RESUMEN
Several somatic mutations specific to aldosterone-producing adenomas (APAs) have been described. A small proportion of adrenocortical carcinomas (ACCs) are associated with hyperaldosteronism, either primary aldosteronism or hyperreninemic hyperaldosteronism. However, it is unknown whether they harbor mutations of the same spectrum as APAs. The objective of this study is to describe the clinical phenotype and molecular genotype of ACCs with hyperaldosteronism, particularly the analysis for common APA-associated genetic changes. Patients were identified by retrospective chart review at a specialized referral center and by positive staining for CYP11B2 of tissue microarrays. Twenty-five patients with ACC and hyperaldosteronism were initially identified by retrospective chart review, and tissue for further analysis was available on 13 tumors. Seven patients were identified by positive staining for CYP11B2 in a tissue microarray, of which two were already identified in the initial chart review. Therefore, a total number of 18 patients with a diagnosis of ACC and features of either primary aldosteronism or hyperreninemic hyperaldosteronism were therefore included in the final study. Mutational status for a select list of oncogenes, tumor suppressor genes and genes known to carry mutations in APAs were analyzed by next-generation sequencing. Review of clinical data suggested autonomous aldosterone production in the majority of cases, while for some cases, hyperreninemic hyperaldosteronism was the more likely mechanism. The mutational landscape of ACCs associated with hyperaldosteronism was not different from ACCs with a different hormonal phenotype. None of the ACCs harbored mutations of known APA-associated genes, suggesting an alternative mechanism conferring aldosterone production.
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Carcinoma Corticosuprarrenal/sangre , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Hiperaldosteronismo/etiología , Adulto , Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad , Mutación , Estudios Retrospectivos , Adulto JovenRESUMEN
BACKGROUND: Integrated molecular profiling has identified intrinsic expression-based bladder cancer molecular subtypes. Despite frequent histological diversity, robustness of subtypes in paired conventional (urothelial) and squamous components of the same bladder tumor has not been reported. OBJECTIVE: To assess the impact of histological heterogeneity on expression-based bladder cancer subtypes. DESIGN, SETTING, AND PARTICIPANTS: We performed clinically applicable, targeted DNA and/or RNA sequencing (multiplexed DNA and RNA sequencing [mxDNAseq and mxRNAseq, respectively]) on 112 formalin-fixed paraffin-embedded (FFPE) bladder cancer samples, including 12 cases with paired urothelial/squamous components and 21 bladder cancer cell lines. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Unsupervised hierarchical and consensus clustering of target gene expression enabled derivation of basal/luminal molecular subtyping. RESULTS AND LIMITATION: Across 21 bladder cancer cell lines, our custom mxRNAseq panel was highly concordant with whole transcriptome sequencing, and assessed targets robustly determined expression-based basal/luminal subtypes from The Cancer Genome Atlas data (in silico) and internally sequenced FFPE tissues. Frequent deleterious TP53 (56%) and activating hotspot PIK3CA (30%) somatic mutations were seen across 69 high-quality tissue samples. Potentially targetable focal ERBB2 (6%) or EGFR (6%) amplifications were also identified, and a novel subgene copy-number detection approach is described. Combined DNA/RNA analysis showed that focally amplified samples exhibit outlier EGFR and ERBB2 expression distinct from subtype-intrinsic profiles. Critically, paired urothelial and squamous components showed divergent basal/luminal status in three of 12 cases (25%), despite identical putatively clonal prioritized somatic genomic alterations. Limitations include lack of profiled paired normal tissues for formal somatic alteration determination, and the need for formal analytical and clinical validation. CONCLUSIONS: Our results support the feasibility of clinically relevant integrative bladder cancer profiling and challenge the intrinsic nature of expression subtypes in histologically diverse bladder cancers. PATIENT SUMMARY: A targeted RNA sequencing assay is capable of assessing gene expression-based subtypes in individual components of clinical bladder cancer tissue specimens. Different histological components of the same tumor may yield divergent expression profiles, suggesting that expression-based subtypes should be interpreted with caution in heterogeneous cancers.
