A novel small molecule-based multi-targeting approach for the selective therapy of epidermal growth factor receptor (EGFR)- or Her2-expressing carcinomas

The chemotherapy of solid tumours is hampered by two major obstacles: (1) the lack of selectivity of the current drugs and (2) the reduced sensitivity to tumour drugs. Recent advances in cancer biology have led to the identification of novel molecular targets responsible for aggressive proliferation, drug resistance and invasiveness. Targeting one such factor, epidermal growth factor receptor (EGFR), has proven an effective strategy to block the progression of solid tumours. However, the potency of the current clinical drugs directed at EGFR is mitigated by their reversible cytostatic activity and reduced cytotoxicity. Here we describe the design and synthesis of a new class of agents termed "cascade release combi-molecule" (I-TZ), "programmed" to hammer the receptor by releasing multiple bioactive metabolites (I-TZ', I-TZ") in a step-wise fashion and a reactive DNA damaging species (TZ), with the purpose of promoting sustained inhibition of EGFR tyrosine kinase (TK) and drug-induced cytotoxicity. This thesis describes the synthesis of one such molecule, RB24 (I-TZ), and demonstrates that it was capable of being hydrolyzed to its various sub-components with an overall half-life of 42 min under physiological conditions. The I-TZ induced sustained inhibition of the EGFR TK and strong antiproliferative activity in human solid tumour cells. It also inflicted higher levels of DNA damage to cancer cells transfected with the EGFR or HER2 gene. This selectivity was based on a novel mechanism of targeting described as a bystander effect, whereby following distribution of the I-TZ in the perinuclear region, the alkylating species (TZ) was subsequently released towards the nucleus. The I-TZ was significantly more potent than its clinical counterpart, TemodaI™. Further investigation of the mechanism underlying its superior potency show that blockade of EGFR-mediated signaling led to down-regulation of: (1) MAPK mitogenic signaling, (2) the BER protein, XRCC1 and (3) anti-apoptotic signaling mediated by Bad. Further down-regulation of XRCCI, with a MEK1 inhibitor, led to the discovery of a significant synergistic antiproliferative effect and a novel formula to further enhance the potency of the I-TZ in refractory tumours. Furthermore, the in vivo activity of the cascade release principle was proven in a prostate cancer xenograft study using a more water-soluble analogue of RB24. Thus, we have developed a novel model for the therapy of growth factor receptor-expressing tumours from the design and synthesis of the probe drugs, to the elucidation of their mechanism of action and the demonstration of the in vivo efficacy of the proposed strategy.