Comprehensive NGS Panel Validation for the Identification of Actionable Alterations in Adult Solid Tumors.

The increasing identification of driver oncogenic alterations and progress of targeted therapies addresses the need of comprehensive alternatives to standard molecular methods. The translation into clinical practice of next-generation sequencing (NGS) panels is actually challenged by the compliance of high quality standards for clinical accreditation. Herein, we present the analytical and clinical feasibility study of a hybridization capture-based NGS panel (Action OncoKitDx) for the analysis of somatic mutations, copy number variants (CNVs), fusions, pharmacogenetic SNPs and Microsatellite Instability (MSI) determination in formalin-fixed paraffin-embedded (FFPE) tumor samples. A total of 64 samples were submitted to extensive analytical validation for the identification of previously known variants. An additional set of 166 tumor and patient-matched normal samples were sequenced to assess the clinical utility of the assay across different tumor types. The panel demonstrated good specificity, sensitivity, reproducibility, and repeatability for the identification of all biomarkers analyzed and the 5% limit of detection set was validated. Among the clinical cohorts, the assay revealed pathogenic genomic alterations in 97% of patient cases, and in 82.7%, at least one clinically relevant variant was detected. The validation of accuracy and robustness of this assay supports the Action OncoKitDx's utility in adult solid tumors.

Gastrointestinal stromal tumors (GISTs): role of CD 117 and PDGFRA Golgi-like staining pattern in the recognition of mutational status.

AIMS: to determine whether potential correlations between CD117 and PDGFRA might serve as an indication for targeted therapies. MATERIAL AND METHODS: immunohistochemical expression of CD117 and PDGFRA was evaluated in 99 paraffin-embedded GISTs in conjunction with KIT and PDGFRA mutational status. RESULTS: CD117-positive staining was noted in 93 out of 99 cases. The predominant staining pattern was cytoplasmic, either with or without membrane accentuation; in 44.5% of cases, a clear Golgi-like pattern was evident. Correlations were found between KIT mutation and both CD117 expression (p = 0.006) and Golgi-like pattern (p = 0.026). Cytoplasmic PDGFRA-positive staining was detected in 87% of cases, both with and without membrane accentuation; in 8% cases an evident Golgi-like staining pattern was observed. A significant correlation was noted between PDGFRA mutations and Golgi-like staining pattern (p = 0.001). Moreover, 95% of PDGFRA-positive GISTs were also CD117-positive, suggesting that expression of the two markers is not mutually exclusive; most of these had mutations in KIT exon 11. PDGFRA-positive/CD117-negative tumors had mutations in PDGFRA, mainly in exon 18. PDGFRA-negative/CD117-negative staining was observed in 15% of cases, all of which displayed mutations in KIT exon 11. CD117-positive/PDGFRA-negative cases were characterized by mutations in KIT, mainly in exon 11. CONCLUSIONS: CD117 and PDGFRA staining are not exclusive, and the presence of a Golgi-like staining pattern for either, whilst not pathognomonic, is highly suggestive of KIT and PDGFRA mutated GISTs, respectively, and may be used with some reservations as an alternative indication for prescribing targeted therapies.

Tumores del estroma gastrointestinal (GISTs): patrón de tinción tipo Golgi de CD 117 Y PDGFRA en el reconocimiento del estado mutacional / Gastrointestinal stromal tumors (GISTs): role of CD 117 and PDGFRA Golgi-like staining pattern in the recognition of mutational status

