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Table S1. Demographic and pathological features of human patients with angiosarcoma Table S2. Demographic and pathological features of dogs with HSA Table S3. Primary antibodies for immunohistochemical evaluation Table S4. Gene ontology and biological functions of fusion partner genes in human angiosarcomas Table S5. Gene ontology and biological functions of fusion partner genes in canine HSAs Table S6. Detection of fusion genes in canine HSAs by RT-PCR Table S7. Biological functions associated with 490 upregulated genes in human angiosarcomas Table S8. Association of frequency between fusion genes, TP53 and PIK3CA mutations Table S9. Association of fusion genes with dog signalment and tumor histology Table S10. Association of frequency between fusion genes, TP53 and PIK3CA mutations Table S11. 41 differentially expressed genes in PF (n = 4) compared to P (n = 9) group Table S12. 212 differentially expressed genes in PF (n = 4) compared to PK (n = 6) group Table S13. 377 differentially expressed genes in PF (n = 4) compared to PFK (n = 4) group Table S14. Immunohistochemical features of human angiosarcomas Table S15. Immunohistochemical features of canine HSAs

(Ai) Box-plot shows the association of copy number of S100A4 gene to Gleason Score of prostate tumors in CaP patients. The graph was generated from the analysis of TCGA clinical data using Oncomine platform. (Aii) Image shows the detection of S100A4 protein in human serum as assessed by immune-Slot blot analyses. The immunoblot data shown here is a representative of experiments conducted three independent times. (Bi) The gel image shows the cloning of s100A4 in pET28a vector using NdeI and Xho I site. (Bii) Immunoblot image shows the detection of S100A4 for different elution fractions of S100A4 protein overexpressed in BL21DE3. (C) Histogram shows the effect of mab6B12 therapy on the rate of proliferation of human CaP cells as assessed by 3[H]thymidine uptake assay. (D) Graph shows the Osterix expression in Bone-mets in CaP patients from Met-CaP Provisional Project 2019 Clinical study as analyzed by cBioportal platform. (E) Heat map shows the gene expression profile of CD4+ T cells Co-cultured with TRAMP-C2 cells in presence and absence of S100A4 protein and mab6B12 antibody.

Supplementary Data showing (i) expression of COMMD3 in PDX models and (ii) association of MYC to COMMD3 in prostate cancer

Sporadic angiosarcomas are aggressive vascular sarcomas whose rarity and genomic complexity present significant obstacles in deciphering the pathogenic significance of individual genetic alterations. Numerous fusion genes have been identified across multiple types of cancers, but their existence and significance remain unclear in sporadic angiosarcomas. In this study, we leveraged RNA-sequencing data from 13 human angiosarcomas and 76 spontaneous canine hemangiosarcomas to identify fusion genes associated with spontaneous vascular malignancies. Ten novel protein-coding fusion genes, including TEX2-PECAM1 and ATP8A2-FLT1, were identified in seven of the 13 human tumors, with two tumors showing mutations of TP53. HRAS and NRAS mutations were found in angiosarcomas without fusions or TP53 mutations. We found 15 novel protein-coding fusion genes including MYO16-PTK2, GABRA3-FLT1, and AKT3-XPNPEP1 in 11 of the 76 canine hemangiosarcomas; these fusion genes were seen exclusively in tumors of the angiogenic molecular subtype that contained recurrent mutations in TP53, PIK3CA, PIK3R1, and NRAS. In particular, fusion genes and mutations of TP53 cooccurred in tumors with higher frequency than expected by random chance, and they enriched gene signatures predicting activation of angiogenic pathways. Comparative transcriptomic analysis of human angiosarcomas and canine hemangiosarcomas identified shared molecular signatures associated with activation of PI3K/AKT/mTOR pathways. Our data suggest that genome instability induced by TP53 mutations might create a predisposition for fusion events that may contribute to tumor progression by promoting selection and/or enhancing fitness through activation of convergent angiogenic pathways in this vascular malignancy.Implications:This study shows that, while drive events of malignant vasoformative tumors of humans and dogs include diverse mutations and stochastic rearrangements that create novel fusion genes, convergent transcriptional programs govern the highly conserved morphologic organization and biological behavior of these tumors in both species.

S1. Representative putative inter-chromosomal fusion transcripts in human AS and canine HSA. SCLT1-NIPBL fusion in human angiosarcoma. S2. Fusion genes and their association with tumor content and sequencing depth in canine HSA tissues. S3. A, Primers designed for amplification of SCLT1-NIPBL fusion transcripts identified in human angiosarcoma. B, SCLT1-NIPBL fusion gene amplification by quantitative RT-PCR. Expression value represents 2(-â^†Ct)(Fusion gene/GAPDH). C, Primers designed for amplification of four fusion transcripts identified in canine hemangiosarcomas. D, Fusion gene amplification was confirmed in canine tumors by quantitative RT-PCR. E, Electrophoresis (1% agarose gel) of DNA product by RTPCR was done. S4. Genetic alterations of fusion partner genes identified in human AS. S5. Cell type enrichment analysis in ASs and TCGA sarcomas. S6. Three distinct molecular groups of canine HSAs. S7. Kaplan-Meier probability of survival for dogs with angiogenic and inflammatory HSA. S8. Association of the fusion events with demographic and histopathologic features in canine HSA. S9. Gene pathways associated with commonly enriched genes in tumors with fusion genes and TP53 mutations in canine HSA. S10. Violin plots display IHC scores of p53, p-p53 (Ser15), and p-p53 (Ser20) in human AS and canine HSA. S11. Violin plots present IHC scores of AKT and p-AKT (Thr308) in human AS and canine HSA. S12. IHC scores of mTOR and p-mTOR (Ser2448) in human AS.