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Biomarcadores de Tumor/genética , Carcinoma de Células Escamosas/genética , ADN de Neoplasias/genética , Heterogeneidad Genética , ARN Neoplásico/genética , Neoplasias de la Vejiga Urinaria/genética , Vejiga Urinaria/metabolismo , Urotelio/metabolismo , Biomarcadores de Tumor/metabolismo , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patología , Línea Celular Tumoral , Variaciones en el Número de Copia de ADN , Análisis Mutacional de ADN , ADN de Neoplasias/metabolismo , Amplificación de Genes , Perfilación de la Expresión Génica/métodos , Predisposición Genética a la Enfermedad , Genoma Humano , Genómica/métodos , Humanos , Mutación , Fenotipo , Valor Predictivo de las Pruebas , ARN Neoplásico/metabolismo , Reproducibilidad de los Resultados , Análisis de Secuencia de ARN , Transcriptoma , Vejiga Urinaria/patología , Neoplasias de la Vejiga Urinaria/metabolismo , Neoplasias de la Vejiga Urinaria/patología , Urotelio/patologíaAsunto(s)
Antígeno B7-H1/genética , Carcinoma de Células Escamosas , Receptor de Muerte Celular Programada 1/genética , Neoplasias de la Vejiga Urinaria , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patología , Regulación Neoplásica de la Expresión Génica , Humanos , Inmunohistoquímica , Vejiga Urinaria/patología , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/patologíaRESUMEN
BACKGROUND: Prognostic biomarkers for localized prostate cancer (PCa) could improve personalized medicine. Our group previously identified a panel of differentially methylated CpGs in primary tumor tissue that predict disease aggressiveness, and here we further validate these biomarkers. METHODS: Pyrosequencing was used to assess CpG methylation of eight biomarkers previously identified using the HumanMethylation450 array; CpGs with strongly correlated (r >0.70) results were considered technically validated. Logistic regression incorporating the validated CpGs and Gleason sum was used to define and lock a final model to stratify men with metastatic-lethal versus non-recurrent PCa in a training dataset. Coefficients from the final model were then used to construct a DNA methylation score, which was evaluated by logistic regression and Receiver Operating Characteristic (ROC) curve analyses in an independent testing dataset. RESULTS: Five CpGs were technically validated and all were retained (P < 0.05) in the final model. The 5-CpG and Gleason sum coefficients were used to calculate a methylation score, which was higher in men with metastatic-lethal progression (P = 6.8 × 10-6 ) in the testing dataset. For each unit increase in the score there was a four-fold increase in risk of metastatic-lethal events (odds ratio, OR = 4.0, 95%CI = 1.8-14.3). At 95% specificity, sensitivity was 74% for the score compared to 53% for Gleason sum alone. The score demonstrated better prediction performance (AUC = 0.91; pAUC = 0.037) compared to Gleason sum alone (AUC = 0.87; pAUC = 0.025). CONCLUSIONS: The DNA methylation score improved upon Gleason sum for predicting metastatic-lethal progression and holds promise for risk stratification of men with aggressive tumors. This prognostic score warrants further evaluation as a tool for improving patient outcomes.
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Solitary fibrous tumors (SFTs) of the head and neck are uncommon. Lesions previously diagnosed in the head and neck as hemangiopericytomas (HPCs), giant cell angiofibromas (GCAs), and orbital fibrous histiocytomas (OFHs) are now recognized as within the expanded spectrum of SFTs. To better understand the clinicopathologic profile of head and neck SFTs, we performed a multi-institutional study of 88 examples. There was no sex predilection (F:M ratio 1.2), and the median patient age was 52 years (range: 15 to above 89 y). The sinonasal tract and orbit were the most common sites involved (30% and 25%), followed by the oral cavity and salivary glands (15% and 14%). Original diagnoses included HPC (25%), SFT (67%), and OFH (6%), with 1 SFT and 1 OFH noted as showing GCA-like morphology. On review, the predominant histologic pattern was classic SFT-like in 53% and cellular (former HPC-like) in 47%; lipomatous differentiation (8%) and GCA-like pattern (7%) were less prevalent. Subsets demonstrated nuclear atypia (23%), epithelioid morphology (15%), or coagulative necrosis (6%). Infiltrative growth (49%) and osseous invasion (82%) were prevalent among evaluable cases. Of the 48 SFTs with follow-up (median: 43 mo), 19 showed recurrence (40%). Of these, 4 patients were alive with disease and 4 dead of disease. Size and mitotic rate were negative prognosticators using a joint prognostic proportional hazards regression model. Three patients experienced metastasis, to lungs, parotid, bone, and skull base, including one case showing overtly sarcomatous "dedifferentiation." As a group, SFTs present in a wide anatomic and morphologic spectrum in the head and neck. Only rare examples metastasize or cause death from disease. However, the fairly high local recurrence rate underscores their aggressive potential and highlights the importance of prospective recognition.
