Discovery and pharmacophoric characterization of chemokine network inhibitors using phage-display, saturation mutagenesis and computational modelling.

CC and CXC-chemokines are the primary drivers of chemotaxis in inflammation, but chemokine network redundancy thwarts pharmacological intervention. Tick evasins promiscuously bind CC and CXC-chemokines, overcoming redundancy. Here we show that short peptides that promiscuously bind both chemokine classes can be identified from evasins by phage-display screening performed with multiple chemokines in parallel. We identify two conserved motifs within these peptides and show using saturation-mutagenesis phage-display and chemotaxis studies of an exemplar peptide that an anionic patch in the first motif and hydrophobic, aromatic and cysteine residues in the second are functionally necessary. AlphaFold2-Multimer modelling suggests that the peptide occludes distinct receptor-binding regions in CC and in CXC-chemokines, with the first and second motifs contributing ionic and hydrophobic interactions respectively. Our results indicate that peptides with broad-spectrum anti-chemokine activity and therapeutic potential may be identified from evasins, and the pharmacophore characterised by phage display, saturation mutagenesis and computational modelling.

New Paradigms to Assess Consequences of Long-Term, Low-Dose Curcumin Exposure in Lung Cancer Cells.

Curcumin has been investigated extensively for cancer prevention, but it has been proposed that long-term treatments may promote clonal evolution and gain of cellular resistance, potentially rendering cancer cells less sensitive to future therapeutic interventions. Here, we used long-term, low-dose treatments to determine the potential for adverse effects in non-small cell lung cancer (NSCLC) cells. IC50s for curcumin, cisplatin, and pemetrexed in A549, PC9, and PC9ER NSCLC cells were evaluated using growth curves. IC50s were subsequently re-assessed following long-term, low-dose curcumin treatment and a three-month treatment withdrawal period, with a concurrent assessment of oncology-related protein expression. Doublet cisplatin/pemetrexed-resistant cell lines were created and the IC50 for curcumin was determined. Organotypic NSCLC-fibroblast co-culture models were used to assess the effects of curcumin on invasive capacity. Following long-term treatment/treatment withdrawal, there was no significant change in IC50s for the chemotherapy drugs, with chemotherapy-resistant cell lines exhibiting similar sensitivity to curcumin as their non-resistant counterparts. Curcumin (0.25-0.5 µM) was able to inhibit the invasion of both native and chemo-resistant NSCLC cells in the organotypic co-culture model. In summary, long-term curcumin treatment in models of NSCLC neither resulted in the acquisition of pro-carcinogenic phenotypes nor caused resistance to chemotherapy agents.

The role of stromal fibroblasts in lung carcinogenesis: A target for chemoprevention?

The tumour microenvironment plays an essential role in the development and spread of cancers. Tumour cells interact with the surrounding extracellular matrix (ECM), embedded within which, are a variety of non-cancer cells including cells of the vasculature, immune system and fibroblasts. The essential role of fibroblasts in the cultivation and maintenance of an environment in which tumour cells are able to maintain their aggressive phenotypic traits is becoming increasingly well documented. Cancer-associated fibroblasts are able to secrete a vast array of ECM-modulating factors, meaning that they have potential for a functional role in every step of the carcinogenic process. In particular, they are likely to have a role in early tumour-initiating inflammatory events, and so may provide a potential target for chemopreventive intervention. This review summarises the known interactions between lung tumour cells and surrounding reactive fibroblasts, highlighting the need to further investigate cancer-associated fibroblasts as therapeutic targets in lung cancer chemoprevention strategies.