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Patient demographic are described in supplementary Table 1

Supplementary Figures

BACKGROUND: Hormonal therapy targeting the androgen receptor inhibits prostate cancer (PCa), but the tumor eventually recurs as castration-resistant prostate cancer (CRPC). OBJECTIVE: To understand the mechanisms by which subclones within early PCa develop into CRPC. DESIGN, SETTING, AND PARTICIPANTS: We isolated epithelial cells from fresh human PCa cases, including primary adenocarcinoma, locally recurrent CRPC, and metastatic CRPC, and utilized single-cell RNA sequencing to identify subpopulations destined to become either CRPC-adeno or small cell neuroendocrine carcinoma (SCNC). OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: We revealed dynamic transcriptional reprogramming that promotes disease progression among 23 226 epithelial cells using single-cell RNA sequencing, and validated subset-specific progression using immunohistochemistry and large cohorts of publically available genomic data. RESULTS AND LIMITATIONS: We identified a small fraction of highly plastic CRPC-like cells in hormone-naïve early PCa and demonstrated its correlation with biochemical recurrence and distant metastasis, independent of clinical characteristics. We show that progression toward castration resistance was initiated from subtype-specific lineage plasticity and clonal expansion of pre-existing neuroendocrine and CRPC-like cells in early PCa. CONCLUSIONS: CRPC-like cells are present early in the development of PCa and are not exclusively the result of acquired evolutionary selection during androgen deprivation therapy. The lethal CRPC and SCNC phenotypes should be targeted earlier in the disease course of patients with PCa. PATIENT SUMMARY: Here, we report the presence of pre-existing castration-resistant prostate cancer (CRPC)-like cells in primary prostate cancer, which represents a novel castration-resistant mechanism different from the adaptation mechanism after androgen deprivation therapy (ADT). Patients whose tumors harbor increased pre-existing neuroendocrine and CRPC-like cells may become rapidly resistant to ADT and may require aggressive early intervention.

Demographics

Signed-Rank Tests of V3-V2 Difference

Purpose: To determine the immunologic effects of neoadjuvant chemotherapy plus ipilimumab in early-stage non–small cell lung cancer (NSCLC) patients.Experimental Design: This is a single-arm chemotherapy plus phased ipilimumab phase II study of 24 treatment-naïve patients with stage IB–IIIA NSCLC. Patients received neoadjuvant therapy consisting of 3 cycles of paclitaxel with either cisplatin or carboplatin and ipilimumab included in the last 2 cycles.Results: Chemotherapy alone had little effect on immune parameters in PBMCs. Profound CD28-dependent activation of both CD4 and CD8 cells was observed following ipilimumab. Significant increases in the frequencies of CD4+ cells expressing activation markers ICOS, HLA-DR, CTLA-4, and PD-1 were apparent. Likewise, increased frequencies of CD8+ cells expressing the same activation markers, with the exception of PD-1, were observed. We also examined 7 resected tumors and found higher frequencies of activated tumor-infiltrating lymphocytes than those observed in PBMCs. Surprisingly, we found 4 cases of preexisting tumor-associated antigens (TAA) responses against survivin, PRAME, or MAGE-A3 present in PBMC at baseline, but neither increased frequencies nor the appearance of newly detectable responses following ipilimumab therapy. Ipilimumab had little effect on the frequencies of circulating regulatory T cells and MDSCs.Conclusions: This study did not meet the primary endpoint of detecting an increase in blood-based TAA T-cell responses after ipilimumab. Collectively, these results highlight the immune activating properties of ipilimumab in early-stage NSCLC. The immune profiling data for ipilimumab alone can contribute to the interpretation of immunologic data from combined immune checkpoint blockade immunotherapies. Clin Cancer Res; 23(24); 7474–82. ©2017 AACR.

List of antibodies used for flow cytometry analysis

Spaghetti plots of MDSC and Tregs by visits; stratified by overall response.

Spaghetti plots of CD4 and CD8 T cells by visits; stratified by overall response.

Summary Gastrointestinal symptoms are commonly reported in patients with 22q11.2 deletion syndrome or DiGeorge syndrome (DGS) in addition to the dominant cardiac manifestations and immunodeficiency. But literature providing specific morphologic details of the gastrointestinal tract pathology is very limited. Here, we provide the first comprehensive morphologic description of the luminal gastrointestinal tract changes in patients with DGS. Cytogenetically confirmed DGS patients were identified, clinical and laboratory data were reviewed to determine the severity of immunodeficiency, and patients were stratified into mildly immunocompromised, that is, partial DiGeorge anomaly or severely immunosuppressed, that is, complete DiGeorge anomaly groups. Gastrointestinal tract biopsies from these patients were retrospectively reviewed and compared with those from controls without the history of DGS. Patients with immunosuppressed DGS showed a near complete absence of plasma cells in the stomach, duodenum, and colon lamina propria by hematoxylin and eosin evaluation. Immunohistochemistry for CD138 used to highlight plasma cells confirmed this finding. The notable absence of plasma cells adds to the existing knowledge of the pathophysiology underlying DGS and expands the differential diagnostic considerations for this finding, which has been previously described in common variable immunodeficiency. It also provides a useful morphologic marker observable by the readily accessible light microscopy. Second, patients with DGS showed a mild increase in epithelial cell apoptosis in their colon. This finding is significant because of its overlap with morphologic features of gastrointestinal graft versus host disease as thymus transplantation is being used as a treatment option for patients with complete DGS.

PURPOSE Comprehensive genomic profiling to inform targeted therapy selection is a central part of oncology care. However, the volume and complexity of alterations uncovered through genomic profiling make it difficult for oncologists to choose the most appropriate therapy for their patients. Here, we present a solution to this problem, The Molecular Registry of Tumors (MRT) and our Molecular Tumor Board (MTB). PATIENTS AND METHODS MRT is an internally developed system that aggregates and normalizes genomic profiling results from multiple sources. MRT serves as the foundation for our MTB, a team that reviews genomic results for all Duke University Health System cancer patients, provides notifications for targeted therapies, matches patients to biomarker-driven trials, and monitors the molecular landscape of tumors at our institution. RESULTS Among 215 patients reviewed by our MTB over a 6-month period, we identified 176 alterations associated with therapeutic sensitivity, 15 resistance alterations, and 51 alterations with potential germline implications. Of reviewed patients, 17% were subsequently treated with a targeted therapy. For 12 molecular therapies approved during the course of this work, we identified between two and 71 patients who could qualify for treatment based on retrospective MRT data. An analysis of 14 biomarker-driven clinical trials found that MRT successfully identified 42% of patients who ultimately enrolled. Finally, an analysis of 4,130 comprehensive genomic profiles from 3,771 patients revealed that the frequency of clinically significant therapeutic alterations varied from approximately 20% to 70% depending on the tumor type and sequencing test used. CONCLUSION With robust informatics tools, such as MRT, and the right MTB structure, a precision cancer medicine program can be developed, which provides great benefit to providers and patients with cancer.