Characteristics of the patients who underwent radical nephroureterectomy for UTUC.

Supplemental Tables

Figure S1. CD133 expression correlated with PSA expression. Figure S2. CD133 expression in xenografts. Figure S3. Characterization of labeled HA10. Table S1. Ex vivo biodistribution data.

Purpose:Metastasis is the major cause of mortality in prostate cancer patients. Factors such as genetic makeup and race play critical role in the outcome of therapies. This study was conducted to investigate the relevance of BMI1 in metastatic prostate cancer disease in Caucasian and African-Americans.Experimental Design:We employed race-specific prostate cancer models, clinical specimens, clinical data mining, gene-microarray, transcription-reporter assay, chromatin-immunoprecipitation (ChIP), IHC, transgenic-(tgfl/fl) zebrafish, and mouse metastasis models.Results:BMI1 expression was observed to be elevated in metastatic tumors (lymph nodes, lungs, bones, liver) of Caucasian and African-American prostate cancer patients. The comparative analysis of stage III/IV tumors showed an increased BMI1 expression in African-Americans than Caucasians. TCGA and NIH/GEO clinical data corroborated to our findings. We show that BMI1 expression (i) positively correlates to metastatic (MYC, VEGF, cyclin D1) and (ii) negative correlates to tumor suppressor (INKF4A/p16, PTEN) levels in tumors. The correlation was prominent in African-American tumors. We show that BMI1 regulates the transcriptional activation of MYC, VEGF, INKF4A/p16, and PTEN. We show the effect of pharmacological inhibition of BMI1 on the metastatic genome and invasiveness of tumor cells. Next, we show the anti-metastatic efficacy of BMI1-inhibitor in transgenic zebrafish and mouse metastasis models. Docetaxel as monotherapy has poor outcome on the growth of metastatic tumors. BMI1 inhibitor as an adjuvant improved the taxane therapy in race-based in vitro and in vivo models.Conclusions:BMI1, a major driver of metastasis, represents a promising therapeutic target for treating advanced prostate cancer in patients (including those belonging to high-risk group).

Oncoprints and Venn diagrams for matched-pair comparisons between UCB and UTUC samples in patients with a history of a prior bladder tumor before resection of the UTUC. The arrows show the time between sample collection in months.

Supplemental figures and box plots

Supplementary Figure 2. Inhibition of colony formation by BMI1 inhibitor and Docetaxel combination in African American and Caucasian CaP models on Colony-formation assay as described in material and methods.

Supplemental legend

Supplementary Methods

Supplementary Table 1. Sequence of primers used for ChIP analysis of PTEN, INK4A/p16 and GAPDH promoters.

Purpose:Aggressive variant prostate cancer (AVPC) is a nonandrogen receptor–driven form of disease that arises in men in whom standard-of-care therapies have failed. Therapeutic options for AVPC are limited, and the development of novel therapeutics is significantly hindered by the inability to accurately quantify patient response to therapy by imaging. Imaging modalities that accurately and sensitively detect the bone and visceral metastases associated with AVPC do not exist.Experimental Design:This study investigated the transmembrane protein CD133 as a targetable cell surface antigen in AVPC. We evaluated the expression of CD133 by microarray and IHC analysis. The imaging potential of the CD133-targeted IgG (HA10 IgG) was evaluated in preclinical prostate cancer models using two different imaging modalities: near-infrared and PET imaging.Results:Evaluation of the patient data demonstrated that CD133 is overexpressed in a specific phenotype of AVPC that is androgen receptor indifferent and neuroendocrine differentiated. In addition, HA10 IgG was selective for CD133-expressing tumors in all preclinical imaging studies. PET imaging with [89Zr]Zr-HA10 IgG revealed a mean %ID/g of 24.30 ± 3.19 in CD133-positive metastatic lesions as compared with 11.82 ± 0.57 in CD133-negative lesions after 72 hours (P = 0.0069). Ex vivo biodistribution showed similar trends as signals were increased by nearly 3-fold in CD133-positive tumors (P < 0.0001).Conclusions:To our knowledge, this is the first study to define CD133 as a targetable marker of AVPC. Similarly, we have developed a novel imaging agent, which is selective for CD133-expressing tumors, resulting in a noninvasive PET imaging approach to more effectively detect and monitor AVPC.

