Landscape Phage-Targeted Drug Delivery to Breast Cancer Cells
Type of Degreedissertation
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The prevalence of breast cancer is still a public health burden worldwide. Recently, there has been significant progress in the management of breast cancer with the two FDA-approved cancer nanomedicines employing the concept of passive targeting. However, their delivery to the tumor cells can be hindered by insufficient permeation of nanomedicines into solid tumors, such as breast cancer, thereby, inhibiting bystander effects. Therapeutic efficacy of nanomedicines can be increased by active targeting of drug loaded nanocarriers. To this end, there is intense research effort in the quest for physiologically stable and cancer-specific ligands for targeting of nanocarriers to the cancer-specific cellular receptors. Although, a pleothora of ligands have been developed for targeting of drugs to the site of disease, only a few have been successfully used because of the need of their conjugation to drug carriers. Accordingly, we proposed that using landscape phage coat proteins for targeting of drug-loaded nanocarriers can enhance drug delivery to breast cancer cells. Screening of multibillion landscape phage library can generate peptide ligands targeting cancer-specific receptors. We hypothesize that spontaneous insertion of isolated phage proteins into drug loaded nanocarriers such as liposomes can enhance their cancer-specific cytotoxicity and exclude the need for complex bioconjugations and derivitization procedures required for targeting. We attempted to this hypothesis by screening two landscape phage libraries. Consequently, 132 phage probes specific for the breast cancer cell line, MCF-7 were generated. Coat proteins of selected phages were isolated and inserted into drug loaded liposome (Doxil™). Two phage coat proteins (with fusion peptides, DWRGDSMDS and GSDWMLGQD) were isolated and inserted into Doxil™. The cytotoxicity of the targeted Doxil™ was compared with non-targeted Doxil™. Our hypothesis was proven further by spontaneous insertion of phage protein (with the fusion peptide DMPGTVLP) into siRNA encapsulated liposomes and applied them to MCF-7 cells. The phage-targeted siRNA-liposome demonstrated significant down-regulation of PRDM14 gene expression and protein synthesis in the target MCF-7 cells in comparison with non-targeted siRNA. To determine the molecular mechanism of phage selectivity for breast cancer cells, selected phages were used as affinity matrixes in affinity chromatography to identify their counterpart receptors. Three phages DWRGDSMDS, GSDWMLGQD and DMPGTVLP were identified as ligands for nucleolin. We believe that the novel drug-targeting technique developed in the scope of this dissertation will allow significantly enhanced therapeutic efficacy of modern anti-cancer nanomedicines.