Inhibition of tumour growth by Raf265 via blockade of Raf/MEK/ERK pathway in colorectal cancer

published_or_final_versionThe 15th Annual Research Conference of the Department of Medicine, The University of Hong Kong, Hong Kong, 16 January 2010. In Hong Kong Medical Journal, 2010, v. 16, suppl. 1, p. 16, abstract no. 1

Emergence of CD26+ cancer stem cells with metastatic properties in colorectal carcinogenesis

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Preclinical analysis of the anti-tumor and anti-metastatic effects of Raf265 on colon cancer cells and CD26(+) cancer stem cells in colorectal carcinoma

© 2015 Chow et al.Background: In colorectal carcinoma (CRC), activation of the Raf/MEK/ERK signaling pathway is commonly observed. In addition, the commonly used 5FU-based chemotherapy in patients with metastatic CRC was found to enrich a subpopulation of CD26+ cancer stem cells (CSCs). As activation of the Raf/MEK/ERK signaling pathway was also found in the CD26+ CSCs and therefore, we hypothesized that an ATP-competitive pan-Raf inhibitor, Raf265, is effective in eliminating the cancer cells and the CD26+ CSCs in CRC patients. Methods: HT29 and HCT116 cells were treated with various concentrations of Raf265 to study the anti-proliferative and apoptotic effects of Raf265. Anti-tumor effect was also demonstrated using a xenograft model. Cells were also treated with Raf265 in combination with 5FU to demonstrate the anti-migratory and invasive effects by targeting on the CD26+ CSCs and the anti-metastatic effect of the combined treatment was shown in an orthotopic CRC model. Results: Raf265 was found to be highly effective in inhibiting cell proliferation and tumor growth through the inhibition of the RAF/MEK/ERK signaling pathway. In addition, anti-migratory and invasive effect was found with Raf265 treatment in combination with 5FU by targeting on the CD26+ cells. Finally, the anti-tumor and anti-metastatic effect of Raf265 in combination with 5FU was also demonstrated. Conclusions: This preclinical study demonstrates the anti-tumor and anti-metastatic activity of Raf265 in CRC, providing the basis for exploiting its potential use and combination therapy with 5FU in the clinical treatment of CRC.published_or_final_versio

Identification of microRNA 885-5p as a novel regulator of tumor metastasis by targeting CPEB2 in colorectal cancer

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Comparative cardiovascular risk in users versus non-users of xanthine oxidase inhibitors and febuxostat versus allopurinol users

OBJECTIVES: The aim of this study is to determine major adverse cardiovascular events (MACE) and all-cause mortality comparing between xanthine oxidase inhibitors (XOIs) and non-XOI users, and between allopurinol and febuxostat. METHODS: This is a retrospective cohort study of gout patients prescribed anti-hyperuricemic medications between 2013 and 2017 using a territory-wide administrative database. XOI users were matched 1:1 to XOI non-users using propensity scores. Febuxostat users were matched 1:3 to allopurinol users. Subgroup analyses were conducted based on colchicine use. RESULTS: Of the 13 997 eligible participants, 3607 (25.8%) were XOI users and 10 390 (74.2%) were XOI non-users. After propensity score matching, compared with non-users (n = 3607), XOI users (n = 3607) showed similar incidence of MACE (hazard ratio [HR]: 0.997, 95% CI, 0.879, 1.131; P>0.05) and all-cause mortality (HR = 0.972, 95% CI 0.886, 1.065, P=0.539). Febuxostat (n = 276) users showed a similar risk of MACE compared with allopurinol users (n = 828; HR: 0.672, 95% CI, 0.416, 1.085; P=0.104) with a tendency towards a lower risk of heart failure-related hospitalizations (HR = 0.529, 95% CI 0.272, 1.029; P=0.061). Concurrent colchicine use reduced the risk for all-cause mortality amongst XOI users (HR = 0.671, 95% 0.586, 0.768; P<0.001). CONCLUSION: In gout patients, XOI users showed similar risk of MACE and all-cause mortality compared with non-users. Compared with allopurinol users, febuxostat users showed similar MACE and all-cause mortality risks but lower heart failure-related hospitalizations

Prognostic significance of CD26 in patients with colorectal cancer

Background: CD26, dipeptidyl peptidase IV, was discovered firstly as a membrane-associated peptidase on the surface of leukocyte. We previously demonstrated that a subpopulation of CD26+ cells were associated with the development of distant metastasis, enhanced invasiveness and chemoresistance in colorectal cancer (CRC). In order to understand the clinical impact of CD26, the expression was investigated in CRC patient's specimens. This study investigated the prognostic significance of tumour CD26 expression in patients with CRC. Examination of CD26+ cells has significant clinical impact for the prediction of distant metastasis development in colorectal cancer, and could be used as a selection criterion for further therapy. Methods: Tumour CD26 expression levels were studied by immunohistochemistry using Formalin-fixed paraffin embedded (FFPE) tissues in 143 patients with CRC. Tumour CD26 expression levels were correlated with clinicopathological features of the CRC patients. The prognostic significance of tumour tissue CD26 expression levels was assessed by univariate and multivariate analyses. Result: CD26 expression levels in CRC patients with distant metastasis were significantly higher than those in non-metastatic. High expression levels of CD26 were significantly associated with advanced tumour staging. Patients with a high CD26 expression level had significantly worse overall survival than those with a lower level (p<0.001). Conclusions: The expression of CD26 was positively associated with clinicopathological correlation such as TNM staging, degree of differentiation and development of metastasis. A high CD26 expression level is a predictor of poor outcome after resection of CRC. CD26 may be a useful prognostic marker in patients with CRC. © 2014 Lam et al.published_or_final_versio