Purpose:To investigate genomic differences between urothelial carcinomas of the upper tract (UTUC) and bladder (UCB), with a focus on defining the clonal relatedness of temporally distinct tumors.Experimental Design:We prospectively sequenced tumors and matched germline DNA using targeted next-generation sequencing methods. The cohort included 195 UTUC patients and 454 UCB patients. For a subgroup of 29 patients with UTUC and a history of a subsequent UCB, both tumors were analyzed to assess their clonal relatedness.Results:With the progression to higher UTUC clinical state, there were fewer alterations in the RTK/RAS pathway but more alterations in TP53/MDM2. Compared with UCB, TP53, RB1, and ERBB2 were less frequently altered in UTUC (26% vs. 46%, 3% vs. 20%, 8% vs. 19%, respectively; Q < 0.001), whereas FGFR3 and HRAS were more frequently altered (40% vs. 26%, 12% vs. 4%, respectively; Q < 0.001). On the basis of an integrated analysis of tumor mutational burden, MSIsensor score and mutational signature, 7.2% of UTUC tumors were classified as MSI-high/MMR-deficient (MSI-H/dMMR). The risk of bladder recurrence after UTUC was significantly associated with mutations in FGFR3, KDM6A, CCND1, and TP53. Comparison of UCB with corresponding UTUC tumors from the same patient supports their clonal relatedness.Conclusions:UTUC and UCB exhibit significant differences in the prevalence of common genomic alterations. In individual patients with a history of both tumors, UCB and UTUC were always clonally related. Genomic characterization of UTUC provides information regarding the risk of bladder recurrence and can identify tumors associated with Lynch syndrome.

Purpose:Neuroendocrine (NE) bladder carcinoma is a rare and aggressive variant. Molecular subtyping studies have found that 5% to 15% of muscle-invasive bladder cancer (MIBC) have transcriptomic patterns consistent with NE bladder cancer in the absence of NE histology. The clinical implications of this NE-like subtype have not been explored in depth.Experimental Design:Transcriptome-wide expression profiles were generated for MIBC collected from 7 institutions and clinical-use of Decipher Bladder. Using unsupervised clustering, we generated a clustering solution on a prospective training cohort (PTC; n = 175), developed single-sample classifiers to predict NE tumors, and evaluated the resultant models on a testing radical cystectomy (RC) cohort (n = 225). A random forest model was finalized and applied to 5 validation cohorts (n = 1302). Uni- and multivariable survival analyses were used to characterize clinical outcomes.Results:In the training cohort (PTC), hierarchical clustering using an 84-gene panel showed a cluster of 8 patients (4.6%) with highly heterogeneous expression of NE markers in the absence of basal or luminal marker expression. NE-like tumors were identified in 1% to 6.6% of cases in validation cohorts. Patients with NE-like tumors had significantly worse 1-year progression-free survival (65% NE-like vs. 82% overall; P = 0.046) and, after adjusting for clinical and pathologic factors, had a 6.4-fold increased risk of all-cause mortality (P = 0.001). IHC confirmed the neuronal character of these tumors.Conclusions:A single-patient classifier was developed that identifies patients with histologic urothelial cancer harboring a NE transcriptomic profile. These tumors represent a high-risk subgroup of MIBC, which may require different treatment.

Well-done cooked red meat consumption is linked to aggressive prostate cancer (PC) risk. Identifying mutation-inducing DNA adducts in the prostate genome can advance our understanding of chemicals in meat that may contribute to PC. 2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), a heterocyclic aromatic amine (HAA) formed in cooked meat, is a potential human prostate carcinogen. PhIP was measured in the hair of PC patients undergoing prostatectomy, bladder cancer patients under treatment for cystoprostatectomy, and patients treated for benign prostatic hyperplasia (BPH). PhIP hair levels were above the quantification limit in 123 of 205 subjects. When dichotomizing prostate pathology biomarkers, the geometric mean PhIP hair levels were higher in patients with intermediate and elevated-risk prostate-specific antigen values than lower-risk values < 4 ng/mL (p = 0.03). PhIP hair levels were also higher in patients with intermediate and high-risk Gleason scores ≥7 compared to lower-risk Gleason score 6 and BPH patients (p = 0.02). PC patients undergoing prostatectomy had higher PhIP hair levels than cystoprostatectomy or BPH patients (p = 0.02). PhIP-DNA adducts were detected in 9.4% of the patients assayed; however, DNA adducts of other carcinogenic HAAs, and benzo[a]pyrene formed in cooked meat, were not detected. Prostate specimens were also screened for 10 oxidative stress-associated lipid peroxidation (LPO) DNA adducts. Acrolein 1,N(2)–propano-2′-deoxyguanosine adducts were detected in 54.5% of the patients; other LPO adducts were infrequently detected. Acrolein adducts were not associated with prostate pathology biomarkers, although DNA adductomic profiles differed between PC patients with low and high-grade Gleason scores. Many DNA adducts are of unknown origin; however, dG adducts of formaldehyde and a series of purported 4-hydroxy-2-alkenals were detected at higher abundance in a subset of patients with elevated Gleason scores. The PhIP hair biomarker and DNA adductomics data support the paradigm of well-done cooked meat and oxidative stress in aggressive PC risk.