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Neoplasias de Cabeza y Cuello/patología , Tumores Fibrosos Solitarios/secundario , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/análisis , Proliferación Celular , Progresión de la Enfermedad , Supervivencia sin Enfermedad , Femenino , Neoplasias de Cabeza y Cuello/química , Neoplasias de Cabeza y Cuello/mortalidad , Neoplasias de Cabeza y Cuello/terapia , Humanos , Inmunohistoquímica , Estimación de Kaplan-Meier , Masculino , Persona de Mediana Edad , Índice Mitótico , Invasividad Neoplásica , Recurrencia Local de Neoplasia , Modelos de Riesgos Proporcionales , Estudios Retrospectivos , Factores de Riesgo , Tumores Fibrosos Solitarios/química , Tumores Fibrosos Solitarios/mortalidad , Tumores Fibrosos Solitarios/terapia , Factores de Tiempo , Tomografía Computarizada por Rayos X , Resultado del Tratamiento , Estados Unidos , Adulto JovenRESUMEN
OBJECTIVES: To use a non-biased assay for circulating tumour cells (CTCs) in patients with prostate cancer (PCa) in order to identify non-traditional CTC phenotypes potentially excluded by conventional detection methods that are reliant on antigen- and/or size-based enrichment. PATIENTS AND METHODS: A total of 41 patients with metastatic castration-resistant PCa (mCRPC) and 20 healthy volunteers were analysed on the Epic CTC platform, via high-throughput imaging of DAPI expression and CD45/cytokeratin (CK) immunofluorescence (IF) on all circulating nucleated cells plated on glass slides. To confirm the PCa origin of CTCs, IF was used for androgen receptor (AR) expression and fluorescence in situ hybridization was used for PTEN and ERG assessment. RESULTS: Traditional CTCs (CD45- /CK+ /morphologically distinct) were identified in all patients with mCRPC and we also identified CTC clusters and non-traditional CTCs in patients with mCRPC, including CK- and apoptotic CTCs. Small CTCs (≤white blood cell size) were identified in 98% of patients with mCRPC. Total, traditional and non-traditional CTCs were significantly increased in patients who were deceased vs alive after 18 months; however, only non-traditional CTCs were associated with overall survival. Traditional and total CTC counts according to the Epic platform in the mCRPC cohort were also significantly correlated with CTC counts according to the CellSearch system. CONCLUSIONS: Heterogeneous non-traditional CTC populations are frequent in mCRPC and may provide additional prognostic or predictive information.
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Metástasis de la Neoplasia/patología , Células Neoplásicas Circulantes/patología , Neoplasias de la Próstata Resistentes a la Castración/sangre , Neoplasias de la Próstata Resistentes a la Castración/patología , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/sangre , Progresión de la Enfermedad , Humanos , Hibridación Fluorescente in Situ , Masculino , Persona de Mediana Edad , Metástasis de la Neoplasia/genética , Fosfohidrolasa PTEN/sangre , Fosfohidrolasa PTEN/genética , Fenotipo , Antígeno Prostático Específico/sangre , Antígeno Prostático Específico/genética , Neoplasias de la Próstata Resistentes a la Castración/genética , Receptores Androgénicos/genéticaRESUMEN
Non-Hodgkin lymphoma of the orbit and ocular adnexa is the most common primary orbital malignancy. Treatments for low- (extra-nodal marginal zone and follicular lymphomas) and high-grade (diffuse large B-cell lymphoma) are associated with local and vision-threatening toxicities. High-grade lymphomas relapse frequently and exhibit poor survival rates. Despite advances in genomic profiling and precision medicine, orbital and ocular adnexal lymphomas remain poorly characterized molecularly. We performed targeted next-generation sequencing (NGS) profiling of 38 formalin-fixed, paraffin-embedded orbital and ocular adnexal lymphomas obtained from a single-center using a panel targeting near-term, clinically relevant genes. Potentially actionable mutations and copy number alterations were prioritized based on gain- and loss-of-function analyses, and catalogued, approved, and investigational therapies. Of 36 informative samples, including marginal zone lymphomas (n=20), follicular lymphomas (n=9), and diffuse large B-cell lymphomas (n=7), 53% harbored a prioritized alteration (median=1, range 0-5/sample). MYD88 was the most frequently altered gene in our cohort, with potentially clinically relevant hotspot gain-of-function mutations identified in 71% of diffuse large B-cell lymphomas and 25% of marginal zone lymphomas. Prioritized alterations in epigenetic modulators were common and included gain-of-function EZH2 and loss-of-function ARID1A mutations (14% of diffuse large B-cell lymphomas and 22% of follicular lymphomas contained alterations in each of these two genes). Single prioritized alterations were also identified in the histone methyltransferases KMT2B (follicular lymphoma) and KMT3B (diffuse large B-cell lymphoma). Loss-of-function mutations and copy number alterations in the tumor suppressors TP53 (diffuse large B-cell and follicular lymphoma), CDKN2A (diffuse large B-cell and marginal zone lymphoma), PTEN (diffuse large B-cell lymphoma), ATM (diffuse large B-cell lymphoma), and NF1 (diffuse large B-cell lymphoma), and gain-of-function mutations in the oncogenes HRAS (follicular lymphoma) and NRAS (diffuse large B-cell lymphoma) were also observed. Together, our study demonstrates that NGS can be used to profile routine formalin-fixed, paraffin-embedded orbital and ocular adnexal lymphomas for identification of somatic-driving alterations and nomination of potential therapeutic strategies.
