Biology subjects

Campbell, F.

Publications and source records attributed to Campbell, F..

4 recordsLinked to original sources

bioRxiv2025

Single VLP lipid-mixing measurements confirm off-pathway state in dengue virus fusion mechanism

Dengue virus (DENV) is the causative agent of dengue fever and exerts a substantial healthcare burden worldwide. Like other flaviviruses, DENV must undergo membrane fusion with the host cell in order to initiate infection. This membrane fusion occurs following acidification during endocytosis and is pH dependent. Here, we interrogate whether the mechanism of DENV fusion contains an off-pathway state, such has been reported previously for two other flaviviruses - Zika virus and West Nile virus. To do this, we utilize single particle lipid mixing measurements of DENV virus-like particles (VLPs) to tethered liposomes, together with computational modeling inspired by chemical kinetics. By observing and then modeling the pH dependence of single VLP fusion kinetics, we provide evidence that the DENV fusion mechanism must contain an off-pathway state. Measuring the proportion of VLPs undergoing hemi-fusion over time, we also demonstrate that the off-pathway state appears to be slowly reversible over tens of minutes, at least for some virions. Additionally, we find that late endosomal anionic lipids do not appear to influence the off-pathway mechanism to any great extent. In conjunction with the prior reports on Zika virus and West Nile virus, this work indicates that an off-pathway fusion state may be a feature of flavivirus fusion more broadly. We also note that the platform and mechanistic model described in this study may be useful in elucidating the mechanism of action of small molecule inhibitors of flavivirus fusion developed by our group and others. Statement of SignificanceDengue virus (DENV) causes dengue fever and infects an estimated hundreds of millions of people annually. To date, there are no specific antiviral drugs for DENV and limited vaccination options, highlighting the need to better understand this important pathogen. In this report, we investigate the mechanism of DENV membrane fusion, an early step in the viral infectious cycle, using a mix of experimental techniques and computer simulations. We find strong evidence that the DENV fusion mechanism contains an off-pathway state, in which it can get stalled prior to membrane fusion. Understanding this off-pathway state could be an avenue to develop antiviral strategies against DENV and other related viruses.

biophysics↗

bioRxiv2019

Customizable, engineered substrates for rapid screening of cellular cues

Biophysical cues robustly direct cell responses and are thus important tools for in vitro and translational biomedical applications. High throughput platforms exploring substrates with varying physical properties are therefore valuable, however, currently existing platforms are limited in throughput, the biomaterials used, the capability to segregate between different cues and the assessment of dynamic cellular responses. Here we present a multiwell array (3x8) using a substrate engineered with patterns that present topography or rigidity cues welded to a bottomless plate with a 96-well format. Both the patterns on the engineered substrate and the well plate format can be easily customized, permitting systematic and efficient screening of biophysical cues. Here, we demonstrate three multiwell arrays patterned with a variety of topographical and mechanical cues (nano-grooves, soft pillars and nano pillars) tested with three different cell types. Using the multiwell array, we were able to measure cell functionality using analytical modalities such as live microscopy, qPCR and fluorescent immunochemistry. Cardiomyocytes cultured on 5{micro}m grooves showed less variation in electrophysiology and contractile function. Nanopillars with 127 nm height, 100 nm diameter and 300 nm pitch showed improved chondrogenic maintenance from matrix deposition and chondrogenic gene expression. High aspect ratio pillars with an elastic shear modulus of 16 kPa mimicking the cortical bone altered cell adhesion, morphology, and increased expression of osteogenic genes. We have demonstrated the bespoke, controlled and high-throughput properties of the multiwell array that are currently unparalleled in the field today.

bioengineering↗

bioRxiv2017

Macrophage-derived granulin drives resistance to immune checkpoint inhibition in metastatic pancreatic cancer

The ability of disseminated cancer cells to evade the immune response is a critical step for efficient metastatic progression. Protection against an immune attack is often provided by the tumour microenvironment that suppresses and/or excludes cytotoxic CD8+ T cells. Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive metastatic disease with unmet needs, yet the immuno-protective role of the metastatic tumour microenvironment in pancreatic cancer is not completely understood. In this study we find that macrophage-derived granulin contributes to cytotoxic CD8+ T cell exclusion in metastatic livers. Mechanistically, we find that granulin expression by macrophages is induced in response to colony stimulating factor-1. Genetic depletion of granulin reduces the formation a fibrotic stroma, thereby allowing T cell entry at the metastatic site. While metastatic PDAC tumours are largely resistant to anti-PD-1 therapy, blockade of PD-1 in granulin depleted tumours restored the anti-tumour immune defence and dramatically decreased metastatic tumour burden. These findings suggest that targeting granulin may serve as a potential therapeutic strategy to restore CD8+ T cell infiltration in metastatic PDAC, thereby converting PDAC metastatic tumours, which are refractory to immune checkpoint inhibitors, into tumours that respond to immune checkpoint inhibition therapies.

cancer biology↗

bioRxiv2017

Blockade of insulin-like growth factors increases efficacy of paclitaxel in metastatic breast cancer.

Breast cancer remains the leading cause of cancer death in women due to metastasis and the development of resistance to established therapies. Macrophages are the most abundant immune cells in the breast tumor microenvironment and can both inhibit and support cancer progression. Thus, gaining a better understanding of how macrophages support cancer could lead to the development of more effective therapies. In this study, we find that breast cancer associated macrophages express high levels of insulin-like growth factors 1 and 2 (IGFs) and are the main source of IGFs within both primary and metastatic tumours. 75% of breast cancer patients show activation of Insulin/IGF-1 receptor signaling and this correlates with increased macrophage infiltration and advanced tumor stage. In patients with invasive breast cancer, activation of Insulin/IGF-1 receptors increased to 87%. Blocking IGF in combination with paclitaxel, a chemotherapeutic agent commonly used to treat breast cancer, showed a significant reduction in tumor cell proliferation and lung metastasis in a pre-clinical breast cancer model compared to paclitaxel monotherapy. Our findings provide the rationale for further developing the combination of paclitaxel with IGF blockers for the treatment of invasive breast cancer, and Insulin/IGF1R activation and IGF+ stroma cells as potential biomarker candidates for further evaluation.

cancer biology↗