Pseudolymphomatous infiltrates associated with angiosarcoma are a rarely reported phenomenon. Recognition of this reactive process is critical to making an accurate diagnosis, both in diagnosing the angiosarcoma and in avoiding an incorrect diagnosis of lymphoma. Here, we present a novel histopathologic pattern, angiosarcoma with a prominently intravascular atypical lymphoid component, mimicking intravascular T-cell lymphoma. Interestingly, serial biopsies in this case revealed a progressive increase in lymphocyte density and intravascular component over time. Despite prior reports of improved progression-free survival and overall survival of patients with pseudolymphomatous angiosarcoma, this patient showed rapid disease progression.

Objectives We aimed to better understand the histologic changes in vaginectomy specimens in transgender and gender-diverse (TGD) individuals after prolonged androgen administration. Methods After obtaining institutional review board approval, we reviewed clinical records for all TGD individuals who underwent vaginal tissue resection at our institution between January 2002 and July 2020. Results Ten transgender males who underwent vaginectomy for gender affirmation were identified. All patients had been assigned female gender at birth, and the median age at surgery was 41 years (range, 22-74 years). All 10 patients had received androgen for 2 to 10 years preoperatively. The corresponding pathology specimens were examined grossly and microscopically, including with immunohistochemical stains for NKX3.1, prostate-specific antigen (PSA), p501s, and androgen receptor (AR). No gross lesions were identified. Microscopically, prostate-like glands (8/10), urothelial metaplasia (4/10), and vaginal atrophy (8/10) were identified. Seven cases with prostate-like glands showed positive staining with PSA, NKX3.1, p501s, and AR in both squamous and glandular components. Conclusions Recognition of these androgen-related changes enables pathologist to avoid the overdiagnosis of dysplasia. Long-term follow-up is needed to thoroughly understand any potential future implications of these androgen-related changes.

A novel software has been created to comprehensively characterize covalent modifications of DNA through mass spectral analysis of enzymatically hydrolyzed DNA using the neutral loss of 2'-deoxyribose, a nearly universal MS2 fragmentation process of protonated 2'-deoxyribonucleosides. These covalent modifications termed DNA adducts form through xenobiotic exposures or by reaction with endogenous electrophiles and can induce mutations during cell division and initiate carcinogenesis. DNA adducts are typically present at trace levels in the human genome, requiring a very sensitive and comprehensive data acquisition and analysis method. Our software, wSIM-City, was created to process mass spectral data acquired by a wide selected ion monitoring (wSIM) with gas-phase fractionation and coupled to wide MS2 fragmentation. This untargeted approach can detect DNA adducts at trace levels as low as 1.5 adducts per 109 nucleotides. This level of sensitivity is sufficient for comprehensive analysis and characterization of DNA modifications in human specimens.

Sporadic angiosarcomas are aggressive vascular sarcomas whose rarity and genomic complexity present significant obstacles in deciphering the pathogenic significance of individual genetic alterations. Numerous fusion genes have been identified across multiple types of cancers, but their existence and significance remain unclear in sporadic angiosarcomas. In this study, we leveraged RNA-sequencing data from 13 human angiosarcomas and 76 spontaneous canine hemangiosarcomas to identify fusion genes associated with spontaneous vascular malignancies. Ten novel protein-coding fusion genes, including TEX2-PECAM1 and ATP8A2-FLT1, were identified in seven of the 13 human tumors, with two tumors showing mutations of TP53. HRAS and NRAS mutations were found in angiosarcomas without fusions or TP53 mutations. We found 15 novel protein-coding fusion genes including MYO16-PTK2, GABRA3-FLT1, and AKT3-XPNPEP1 in 11 of the 76 canine hemangiosarcomas; these fusion genes were seen exclusively in tumors of the angiogenic molecular subtype that contained recurrent mutations in TP53, PIK3CA, PIK3R1, and NRAS. In particular, fusion genes and mutations of TP53 cooccurred in tumors with higher frequency than expected by random chance, and they enriched gene signatures predicting activation of angiogenic pathways. Comparative transcriptomic analysis of human angiosarcomas and canine hemangiosarcomas identified shared molecular signatures associated with activation of PI3K/AKT/mTOR pathways. Our data suggest that genome instability induced by TP53 mutations might create a predisposition for fusion events that may contribute to tumor progression by promoting selection and/or enhancing fitness through activation of convergent angiogenic pathways in this vascular malignancy. Implications: This study shows that, while drive events of malignant vasoformative tumors of humans and dogs include diverse mutations and stochastic rearrangements that create novel fusion genes, convergent transcriptional programs govern the highly conserved morphologic organization and biological behavior of these tumors in both species.