The clinicopathological significance of miR-133a in colorectal cancer

This study determined the expression of microRNA-133a (MiR-133a) in colorectal cancer (CRC) and adjacent normal mucosa samples and evaluated its clinicopathological role in CRC. The expression of miR-133a in 125 pairs of tissue samples was analyzed by quantitative real-time polymerase chain reaction (qRT-PCR) and correlated with patient's clinicopathological data by statistical analysis. Endogenous expression levels of several potential target genes were determined by qRT-PCR and correlated using Pearson's method. MiR-133a was downregulated in 83.2% of tumors compared to normal mucosal tissue. Higher miR-133a expression in tumor tissues was associated with development of distant metastasis, advanced Dukes and TNM staging, and poor survival. The unfavorable prognosis of higher miR-133a expression was accompanied by dysregulation of potential miR-133a target genes, LIM and SH3 domain protein 1 (LASP1), Caveolin-1 (CAV1), and Fascin-1 (FSCN1). LASP1 was found to possess a negative correlation (γ=-0.23), whereas CAV1 exhibited a significant positive correlation (γ=0.27), and a stronger correlation was found in patients who developed distant metastases (γ=0.42). In addition, a negative correlation of FSCN1 was only found in nonmetastatic patients. In conclusion, miR-133a was downregulated in CRC tissues, but its higher expression correlated with adverse clinical characteristics and poor prognosis. © 2014 Timothy Ming-Hun Wan et al.published_or_final_versio

Influences of Excluded Volume of Molecules on Signaling Processes on Biomembrane

We investigate the influences of the excluded volume of molecules on biochemical reaction processes on 2-dimensional surfaces using a model of signal transduction processes on biomembranes. We perform simulations of the 2-dimensional cell-based model, which describes the reactions and diffusion of the receptors, signaling proteins, target proteins, and crowders on the cell membrane. The signaling proteins are activated by receptors, and these activated signaling proteins activate target proteins that bind autonomously from the cytoplasm to the membrane, and unbind from the membrane if activated. If the target proteins bind frequently, the volume fraction of molecules on the membrane becomes so large that the excluded volume of the molecules for the reaction and diffusion dynamics cannot be negligible. We find that such excluded volume effects of the molecules induce non-trivial variations of the signal flow, defined as the activation frequency of target proteins, as follows. With an increase in the binding rate of target proteins, the signal flow varies by i) monotonically increasing; ii) increasing then decreasing in a bell-shaped curve; or iii) increasing, decreasing, then increasing in an S-shaped curve. We further demonstrate that the excluded volume of molecules influences the hierarchical molecular distributions throughout the reaction processes. In particular, when the system exhibits a large signal flow, the signaling proteins tend to surround the receptors to form receptor-signaling protein clusters, and the target proteins tend to become distributed around such clusters. To explain these phenomena, we analyze the stochastic model of the local motions of molecules around the receptor.Comment: 31 pages, 10 figure

Semiquantitative immunoblots of membrane protein-epidermal growth factor

Membrane proteins or cytokines are sometimes difficult to isolate and purify. Our group recently concentrated on epidermal growth factor (EGF) protein expression studies. Mature EGF was initially identified from mouse submaxillary gland extract as a stimulator of eyelid opening and incisor eruption when injected into newborn mice and rats. The EGF precursor is a transmembrane protein with eight additional EGF-like repeats. Our previous study has shown that the EGF precursor without these eight EGF-like repeats (hEGF) was biologically active. Here, we introduce a modified method for rapid detection of hEGF. The membranous protein was directly extracted from various organs of transgenic mice (including the submandibular gland, kidney, liver, heart, and testis) with two different buffers and easily detected by semiquantitative immunoblotting.link_to_subscribed_fulltex

Expression of epidermal growth factor in transgenic mice causes growth retardation

The epidermal growth factor (EGF) family of peptides signals through the erbB family of receptor tyrosine kinases and plays important roles in development and tumorigenesis. Both EGF and transforming growth factor (TGF)-α only bind to erbB1 and activate it. The precursor of EGF is distinct from that of TGF-α in having eight additional EGF-like repeats. We have recently shown that the EGF precursor without these repeats is biologically active and leads to hypospermatogenesis in transgenic mice. Here we present evidence that the growth of transgenic mice widely expressing this engineered EGF precursor is also stunted. These mice were consistently born at half the normal weight and reached almost 80% of normal weight at adulthood. The mechanism involved a reduction of serum insulin-like growth factor-binding protein-3. Chondrocyte development in the growth plate was affected, and osteoblasts accumulated in the endosteum and periosteum. Besides these novel findings on the in vivo effects of EGF on bone development, we observed no sign of tumor formation in our transgenic animals. In contrast to previous reports on TGF-α transgenic mice, we show that the biological functions of EGF and TGF-α are clearly distinct.link_to_subscribed_fulltex