STUDIES ON THE MOLECULAR MECHANISM AND TARGETED DELIVERY OF PLATINUM-ACRIDINE ANTICANCER AGENTS

Abstract

Platinum-acridines (PAs) were developed as a class of DNA-targeted hybrid anticancer agents to treat chemoresistant cancers that do not respond to conventional chemotherapies. The three projects in this dissertation were designed to address the following questions: 1. Are PAs chemically compatible with targeted delivery as payloads in bioconjugates? 2. Can chemical biology tools be developed to detect covalent adducts formed by PAs in potential target genes in genomic DNA? 3. How do variations in the molecular makeup of cancer cells affect the spectrum of activity of PAs? Project 1: A combination of strain-promoted click chemistry and maleimide Michael addition chemistry was used to attach azide-modified PAs to a protein carrier. The structure and functionality of the payload was validated in human serum albumin (HSA) using ESI-TOF MS of intact bioconjugate, MS/MS of tryptic digests, 2-D HSQC heteronuclear NMR spectroscopy, and confocal microscopy. The results suggest that the technique may have broad utility for targeted delivery of highly cytotoxic PAs. Project 2: An azide-functionalized PA derivative (APA) was also used to develop a chemical biology tool that allows the biotinylation, affinity capture, and chemically retrieval of platinum-damaged sequences in genomic DNA. Using this Adduct Detection after Post-Labeling Technique (ADAPT) in combination with quantitative PCR (qPCR) it was possible to identify PA adducts in ribosomal DNA (rDNA). These findings suggest that rDNA in the cell’s nucleolus may play an important role in the mechanism of PA-mediated cell death. Project 3: Using NCI-60 cancer screening data in conjunction with Pearson correlation analysis (COMPARE), human multidrug and toxin extrusion protein 1 (hMATE1, SLC47A1) was identified as the dominant predictor of chemosensitivity of cancer cells to PAs (p < 10-5). The crucial role of hMATE1 was validated in A549 lung cancer cells, which express high levels of the membrane transporter, using transporter inhibition assays, transient gene knockdown by RNAi, and cell viability screening. HCT-116 colon cancer cells, in which hMATE1 is epigenetically repressed, can be sensitized by priming the cells with epigenetic drugs. These results suggest that hMATE1 may have applications as a pan-cancer molecular marker to identify tumors that are likely to respond to PAs.