Background Prostate cancer is a clinically and molecularly heterogeneous disease, with highest incidence and mortality among men of African ancestry. To date, prostate cancer patient-derived xenograft (PCPDX) models to study this disease have been difficult to establish because of limited specimen availability and poor uptake rates in immunodeficient mice. Ancestrally diverse PCPDXs are even more rare, and only six PCPDXs from self-identified African American patients from one institution were recently made available. Methods In the present study, we established a PCPDX from prostate cancer tissue from a patient of estimated 90% West African ancestry with metastatic castration resistant disease, and characterized this model's pathology, karyotype, hotspot mutations, copy number, gene fusions, gene expression, growth rate in normal and castrated mice, therapeutic response, and experimental metastasis. Results This PCPDX has a mutation in TP53 and loss of PTEN and RB1. We have documented a 100% take rate in mice after thawing the PCPDX tumor from frozen stock. The PCPDX is castrate- and docetaxel-resistant and cisplatin-sensitive, and has gene expression patterns associated with such drug responses. After tail vein injection, the PCPDX tumor cells can colonize the lungs of mice. Conclusion This PCPDX, along with others that are established and characterized, will be useful pre-clinically for studying the heterogeneity of prostate cancer biology and testing new therapeutics in models expected to be reflective of the clinical setting.

Acquired resistance remains a major challenge for therapies targeting oncogene activated pathways. KRAS is the most frequently mutated oncogene in human cancers, yet strategies targeting its downstream signaling kinases have failed to produce durable treatment responses. Here, we developed multiple models of acquired resistance to dual-mechanism ERK/MAPK inhibitors across KRAS-mutant pancreatic, colorectal, and lung cancers, and then probed the long-term events enabling survival against this novel class of drugs. These studies revealed that resistance emerges secondary to large-scale transcriptional adaptations that are diverse and tumor-specific. Transcriptional reprogramming extends beyond the well-established early response, and instead represents a dynamic, evolved population-level process that is refined to attain a stably resistant phenotype. Mechanistic and translational studies reveal that resistance to dual-mechanism ERK/MAPK inhibition is broadly susceptible to manipulation of the epigenetic machinery, and that Mediator kinase, in particular, can be co-targeted at a bottleneck point to prevent diverse, tumor-specific resistance programs.

Background: Limited information is available about treatments and clinical outcomes among men with metastatic castration-resistant prostate cancer (mCRPC) whose tumors harbor DNA damage repair (DDR) mutations. Methods: This retrospective chart review study included men diagnosed with mCRPC with ≥1 DDR mutation detected by tumor comprehensive genomic profiling who began a line of therapy on or after July 1, 2014, at oncology centers in the American Association for Cancer Research (AACR) Project Genomics Evidence Neoplasia Information Exchange (GENIE) consortium. The index date was the initiation of a new therapy in this period, in patients who developed castration-resistant disease and progressed while receiving androgen deprivation therapy. Data on all available lines of therapy post-index were collected. Outcomes reported include time to treatment discontinuation for any reason (TTTD) and overall survival (OS), which were assessed using Kaplan-Meier analyses. Results for tumor response-related endpoints are forthcoming. Results: A total of 138 patients with mCRPC met the inclusion criteria. The mean age at index was 68 years, and 76% of patients were White. The median follow-up time was 27.1, 15.9, 11.3, and 9.4 months for 1L, 2L, 3L, and 4L+ therapy, respectively. Common sites of metastases included bone (74%), lymph nodes (69%), and lungs (10%). Common tumor DDR mutations included CDK12 (48%), BRCA2 (23%), ATM (21%), and BRCA1 (8%). Assessment of germline mutations was conducted in 114 (83%) patients; the most common was BRCA2 (8%). First-line (1L) treatment information was available for 119 (86%) patients, 2L for 101 (73%), and 3L for 75 (54%) patients. Novel hormonal therapy (NHT; 55%) and taxane chemotherapy (20%) were most common in 1L. Enzalutamide (35%) and abiraterone acetate (14%) were the most frequently used NHTs. Compared to 1L, the proportion of patients receiving NHTs was lower in 2L (33%) and 3L (17%), while the proportion of patients receiving taxanes was higher (36% in 2L and 31% in 3L). In 3L, poly ADP-ribose polymerase (PARP) inhibitors were used in 21% of patients. Median TTTD was 5.6 months (95% CI: 4.2, 7.2) in 1L, 3.9 months (95% CI: 3.0, 4.6) in 2L, and 4.0 months (95% CI: 3.3, 4.7) in 3L. Median OS was 36.3 months (95% CI: 30.7, 47.8) from the start of 1L, 21.1 months (95% CI: 15.6, 29.3) from the start of 2L, and 14.1 months (95% CI: 9.5, 20.5) from the start of 3L. Conclusions: These findings provide real-world insights about treatment patterns and clinical outcomes among patients with mCRPC harboring DDR mutations. Such insight may provide a helpful benchmark for research aimed at improving clinical outcomes, particularly in later lines of therapy. Citation Format: David B. Solit, Niharika B. Mettu, Shannon J. McCall, Mallika Dhawan, Priyanka J. Bobbili, Maral DerSarkissian, Jasmina Ivanova, Bhakti Arondekar, Jane Chang, Alexander Niyazov, Jocelyn Lee, Risha Huq, Michelle Green, Michelle Turski, Aruna Muthukumar, Tracy Guo, Mei Sheng Duh, William K. Oh. Treatment patterns and overall survival among men with metastatic castration-resistant prostate cancer harboring DDR mutations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 930.

Recent advances in next-generation sequencing have allowed for an increase in molecular tumor profiling. We sought to assess the actionability and clinical utilization of molecular tumor profiling results obtained via Foundation Medicine tumor sequencing tests in uterine and ovarian cancers. We performed a single-institution retrospective chart review to obtain demographic and clinical information in patients with uterine and ovarian cancer whose tumors were submitted to Foundation Medicine for molecular tumor profiling over a 7-year period. Alterations identified on testing were stratified according to the OncoKB database actionability algorithm. Descriptive statistics were primarily used to analyze the data. Tumors from 185 women with gynecologic cancer were submitted for molecular tumor profiling between 2013 and 2019. The majority of tests (144/185; 78%) were ordered after a diagnosis of recurrence. In 60 (32%), no actionable molecular alteration was identified. Thirteen (7%) identified an alteration that directed to a US Food and Drug Administration-approved therapy in that tumor type, while 112 (61%) had alterations with investigational or hypothetical treatment implications. In patients with any actionable finding, treatment was initiated in 27 (15%) based on these results. The majority of uterine and ovarian cancers (93%) did not have molecular alterations with corresponding Food and Drug Administration-approved treatments. Even in patients with a potentially actionable alteration, gynecologic oncologists were more likely to choose an alternative therapy. Further investigation is warranted to determine which patients with uterine and ovarian cancer are most likely to benefit from molecular tumor profiling and the ideal timing of testing. The potential to identify effective therapeutic options in a minority of patients needs to be balanced with the current limited clinical applicability of these results in most cases.