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Biomarcadores de Tumor/genética , Neoplasias del Ojo/genética , Perfilación de la Expresión Génica , Linfoma/genética , Anciano , Anciano de 80 o más Años , Inhibidor p16 de la Quinasa Dependiente de Ciclina , Inhibidor p18 de las Quinasas Dependientes de la Ciclina/genética , Proteínas de Unión al ADN , Proteína Potenciadora del Homólogo Zeste 2/genética , Neoplasias del Ojo/patología , Femenino , Genómica , N-Metiltransferasa de Histona-Lisina/genética , Humanos , Linfoma/patología , Masculino , Persona de Mediana Edad , Mutación , Proteínas Nucleares/genética , Fosfohidrolasa PTEN/genética , Proteínas Proto-Oncogénicas p21(ras)/genética , Factores de Transcripción/genéticaRESUMEN
The presence of two or more prostate cancer foci separated by intervening benign tissue in a single core is a well-recognized finding on prostate biopsy. Cancer involvement can be measured by including intervening benign tissue or only including the actual cancer involved area. Importantly, this parameter is a common enrollment criterion for active surveillance protocols. We hypothesized that spatially distinct prostate cancer foci in biopsies may arise from separate clones, impacting cancer involvement assessment. Hence, we used dual ERG/SPINK1 immunohistochemistry to determine the frequency of separate clones-when separate tumor foci showed discordant ERG and/or SPINK1 status-in discontinuously involved prostate biopsy cores from two academic institutions. In our cohort of 97 prostate biopsy cores with spatially discrete tumor foci (from 80 patients), discontinuous cancer involvement including intervening tissue ranged from 20 to 100% and Gleason scores ranged from 6 to 9. Twenty-four (25%) of 97 discontinuously involved cores harbored clonally distinct cancer foci by discordant ERG and/or SPINK1 expression status: 58% (14/24) had one ERG(+) focus, and one ERG(-)/SPINK1(-) focus; 29% (7/24) had one SPINK1(+) focus and one ERG(-)/SPINK1(-) focus; and 13% (3/24) had one ERG(+) focus and one SPINK1(+) focus. ERG and SPINK1 overexpression were mutually exclusive in all tumor foci. In summary, our results show that ~25% of discontinuously involved prostate biopsy cores showed tumor foci with discordant ERG/SPINK1 status, consistent with multiclonal disease. The relatively frequent presence of multiclonality in discontinuously involved prostate biopsy cores warrants studies on the potential clinical impact of clonality assessment, particularly in cases where tumor volume in a discontinuous core may impact active surveillance eligibility.
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Adenocarcinoma/química , Biomarcadores de Tumor/análisis , Proteínas Portadoras/análisis , Inmunohistoquímica , Neoplasias de la Próstata/química , Transactivadores/análisis , Adenocarcinoma/patología , Biopsia con Aguja Gruesa , Células Clonales , Humanos , Masculino , Michigan , Clasificación del Tumor , Ciudad de Nueva York , Valor Predictivo de las Pruebas , Neoplasias de la Próstata/patología , Regulador Transcripcional ERG , Inhibidor de Tripsina Pancreática de KazalRESUMEN
Solitary fibrous tumor (SFT), a mesenchymal neoplasm with widespread anatomic distribution, can be diagnostically challenging in limited samples. We recently encountered an aspirate of a pancreatic mass, incorrectly interpreted as metastatic renal cell carcinoma based on strong PAX8 expression by immunohistochemistry (IHC). After resection, morphologic features with additional IHC (CD34 positivity) correctly identified this lesion as a SFT. PAX8 and PAX2 are commonly used as renal tumor markers; however, no series has investigated PAX8 or PAX2 expression in SFT. IHC for PAX8 and PAX2 was performed on 41 SFTs (biopsy and resections) from varying sites. Eight were histologically malignant and eight were recurrences of previous resections. PAX8 staining was observed at least focally in 26.8% (11 of 41) SFT cases; additionally, PAX2 was positive in 12.2% (5 of 41 cases) of SFTs. For PAX8 and PAX2 positive cases 45.6% and 40%, respectively, showed diffuse expression. No correlation was found between PAX8/PAX2 positivity and age, tumor size, site, malignancy, or recurrence. In conclusion, a substantial minority of SFTs express PAX8 and PAX2 via IHC. This presents a diagnostic pitfall when evaluating possible metastases from the kidney, particularly when primary tumors show sarcomatoid or spindle cell morphologies.
