imaging patterns were observed (solid, heterogeneous or cystic metastases), which rarely exhibited calcifications (2%). All metastases showed hyperenhancement in AP and PVP. Inter-reader agreement was substantial (Fleiss’ κ 0.6–0.8, p

Background Recommendations against prostate-specific antigen (PSA) testing in 2012 have increased advanced-stage diagnosis and prostate cancer–specific mortality rates. Objective To present the position of the European Association of Urology (EAU) in 2021 and provide recommendations for the use of PSA testing as part of a risk-adapted strategy for the early detection of prostate cancer. Evidence acquisition The authors combined their review of relevant literature, including the EAU prostate cancer guidelines 2021 update, with their own knowledge to provide an expert opinion, representing the EAU’s position in 2021. Evidence synthesis The EAU has developed a risk-adapted early prostate cancer detection strategy for well-informed men based on PSA testing, risk calculators, and multiparametric magnetic resonance imaging, which can differentiate significant from insignificant prostate cancer. This approach largely avoids the overdiagnosis/overtreatment of men unlikely to experience disease-related symptoms during their lifetime and facilitates an early diagnosis of men with significant cancer to receive active treatment. It also reduces advanced-stage diagnosis, thereby potentially reducing prostate cancer–specific mortality and improving quality of life. Education is required among urologists, general practitioners, radiologists, policy makers, and healthy men, including endorsement by the European Commission to adapt the European Council’s screening recommendations in its 2022 plan and requests to individual countries for its incorporation into national cancer plans. Conclusions This risk-adapted approach for the early detection of prostate cancer will reverse current unfavourable trends and ultimately save lives. Patient summary The European Association of Urology has developed a patient information leaflet and algorithm for the early diagnosis of prostate cancer. It can identify men who do not need magnetic resonance imaging or a biopsy and those who would not show any symptoms versus those with more aggressive disease who require further tests/treatment. We need to raise awareness of this algorithm to ensure that all well-informed men at risk of significant prostate cancer are offered a prostate-specific antigen test. Take Home Message A risk-adapted early prostate cancer detection strategy, incorporating prostate-specific antigen testing, multiparametric magnetic resonance imaging, risk calculators, and biomarkers, will avoid overdiagnosis/overtreatment of insignificant cancers and ensure early detection and treatment of significant cancers, thereby improving quality of life and reducing prostate cancer–related deaths.

T cell recognition of human leukocyte antigen (HLA)-presented tumor-associated peptides is central for cancer immune surveillance. Mass spectrometry (MS)-based immunopeptidomics represents the only unbiased method for the direct identification and characterization of naturally presented tumor-associated peptides, a key prerequisite for the development of T cell-based immunotherapies. This study reports on the de novo implementation of ion mobility separation-based timsTOF MS for next-generation immunopeptidomics, enabling high-speed and sensitive detection of HLA-presented peptides. A direct comparison of timsTOF-based with state-of-the-art immunopeptidomics using orbitrap technology showed significantly increased HLA peptide identifications from benign and malignant primary samples of solid tissue and hematological origin. First application of timsTOF-based immunopeptidomics for tumor antigen discovery enabled (i) the expansion of benign reference immunopeptidome databases with > 150,000 HLA-presented peptides from 94 primary benign tissue samples, (ii) the refinement of previously described tumor antigens, and (iii) the identification of a vast array of novel tumor antigens, comprising low abundant neoepitopes, that might serve as targets for future cancer immunotherapy development.

Background: The retzius-sparing approach for robotic-assisted radical prostatectomy (RARP) has been increasingly adopted. Symptomatic lymphoceles are a widespread complication of RARP with pelvic lymph node dissection. Here, we present a new technique, the peritoneal purse-string suture (PPSS), that seems to reduce the rate of symptomatic lymphoceles following retzius-sparing RARP with extended pelvic lymph node dissection (ePLND). Methods: The radical prostatectomy and bilateral lymphadenectomy are performed through three separate peritoneal openings. The PPSS uses a single suture in a way similar to a purse-string suture; the openings of both lymphadenectomy fields are widened, and the rectovesical opening from the prostatectomy is simultaneously closed. This report retrospectively evaluates the perioperative and postoperative outcomes of two consecutive patient cohorts undergoing RARP with ePLND by a single surgeon between May 2015 and June 2019, one cohort prior to introducing PPSS as control (n = 145) and the other after introducing PPSS (n = 91). Results: The two study groups were comparable on baseline characteristics, except ASA. There were no Clavien–Dindo grade IV-V complications, and comparable rates of grade I-III complications. The difference in postoperative lymphocele formation was 22% in PPSS versus 27% in the control group (p = 0.33). The rate of symptomatic lymphoceles was significantly lower in the PPSS group (3% vs. 10%, p = 0.047). Conclusion: The PPSS is a feasible procedure that reduces symptomatic lymphoceles in patients undergoing RARP with a retzius-sparing approach.

BackgroundThe clinical utility of molecular profiling and targeted therapies for neuro-oncology patients outside of clinical trials is not established. We aimed at investigating feasibility and clinical utility of molecular profiling and targeted therapy in adult patients with advanced tumors in the nervous system within a prospective observational study.Methodsmolecular tumor board (MTB)@ZPM (NCT03503149) is a prospective observational precision medicine study for patients with advanced tumors. After inclusion of patients, we performed comprehensive molecular profiling, formulated ranked biomarker-guided therapy recommendations based on consensus by the MTB, and collected prospective clinical outcome data.ResultsHere, we present initial data of 661 adult patients with tumors of the nervous system enrolled by December 31, 2021. Of these, 408 patients were presented at the MTB. Molecular-instructed therapy recommendations could be made in 380/408 (93.1%) cases and were prioritized by evidence levels. Therapies were initiated in 86/380 (22.6%) cases until data cutoff. We observed a progression-free survival ratio >1.3 in 31.3% of patients.ConclusionsOur study supports the clinical utility of biomarker-guided therapies for neuro-oncology patients and indicates clinical benefit in a subset of patients. Our data might inform future clinical trials, translational studies, and even clinical care.