S100A4 oncoprotein plays a critical role during prostate cancer progression and induces immunosuppression in host tissues. We hypothesized that S100A4-regulated oncogenic activity in immunosuppressed prostate tumors promotes growth of neoplastic cells, which are likely to become aggressive. In the current study, we investigated whether biopsy-S100A4 gene alteration independently predicts the outcome of disease in patients and circulatory-S100A4 is druggable target for treating immunosuppressive prostate cancer. Aided by DECIPHER-genomic test, we show biopsy-S100A4 overexpression as predictive of (i) poor ADT response and (ii) high risk of mortality in 228 radical prostatectomy-treated patients. Furthermore, analysis of tumor genome data of more than 1,000 patients with prostate cancer (PRAD/SU2C/FHCRC studies) validated the association of S100A4-alteration to poor survival and metastasis. We show that increased serum-S100A4 levels are associated to the prostate cancer progression in patients. The prerequisite for metastasis is the escape of tumor cells via vascular system. We show that extracellular-S100A4 protein as a growth factor induces vascular transmigration of prostate cancer cells and bone demineralization thus forms an ideal target for therapies for treating prostate cancer. By employing surface plasmon resonance and isothermal titration calorimetry, we show that mab6B12 antibody interacts with and neutralizes S100A4 protein. When tested for therapeutic efficacy, the mab6B12 therapy reduced the (i) osteoblastic demineralization of bone-derived MSCs, (ii) S100A4-target (NFκB/MMP9/VEGF) levels in prostate cancer cells, and (iii) tumor growth in a TRAMPC2 syngeneic mouse model. The immuno-profile analysis showed that mAb6B12-therapy (i) shifted Th1/Th2 balance (increased Stat4+/T-bet+ and decreased GATA2+/CD68+/CD45+/CD206+ cells); (ii) modulated cytokine levels in CD4+ T cells; and (iii) decreased levels of IL5/6/12/13, sTNFR1, and serum-RANTES. We suggest that S100A4-antibody therapy has clinical applicability in treating immunosuppressive prostate cancer in patients.

The accurate identification of different components in testicular germ cell tumors (GCT) is essential for tailoring treatment and informing the clinical prognosis. PRAME (preferentially expressed antigen in melanoma), a member in the family of cancer testis antigens, plays critical roles in regulating pluripotency and suppressing somatic/germ cell differentiation in seminomas (SEM). To investigate the potential diagnostic value of PRAME in testicular GCT, here we comparatively examined the expression patterns of PRAME and SOX17 by immunohistochemistry in both pure and mixed GCT. Tissue microarrays constructed from 66 pure or mixed GCT were examined, including 25 seminomas (13 pure and 12 mixed), 35 embryonal carcinomas (EC; 7 pure and 28 mixed), 23 teratomas (TER; 10 pure and 13 mixed), 15 yolk sac tumors (YST; 1 pure and 14 mixed), and 5 choriocarcinomas (CC; 1 pure and 4 mixed), with 11 germ cell neoplasia in situ (GCNIS) and 6 normal testicular tissue as controls. The expression levels of PRAME or SOX17 were evaluated by a scoring system counting for intensity and extent of staining. PRAME nuclear expression was present in 92% (23/25) of SEM, including all 13 pure SEM, and 10 out of 12 seminomatous component of mixed GCT. In contrast, all EC and TER were completely negative for PRAME, and focal expression was demonstrated in 33.3% of YST and 20% of CC. As for SOX17, 96% of SEM and 73% of YST stained positively, whereas EC and CC were negative. Focal nuclear positivity was identified in the epithelial cell component of 17.4% (4/23) of TER. We found the sensitivity of PRAME to detect SEM to be comparable to SOX17, although SOX17 staining is more diffuse and stronger in the majority of cases. The specificity of PRAME for SEM appeared to be superior to that of SOX17 (92% versus 81%). In conclusion, PRAME is preferentially expressed in SEM or within the seminomatous component of mixed GCT with only focal variable expression in YST and CC, but shows no expression in EC and TER. These findings suggest that PRAME can be explored as a diagnostic marker for SEM.

Background There is a need to develop novel therapies which could be beneficial to patients with prostate cancer (CaP) including those who are predisposed to poor outcome, such as African-Americans. This study investigates the role of ROBO1-pathway in predicting outcome and race-based disparity in patients with CaP. Methods and results Aided by RNA sequencing-based DECIPHER-testing and immunohistochemical (IHC) analysis of tumors we show that ROBO1 is lost during the progressive stages of CaP, a prevalent feature in African-Americans. We show that the loss of ROBO1 predicts high-risk of recurrence, metastasis and poor outcome of androgen-deprivation therapy in radical prostatectomy-treated patients. These data identified an aggressive ROBO1deficient /DOCK1+ve sub-class of CaP. Combined genetic and IHC data showed that ROBO1 loss is accompanied by DOCK1/Rac1 elevation in grade-III/IV primary-tumors and Mets. We observed that the hypermethylation of ROBO1-promoter contributes to loss of expression that is highly prevalent in African-Americans. Because of limitations in restoring ROBO1 function, we asked if targeting the DOCK1 could be an ideal strategy to inhibit progression or treat ROBO1deficient metastatic-CaP. We tested the pharmacological efficacy of CPYPP, a selective inhibitor of DOCK1 under in vitro and in vivo conditions. Using ROBO1-ve and ROBO1+ve CaP models, we determined the median effective concentration of CPYPP for growth. DOCK1-inhibitor treatment significantly decreased the (a) Rac1-GTP/β-catenin activity, (b) transmigration of ROBO1deficient cells across endothelial lining, and (c) metastatic spread of ROBO1deficient cells through the vasculature of transgenicfl Zebrafish model. Conclusion We suggest that ROBO1 status forms as predictive biomarker of outcome in high-risk populations such as African-Americans and DOCK1-targeting therapy has a clinical potential for treating metastatic-CaP.