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Núcleo Celular/metabolismo , Factor de Transcripción PAX2/metabolismo , Factores de Transcripción Paired Box/metabolismo , Tumores Fibrosos Solitarios/metabolismo , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/metabolismo , Femenino , Humanos , Inmunohistoquímica , Neoplasias Renales/metabolismo , Masculino , Persona de Mediana Edad , Recurrencia Local de Neoplasia/metabolismo , Factor de Transcripción PAX8 , TranscriptomaRESUMEN
Expression of the costimulatory ligand PDL1 (B7-H1) on tumors allows escape from antitumor immunity. PDL1 expression has been reported in urothelial carcinoma (UC), suggesting it could be used as an immunotherapy target. We investigated the impact of cytotoxic neoadjuvant chemotherapy (NAC) on PDL1 expression in UC. Immunohistochemical staining for PDL1 was performed using an anti-B7-H1 monoclonal antibody (5H1) on a tissue microarray with matched pre-NAC and post-NAC UC samples from 40 patients treated between 1999 and 2011. PDL1 expression was significantly higher in post-NAC specimens than in matched pre-NAC specimens (p=0.0235, Wilcoxen's signed rank test). PDL1 expression in pre-NAC tissue did not correlate with clinical or pathologic stage but was associated with recurrence free survival. The increase in PDL1 expression following NAC in our limited cohort has potential to guide optimal combination and sequencing of immune and cytotoxic therapies in UC patients. PATIENT SUMMARY: We investigated the relationship between levels of the drug target PDL1 before and after chemotherapy in patients with bladder cancer. We found that levels of PDL1 increased following chemotherapy. We conclude that this information may help doctors in optimizing the sequence of therapies in bladder cancer.
RESUMEN
Penile squamous cell carcinoma (PeSCCA) is a rare malignancy for which there are limited treatment options due to a poor understanding of the molecular alterations underlying disease development and progression. Therefore, we performed comprehensive, targeted next-generation sequencing to identify relevant somatic genomic alterations in a retrospective cohort of 60 fixed tumor samples from 43 PeSCCA cases (including 14 matched primary/metastasis pairs). We identified a median of two relevant somatic mutations and one high-level copy-number alteration per sample (range, 0-5 and 0-6, respectively). Expression of HPV and p16 was detectable in 12% and 28% of patients, respectively. Furthermore, advanced clinical stage, lack of p16 expression, and MYC and CCND1 amplifications were significantly associated with shorter time to progression or PeSCCA-specific survival. Notably, four cases harbored EGFR amplifications and one demonstrated CDK4 amplification, genes for which approved and investigational targeted therapies are available. Importantly, although paired primary tumors and lymph node metastases were largely homogeneous for relevant somatic mutations, we identified heterogeneous EGFR amplification in primary tumor/lymph node metastases in 4 of 14 cases, despite uniform EGFR protein overexpression. Likewise, activating HRAS mutations occurred in 8 of 43 cases. Taken together, we provide the first comprehensive molecular PeSCCA analysis, which offers new insight into potential precision medicine approaches for this disease, including strategies targeting EGFR.
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Carcinoma de Células Escamosas/genética , Neoplasias del Pene/genética , Transcriptoma , Adulto , Anciano , Anciano de 80 o más Años , Carcinoma de Células Escamosas/mortalidad , Carcinoma de Células Escamosas/virología , Quinasa 4 Dependiente de la Ciclina/genética , Análisis Mutacional de ADN , Supervivencia sin Enfermedad , Amplificación de Genes , Genes erbB-1/genética , Genómica , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Inmunohistoquímica , Estimación de Kaplan-Meier , Masculino , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa Multiplex , Infecciones por Papillomavirus/complicaciones , Infecciones por Papillomavirus/epidemiología , Neoplasias del Pene/mortalidad , Neoplasias del Pene/virología , Estudios RetrospectivosRESUMEN
IMPORTANCE: High-grade serous carcinoma (HGSC) is the most prevalent and lethal form of ovarian cancer. HGSCs frequently arise in the distal fallopian tubes rather than the ovary, developing from small precursor lesions called serous tubal intraepithelial carcinomas (TICs, or more specifically, STICs). While STICs have been reported to harbor TP53 mutations, detailed molecular characterizations of these lesions are lacking. OBSERVATIONS: We performed targeted next-generation sequencing (NGS) on formalin-fixed, paraffin-embedded tissue from 4 women, 2 with HGSC and 2 with uterine endometrioid carcinoma (UEC) who were diagnosed as having synchronous STICs. We detected concordant mutations in both HGSCs with synchronous STICs, including TP53 mutations as well as assumed germline BRCA1/2 alterations, confirming a clonal association between these lesions. Next-generation sequencing confirmed the presence of a STIC clonally unrelated to 1 case of UEC, and NGS of the other tubal lesion diagnosed as a STIC unexpectedly supported the lesion as a micrometastasis from the associated UEC. CONCLUSIONS AND RELEVANCE: We demonstrate that targeted NGS can identify genetic alterations in minute lesions, such as TICs, and confirm TP53 mutations as early driving events for HGSC. Next-generation sequencing also demonstrated unexpected associations between presumed STICs and synchronous carcinomas, providing evidence that some TICs are actually metastases rather than HGSC precursors.