Intrahepatic cholangiocarcinoma (ICC) remains a deadly malignancy lacking systemic therapies for advanced disease. Recent advancements include selective FGFR1–3 inhibitors for the 15% of ICC patients harboring fusions, although survival is limited by poor response and resistance. Herein we report generation of a patient-derived FGFR2 fusion-positive ICC model system consisting of a cell line, organoid, and xenograft, which have undergone complete histologic, genomic, and phenotypic characterization, including testing standard-of-care systemic therapies. Using these FGFR2 fusion-positive ICC models, we conducted an unbiased high-throughput small molecule screen to prioritize combination strategies with FGFR inhibition, from which HDAC inhibition together with pemigatinib was validated in vitro and in vivo as a synergistic therapy for ICC. Additionally, we demonstrate broad utility of the FGFR/HDAC combination for other FGFR fusion-positive solid tumors. These data are directly translatable and justify early phase trials to establish dosing, safety, and therapeutic efficacy of this synergistic combination.

Background Immune checkpoint inhibitors are now standard of care for many patients with metastatic renal cell carcinoma (mRCC) and metastatic urothelial carcinoma (mUC). Given real-world limitations in programmed death-ligand 1 (PD-L1) testing, concordance studies between PD-L1 assays are needed. We undertook comparisons of Dako 28-8 and Ventana SP142 assays in mRCC and Dako 22C3 and Ventana SP263 assays in mUC. Patients and Methods Thirty-two patients with mRCC and 18 patients with mUC who had received immune checkpoint inhibitor therapy were identified. Formalin-fixed paraffin-embedded tumor samples for patients with mRCC were evaluated with Dako 28-8 and Ventana SP142 PD-L1 immunohistochemistry assays. For patients with mUC, formalin-fixed paraffin-embedded tumor samples were evaluated with Dako 22C3 and Ventana SP263 PD-L1 immunohistochemistry assays. Results The majority (29/32; 91%) of mRCC cases were concordant between assays. The majority (17/18; 94%) of mUC cases were also concordant between assays. Conclusions There was strong concordance between PD-L1 assays chosen for comparison in both mRCC and mUC, with similar performance characteristics. One limitation is the small number of cases in this study; larger comparison studies are needed for this biomarker in mRCC and mUC.

The diagnosis of invasive adenocarcinoma of the gallbladder can sometimes be challenging. The presence of true desmoplastic reaction facilitates the diagnosis of invasion. However, desmoplasia-like changes can be observed in benign gallbladder conditions, and recognition of desmoplasia may be challenging based on morphology. In this study, we tested the expression pattern of microfibril-associated protein 5 (MFAP5), a promising immunohistochemical marker for desmoplasia, in benign gallbladders with desmoplasia-like reaction and gallbladders with invasive adenocarcinoma. We also evaluated the diagnostic utility of MFAP5 in challenging cases with an interobserver agreement study. The results showed that all benign cases retained intact/positive MFAP5 staining pattern in periglandular connective tissue, whereas 79.3% (23 out of 29) of cases of adenocarcinomas demonstrated diffuse and complete loss of MFAP5 staining in the tumor stroma. Interobserver agreement was improved by 2.66 times when images of MFAP5 immunohistochemistry were provided. In conclusion, MFAP5 expression is downregulated in the desmoplastic stroma of gallbladder adenocarcinoma and may provide a useful diagnostic marker in difficult cases.

The Project Baseline Health Study (PBHS) was launched to map human health through a comprehensive understanding of both the health of an individual and how it relates to the broader population. The study will contribute to the creation of a biomedical information system that accounts for the highly complex interplay of biological, behavioral, environmental, and social systems. The PBHS is a prospective, multicenter, longitudinal cohort study that aims to enroll thousands of participants with diverse backgrounds who are representative of the entire health spectrum. Enrolled participants will be evaluated serially using clinical, molecular, imaging, sensor, self-reported, behavioral, psychological, environmental, and other health-related measurements. An initial deeply phenotyped cohort will inform the development of a large, expanded virtual cohort. The PBHS will contribute to precision health and medicine by integrating state of the art testing, longitudinal monitoring and participant engagement, and by contributing to the development of an improved platform for data sharing and analysis.

PURPOSE National Cancer Institute (NCI)–sponsored clinical trial network studies frequently require biopsy specimens for pharmacodynamic and molecular biomarker analyses, including paired pre- and post-treatment samples. The purpose of this meeting of NCI-sponsored investigators was to identify local institutional standard procedures found to ensure quantitative and qualitative specimen adequacy. METHODS NCI convened a conference on best biopsy practices, focusing on the clinical research community. Topics discussed were (1) criteria for specimen adequacy in the personalized medicine era, (2) team-based approaches to ensure specimen adequacy and quality control, and (3) risk considerations relevant to academic and community practitioners and their patients. RESULTS AND RECOMMENDATIONS Key recommendations from the convened consensus panel included (1) establishment of infrastructure for multidisciplinary biopsy teams with a formalized information capture process, (2) maintenance of standard operating procedures with regular team review, (3) optimization of tissue collection and yield methodology, (4) incorporation of needle aspiration and other newer techniques, and (5) commitment of stakeholders to use of guideline documents to increase awareness of best biopsy practices, with the goal of universally improving tumor biopsy practices.

Simple Summary The X-linked inhibitor of apoptosis protein (XIAP) is considered the most potent inhibitor of cell death, and it is well established that XIAP promotes resistance to chemotherapy, radiation, and anti-cancer immune responses. This study evaluates the correlations between XIAP expression and clinicopathological features, including disease-free survival (DFS) and pathological complete response (pCR) to chemotherapy, in more than 2300 invasive primary breast cancer samples. We found a significant association of XIAP expression with younger patients’ age (≤50 years), pathological ductal type, lower tumor grade, node-positive status, and PAM50 luminal B subtype. Analysis of molecular subtypes revealed a stronger prognostic value in HR+/HER2− tumors. Higher XIAP expression was associated with shorter DFS and lower pCR rate to chemotherapy in both uni- and multivariate analyses. All these correlations were observed at both the RNA and protein level, indicating the potential of XIAP as a promising therapeutic target in primary invasive breast cancer. Abstract XIAP, the most potent inhibitor of cell death pathways, is linked to chemotherapy resistance and tumor aggressiveness. Currently, multiple XIAP-targeting agents are in clinical trials. However, the characterization of XIAP expression in relation to clinicopathological variables in large clinical series of breast cancer is lacking. We retrospectively analyzed non-metastatic, non-inflammatory, primary, invasive breast cancer samples for XIAP mRNA (n = 2341) and protein (n = 367) expression. XIAP expression was analyzed as a continuous value and correlated with clinicopathological variables. XIAP mRNA expression was heterogeneous across samples and significantly associated with younger patients’ age (≤50 years), pathological ductal type, lower tumor grade, node-positive status, HR+/HER2− status, and PAM50 luminal B subtype. Higher XIAP expression was associated with shorter DFS in uni- and multivariate analyses in 909 informative patients. Very similar correlations were observed at the protein level. This prognostic impact was significant in the HR+/HER2− but not in the TN subtype. Finally, XIAP mRNA expression was associated with lower pCR rate to anthracycline-based neoadjuvant chemotherapy in both uni- and multivariate analyses in 1203 informative patients. Higher XIAP expression in invasive breast cancer is independently associated with poorer prognosis and resistance to chemotherapy, suggesting the potential therapeutic benefit of targeting XIAP.