Gene rearrangement is reported to be associated to the aggressive phenotype and poor prognosis in prostate cancer. We identified a gene fusion between a transcription repressor (BMI1) and transcriptional factor (COMMD3) in human prostate cancer. We show that COMMD3:BMI1 fusion expression is significantly increased in prostate cancer disease in an order: normal tissue < primary < metastatic tumors (Mets). Although elevated TMPRSS-ERG/ETV fusion is reported in prostate cancer, we identified a subtype of Mets exhibiting low TMPRSS:ETV and high COMMD3:BMI1. We delineated the mechanism and function of COMMD3 and COMMD3:BMI1 in prostate cancer. We show that COMMD3 level is elevated in prostate cancer cell models, PDX models (adenocarcinoma, NECaP), and Mets. The analysis of TCGA/NIH/GEO clinical data showed a positive correlation between increased COMMD3 expression to the disease recurrence and poor survival in prostate cancer. We show that COMMD3 drives proliferation of normal cells and promotes migration/invasiveness of neoplastic cells. We show that COMMD3:BMI1 and COMMD3 regulate C-MYC transcription and C-MYC downstream pathway. The ChIP analysis showed that COMMD3 protein is recruited at the promoter of C-MYC gene. On the basis of these data, we investigated the relevance of COMMD3:BMI1 and COMMD3 as therapeutic targets using in vitro and xenograft mouse models. We show that siRNA-mediated targeting of COMMD3:BMI1 and COMMD3 significantly decreases (i) C-MYC expression in BRD/BET inhibitor–resistant cells, (ii) proliferation/invasion in vitro, and (iii) growth of prostate cancer cell tumors in mice. The IHC analysis of tumors confirmed the targeting of COMMD3-regulated molecular pathway under in vivo conditions. We conclude that COMMD3:BMI1 and COMMD3 are potential progression biomarkers and therapeutic targets of metastatic prostate cancer.

Endocrine therapies for prostate cancer inhibit the androgen receptor (AR) transcription factor. In most cases, AR activity resumes during therapy and drives progression to castration-resistant prostate cancer (CRPC). However, therapy can also promote lineage plasticity and select for AR-independent phenotypes that are uniformly lethal. Here, we demonstrate the stem cell transcription factor Krüppel-like factor 5 (KLF5) is low or absent in prostate cancers prior to endocrine therapy, but induced in a subset of CRPC, including CRPC displaying lineage plasticity. KLF5 and AR physically interact on chromatin and drive opposing transcriptional programs, with KLF5 promoting cellular migration, anchorage-independent growth, and basal epithelial cell phenotypes. We identify ERBB2 as a point of transcriptional convergence displaying activation by KLF5 and repression by AR. ERBB2 inhibitors preferentially block KLF5-driven oncogenic phenotypes. These findings implicate KLF5 as an oncogene that can be upregulated in CRPC to oppose AR activities and promote lineage plasticity., While many treatments for prostate cancer suppress the androgen receptor it becomes reactivated during disease progression. Here, the authors show that a KLF5 transcriptional programme is also activated during treatment and promotes migration and the appearance of a basal cell phenotype.