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Biomarcadores de Tumor/genética , Carcinoma in Situ/genética , Análisis Mutacional de ADN , Neoplasias de las Trompas Uterinas/genética , Pruebas Genéticas/métodos , Secuenciación de Nucleótidos de Alto Rendimiento , Mutación , Neoplasias Quísticas, Mucinosas y Serosas/genética , Neoplasias Ováricas/genética , Proteína BRCA1/genética , Proteína BRCA2/genética , Carcinoma in Situ/patología , Neoplasias de las Trompas Uterinas/secundario , Femenino , Predisposición Genética a la Enfermedad , Humanos , Micrometástasis de Neoplasia , Neoplasias Quísticas, Mucinosas y Serosas/secundario , Neoplasias Ováricas/patología , Fenotipo , Valor Predictivo de las Pruebas , Proteína p53 Supresora de Tumor/genéticaRESUMEN
Next-generation sequencing (NGS) has enabled genome-wide personalized oncology efforts at centers and companies with the specialty expertise and infrastructure required to identify and prioritize actionable variants. Such approaches are not scalable, preventing widespread adoption. Likewise, most targeted NGS approaches fail to assess key relevant genomic alteration classes. To address these challenges, we predefined the catalog of relevant solid tumor somatic genome variants (gain-of-function or loss-of-function mutations, high-level copy number alterations, and gene fusions) through comprehensive bioinformatics analysis of >700,000 samples. To detect these variants, we developed the Oncomine Comprehensive Panel (OCP), an integrative NGS-based assay [compatible with <20 ng of DNA/RNA from formalin-fixed paraffin-embedded (FFPE) tissues], coupled with an informatics pipeline to specifically identify relevant predefined variants and created a knowledge base of related potential treatments, current practice guidelines, and open clinical trials. We validated OCP using molecular standards and more than 300 FFPE tumor samples, achieving >95% accuracy for KRAS, epidermal growth factor receptor, and BRAF mutation detection as well as for ALK and TMPRSS2:ERG gene fusions. Associating positive variants with potential targeted treatments demonstrated that 6% to 42% of profiled samples (depending on cancer type) harbored alterations beyond routine molecular testing that were associated with approved or guideline-referenced therapies. As a translational research tool, OCP identified adaptive CTNNB1 amplifications/mutations in treated prostate cancers. Through predefining somatic variants in solid tumors and compiling associated potential treatment strategies, OCP represents a simplified, broadly applicable targeted NGS system with the potential to advance precision oncology efforts.
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Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Mutación/genética , Neoplasias/genética , Anciano , Quinasa de Linfoma Anaplásico , Biología Computacional/métodos , Análisis Mutacional de ADN/métodos , ADN de Neoplasias/genética , Receptores ErbB/genética , Femenino , Humanos , Masculino , Persona de Mediana Edad , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas B-raf/genética , Proteínas Proto-Oncogénicas p21(ras) , Proteínas Tirosina Quinasas Receptoras/genética , Estudios Retrospectivos , Serina Endopeptidasas/genética , Transactivadores/genética , Regulador Transcripcional ERG , beta Catenina/genética , Proteínas ras/genéticaRESUMEN
UNLABELLED: Phyllodes tumors are rare fibroepithelial tumors with variable clinical behavior accounting for a small subset of all breast neoplasms, yet little is known about the genetic alterations that drive tumor initiation and/or progression. Here, targeted next-generation sequencing (NGS) was used to identify somatic alterations in formalin-fixed paraffin-embedded (FFPE) patient specimens from malignant, borderline, and benign cases. NGS revealed mutations in mediator complex subunit 12 (MED12) affecting the G44 hotspot residue in the majority (67%) of cases spanning all three histologic grades. In addition, loss-of-function mutations in p53 (TP53) as well as deleterious mutations in the tumor suppressors retinoblastoma (RB1) and neurofibromin 1 (NF1) were identified exclusively in malignant tumors. High-level copy-number alterations (CNA) were nearly exclusively confined to malignant tumors, including potentially clinically actionable gene amplifications in IGF1R and EGFR. Taken together, this study defines the genomic landscape underlying phyllodes tumor development, suggests potential molecular correlates to histologic grade, expands the spectrum of human tumors with frequent recurrent MED12 mutations, and identifies IGF1R and EGFR as potential therapeutic targets in malignant cases. IMPLICATIONS: Integrated genomic sequencing and mutational profiling provides insight into the molecular origin of phyllodes tumors and indicates potential druggable targets in malignant disease. Visual Overview: http://mcr.aacrjournals.org/content/early/2015/04/02/1541-7786.MCR-14-0578/F1.large.jpg.
