Applications are invited for a fully-funded BBSRC-DTP industrial case doctoral studentship commencing on 1st October 2021. The successful applicant will receive a first class training experience across multiple disciplines within academic and industrial organisations. The project is partnered by AstraZeneca (AZ), a world-leading pharmaceutical company specialising in drug delivery. Particularly challenging, and the focus of this studentship, is delivery of brain cancer therapies that cross the blood brain barrier and evade drug-resistance mechanisms. Through development of biocompatible protein-based nano-sized drug delivery structures (DDS), it is possible to deliver therapeutic agents able to target specific sites, overcome drug resistance and confer reduced systemic toxicity. However, interactions between DDS / encapsulated agents and cells / tissues, and how drug-resistance can be by-passed is poorly understood. The biocompatible protein apoferritin (AFt), amenable to protein engineering and manipulation by synthetic biology will be used as a DDS. Previously, we have demonstrated AFt delivery of near-infrared PbS quantum dots (QDs), anti-cancer agents (e.g. EGFR tyrosine kinase inhibitor gefitinib, imidazotetrazine- and benzothiazole analogues) to carcinoma cell lines. By exploiting cancer cells` enhanced expression of transferrin receptor (TfR1) and the intrinsic binding properties of AFt, AFt-encapsulation confers a significant degree of cancer-selectivity. Our hypothesis is that AFt-encapsulated cargo is endocytosed, and as pH falls in the late endosome and acidic lysosome, the AFt nanocage disassembles releasing cargo to exert a therapeutic effect. We have established in 2D cell culture that AFt-encapsulation of temozolomide (TMZ) is able to overcome resistance to this methylating agent conferred by MGMT expression, DNA-mismatch repair (MMR) deficiency or p-glycoprotein (p-gp) efflux.
In this project you will determine mechanisms by which AFt-delivery overcomes drug-resistance by studying AFt-uptake, -trafficking though tumour cells and cargo delivery. The successful applicant will investigate: i) Fluorescence properties of imaging agents and molecular characteristics of therapeutic agents to image and locate AFt-cargo within cells. ii) Activity in carcinoma cell lines possessing inherent and acquired resistance to anticancer agents through clinically-relevant mechanisms including drug efflux-, DNA repair, drug transport mechanisms. iii) Penetration of, and sensitivity to AFt-encapsulated therapeutic and imaging agents in tumour spheroids - a more patient-relevant in vitro model of disease. iv) Mechanisms i) of drug action; ii) by which drug-resistance is surmounted, through combination of morphological and compositional analyses of uptake, penetration and activity of AFt-encapsulated agents. Training in techniques adopted throughout these studies will include: i) AFt purification, AFt-encapsulation of imaging and therapeutic agents via pH-mediated dis-/re-assembly and nanoreactor methods, and characterisation (native PAGE, DLS). ii) Cell and spheroid culture coupled with in vitro cytotoxicity assays. iii) Pharmacokinetic analyses of intracellular cargo delivery (HPLC). iv) Imaging and microscopy techniques including SEM, cryo TEM. v) Analytical mass spectrometry (MS) techniques e.g. ToFSIMS, OrbiSIMS.