Introduction Neurofibromatosis-1 (NF-1) is a genetic condition known to be associated with pheochromocytoma. We report a case of giant cystic pheochromocytoma in a patient with NF-1 and hypertensionincidentally found to have gastrointestinal stromal tumor (GIST) at operation. Clinical case: A 60-years old woman presented with a 5-week history of left sided abdominal lump. Her history is significant for NF-1, flank surgery age 5 and hypertension managed by amlodipine. Examination revealed a large nontender left upper quadrant abdominal mass crossing the midline. Computed tomography showed a20. 0×13.1×21.1 cm septate cystic and/or necrotic lesion arising from the left retroperitoneum versus adrenal gland, displacing the kidney anteromedially. Plasma free metanephrine was 3.38 nM (0-0.49) normetanephrine 3.47 nM (0-0.89),urine metanephrine 2605 mcg/day (36-229), urine normetanephrine 1215 mcg/day (95-650). There were no prior images or screens for pheochromocytoma. After preoperative blockade with phenoxybenzamine, open left adrenalectomy was performed. Intraoperative blood pressure fluctuated significantly. Exophytic firm nodules were incidentally noted and removed from the jejunum. Pathology revealed a22.5×21. 0×11.9 cm pheochromocytoma weighing 3058.9 g and five 0.2-0.7 cm gastrointestinal stromal tumors, spindle type. One-month postoperative metanephrine was 0.16 with normetanephrine 0.53 nM. The blood pressure remained normal without antihypertensives. Discussion Giant cystic pheochromocytomas are rare and may be silent. In populations with NF-1 the prevalence of pheochromocytoma is 0.1-7.7% and GIST 1.2% 1–3 . Their co-existence is seen less frequently. Recommendations for pheochromocytoma screening in NF-1 vary, from only hypertensive or symptomatic patients to routinely every 3 years 2–5 . There is no screening test for GIST 1 . Our patient presented at age 60 with the rare combination of clinically giant unilateral cystic pheochromocytoma, with incidental GIST in the setting of NF-1 and hypertension . Conclusion Routine screening for pheochromocytoma in patients with NF-1, especially those with hypertension may lead to earlier diagnosis and treatment. References: Landry JP, Schertz KL, Chiang YJ, et al. Comparison of Cancer Prevalence in Patients with Neurofibromatosis Type 1 at an Academic Cancer Center vs in the General Population from 1985 to 2020. JAMA Netw Open. 2021;4(3): 1-15. Gruber LM, Erickson D, Babovic-Vuksanovic D, Thompson GB, Young WF, Bancos I. Pheochromocytoma and paraganglioma in patients with neurofibromatosis type 1. Clin Endocrinol (Oxf). 2017;86(1): 141-149. Képénékian L, Mognetti T, Lifante J-C, et al. Interest of systematic screening of pheochromocytoma in patients with neurofibromatosis type 1. Eur J Endocrinol. 2016;175(4): 335-344. Chen H, Sippel RS, O'Dorisio MS, et al. The North American Neuroendocrine Tumor Society consensus guideline for the diagnosis and management of neuroendocrine tumors: pheochromocytoma, paraganglioma, and medullary thyroid cancer. Pancreas. 2010;39(6): 775-783. Lenders JWM, Duh Q-Y, Eisenhofer G, et al. Pheochromocytoma and paraganglioma: an endocrine society clinical practice guideline. J Clin Endocrinol Metab. 2014;99(6): 1915-1942. Presentation: No date and time listed

Background Percutaneous needle biopsy of renal masses has been increasingly utilized to aid the diagnosis and guide management. It is generally considered as a safe procedure. However, tumor seeding along the needle tract, one of the complications, theoretically poses potential risk of tumor spread by seeded malignant cells. Prior studies on the frequency of needle tract seeding in renal tumor biopsies are limited and clinical significance of biopsy-associated tumor seeding remains largely controversial. Methods Here we investigated the frequencies of biopsy needle tract tumor seeding at our institution by reviewing the histology of renal cell carcinoma nephrectomy specimens with a prior biopsy within the last seventeen years. Biopsy site changes were recognized as a combination of foreign body reaction, hemosiderin deposition, fibrosis and fat necrosis. The histologic evidence of needle tract tumor seeding was identified as clusters of tumor cells embedded in perinephric tissue spatially associated with the biopsy site. In addition, association between parameters of biopsy techniques and tumor seeding were investigated. Results We observed needle tract tumor seeding to perinephric tissue in six out of ninety-eight (6 %) renal cell carcinoma cases including clear cell renal cell carcinoma, papillary renal cell carcinoma, chromophobe, and clear cell papillary renal cell carcinoma. The needle tract tumor seeding was exclusively observed in papillary renal cell carcinomas (6/28, 21 %) that were unifocal, small-sized (≤ 4 cm), confined to the kidney and had type 1 features. No recurrence or metastasis was observed in the papillary renal cell carcinoma cases with tumor seeding or the stage-matched cases without tumor seeding. Conclusions Our study demonstrated a higher than reported frequency of needle tract tumor seeding. Effective communication between pathologists and clinicians as well as documentation of tumor seeding is recommended. Further studies with a larger patient cohort and longer follow up to evaluate the impact of needle tract tumor seeding on long term prognosis are needed. This may also help reach a consensus on appropriate pathologic staging of renal cell carcinoma when the only site of perinephric fat invasion is within a biopsy needle tract.

Objectives Current protocols for processing multiple prostate biopsy cores per case are uneconomical and cumbersome. Tissue fragmentation and loss compromise cancer diagnosis. We sought to study an alternate method to improve processing and diagnosis of prostate cancer. Methods Two sets of sextant biopsy specimens from near-identical locations were obtained ex vivo from 48 prostate specimens. One set was processed in the standard fashion while the other was processed using the BxChip, a proprietary biomimetic matrix that accommodates six cores on a single chip. Parameters including grossing, embedding, sectioning and reading time, length of tissue, and degree of fragmentation were compared. Results A significant reduction (more than threefold) in preanalytical and analytical time was observed using the multiplex method. Nonlinear fragmentation was absent, in contrast to standard processing. Conclusions The BxChip reduced tissue fragmentation and increased efficiency of prostate biopsy diagnosis. It also resulted in overall cost savings and significantly increased tissue length.

