RESUMEN
OBJECTIVE: Whether gastric metaplasia (GM) of the oesophagus should be considered as Barrett's oesophagus (BO) is controversial. Given concern intestinal metaplasia (IM) may be missed due to sampling, the UK guidelines include GM as a type of BO. Here, we investigated whether the risk of misdiagnosis and the malignant potential of GM warrant its place in the UK surveillance. DESIGN: We performed a thorough pathology and endoscopy review to follow clinical outcomes in a novel UK cohort of 244 patients, covering 1854 person years of follow-up. We complemented this with a comparative genomic analysis of 160 GM and IM specimens, focused on early molecular hallmarks of BO and oesophageal adenocarcinoma (OAC). RESULTS: We found that 58 of 77 short-segment (<3 cm) GM (SS-GM) cases (75%) continued to be observed as GM-only across a median of 4.4 years of follow-up. We observed that disease progression in GM-only cases and GM+IM cases (cases with reported GM on some occasions, IM on others) was significantly lower than in the IM-only cases (Kaplan-Meier, p=0.03). Genomic analysis revealed that the mutation burden in GM is significantly lower than in IM (p<0.01). Moreover, GM does not bear the mutational hallmarks of OAC, with an absence of associated signatures and driver gene mutations. Finally, we established that GM found adjacent to OAC is evolutionarily distant from cancer. CONCLUSION: SS-GM is a distinct entity from SS-IM and the malignant potential of GM is lower than IM. It is questionable whether SS-GM warrants inclusion in BO surveillance.
Asunto(s)
Adenocarcinoma , Esófago de Barrett , Neoplasias Esofágicas , Humanos , Esófago de Barrett/diagnóstico , Esófago de Barrett/genética , Esófago de Barrett/complicaciones , Neoplasias Esofágicas/diagnóstico , Neoplasias Esofágicas/genética , Metaplasia , Endoscopía GastrointestinalAsunto(s)
Adenocarcinoma/cirugía , Biomarcadores de Tumor/sangre , ADN Tumoral Circulante/sangre , Resección Endoscópica de la Mucosa , Neoplasias Esofágicas/cirugía , Esofagectomía , Medicina de Precisión , Secuenciación Completa del Genoma , Adenocarcinoma/sangre , Adenocarcinoma/genética , Adenocarcinoma/mortalidad , Biomarcadores de Tumor/genética , ADN Tumoral Circulante/genética , Toma de Decisiones Clínicas , Supervivencia sin Enfermedad , Resección Endoscópica de la Mucosa/efectos adversos , Resección Endoscópica de la Mucosa/mortalidad , Neoplasias Esofágicas/sangre , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/mortalidad , Esofagectomía/efectos adversos , Esofagectomía/mortalidad , Humanos , Recurrencia Local de Neoplasia , Neoplasia Residual , Valor Predictivo de las Pruebas , Reproducibilidad de los Resultados , Estudios Retrospectivos , Factores de TiempoRESUMEN
Two-pore domain potassium (K2P) channels carry background (or leak) potassium current and play a key role in regulating resting membrane potential and cellular excitability. Accumulating evidence points to a role for K2Ps in human pathophysiologies, most notably in pain and migraine, making them attractive targets for therapeutic intervention. However, there remains a lack of selective pharmacological tools. The aim of this work was to apply a "target class" approach to investigate the K2P superfamily and identify novel activators across all the described subclasses of K2P channels. Target class drug discovery allows for the leveraging of accumulated knowledge and maximizing synergies across a family of targets and serves as an additional approach to standard target-based screening. A common assay platform using baculovirus (BacMam) to transiently express K2P channels in mammalian cells and a thallium flux assay to determine channel activity was developed, allowing the simultaneous screening of multiple targets. Importantly, this system, by allowing precise titration of channel function, allows optimization to facilitate the identification of activators. A representative set of channels (THIK-1, TWIK-1, TREK-2, TASK-3, and TASK-2) were screened against a library of Food and Drug Administration (FDA)-approved compounds and the LifeArc Index Set. Activators were then analyzed in concentration-response format across all channels to assess selectivity. Using the target class approach to investigate the K2P channels has enabled us to determine which of the K2Ps are amenable to small-molecule activation, de-risk multiple channels from a technical point of view, and identify a diverse range of previously undescribed pharmacology.
Asunto(s)
Ensayos Analíticos de Alto Rendimiento , Canales de Potasio de Dominio Poro en Tándem/genética , Bibliotecas de Moléculas Pequeñas/farmacología , Baculoviridae/genética , Baculoviridae/metabolismo , Línea Celular Tumoral , Clonación Molecular , Descubrimiento de Drogas/métodos , Células Epiteliales/citología , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Expresión Génica , Vectores Genéticos/química , Vectores Genéticos/metabolismo , Humanos , Transporte Iónico , Potasio/metabolismo , Canales de Potasio de Dominio Poro en Tándem/agonistas , Canales de Potasio de Dominio Poro en Tándem/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Talio/metabolismoRESUMEN
Esophageal adenocarcinoma (EAC) is a poor-prognosis cancer type with rapidly rising incidence. Understanding of the genetic events driving EAC development is limited, and there are few molecular biomarkers for prognostication or therapeutics. Using a cohort of 551 genomically characterized EACs with matched RNA sequencing data, we discovered 77 EAC driver genes and 21 noncoding driver elements. We identified a mean of 4.4 driver events per tumor, which were derived more commonly from mutations than copy number alterations, and compared the prevelence of these mutations to the exome-wide mutational excess calculated using non-synonymous to synonymous mutation ratios (dN/dS). We observed mutual exclusivity or co-occurrence of events within and between several dysregulated EAC pathways, a result suggestive of strong functional relationships. Indicators of poor prognosis (SMAD4 and GATA4) were verified in independent cohorts with significant predictive value. Over 50% of EACs contained sensitizing events for CDK4 and CDK6 inhibitors, which were highly correlated with clinically relevant sensitivity in a panel of EAC cell lines and organoids.
Asunto(s)
Adenocarcinoma/genética , Biomarcadores de Tumor/genética , Neoplasias Esofágicas/genética , Estudios de Cohortes , Variaciones en el Número de Copia de ADN/genética , Exoma/genética , Femenino , Perfilación de la Expresión Génica/métodos , Regulación Neoplásica de la Expresión Génica/genética , Genómica/métodos , Humanos , Masculino , Mutación/genéticaRESUMEN
Two-pore domain potassium channels (K2Ps) are characterized by their four transmembrane domain and two-pore topology. They carry background (or leak) potassium current in a variety of cell types. Despite a number of important roles there is currently a lack of pharmacological tools with which to further probe K2P function. We have developed a cell-based thallium flux assay, using baculovirus delivered TASK3 (TWIK-related acid-sensitive K+ channel 3, KCNK9, K2P9.1) with the aim of identifying novel, selective TASK3 activators. After screening a library of 1000 compounds, including drug-like and FDA approved molecules, we identified Terbinafine as an activator of TASK3. In a thallium flux assay a pEC50 of 6.2 ( ±0.12) was observed. When Terbinafine was screened against TASK2, TREK2, THIK1, TWIK1 and TRESK no activation was observed in thallium flux assays. Several analogues of Terbinafine were also purchased and structure activity relationships examined. To confirm Terbinafine's activation of TASK3 whole cell patch clamp electrophysiology was carried out and clear potentiation observed in both the wild type channel and the pathophysiological, Birk-Barel syndrome associated, G236R TASK3 mutant. No activity at TASK1 was observed in electrophysiology studies. In conclusion, we have identified the first selective activator of the two-pore domain potassium channel TASK3.
