Biology subjects

Awanis, G.

Publications and source records attributed to Awanis, G..

4 recordsLinked to original sources

bioRxiv2026

A three-dimensional imaging pipeline for the detection of vasculogenic mimicry in mouse tumor models

Vasculogenic mimicry (VM) describes the ability for tumor cells to mimic endothelial cells and form tumor-lined blood vessels. Although VM is associated with poor prognosis, resistance to therapeutics and metastasis, its presence, relevance and functionality have been debated, largely due to the techniques employed to determine its presence. Dual CD31 and periodic acid-Schiff (PAS) histological staining, the method widely used to detect VM within tumor tissue, is hampered by issues with artifacts of 2D sections and does not assess active blood flow through vessels and hence cannot confirm vessel functionality. Here, we present a 3D imaging pipeline to detect functionally perfused VM and host vessels within mouse tumor models. Mice tumor models are intravenously injected with fluorescently-labelled tomato lectin which binds to the lumen of functional blood vessels. These tumors are thickly sliced, optically-cleared, stained with endothelial markers and imaged using light-sheet microscopy. We apply this imaging modality to image tumors from six breast cancer patient-derived tissue xenograft models, an orthotopic mammary cancer model and two small cell lung cancer circulating-tumor cell explant models, where VM vessels, mosaic vessels and host-VM vessel junctions can be detected. To our knowledge, this is the largest collection of 3D VM vessel images to date, providing convincing evidence of the presence and functionality of VM within tumors and defining the level of evidence necessary to detect VM in vivo.

cancer biology↗

bioRxiv2026

Epidermal Growth Factor/c-Met receptor signalling crosstalk drives tunneling nanotube formation in A549 lung adenocarcinoma cells

Tunneling nanotubes (TNTs) are actin-based cytoplasmic connections that can mediate intercellular transfer of various cellular cargo and have been implicated in cancer progression and chemoresistance. However, the signalling mechanisms driving their formation remain poorly understood. Given the frequent dysregulation of EGFR and c-Met signalling in non-small cell lung cancer (NSCLC), and prior evidence of TNTs in lung adenocarcinoma patient samples, we investigated the role of EGFR and c-Met receptor signalling crosstalk in TNT induction in A549 lung adenocarcinoma cells. Stimulation with EGF, HGF, or in combination induced a concentration dependent increase in the formation of TNTs. TNTs exhibited typical characteristics, including F-actin expression, non-adherence to the substratum and facilitated intercellular trafficking of lysosomes, mitochondria, and lipid vesicles. EGFR was identified as a novel component of TNTs, but had little co-localisation with the c-Met receptor. Co-stimulation with HGF and EGF did not produce consistent additive or synergistic effects on TNT formation, suggesting shared downstream signalling. Furthermore, although EGFR and c-Met inhibition blocked EGF- and HGF-induced TNTs respectively, inhibition of both receptors was required to suppress TNTs following dual HGF/EGF treatment. Interestingly, blocking the EGF receptor alongside c-Met resulted in a more potent inhibition of HGF-induced TNTs, indicating crosstalk. Furthermore, inhibition of downstream MEK and PI3K pathways reduced HGF- or EGF- induced TNT formation, but dual inhibition was required to completely block TNT formation in HGF+EGF co-stimulated cells. These findings reveal a novel convergence of EGFR and c-Met and their downstream MAPK/PI3K pathways in TNT regulation, which can have important clinical implications in combinatorial receptor and cell signalling pathway targeting in NSCLC.

cancer biology↗

bioRxiv2022

Hepatocyte Growth Factor and β1-integrin signalling axis drives tunneling nanotube formation in A549 lung adenocarcinoma cells

Tunneling nanotubes (TNTs) are thin cytoplasmic protrusions involved in long-distance cellular communication. The presence of TNTs has been found in vivo and in vitro studies in non-small cell lung cancer (NSCLC). Cancer cells transport a range of organelles and signalling molecules along TNTs, to confer a survival phenotype for the recipient cell, contributing toward chemoresistance and malignancy. Despite its important role in cancer progression, the molecular mechanisms underlying TNT formation is not well defined. Within the tumour microenvironment (TME) of NSCLC, hepatocyte growth factor (HGF) and its receptor, c-Met, are mutationally upregulated causing growth, and invasion. In this study, we report a novel crosstalk between HGF/c-Met and {beta}1-integrin involved in the formation of functional TNTs in A549 cells. Through pharmacological inhibitor studies, we discovered Arp2/3 complex, MAPK and PI3K pathways were activated downstream of this crosstalk signalling axis. Furthermore, paxillin was recruited during this key process, localising at the protrusion site of HGF-induced TNTs, and therefore serving as the central link between the upstream and downstream regulators involved. Overall, these results demonstrate a novel strategy to inhibit TNT formation in NSCLC through targeting the HGF/c-Met and {beta}1-integrin signalling axis, thus highlighting the importance of personalised multi-drug targeting in NSCLC.

cancer biology↗

bioRxiv2022

M1 and M2 macrophages differentially regulate colonic crypt renewal.

The colonic epithelium is the most rapidly renewing tissue in the body and is organized into a single cell layer of invaginations called crypts. Crypt renewal occurs through Lgr5+ gut stem cells situated at the crypt base, which divide, produce daughter cells that proliferate, migrate, differentiate into all the cells required for normal gut function (eg. Goblet cells, enterocytes), and are finally shed into the crypt lumen. In health this rapid renewal helps maintain barrier function next to the hostile gut luminal environment that contains microbes and food. In parallel, the peri-cryptal lamina propria hosts the largest monocyte-derived macrophage population in the human body. Different macrophage phenotypes have been associated with intestinal health/intact barrier function, namely M2 compared to M1 macrophages that indicate inflammation/compromised barrier function. However, the direct effect of different macrophage subtypes have on colonic crypt renewal is not well understood. In this study we have utilized a reductionist 3D in vitro co-culture model to determine the regulatory capacity of M1 and M2 macrophages on colonic crypt renewal. We show that colonic crypt proliferation is increased in the presence of M1 or M2 macrophages, while we further demonstrate that a decrease in goblet and tuft cell expression as well as an increase in Lgr5+ stem cell numbers is only achieved through M1-crypt crosstalk in a contact dependent manner.

cell biology↗