Probes for KRAS

Development of cribriform morphology (CM) heralds malignant change in human colon but lack of mechanistic understanding hampers preventive therapy. This study investigated CM pathobiology in three-dimensional (3D) Caco-2 culture models of colorectal glandular architecture, assessed translational relevance and tested effects of 1,25(OH)2D3,theactive form of vitamin D. CM evolution was driven by oncogenic perturbation of the apical polarity (AP) complex comprising PTEN, CDC42 and PRKCZ (phosphatase and tensin homolog, cell division cycle 42 and protein kinase C zeta). Suppression of AP genes initiated a spatiotemporal cascade of mitotic spindle misorientation, apical membrane misalignment and aberrant epithelial configuration. Collectively, these events promoted "Swiss cheese-like" cribriform morphology (CM) comprising multiple abnormal "back to back" lumens surrounded by atypical stratified epithelium, in 3D colorectal gland models. Intestinal cancer driven purely by PTEN-deficiency in transgenic mice developed CM and in human CRC, CM associated with PTEN and PRKCZ readouts. Treatment of PTEN-deficient 3D cultures with 1,25(OH)2D3 upregulated PTEN, rapidly activated CDC42 and PRKCZ, corrected mitotic spindle alignment and suppressed CM development. Conversely, mutationally-activated KRAS blocked1,25(OH)2D3 rescue of glandular architecture. We conclude that 1,25(OH)2D3 upregulates AP signalling to reverse CM in a KRAS wild type (wt), clinically predictive CRC model system. Vitamin D could be developed as therapy to suppress inception or progression of a subset of colorectal tumors.

Abstract

BACKGROUND & AIMS:

The present study aimed to elucidate the clinicopathological significance of IL-6 and IL-33 expression in intrahepatic cholangiocarcinomas (iCCAs) and perihilar cholangiocarcinomas (pCCAs).

METHODS:

IL-6 and IL-33 mRNA expression was examined in iCCAs (n=55) and pCCAs (n=32) using quantitative real-time PCR and a highly sensitive in situ hybridization protocol (RNAscope™ ), and expression values were correlated with clinicopathological features. According to a recently proposed classification scheme, iCCAs were separated into small- (n=33) and large-duct types (n=22).

RESULTS:

IL-6 and IL-33 expression levels were higher in large-duct iCCAs and pCCAs than in small-duct iCCAs, with a positive correlation between the values of these cytokines. In double in situ hybridization/immunostaining, IL-6 mRNA was expressed in actin-positive (myo)fibroblasts, while IL-33 was mainly produced by CD31-positive endothelial cells. Based on the average expression value as a cut-off point, cases were classified as IL-6high and IL-6low or IL-33high and IL-33low . In the combined cohort of large-duct iCCAs and pCCAs, IL-6high and IL-6low cholangiocarcinomas shared many features, while IL-33high cases had less aggressive characteristics than IL-33low cases as evidenced by lower tumour marker concentrations, smaller tumour sizes, less common vascular invasion, lower pT stages, and higher lymphocyte-to-monocyte ratios in blood. KRAS mutations were slightly less common in IL-33high cases than in IL-33low cancers (9% vs 29%; p=0.061). The strong expression of IL-33 in tissue appeared to be an independent favourable prognostic factor.

CONCLUSIONS:

IL-33high cholangiocarcinomas may represent a unique, less aggressive carcinogenetic process of the large bile ducts.