Context.— Prostate cancer is a common malignancy, and accurate diagnosis typically requires histologic review of multiple prostate core biopsies per patient. As pathology volumes and complexity increase, new tools to improve the efficiency of everyday practice are keenly needed. Deep learning has shown promise in pathology diagnostics, but most studies silo the efforts of pathologists from the application of deep learning algorithms. Very few hybrid pathologist–deep learning approaches have been explored, and these typically require complete review of histologic slides by both the pathologist and the deep learning system. Objective.— To develop a novel and efficient hybrid human–machine learning approach to screen prostate biopsies. Design.— We developed an algorithm to determine the 20 regions of interest with the highest probability of malignancy for each prostate biopsy; presenting these regions to a pathologist for manual screening limited the initial review by a pathologist to approximately 2% of the tissue area of each sample. We evaluated this approach by using 100 biopsies (29 malignant, 60 benign, 11 other) that were reviewed by 4 pathologists (3 urologic pathologists, 1 general pathologist) using a custom-designed graphical user interface. Results.— Malignant biopsies were correctly identified as needing comprehensive review with high sensitivity (mean, 99.2% among all pathologists); conversely, most benign prostate biopsies (mean, 72.1%) were correctly identified as needing no further review. Conclusions.— This novel hybrid system has the potential to efficiently triage out most benign prostate core biopsies, conserving time for the pathologist to dedicate to detailed evaluation of malignant biopsies.

X-ray transmission imaging has been used in a variety of applications for high-resolution measurements based on shape and density. Similarly, X-ray diffraction (XRD) imaging has been used widely for molecular structure-based identification of materials. Combining these X-ray methods has the potential to provide high-resolution material identification, exceeding the capabilities of either modality alone. However, XRD imaging methods have been limited in application by their long measurement times and poor spatial resolution, which has generally precluded combined, rapid measurements of X-ray transmission and diffraction. In this work, we present a novel X-ray fan beam coded aperture transmission and diffraction imaging system, developed using commercially available components, for rapid and accurate non-destructive imaging of industrial and biomedical specimens. The imaging system uses a 160 kV Bremsstrahlung X-ray source while achieving a spatial resolution of ≈ 1 × 1 mm2 and a spectral accuracy of > 95% with only 15 s exposures per 150 mm fan beam slice. Applications of this technology are reported in geological imaging, pharmaceutical inspection, and medical diagnosis. The performance of the imaging system indicates improved material differentiation relative to transmission imaging alone at scan times suitable for a variety of industrial and biomedical applications.

INTRODUCTION: Aggregation of Barretts esophagus (BE) and esophageal adenocarcinoma (EAC) in families has been termed familial Barretts esophagus (FBE). Analysis of single, large FBE families can enable the identification of genetic susceptibility to complex diseases such as BE and EAC. METHODS: Phenotypes of BE and EAC were ascertained in a large FBE family with 7 affected members: 4 men with EAC, 1 man with BE and high grade dysplasia, and 2 women with non-dysplastic BE by review of endoscopy and surgical pathology reports. Whole exome sequencing was performed on germline DNA from 3 affected members to identify variants in coding genes that segregated with disease. Formalin fixed paraffin embedded tissue from an affected family member as well as non-familial subjects with BE and EAC was examined with regular histology and immunohistochemistry. The CAV3 gene with a variant segregating in the family was further characterized in a porcine model of esophageal injury using immunofluorescence. RESULTS: Using a whole exome sequencing approach on an exceptional FBE family we identified a segregating nonsense mutation in the gene Caveolin-3 (CAV3). Histologic examination of a formalin fixed paraffin embedded (FFPE) esophagectomy specimen from an individual carrying the CAV3 null mutation revealed esophageal submucosal glands (ESMG) that showed acinar metaplasia with marked atypia and absence of myoepithelial cells, distinctly different from acinar metaplasia seen in ESMG of non-familial subjects with BE and high grade dysplasia. Immunofluorescence studies of ESMG in porcine esophagus revealed the presence of CAV3 in selected cells in a distribution that was consistent with myoepithelial cells. Experimental injury of the porcine esophagus using radiofrequency ablation revealed that CAV3 expression increased markedly within ESMGs, ESMG ductal epithelium, and overlying healing neosquamous epithelium 10 days after injury. CONCLUSIONS: We theorize that CAV3 expression, perhaps through myoepithelial cells within ESMGs, controls the differentiation and proliferation of squamous epithelial precursor cells in response to injury. Furthermore, the truncating nonsense CAV3 mutation discovered in a family disrupts normal squamous healing and the organization of ESMGs, making affected family members susceptible to the proliferation and development of metaplastic columnar BE and EAC.

The lack of validated, distributed comprehensive genomic profiling assays for patients with cancer inhibits access to precision oncology treatment. To address this, we describe elio tissue complete, which has been FDA-cleared for examination of 505 cancer-related genes. Independent analyses of clinically and biologically relevant sequence changes across 170 clinical tumor samples using MSK-IMPACT, FoundationOne, and PCR-based methods reveals a positive percent agreement of >97%. We observe high concordance with whole-exome sequencing for evaluation of tumor mutational burden for 307 solid tumors (Pearson r = 0.95) and comparison of the elio tissue complete microsatellite instability detection approach with an independent PCR assay for 223 samples displays a positive percent agreement of 99%. Finally, evaluation of amplifications and translocations against DNA- and RNA-based approaches exhibits >98% negative percent agreement and positive percent agreement of 86% and 82%, respectively. These methods provide an approach for pan-solid tumor comprehensive genomic profiling with high analytical performance.

