Our laboratory has a strong interest in pharmacogenetics. We have been active in studying how germline genetic variants can alter pharmacokinetics, response, and toxicity of various anticancer agents, thereby contributing to inter-individual variation in clinical outcomes in therapies with an already narrow therapeutic window. We have established a molecular link between these polymorphisms and their phenotype as it relates to drug treatment. Most of our work has been focused on genetic variations in drug metabolism and transporting candidate genes such as ABCB1 (P-glycoprotein, MDR1), ABCG2 (BCRP), SLCO1B3 (OATP1B3, OATP8), CYP3A4, CYP3A5, CYP1B1, CYP2C19, CYP2D6, UGT1A1, UGT1A9 and several others. We are also interested in non-candidate gene approaches where large numbers of polymorphisms are explored to establish a relationship with clinical outcome, and experiments are conducted to validate potential causative alleles resulting from exploratory scanning. We have worked with Affymetrix to beta-test the DMET chip that contains 1,256 genetic variations in 170 drug disposition genes, and are currently establishing a clinical trial where patients treated at the NCI will be genotyped with the DMET chip to explore potential links between these genes and various treatments of several cancers. We are currently making progress in validating the results from the initial DMET chip experiments. While many of these studies have been conducted in order to explain some of the genetic influence on pharmacokinetic variability, we also have a strong interest in clarifying genetic markers of pharmacodynamics and therapeutic outcome of several major anticancer agents since this field has been rather poorly studied. DMET The anticancer agent docetaxel exhibits significant inter-individual variation in its pharmacokinetic and toxicity profile. Thalidomide is an active anticancer agent and also exhibits wide pharmacologic variation. Past pharmacogenetic research has not explained this variation. Patients with prostate cancer enrolled in a randomized phase II trial using docetaxel and thalidomide versus docetaxel alone were genotyped using the Affymetrix DMET 1.0 platform, which tests for 1,256 genetic variations in 170 drug disposition genes. Genetic polymorphisms were analyzed for associations with clinical response and toxicity. Ten SNPs in three genes were potentially associated with response to therapy: PPARdelta, SULT1C2, and CHST3. Eleven SNPs in eight genes were associated with toxicities to treatment: SPG7, CHST3, CYP2D6, NAT2, ABCC6, ATP7A, CYP4B1, and SLC10A2. Genotyping results between DMET and direct sequencing showed greater than 96% concordance. These findings highlight the role that non-CYP450 metabolizing enzymes and transporters may play in the pharmacology of docetaxel and thalidomide. XRCC1 polymorphism and radiation: Genetic polymorphisms in XRCC1 associated with radiation therapy in prostate cancer: Radiation therapy is a potentially curative, important treatment option in localized prostate cancer. However, at 8 years after radiation therapy, even in the best risk subset of patients, approximately 10% of patients will experience clinical disease recurrence. Herein, we investigated five molecular markers of DNA repair. 513 patients with castrate-resistant prostate cancer (CRPC), including 284 patients who received radiotherapy, 229 patients without radiotherapy, and 152 healthy individuals were genotyped for 5 polymorphisms in DNA excision repair genes: ERCC1 N118N (500C greater than T), XPD K751Q (2282A greater than C), XRCC1 R194W (685C greater than T), XRCC1 R399Q (1301G greater than A) and PARP1 V762A (2446T greater than C). The polymorphisms evaluated did not show differences between the patient group and the healthy controls, nor did they show a trend toward an association with survival. However, in the radiation treated subgroup, the median survival time was associated with the XRCC1 haplotype. The median survival time was 11.75 years for patients with the R399Q AA/R194W CC haplotype, 12.17 years for patients with the R399Q AG/R194W CC haplotype, 66.65 years for patients with the R399Q AG/R194W CT haplotype, and 6.21 years for patients with the R399Q GG/R194W CT haplotype (p = 0.034). This association was not found when all patients were investigated. We conclude that the genetic polymorphisms in XRCC1 may affect the outcome in patients who received radiotherapy for localized prostate cancer. ERCC1 polymorphism and platinum-base chemotherapy: ERCC1 has been considered a prognostic marker for platinum-based chemotherapy and many genetic association studies focused on ERCC1 N118N polymorphism. To verify this functional implication, we introduced the variant allele into the open reading frame (ORF) clone of ERCC1 and assayed for platinum sensitivity of the transfected ERCC1 deficient UV-20 cell line. The cells transfected with the ERCC1 clone exhibited significant higher resistance to platinum-containing drugs, including cisplatin, carboplatin and oxaliplatin. Studies are currently underway to futher elucidate elucidate the functional significance of this polymorphism. VEGFR2 polymorphism and bevacizumab/sorafenib: Hypertension (HT) and hand-foot skin reactions (HFSR) may be related to the activity of bevacizumab and sorafenib. We hypothesized that these toxicities would correspond to favorable outcome in these drugs, that HT and HFSR would coincide, and that VEGFR2 genotypic variation would be related to toxicity and clinical outcomes. Toxicity incidence and VEGFR2 H472Q and V297I status were compared to clinical outcomes. Individuals experiencing HT had longer PFS following bevacizumab therapy than those without this toxicity in trials utilizing bevacizumab in patients with prostate cancer (31.5 vs 14.9mo, n=60, P=0.0009), and bevacizumab and sorafenib in patients with solid tumors (11.9 vs 3.7 mo, n=27, P=0.052). HT was also linked to a greater than 5-fold OS benefit after sorafenib and bevacizumab cotherapy (5.7 versus 29.0 mo, P=0.0068). HFSR was a marker for prolonged PFS during sorafenib therapy (6.1 vs 3.7 mo, n=113, P=0.0003). HT was a risk factor for HFSR in patients treated with bevacizumab and/or sorafenib (OR(95%CI) = 3.2(1.5-6.8), P=0.0024). Carriers of variant alleles at VEGFR2 H472Q experienced greater risk of developing HT (OR(95%CI) = 2.3(1.2-4.6), n=170, P=0.0154) and HFSR (OR(95%CI) = 2.7(1.3-5.6), n=170, P=0.0136). This study suggests that HT and HFSR may be markers for favorable clinical outcome, HT development may be a marker for HFSR, and VEGFR2 alleles may be related to the development of toxicities during therapy with bevacizumab and/or sorafenib. Impact of ABCB1 allelic variants on QTc interval prolongation: ABCB1 could decrease the intracardiac concentrations of drugs that cause QT-prolongation and cardiotoxicity. In this study, we demonstrate that mice lacking the ABCB1-type P-glycoprotein have higher intracardiac concentrations of a model ABCB1 substrate, romidepsin, that correspond to changes in QT-prolongation from baseline (deltaQTc) over time. Consistent with this observation, we also demonstrate that patients carrying genetic variants that could lower ABCB1 expression in the cardiac endothelium also have higher deltaQTc immediately following romidepsin administration. To our knowledge, this is the first evidence that ABCB1 can limit intracardiac exposure to a drug that mediates QT-prolongation and suggests that certain commonly inherited polymorphisms in ABCB1 may serve as markers for QT-prolongation following the administration of ABCB1-substrate drugs.
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