The failure of cancer chemotherapy can be attributed in large measure to a lack of tumor selectivity of chemotherapeutic agents. Currently approved antitumor antifolates, methotrexate, pemetrexate, raltitrexed and pralatrexate, are all transported by the ubiquitous, reduced folate carrier (RFC) into both tumor and normal host cells, and hence are nonselective for tumor cells and are consequently toxic. If an agent could selectively attack only cancer cells, a qualitative and not just a quantitative difference between cancer cells and normal cells would be at hand for exploitation. We have discovered compounds that specifically target tumor cells by selective transport by folate receptor (FR) -? and -?. These are folic acid transport systems (almost) exclusively expressed by several tumor cells (e.g., ovarian, non-small cell lung cancer, endometrial, renal, breast, cervical, lung) but nt normal cells. Our compounds, once selectively transported into tumors, target the de novo purine nucleotide biosynthesis pathways dependent on folate, i.e. glycinamide ribonucleotide formyltransferase (GARFTase) or 5-amino-4- imidazolecarboxamide ribonucleotide (AICAR) formyltransferase (AICARFTase). These are both novel enzyme targets, as there are no known clinical agents that target de novo purine biosynthesis as their primary mechanism of action. Antipurine antifolates are cytotoxic independent of p53 status. Selectivity of antipurine agents can result from the loss of purine salvage in a large number of human tumors. Inhibition of AICARFTase causes an accumulation of AICAR (ZMP) that, via AMPK, inhibits mTOR, resulting in an additional mechanism of cytotoxic antitumor activity. The goal of this proposal is to optimize our lead structures for tumor specificity via FR? and -? over RFC, and GARFTase or AICARFTase inhibitory activities.
In Aim 1, we will synthesize novel analogs from 16 series (two in each series) based on structure-activity profiles of our lead compounds.
In Aim 2, we will evaluate compounds from Aim 1 for cytotoxicity in isogenic hamster and human tumor cell line models with established RFC and FR expression, and will identify molecular mechanisms including cellular targets by nucleoside protection, in situ metabolic labeling, analysis of intracellular metabolites, and studies with isolated enzymes. Additional studies will characterize transport properties of the novel analogs with FRs vis ? vis RFC, polyglutamate synthesis, and mechanisms of cell death.
In Aim 3, we will determine in vivo efficacies of the most potent FR-targeted GARFTase or AICARFTase inhibitors by in vivo toxicity and efficacy trials in FR-expressing human tumor implants in SCID mice. Finally, in Aim 4, we will determine X-ray crystal structures of the most potent and selective analogs with FR? and/or -?, as well as GARFTase or AICARFTase. All the Specific Aims will be performed concurrently from year 1 to year 5. Collectively, our studies will afford a molecular understanding of the interactions of nove analogs with FRs and enzyme targets to guide the design of selective analogs. We anticipate advancing one or more of our novel folate analogs with optimized FR-selective antitumor agents to clinical trials to be used alone or in combination with other agents.
A major problem of cancer treatment failure is that the drugs used are not specific for cancer cells and also attack healthy host cells that results in toxicity o the patient. We have discovered highly selective drugs that are transported by folate receptors (FR) expressed on many types of cancer cells and not on normal cells. These drugs are able to kill only the cancer cells. This spares normal cells from the toxicity. Though the drugs we have discovered are selective for tumor cells they are not very potent or they are highly potent but not highly selective. This study is designed to determine the structural basis of both selectivity and potency and provide unique compounds with the best balance of both potency and selectivity ready for clinical trials in humans as cancer chemotherapeutic agents. To our knowledge, this collaboration between an accomplished biochemist specializing in molecular pharmacology of anticancer drugs and an outstanding medicinal chemist with a demonstrated track record of antifolate drug design and a crystallographer with FR crystallization capabilitites represents the only such endeavor capable of generating unique nontoxic compounds for the treatment of cancers.
|Golani, Lalit K; George, Christina; Zhao, Sai et al. (2014) Structure-activity profiles of novel 6-substituted pyrrolo[2,3-d]pyrimidine thienoyl antifolates with modified amino acids for cellular uptake by folate receptors ? and ? and the proton-coupled folate transporter. J Med Chem 57:8152-66|
|Wibowo, Ardian S; Singh, Mirage; Reeder, Kristen M et al. (2013) Structures of human folate receptors reveal biological trafficking states and diversity in folate and antifolate recognition. Proc Natl Acad Sci U S A 110:15180-8|
|Wang, Yiqiang; Cherian, Christina; Orr, Steven et al. (2013) Tumor-targeting with novel non-benzoyl 6-substituted straight chain pyrrolo[2,3-d]pyrimidine antifolates via cellular uptake by folate receptor * and inhibition of de novo purine nucleotide biosynthesis. J Med Chem 56:8684-95|