Background: Osteosarcoma is an aggressive bone cancer in which therapeutic advancements have been limited over the last 30 years, in part due to genomic heterogeneity. The combination of high-throughput drug screening platforms that efficiently pinpoint drug sensitivities with patient-derived cross-species models is an innovative approach to address the critical need to identify novel treatment strategies for osteosarcoma patients. Methods: We performed high-throughput drug screens on patient-derived osteosarcoma cell lines D418 (canine) and 17-3x (human), followed by validation of the top compounds, to identify drug sensitivities and novel therapeutic combinations. Results: High-throughput drug screens using 2100 bioactive compounds show that osteosarcoma cell lines D418 and 17-3x exhibited sensitivity to standard-of-care chemotherapy drugs, inhibitors of XPO1 nuclear export, and proteasome inhibitors. The XPO1 inhibitor, verdinexor (VER), and the proteasome inhibitor, bortezomib (BORT), induced dose-dependent cytotoxicity in multiple osteosarcoma cell lines (D418, IC50VER: 3187 nM, IC50BORT: 2.8 nM; 17-3x IC50VER: 679 nM, IC50BORT: 10.9 nM). In addition, dual XPO1 and proteasome inhibition synergistically reduced cell proliferation in D418 (synergy score=12.89) and 17-3x (synergy score=17.87) cell lines (p Conclusions: Inhibition of XPO1-mediated nuclear export is a promising therapeutic strategy in osteosarcoma. These effects may be further potentiated when used in combination with other agents, such as proteasome inhibitors. Additional drug screening and validation assays are underway to identify novel synergistic agents for use in combination with XPO1 inhibitors in osteosarcoma. Citation Format: Laurie Graves, Gabrielle Rupprecht, Erdem Altunel, Etienne M. Flamant, Sneha Rao, Dharshan Sivara, Alexander L. Lazarides, Sarah M. Hoskinson, Maya U. Sheth, Serene Cheng, So Young Kim, Kathryn E. Ware, Anika Agarwal, Mark M. Cullen, Casey Syal, Laura E. Selmic, Jeffrey I. Everitt, Shannon J. McCall, Cindy Eward, Trinayan Kashyap, Marie Maloof, Christopher J. Walker, Yosef Landesman, Lars Wagner, William C. Eward, David S. Hsu, Jason A. Somarelli. Exportin 1 (XPO1) inhibition alone or in combination as a novel therapeutic strategy in osteosarcoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1061.

The era of COVID-19 has brought about a number of novel challenges for the global biobanking community. To better position the biobanking community to cope with current and future challenges, the International Society for Biological and Environmental Repositories (ISBER) COVID-19 Response Task Force was convened to identify needs and gaps in biobanking tools (existing resources that support good practice), for example, standards, best practices, business, etc. and to make recommendations to benefit the community. Toward these goals, the Task Force assembled a set of questions to explore individual biobanks' experiences, with emphasis on identification of key challenges and approaches, including tools employed. A survey was designed with the use of these questions and administered by ISBER. This article presents a summary of the aggregated data obtained from the survey responses, illustrating some of the major issues encountered and identifying which tools the survey respondents found most useful. In particular, this article focuses on the challenges identified during the early months of the COVID-19 era. Recommendations are provided to support biobank emergency preparedness for the future, address lessons learned, and propose solutions to bridge identified gaps. The analysis and the complete survey dataset will also inform the larger Task Force goal to develop specific tool recommendations., info:eu-repo/semantics/publishedVersion

Recent advances in next-generation sequencing have allowed for an increase in molecular tumor profiling.We sought to assess the actionability and clinical utilization of molecular tumor profiling results obtained via Foundation Medicine tumor sequencing tests in uterine and ovarian cancers.We performed a single-institution retrospective chart review to obtain demographic and clinical information in patients with uterine and ovarian cancer whose tumors were submitted to Foundation Medicine for molecular tumor profiling over a 7-year period. Alterations identified on testing were stratified according to the OncoKB database actionability algorithm. Descriptive statistics were primarily used to analyze the data.Tumors from 185 women with gynecologic cancer were submitted for molecular tumor profiling between 2013 and 2019. The majority of tests (144/185; 78%) were ordered after a diagnosis of recurrence. In 60 (32%), no actionable molecular alteration was identified. Thirteen (7%) identified an alteration that directed to a US Food and Drug Administration-approved therapy in that tumor type, while 112 (61%) had alterations with investigational or hypothetical treatment implications. In patients with any actionable finding, treatment was initiated in 27 (15%) based on these results.The majority of uterine and ovarian cancers (93%) did not have molecular alterations with corresponding Food and Drug Administration-approved treatments. Even in patients with a potentially actionable alteration, gynecologic oncologists were more likely to choose an alternative therapy. Further investigation is warranted to determine which patients with uterine and ovarian cancer are most likely to benefit from molecular tumor profiling and the ideal timing of testing. The potential to identify effective therapeutic options in a minority of patients needs to be balanced with the current limited clinical applicability of these results in most cases.

The COVID-19 era has brought about a number of novel challenges for the global biobanking community. An array of diverse tools (e.g., standards, best practices, and plans) exists to support quality and fitness-for-purpose in biobank operations. The International Society for Biological and Environmental Repositories (ISBER) COVID-19 Response Task Force has set out to identify needs and gaps in these tools and make recommendations for the next generation of available tools, having closely examined the COVID-19-related challenges. While conducting this work to examine the relationships between tools and biobank adaptability, a subgroup of the task force conducted a parallel effort to develop and describe individual COVID-19 era case studies based on a number of operating biobanks. Each case study presents a different combination of implemented tools. Observations and lessons learned from these case studies are provided, and experiences with tool implementation are discussed. This information is supplemented by data relating to tool usefulness that was obtained through an ISBER survey discussed in a companion article. The knowledge gained from this study will be combined with other task force efforts to make recommendations to better position the biobanking community in their response to future emergencies., This study was performed by a Task Force organized by members of the International Society for Biological and Environmental Repositories (ISBER) Standards Advisory Committee. The authors gratefully acknowledge our fellow ISBER COVID-19 tools Task Force members who reviewed this article, developed figures and tables, and provided valuable feedback, particularly Daniel Simeon-Dubach, Rebecca Pugh, Sergey Anisimov, and Yehudit Cohen. Shannon McCall and the Duke University BioRepository & Precision Pathology Center receive funding from the United States National Institutes of Health, National Cancer Institute under UM1CA239755 (The Cooperative Human Tissue Network) and P30CA014236 (Duke University’s Cancer Center Support Grant)., info:eu-repo/semantics/publishedVersion

Colorectal cancer is the third most common cancer in the United States and responsible for over 50,000 deaths each year. Therapeutic options for advanced colorectal cancer are limited, and there remains an unmet clinical need to identify new treatments for this deadly disease. To address this need, we developed a precision medicine pipeline that integrates high-throughput chemical screens with matched patient-derived cell lines and patient-derived xenografts (PDX) to identify new treatments for colorectal cancer. High-throughput screens of 2,100 compounds were performed across six low-passage, patient-derived colorectal cancer cell lines. These screens identified the CDK inhibitor drug class among the most effective cytotoxic compounds across six colorectal cancer lines. Among this class, combined targeting of CDK1, 2, and 9 was the most effective, with IC50s ranging from 110 nmol/L to 1.2 μmol/L. Knockdown of CDK9 in the presence of a CDK2 inhibitor (CVT-313) showed that CDK9 knockdown acted synergistically with CDK2 inhibition. Mechanistically, dual CDK2/9 inhibition induced significant G2–M arrest and anaphase catastrophe. Combined CDK2/9 inhibition in vivo synergistically reduced PDX tumor growth. Our precision medicine pipeline provides a robust screening and validation platform to identify promising new cancer therapies. Application of this platform to colorectal cancer pinpointed CDK2/9 dual inhibition as a novel combinatorial therapy to treat colorectal cancer.