Purpose: Neuroendocrine (NE) bladder carcinoma is a rare and aggressive variant. Molecular subtyping studies have found that 5% to 15% of muscle-invasive bladder cancer (MIBC) have transcriptomic patterns consistent with NE bladder cancer in the absence of NE histology. The clinical implications of this NE-like subtype have not been explored in depth. Experimental Design: Transcriptome-wide expression profiles were generated for MIBC collected from 7 institutions and clinical-use of Decipher Bladder. Using unsupervised clustering, we generated a clustering solution on a prospective training cohort (PTC; n = 175), developed single-sample classifiers to predict NE tumors, and evaluated the resultant models on a testing radical cystectomy (RC) cohort (n = 225). A random forest model was finalized and applied to 5 validation cohorts (n = 1302). Uni- and multivariable survival analyses were used to characterize clinical outcomes. Results: In the training cohort (PTC), hierarchical clustering using an 84-gene panel showed a cluster of 8 patients (4.6%) with highly heterogeneous expression of NE markers in the absence of basal or luminal marker expression. NE-like tumors were identified in 1% to 6.6% of cases in validation cohorts. Patients with NE-like tumors had significantly worse 1-year progression-free survival (65% NE-like vs. 82% overall; P = 0.046) and, after adjusting for clinical and pathologic factors, had a 6.4-fold increased risk of all-cause mortality (P = 0.001). IHC confirmed the neuronal character of these tumors. Conclusions: A single-patient classifier was developed that identifies patients with histologic urothelial cancer harboring a NE transcriptomic profile. These tumors represent a high-risk subgroup of MIBC, which may require different treatment.

A novel software has been created to comprehensively characterize covalent modifications of DNA through mass spectral analysis of enzymatically hydrolyzed DNA using the neutral loss of 2′-deoxyribose, a nearly universal MS(2) fragmentation process of protonated 2′-deoxyribonucleosides. These covalent modifications termed DNA adducts form through xenobiotic exposures or by reaction with endogenous electrophiles and can induce mutations during cell division and initiate carcinogenesis. DNA adducts are typically present at trace levels in the human genome, requiring a very sensitive and comprehensive data acquisition and analysis method. Our software, wSIM-City, was created to process mass spectral data acquired by a wide selected ion monitoring (wSIM) with gas-phase fractionation and coupled to wide MS(2) fragmentation. This untargeted approach can detect DNA adducts at trace levels as low as 1.5 adducts per 10(9) nucleotides. This level of sensitivity is sufficient for comprehensive analysis and characterization of DNA modifications in human specimens.

Context.— Transgender women experience health disparities in all areas of medicine. Within surgical pathology, knowledge gaps relating to the concepts of transgender care exist. Medical transition for transgender women and transfeminine persons may involve hormone therapy and/or surgery to feminize the body. Understanding the common histologic changes in specimens from feminizing surgeries, as well as other specimens from patients on feminizing hormone therapy, will aid surgical pathologists in providing better care to this unique patient population. Objective.— To summarize histologic findings in surgical pathology specimens from transgender women taking feminizing hormones. Data Sources.— A systematic review of the OVID Medline and PubMed databases was performed to identify all studies describing histologic findings in surgical pathology specimens from transgender women from 1946 to 2019. Conclusions.— Much of the literature to date describing histologic findings in transgender women comes from the examination of genitourinary specimens removed during feminizing surgeries. Common benign changes associated with feminizing hormone therapy include the development of acini and lobules in the breast, testicular tubular changes, and squamous metaplasia of the prostate and urethra. Neoplastic cases include breast adenocarcinoma and fibroepithelial lesions, testicular germ cell tumors, prostatic adenocarcinoma, anal squamous cell carcinoma, pituitary adenomas, and meningiomas. Additional studies assessing the findings in other organ systems as well as population-based studies assessing rates of neoplasia are needed. However, future research relies on engagement within the surgical pathology community as well as collaboration with clinicians and patients to achieve optimal results.

Background Chordoma is a rare bone tumor that is typically resistant to chemotherapy and is associated with genetic abnormalities of the T-box transcription factor T (TBXT) gene, which encodes the transcription factor brachyury. Brachyury is felt to be a major contributor to the development of chordomas. Case presentation We describe a 67-year-old woman who developed an undifferentiated pleomorphic sarcoma in her thigh. Despite treatment with standard chemotherapy regimens, she had a rapidly progressive course of disease with pulmonary metastases and passed away 8 months from diagnosis with pulmonary complications. Her medical history was remarkable in that she had a spheno-occipital chordoma at age 39 and later developed multiple other tumors throughout her life including Hodgkin lymphoma and squamous cell carcinoma and basal cell carcinoma of the skin. She had a family history of chordoma and her family underwent extensive genetic study in the past and were found to have a duplication of the TBXT gene. Conclusions Brachyury has been found to associate with tumor progression, treatment resistance, and metastasis in various epithelial cancers, and it might play roles in tumorigenesis and aggressiveness in this patient with multiple rare tumors and germ line duplication of the TBXT gene. Targeting this molecule may be useful for some malignancies.

Pregnancy presents unique obstacles to diagnosis and management of urologic disease. We present a case of a primigravid female with clot retention requiring evacuation in the operating room due to the avulsion of a bladder mass which prolapsed during labor. Tumor pathology demonstrated a low-grade spindle cell lesion positive for progesterone receptor (PR) and high mobility group A2 (HMGA2), suggestive of deep angiomyxoma versus a benign fibroepithelial polyp or inflammatory myofibroblastic tumor.