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Neoplasias de la Mama/genética , Receptores ErbB/genética , Complejo Mediador/genética , Mutación , Tumor Filoide/genética , Receptores de Somatomedina/genética , Neoplasias de la Mama/patología , Variaciones en el Número de Copia de ADN , Femenino , Amplificación de Genes , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Humanos , Neurofibromina 1/genética , Tumor Filoide/patología , Receptor IGF Tipo 1 , Proteína de Retinoblastoma/genética , Análisis de Secuencia de ADN/métodos , Proteína p53 Supresora de Tumor/genéticaRESUMEN
Although multifocal tumors and non-invasive/invasive components are commonly encountered in surgical pathology, their genetic relationship is often poorly characterized. We used next-generation sequencing (NGS) to characterize somatic alterations in a patient with five spatially distinct, high-grade papillary urothelial carcinomas (UCs), with one tumor harboring an underlying invasive component. NGS of 409 cancer-related genes was performed on DNA isolated from formalin-fixed paraffin-embedded (FFPE) blocks representing each papillary tumor (n = 5), the invasive component of one tumor, and matched normal tissue. We identified nine unique non-synonymous somatic mutations across the six UC samples, including five present in each carcinoma sample, consistent with clonal origin and limited intertumoral heterogeneity. Copy number and loss of heterogeneity (LOH) profiles were similar in all six carcinomas; however, the invasive carcinoma component uniquely showed focal CDKN2A loss and chromosome 9 LOH and did not harbor gains of chromosomes 5p or X that were present in the other tumor samples. Phylogenetic analysis supported the invasive component arising from a shared progenitor prior to the outgrowth of cells in the non-invasive tumors. Results were extended to three additional cases of upper tract UC with paired non-invasive/invasive components, which identified driving alterations exclusive to both non-invasive and invasive components. Lastly, we performed targeted RNA sequencing (RNAseq) using a custom bladder cancer panel, which confirmed gene expression signature differences between paired non-invasive/invasive components. The results and approaches presented here may be useful in understanding the clonal relationships in multifocal cancers or paired non-invasive/invasive components from routine FFPE specimens.
Asunto(s)
Carcinogénesis/patología , Carcinoma de Células Transicionales/patología , Progresión de la Enfermedad , Invasividad Neoplásica/patología , Neoplasias de la Vejiga Urinaria/patología , Urotelio/patología , Anciano , Anciano de 80 o más Años , Carcinogénesis/genética , Carcinoma de Células Transicionales/genética , Dosificación de Gen/genética , Genes p16 , Humanos , Pérdida de Heterocigocidad/genética , Masculino , Invasividad Neoplásica/genética , Oncogenes/genética , Filogenia , Neoplasias de la Vejiga Urinaria/genéticaRESUMEN
Inverted urothelial papilloma (IUP) is an uncommon neoplasm of the urinary bladder with distinct morphologic features. Studies regarding the role of human papillomavirus (HPV) in the etiology of IUP have provided conflicting evidence of HPV infection. In addition, little is known regarding the molecular alterations present in IUP or other urothelial neoplasms, which might demonstrate inverted growth pattern like low-grade or high-grade urothelial carcinoma (UCA). Here, we evaluated for the presence of common driving somatic mutations and HPV within a cohort of IUPs, (n = 7) noninvasive low-grade papillary UCAs with inverted growth pattern (n = 5), and noninvasive high-grade papillary UCAs with inverted growth pattern (n = 8). HPV was not detected in any case of IUP or inverted UCA by either in situ hybridization or by polymerase chain reaction. Next-generation sequencing identified recurrent mutations in HRAS (Q61R) in 3 of 5 IUPs, described for the first time in this neoplasm. Additional mutations of Ras pathway members were detected including HRAS, KRAS, and BRAF. The presence of Ras pathway member mutations at a relatively high rate suggests this pathway may contribute to pathogenesis of inverted urothelial neoplasms. In addition, we did not find any evidence supporting a role for HPV in the etiology of IUP.