PurposeOsteosarcoma is a rare but aggressive bone cancer that occurs primarily in children. Like other rare cancers, treatment advances for osteosarcoma have stagnated, with little improvement in survival for the past several decades. Developing new treatments has been hampered by extensive genomic heterogeneity and limited access to patient samples to study the biology of this complex disease.Experimental designTo overcome these barriers, we combined the power of comparative oncology with patient-derived models of cancer and high-throughput chemical screens in a cross-species drug discovery pipeline.ResultsCouplingin vitrohigh-throughput drug screens on low-passage and established cell lines within vivovalidation in patient-derived xenografts we identify the proteasome and CRM1 nuclear export pathways as therapeutic sensitivities in osteosarcoma, with dual inhibition of these pathways inducing synergistic cytotoxicity.ConclusionsThese collective efforts provide an experimental framework and set of new tools for osteosarcoma and other rare cancers to identify and study new therapeutic vulnerabilities.

Objectives Despite disparities in lung cancer incidence and mortality, the molecular landscape of lung cancer in patients of African ancestry remains underexplored, and race-related differences in RNA splicing remain unexplored. Materials and methods We identified differentially spliced genes (DSGs) and differentially expressed genes (DEGs) in biobanked lung squamous cell carcinoma (LUSC) between patients of West African and European ancestry, using ancestral genotyping and Affymetrix Clariom D array. DSGs and DEGs were validated independently using the National Cancer Institute Genomic Data Commons. Associated biological processes, overlapping canonical pathways, enriched gene sets, and cancer relevance were identified using Gene Ontology Consortium, Ingenuity Pathway Analysis, Gene Set Enrichment Analysis, and CancerMine, respectively. Association with LUSC survival was conducted using The Cancer Genome Atlas. Results 4,829 DSGs and 267 DEGs were identified, including novel targets in NSCLC as well as genes identified previously to have relevance to NSCLC. RNA splicing events within 3 DSGs as well as 1 DEG were validated in the independent cohort. 853 DSGs and 29 DEGs have been implicated as potential drivers, oncogenes and/or tumor suppressor genes. Biological processes enriched among DSGs and DEGs included metabolic process, biological regulation, and multicellular organismal process and, among DSGs, ion transport. Overlapping canonical pathways among DSGs included neuronal signaling pathways and, among DEGs, cell metabolism involving biosynthesis. Gene sets enriched among DSGs included KRAS Signaling, UV Response, E2 F Targets, Glycolysis, and Coagulation. 355 RNA splicing events within DSGs and 18 DEGs show potential association with LUSC patient survival. Conclusion These DSGs and DEGs, which show potential biological and clinical relevance, could have the ability to drive novel biomarker and therapeutic development to mitigate LUSC disparities.

BackgroundEsophageal cancer is a deadly cancer with 5-year survival MethodsCases included 452 esophagectomy cases, including 396 with EAC and 56 who underwent esophagectomy for Barrett's esophagus (BE) with high grade dysplasia (HGD). 1,102 thoracic surgery patients with surgical indications other than dysplastic BE or esophageal cancer represented the controls for our analysis. The association of tonsillectomy and HGD/EAC were primarily evaluated by using univariate tests and then verified by logistic regression analysis. Baseline demographics, medical history, and thoracic surgery controls were compared by using χ2 tests or 95% CIs. Significant risk factors were considered as covariates in the multivariate models while evaluating the association between tonsillectomy and HGD/EAC. P-values or odds ratios were estimated with 95% confidence limits to identify significances which was more appropriate.ResultsTonsillectomy was more common in cases than controls and was found to have a significant association with esophageal cancer (19.9% vs. 12.7%; p-value = 0.0003). This significant association persisted after controlling for other known risk factors/covariates.ConclusionA prior history of tonsillectomy was significantly associated with HGD/EAC and may represent an independent risk factor for the development of EAC. However, the underlying biology driving this association remains unclear.

Background Metastatic colorectal cancer (CRC) continues to be a major health problem, and current treatments are primarily for disease control and palliation of symptoms. In this study, we developed a precision medicine strategy to discover novel therapeutics for patients with CRC. Methods Six matched low-passage cell lines and patient-derived xenografts (PDX) were established from CRC patients undergoing resection of their cancer. High-throughput drug screens using a 119 FDA-approved oncology drug library were performed on these cell lines, which were then validated in vivo in matched PDXs. RNA-Seq analysis was then performed to identify predictors of response. Results Our study revealed marked differences in response to standard-of-care agents across patients and pinpointed druggable pathways to treat CRC. Among these pathways co-targeting of fibroblast growth factor receptor (FGFR), SRC, platelet derived growth factor receptor (PDGFR), or vascular endothelial growth factor receptor (VEGFR) signaling was found to be an effective strategy. Molecular analyses revealed potential predictors of response to these druggable pathways. Conclusions Our data suggests that the use of matched low-passage cell lines and PDXs is a promising strategy to identify new therapies and pathways to treat metastatic CRC.

Background Biomarkers predicting immunotherapy response in metastatic renal cell cancer (mRCC) are lacking. PD-L1 immunohistochemistry is a complementary diagnostic for immune checkpoint inhibitors (ICIs) in mRCC, but has shown minimal clinical utility and is not used in routine clinical practice. Methods Tumor specimens from 56 patients with mRCC who received nivolumab were evaluated for PD-L1, cell proliferation (targeted RNA-seq), and outcome. Results For 56 patients treated with nivolumab as a standard of care, there were 2 complete responses and 8 partial responses for a response rate of 17.9%. Dividing cell proliferation into tertiles, derived from the mean expression of 10 proliferation-associated genes in a reference set of tumors, poorly proliferative tumors (62.5%) were more common than moderately (30.4%) or highly proliferative (8.9%) counterparts. Moderately proliferative tumors were enriched for PD-L1 positive (41.2%), compared to poorly proliferative counterparts (11.4%). Objective response for moderately proliferative (29.4%) tumors was higher than that of poorly (11.4%) proliferative counterparts, but not statistically significant (p = .11). When cell proliferation and negative PD-L1 tumor proportion scores were combined statistically significant results were achieved (p = .048), showing that patients with poorly proliferative and PD-L1 negative tumors have a very low response rate (6.5%) compared to moderately proliferative PD-L1 negative tumors (30%). Conclusions Cell proliferation has value in predicting response to nivolumab in clear cell mRCC patients, especially when combined with PD-L1 expression. Further studies which include the addition of progression-free survival (PFS) along with sufficiently powered subgroups are required to further support these findings.

