Concordance analysis between liquid biopsy (ctDNA) and tumor DNA molecular profiles from panel-based next-generation sequencing.

INTRODUCTION: Analysis of circulating tumor DNA (ctDNA), also known as liquid biopsy, has been postulated to be a useful test in the prognostication, molecular profiling, and monitoring of cancer patients. In this series we aimed to analyze the concordance between the mutation status of formalin-fixed paraffin-embedded (FFPE) tumor samples and matched ctDNA, considering tumor molecular profiling as the gold standard technique. METHODS: This retrospective study included cancer patients with complete diagnostics and gene mutations detected in a previous FFPE tumor tissue Next-Generation Sequencing (NGS) study with a matched frozen plasma sample available for an NGS ctDNA assay. RESULTS AND DISCUSSION: Sixty patients were included, 24 with colorectal carcinoma (CRC) and 36 with non-small cell lung cancer (NSCLC). In 27.1% of ctDNA studies a new mutation not previously detected in the matched tumor was found. 11.9% of these ctDNA results had the potential to impact clinical management. Globally, the concordance rate between FFPE tumor samples and ctDNA was 44.4%. When tumors were stratified by stage, the concordance was 76.5%, 70%, 36.4%, and 0% in tumor stages IV, III, II, and I, respectively. ctDNA molecular profiles showed a good concordance rate in advanced stage tumors and identified undetected mutations in tumor tissues. In early tumor stages the concordance was low, casting doubt on the usefulness of ctDNA in these patients.

Canalization technique to obtain deep tissue biopsy of gastrointestinal subepithelial tumors as an alternative to conventional known techniques.

BACKGROUND AND OBJECTIVES: The most accurate technology to detect and diagnose subepithelial tumors (SETs) is the endoscopic ultrasonography (EUS) combined with puncture techniques, such as the endoscopic ultrasonography-guided fine-needle aspiration (EUS-FNA) or the endoscopic ultrasonography-guided fine-needle biopsy. Going further in the improvement of the results of tumor samples obtained endoscopically to diagnose the SETs, the canalization technique guided by miniprobes (MPs) to obtain biopsies of SET could be an alternative to EUS-FNA. The objective of this study is to analyze the results of samples obtained by this procedure. MATERIALS AND METHODS: A multicenter, retrospective study of a review of a database of 32 consecutive patients with a SET in the digestive tract, from 2000 to 2015 was conducted. All patients underwent EUS-performed by MP, to define the size, internal echostructure, and layer of origin of tumor. Once the echostructure was defined, it proceeded to the canalization technique to arrive to the tumor tissue. RESULTS: The average diameter of SETs in this series (32 patients) was about 21.6±11 mm (range: 5-41 mm). The diagnostic accuracy was 28/32, 87.50% (Confidence interval 95%: 76.04%-98.99%), and there were no major complications. All procedures were performed on outpatients, none of which required additional hospitalization. The 50% of patients were operated or endoscopically resected and in all cases, the previous pathological diagnosis was confirmed. CONCLUSIONS: This is a feasible, safe, and effective procedure that allows to access to inside of SET to obtain deep biopsies. Tumor samples obtained by deep biopsy, with prior performing of the canalization technique guided by MP, were sufficient for histopathological and immunohistochemical diagnosis and similar to those obtained with other known methods (FNA Trucut, ProCore®, etc.). However, more prospective comparative studies with a larger number of patients and different specialists carrying out the procedure to reach a higher statistical significance are necessary.

Protocolo de estudio molecular del oncogén HER2/neu en el carcinoma de mama / Molecular protocol for HER2/neu analysis in breast carcinoma

Antecedentes. El oncogén HER2/neu es un determinante biológico predictivo esencial en el cáncer de mama por su papel como diana terapéutica del trastuzumab. No obstante, no se ha llegado aún a consensuar un método de referencia para su estudio. Métodos. En este trabajo, hemos aplicado y comparado, sobre 222 carcinomas de mama recopilados prospectivamente, los dos ensayos de uso más frecuente para determinar el estado de HER2/neu: inmunohistoquímica para expresión de la proteína p185 empleando dos anticuerpos monoclonales (CB11 y TAB250) e hibridación in situ fluorescente (FISH), utilizando secciones de tejido incluido en parafina. Resultados. Los resultados demuestran mayor sensibilidad y especificidad con CB11 (62,5% y 93,4% respectivamente) que con TAB250 (40% y 76,4% respectivamente). El estudio inmunohistoquímico con CB11 es adecuado como prueba inicial para predecir el estado del gen, proporciona un alto valor predictivo negativo (95,5%) en los casos de expresión débil o nula y positivo (96,2%) en los tumores con sobreexpresión. Si la expresión inmunohistoquímica es moderada debe considerarse no concluyente y es indispensable el estudio de FISH. También es recomendable aplicar FISH si hay discordancia entre la expresión de p185 y el perfil morfológico y/o molecular tumoral. Finalmente, aunque la tasa de falsos positivos producidos por estudios inmunohistoquímicos es inferior al 5%, debido a la toxicidad y coste de la terapia con trastuzumab es razonable considerar el empleo sistemático de FISH antes de indicar el tratamiento. Conclusión. En base a nuestros resultados proponemos un protocolo para el estudio molecular de HER2/neu

Background. The HER2/neu proto-oncogene is frequently over-expressed in breast cancer and serves as a biological target for trastuzumab therapy. However, there is no consensus regarding the technical aspects to be used to define HER2/neu status in clinical practice. Methods. The present study was conducted to address this critical issue by prospectively analysing a large cohort of breast cancer patients (n=222) and using a variety of methods. To define HER2/neu expression, detection of its encoded protein (p185) was performed by comparative immunohistochemical (IHC) analysis using two mouse monoclonal antibodies (mAb CB11 and mAb TAB250). To assess HER2/neu gene amplification, fluorescent in situ hybridisation (FISH) assays with gene-specific probes were conducted. All procedures were applied to de-paraffinised tissue sections of breast tumour samples. Results. Results showed that mAb CB11 had increased sensitivity and specificity (62.5% and 93.4%, respectively) compared to mAb TAB250 (40% and 76.4%, respectively) in defining HER2/neu amplification. We conclude that HER2/neu measurement by IHC using mAb CB11 is an appropriate strategy which provides a high negative predictive value (95.5%) for HER2/neu amplification in cases with low or undetectable p185 expression. Conversely, mAb CB11 has a high positive predictive value (96.2%) for HER2/neu amplification in cases with p185 overexpression. However, cases with moderate p185 expression need to be considered as inconclusive. In such cases, it is necessary to use FISH measurement to evaluate HER2/neu amplification. It is also advisable to conduct FISH if there is discordance between p185 expression and the histopathology classification of the lesion, or molecular profile of the tumour. Finally, even though the false positive rate of IHC assay is <5%, the toxicity and cost of trastuzumab therapy suggest that FISH be used systematically prior to implementation of treatment. Conclusion. We suggest the use of a molecular protocol for HER2/neu analysis in this type of tumor