Objetivo: determinar si las posibles correlaciones entre CD117 y PDGFRA podrían servir como una indicación de terapias dirigidas. Material y métodos: la expresión inmunohistoquímica de CD117 y PDGFRA se evaluó en 99 GIST incluidos en parafina en conjunción con el estado mutacional de KIT y PDGFRA Resultados: se observó tinción CD117-positivo en 93 de los 99 casos. El patrón de tinción predominante fue citoplasmático o de membrana; en el 44,5% de los casos, se evidencio patrón de tipo Golgi. Se encontraron correlaciones entre la mutación KIT tanto con la expresión de CD117 (p = 0,006) y con el patrón tipo Golgi (p = 0,026). Se detectó tinción citoplasmática PDGFRA-positiva en el 87% de los casos, con y sin acentuación de membrana, en el 8% se observó patrón de tinción tipo Golgi. Se observó una correlación significativa entre las mutaciones PDGFRA y el patrón de tinción tipo Golgi (p = 0,001). Por otra parte, el 95% de los GIST PDGFRA positivos también fueron CD117-positivo, lo que sugiere que la expresión de los dos marcadores no se excluyen mutuamente, la mayoría de ellos tenían mutaciones en el exón 11 de KIT. Los tumores PDGFRA-positivo/CD117-negativo tenían mutaciones en PDGFRA, principalmente en el exón 18. Se observó tinción PDGFRA-negativo/CD117-negativo en el 15% de los casos, todos los cuales revelaban mutaciones en el exón 11 de KIT. Los casos CD117-positivo/PDGFRA-negativo casos se caracteriza por mutaciones en KIT, principalmente en el exón 11. Conclusiones: las tinciones CD117 y PDGFRA no son excluyentes, y la presencia de un patrón de tinción de Golgi, aunque no es patognomónica, es altamente sugestiva de GIST mutado en KIT y PDGFRA, respectivamente, y se puede utilizar con algunas reservas, como una indicación alternativa para la prescripción de terapias dirigidas(AU)

Aims: determine whether potential correlations between CD117 to and PDGFRA might serve as an indication for targeted therapies. Material and methods: immunohistochemical expression of CD117 and PDGFRA was evaluated in 99 paraffin-embedded GISTs in conjunction with KIT and PDGFRA mutational status. Results: CD117-positive staining was noted in 93 out of 99 cases. The predominant staining pattern was cytoplasmic, either with or without membrane accentuation; in 44.5% of cases, a clear Golgi-like pattern was evident. Correlations were found be - tween KIT mutation and both CD117 expression (p = 0.006) and Golgi-like pattern (p = 0.026). Cytoplasmic PDGFRA-positive staining was detected in 87% of cases, both with and without membrane accentuation; in 8% cases an evident Golgi-like staining pattern was observed. A significant correlation was noted between PDGFRA mutations and Golgi-like staining pattern (p = 0.001). Moreover, 95% of PDGFRA-positive GISTs were also CD117- positive, suggesting that expression of the two markers is not mutually exclusive; most of these had mutations in KIT exon 11. PDGFRA-positive/CD117-negative tumors had mutations in PDGFRA, mainly in exon 18. PDGFRA-negative/CD117-negative staining was observed in 15% of cases, all of which displayed mutations in KIT exon 11. CD117-positive/PDGFRA-negative cases were characterized by mutations in KIT, mainly in exon 11. Conclusions: CD117 and PDGFRA staining are not exclusive, and the presence of a Golgi-like staining pattern for either, whilst not pathognomonic, is highly suggestive of KIT and PDGFRA mutated GISTs, respectively, and may be used with some reservations as an alternative indication for prescribing targeted therapies(AU)

Molecular findings of nasopharyngeal carcinoma in a European population.

OBJECTIVE: To explore biomolecular characteristics of a group of patients with nasopharyngeal carcinoma from European (Spanish) hospitals, addressing the pathogenesis of the tumor and the response to treatment. STUDY DESIGN: Cyclin D1 and p16 expression were evaluated immunohistochemically in 33 tissue samples of nasopharyngeal carcinoma. CCDN1 gene amplification and p16 gene deletion were studied by fluorescence in situ hybridization. Patient clinical data were examined, and tissues were evaluated histologically using hematoxylin-eosin staining. RESULTS: Cyclin D1 overexpression was found in 19 cases, and p16 expression was undetected in 30 cases. An association was observed between impaired p16 expression and cyclin D1 overexpression (p = 0.034). Eleven patients displayed p16 gene deletion and CCDN1 gene amplification. CONCLUSION: Cyclin D1 overexpression and CCDN1 amplification, loss of p16 expression and p16 deletion may be among the genetic alterations involved in the pathogenesis of nasopharyngeal carcinoma.