The diagnosis of invasive adenocarcinoma of the gallbladder can sometimes be challenging. The presence of true desmoplastic reaction facilitates the diagnosis of invasion. However, desmoplasia-like changes can be observed in benign gallbladder conditions, and recognition of desmoplasia may be challenging based on morphology. In this study, we tested the expression pattern of microfibril-associated protein 5 (MFAP5), a promising immunohistochemical marker for desmoplasia, in benign gallbladders with desmoplasia-like reaction and gallbladders with invasive adenocarcinoma. We also evaluated the diagnostic utility of MFAP5 in challenging cases with an interobserver agreement study. The results showed that all benign cases retained intact/positive MFAP5 staining pattern in periglandular connective tissue, whereas 79.3% (23 out of 29) of cases of adenocarcinomas demonstrated diffuse and complete loss of MFAP5 staining in the tumor stroma. Interobserver agreement was improved by 2.66 times when images of MFAP5 immunohistochemistry were provided. In conclusion, MFAP5 expression is downregulated in the desmoplastic stroma of gallbladder adenocarcinoma and may provide a useful diagnostic marker in difficult cases.

The College of American Pathologists (CAP) developed the Biorepository Accreditation Program (BAP) in 2012. This program integrates best practices from the International Society for Biological and Environmental Biorepositories, the National Cancer Institute, the Organisation for Economic Cooperation and Development, the Center for Medicare and Medicaid Services, and the CAP Laboratory Accreditation Program. The goal of this elective program is to provide requirements for standardization in biorepository processes that will result in high-quality specimens that can be used to support research, drug discovery, and personalized medicine. CAP uses a peer inspection model to ensure the inspectors have proper expertise and to promote educational efforts through information sharing. Lead inspectors are comprised of pathologists, PhDs, and managers of biorepositories and they are often supported by CAP staff inspectors. Accreditation is a 3-year continuous cycle of quality with a peer inspection occurring at the start of year 1 and a self-inspection and CAP desk assessment at the start of year 2 and 3. At this time 53 biorepositories are fully CAP BAP accredited and 13 are in the process of obtaining accreditation. There are currently 273 established standards with requirement lists customized based on the scope of activities performed by a biorepository. A total of 90 inspections were completed between May 2012 and December 2016. Sixty-one were initial inspections and 29 were reinspections. A total of 527 deficiencies were identified in the areas of Equipment/Instrumentation (22%), Information Technology (18%), Specimen Handling and QC (15%), Quality Management (16%), Personnel (11%), Safety (10%), Facilities (6%), and Regulatory (2%). Assessment of common deficiencies identifies areas of focus for continuous improvement and educational opportunities. Overall success of the program is high based on the current enrollment of 66 biorepositories, anecdotal participant feedback and increasing national recognition of the BAP in federal documents.

Objectives To characterize the expression of SOX11 and TFE3 proteins in solid-pseudopapillary neoplasms (SPNs) and their histologic mimickers. Methods Immunohistochemistry for SOX11, TFE3, and β-catenin was performed on 31 cases of surgically resected SPNs. Neuroendocrine tumors, acinar cell carcinomas, and pancreatoblastomas served as controls. Results Nuclear immunoreactivity for SOX11 was detected in all SPNs and five of 31 control tumors. Nuclear immunoreactivity for TFE3 was detected in 30 SPNs and three control tumors. Nuclear immunoreactivity for β-catenin was detected in all SPNs and four control tumors. The combination of three markers as immunohistochemical panels resulted in optimal sensitivity and specificity. Conclusions Both SOX11 and TFE3 were overexpressed in SPNs and may be involved in the pathogenesis. Clinically, SOX11 and TFE3 can be potentially used as diagnostic markers in distinguishing indeterminate SPNs from their histologic mimickers.

Purpose: To determine the immunologic effects of neoadjuvant chemotherapy plus ipilimumab in early-stage non–small cell lung cancer (NSCLC) patients. Experimental Design: This is a single-arm chemotherapy plus phased ipilimumab phase II study of 24 treatment-naïve patients with stage IB–IIIA NSCLC. Patients received neoadjuvant therapy consisting of 3 cycles of paclitaxel with either cisplatin or carboplatin and ipilimumab included in the last 2 cycles. Results: Chemotherapy alone had little effect on immune parameters in PBMCs. Profound CD28-dependent activation of both CD4 and CD8 cells was observed following ipilimumab. Significant increases in the frequencies of CD4+ cells expressing activation markers ICOS, HLA-DR, CTLA-4, and PD-1 were apparent. Likewise, increased frequencies of CD8+ cells expressing the same activation markers, with the exception of PD-1, were observed. We also examined 7 resected tumors and found higher frequencies of activated tumor-infiltrating lymphocytes than those observed in PBMCs. Surprisingly, we found 4 cases of preexisting tumor-associated antigens (TAA) responses against survivin, PRAME, or MAGE-A3 present in PBMC at baseline, but neither increased frequencies nor the appearance of newly detectable responses following ipilimumab therapy. Ipilimumab had little effect on the frequencies of circulating regulatory T cells and MDSCs. Conclusions: This study did not meet the primary endpoint of detecting an increase in blood-based TAA T-cell responses after ipilimumab. Collectively, these results highlight the immune activating properties of ipilimumab in early-stage NSCLC. The immune profiling data for ipilimumab alone can contribute to the interpretation of immunologic data from combined immune checkpoint blockade immunotherapies. Clin Cancer Res; 23(24); 7474–82. ©2017 AACR.

Summary: Diverse pathways drive resistance to BRAF/MEK inhibitors in BRAF-mutant melanoma, suggesting that durable control of resistance will be a challenge. By combining statistical modeling of genomic data from matched pre-treatment and post-relapse patient tumors with functional interrogation of >20 in vitro and in vivo resistance models, we discovered that major pathways of resistance converge to activate the transcription factor, c-MYC (MYC). MYC expression and pathway gene signatures were suppressed following drug treatment, and then rebounded during progression. Critically, MYC activation was necessary and sufficient for resistance, and suppression of MYC activity using genetic approaches or BET bromodomain inhibition was sufficient to resensitize cells and delay BRAFi resistance. Finally, MYC-driven, BRAFi-resistant cells are hypersensitive to the inhibition of MYC synthetic lethal partners, including SRC family and c-KIT tyrosine kinases, as well as glucose, glutamine, and serine metabolic pathways. These insights enable the design of combination therapies that select against resistance evolution. : Diverse pathways drive resistance to BRAF/MEK inhibitors in BRAF-mutant melanoma, but by combining statistical modeling of tumor data with functional interrogation of resistance models, Singleton et al. show that these pathways converge to activate MYC. BRAFi-resistant cells are hypersensitive to the inhibition of MYC synthetic lethal partners, informing therapies that select against resistance. Keywords: melanoma, cancer therapeutics, therapeutic resistance, signaling networks, MYC, metabolism, synthetic lethality