Asunto(s)
Biomarcadores de Tumor/genética , Carcinoma de Células Transicionales/genética , Papiloma Invertido/genética , Proteínas Proto-Oncogénicas p21(ras)/genética , Neoplasias de la Vejiga Urinaria/genética , Anciano , Anciano de 80 o más Años , Sustitución de Aminoácidos , Carcinoma de Células Transicionales/patología , Estudios de Cohortes , Femenino , Dosificación de Gen , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Hibridación in Situ , Masculino , Persona de Mediana Edad , Mutación , Papiloma Invertido/patología , Papillomaviridae/genética , Papillomaviridae/aislamiento & purificación , Infecciones por Papillomavirus/virología , Análisis de Secuencia de ADN , Neoplasias de la Vejiga Urinaria/patologíaRESUMEN
Nephrogenic adenoma (NA) is a benign lesion of the urinary tract associated with injury to the urothelium. The varied morphologic patterns of NA make it a potential diagnostic pitfall, because it can mimic closely prostatic adenocarcinoma and urothelial carcinoma. In current practice, an antibody cocktail comprising p63, CK903, and AMACR (PIN-4 cocktail) is frequently utilized to evaluate foci suspicious for prostatic adenocarcinoma. Although the staining characteristics of the individual components of the PIN-4 cocktail have been reported for NA, no study has described the expression patterns for NA when the stains are applied as a cocktail. GATA-3 is an emerging marker of urothelial carcinoma; however, the GATA-3 staining characteristics of NA have yet to be described. Sixty-three NA specimens (M:F=36:27, average age=51.4 y) from various locations in the urinary tract including urinary bladder (n=40), ureter (n=3), and urethra (n=20) were collected from the archives of 2 institutions. Immunohistochemical analysis with the PIN-4 cocktail and GATA-3 antibodies was performed, and the distribution and intensity of staining was recorded for each antibody in each case. PIN-4 cocktail staining revealed AMACR expression in 56% of cases, CK903 expression in 97%, and rare p63 positivity (in only 2 cases). Only 2 NA cases displayed an overall PIN-4 staining pattern compatible with prostate cancer. GATA-3 expression was noted in 40% of NAs. No correlation between AMACR, CK903, or GATA-3 positivity and histologic pattern or anatomic location was identified. Although heterogenous staining patterns were seen within individual cases, use of the PIN-4 cocktail effectively discriminates NA from prostate cancer because of the high frequency of coexpression of AMACR and CK903 within NA. In addition, GATA-3 is not a useful marker in differentiating between NA and urothelial carcinoma.
Asunto(s)
Adenocarcinoma/diagnóstico , Adenoma/diagnóstico , Biomarcadores de Tumor/análisis , Neoplasias Urológicas/diagnóstico , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Niño , Diagnóstico Diferencial , Femenino , Factor de Transcripción GATA3/análisis , Humanos , Inmunohistoquímica , Masculino , Proteínas de la Membrana/análisis , Persona de Mediana Edad , Racemasas y Epimerasas/análisis , Adulto JovenRESUMEN
Salivary duct carcinoma is a highly aggressive salivary gland malignancy that may be misdiagnosed as high-grade mucoepidermoid carcinoma. We utilized tissue microarrays with 78 examples of mucoepidermoid carcinoma and 47 salivary duct carcinomas to evaluate the utility of an immunohistochemical panel consisting of androgen receptor, Her2/neu, p63, and cytokeratin 5/6 in distinguishing these entities. Among all cases in the cohorts, androgen receptor was highly specific for salivary duct carcinoma, while cytokeratin 5/6 and p63 were specific for mucoepidermoid carcinoma. While the rate of unequivocal Her2/neu overexpression among the salivary duct carcinomas was low (8.9 %), discrimination of salivary duct carcinoma was enhanced when this marker was used in combination with androgen receptor due to profound sensitivity. The immunohistochemical panel was particularly efficacious at distinguishing the problematic subset of high-grade mucoepidermoid carcinomas from salivary duct carcinoma. Utilization of this set of immunohistochemical markers allows reliable differentiation of salivary duct and mucoepidermoid carcinoma, a distinction with important prognostic and therapeutic implications.
Asunto(s)
Biomarcadores de Tumor/análisis , Carcinoma Mucoepidermoide/diagnóstico , Conductos Salivales/patología , Neoplasias de las Glándulas Salivales/diagnóstico , Adulto , Anciano , Diagnóstico Diferencial , Femenino , Humanos , Inmunohistoquímica , Masculino , Persona de Mediana Edad , Sensibilidad y Especificidad , Análisis de Matrices TisularesRESUMEN
CONTEXT: In 2006, the department of pathology at our institution established an intramural research funding mechanism to support anatomic pathology research projects for faculty and trainee development. A review committee consisting of faculty members with diverse academic interests evaluated applications; proposals were eligible for a maximum award amount of $30 000 per project with a maximum program cost of $150 000 annually. OBJECTIVE: To report our experience based on a retrospective review of the research proposals submitted to the committee since the inception of the Anatomic Pathology Research Fund and evaluate the outcomes of the funded projects. DESIGN: We retrospectively analyzed all project applications that were received by the committee. Outcome data were collected by reviewing progress reports, abstracts for national and international meetings, PubMed search results, and/or direct communication with investigators. RESULTS: To date, a total of 59 individual projects have been awarded funding, for a total amount of $349 792, with an average award amount of $5381 per project. A total of 26 faculty members have secured funding for projects through this mechanism, and 27 resident and fellow trainees have been engaged in the funded projects. Spanning 11 subspecialty disciplines in anatomic pathology, 32 abstracts (54%) have been presented at national meetings and 26 (44%) have been published in the peer-reviewed literature to date. One project generated data used to secure an extramural (R01) grant. CONCLUSIONS: Our funding mechanism could serve as a model used by other academic departments to support research activities, thereby fostering faculty development through scholarly activities.