Background & Aims Although cells comprising esophageal submucosal glands (ESMGs) represent a potential progenitor cell niche, new models are needed to understand their capacity to proliferate and differentiate. By histologic appearance, ESMGs have been associated with both overlying normal squamous epithelium and columnar epithelium. Our aim was to assess ESMG proliferation and differentiation in a 3-dimensional culture model. Methods We evaluated proliferation in human ESMGs from normal and diseased tissue by proliferating cell nuclear antigen immunohistochemistry. Next, we compared 5-ethynyl-2′-deoxyuridine labeling in porcine ESMGs in vivo before and after esophageal injury with a novel in vitro porcine organoid ESMG model. Microarray analysis of ESMGs in culture was compared with squamous epithelium and fresh ESMGs. Results Marked proliferation was observed in human ESMGs of diseased tissue. This activated ESMG state was recapitulated after esophageal injury in an in vivo porcine model, ESMGs assumed a ductal appearance with increased proliferation compared with control. Isolated and cultured porcine ESMGs produced buds with actively cycling cells and passaged to form epidermal growth factor–dependent spheroids. These spheroids were highly proliferative and were passaged multiple times. Two phenotypes of spheroids were identified: solid squamous (P63+) and hollow/ductal (cytokeratin 7+). Microarray analysis showed spheroids to be distinct from parent ESMGs and enriched for columnar transcripts. Conclusions Our results suggest that the activated ESMG state, seen in both human disease and our porcine model, may provide a source of cells to repopulate damaged epithelium in a normal manner (squamous) or abnormally (columnar epithelium). This culture model will allow the evaluation of factors that drive ESMGs in the regeneration of injured epithelium. The raw microarray data have been uploaded to the National Center for Biotechnology Information Gene Expression Omnibus (accession number: GSE100543)., Graphical abstract

Hormonal therapy targeting androgen receptor (AR) is initially effective to treat prostate cancer (PCa), but it eventually fails. It has been hypothesized that cellular heterogeneity of PCa, consisting of AR(+) luminal tumor cells and AR(−) neuroendocrine (NE) tumor cells, may contribute to therapy failure. Here, we describe the successful purification of NE cells from primary fresh human prostate adenocarcinoma based on the cell surface receptor C-X-C motif chemokine receptor 2 (CXCR2). Functional studies revealed CXCR2 to be a driver of the NE phenotype, including loss of AR expression, lineage plasticity, and resistance to hormonal therapy. CXCR2-driven NE cells were critical for the tumor microenvironment by providing a survival niche for the AR(+) luminal cells. We demonstrate that the combination of CXCR2 inhibition and AR targeting is an effective treatment strategy in mouse xenograft models. Such a strategy has the potential to overcome therapy resistance caused by tumor cell heterogeneity.

Background Resistance to immune checkpoint inhibitors (ICIs) has been linked to local immunosuppression independent of major ICI targets (e.g., PD-1). Clinical experience with response prediction based on PD-L1 expression suggests that other factors influence sensitivity to ICIs in non-small cell lung cancer (NSCLC) patients. Methods Tumor specimens from 120 NSCLC patients from 10 institutions were evaluated for PD-L1 expression by immunohistochemistry, and global proliferative profile by targeted RNA-seq. Results Cell proliferation, derived from the mean expression of 10 proliferation-associated genes (namely BUB1, CCNB2, CDK1, CDKN3, FOXM1, KIAA0101, MAD2L1, MELK, MKI67, and TOP2A), was identified as a marker of response to ICIs in NSCLC. Poorly, moderately, and highly proliferative tumors were somewhat equally represented in NSCLC, with tumors with the highest PD-L1 expression being more frequently moderately proliferative as compared to lesser levels of PD-L1 expression. Proliferation status had an impact on survival in patients with both PD-L1 positive and negative tumors. There was a significant survival advantage for moderately proliferative tumors compared to their combined highly/poorly counterparts (p = 0.021). Moderately proliferative PD-L1 positive tumors had a median survival of 14.6 months that was almost twice that of PD-L1 negative highly/poorly proliferative at 7.6 months (p = 0.028). Median survival in moderately proliferative PD-L1 negative tumors at 12.6 months was comparable to that of highly/poorly proliferative PD-L1 positive tumors at 11.5 months, but in both instances less than that of moderately proliferative PD-L1 positive tumors. Similar to survival, proliferation status has impact on disease control (DC) in patients with both PD-L1 positive and negative tumors. Patients with moderately versus those with poorly or highly proliferative tumors have a superior DC rate when combined with any classification schema used to score PD-L1 as a positive result (i.e., TPS ≥ 50% or ≥ 1%), and best displayed by a DC rate for moderately proliferative tumors of no less than 40% for any classification of PD-L1 as a negative result. While there is an over representation of moderately proliferative tumors as PD-L1 expression increases this does not account for the improved survival or higher disease control rates seen in PD-L1 negative tumors. Conclusions Cell proliferation is potentially a new biomarker of response to ICIs in NSCLC and is applicable to PD-L1 negative tumors. Electronic supplementary material The online version of this article (10.1186/s40425-019-0506-3) contains supplementary material, which is available to authorized users.

[Background] PD-L1 immunohistochemistry (IHC) has been traditionally used for predicting clinical responses to immune checkpoint inhibitors (ICIs). However, there are at least 4 different assays and antibodies used for PD-L1 IHC, each developed with a different ICI. We set to test if next generation RNA sequencing (RNA-seq) is a robust method to determine PD-L1 mRNA expression levels and furthermore, efficacy of predicting response to ICIs as compared to routinely used, standardized IHC procedures., [Methods] A total of 209 cancer patients treated on-label by FDA-approved ICIs, with evaluable responses were assessed for PD-L1 expression by RNA-seq and IHC, based on tumor proportion score (TPS) and immune cell staining (ICS). A subset of serially diluted cases was evaluated for RNA-seq assay performance across a broad range of PD-L1 expression levels., [Results] Assessment of PD-L1 mRNA levels by RNA-seq demonstrated robust linearity across high and low expression ranges. PD-L1 mRNA levels assessed by RNA-seq and IHC (TPS and ICS) were highly correlated (p, [Conclusions] Measurement of PD-L1 mRNA expression by RNA-seq is comparable to PD-L1 expression by IHC both analytically and clinically in predicting ICI response. RNA-seq has the added advantages of being amenable to standardization and avoidance of interpretation bias. PD-L1 by RNA-seq needs to be validated in future prospective ICI clinical studies across multiple histologies., This research was funded by OmniSeq, Inc. (Buffalo, NY).

Historically, surgical pathology reports have been manually transcribed following dictation. More recently, voice recognition technology has been employed with or without the use of templates for common specimens. Required cancer reporting protocols are often combined with both manual transcription and voice recognition technology approaches. Errors observed in both approaches are discussed. Concurrently, there has been an evolution from manual/paper pathology report distribution into the patient’s physical chart to inclusion of the report in the patient’s comprehensive electronic health record. Data that crosses an electronic interface between a laboratory information system and an electronic health record is subject to loss of fidelity or formatting alterations which may render a final product different from what the pathologist originally intended. Inclusion of diagrams and/or photomicrographs with surgical pathology reports is possible in some systems, but can add a layer of complexity to successful electronic report distribution. Finally, some thought must be applied to the distribution of information within an individual pathology report for maximum interpretability. This chapter discusses errors that can arise in the transcription and distribution of diagnostic data in pathology reports. Strategies for assessment and prevention of these errors in real practice situations are provided.