Application of a Combination of a Knowledge-Based Algorithm and 2-Stage Screening to Hypothesis-Free Genomic Data on Irinotecan-Treated Patients for Identification of a Candidate Single Nucleotide Polymorphism Related to an Adverse Effect

<div><p>Interindividual variation in a drug response among patients is known to cause serious problems in medicine. Genomic information has been proposed as the basis for “personalized” health care. The genome-wide association study (GWAS) is a powerful technique for examining single nucleotide polymorphisms (SNPs) and their relationship with drug response variation; however, when using only GWAS, it often happens that no useful SNPs are identified due to multiple testing problems. Therefore, in a previous study, we proposed a combined method consisting of a knowledge-based algorithm, 2 stages of screening, and a permutation test for identifying SNPs. In the present study, we applied this method to a pharmacogenomics study where 109,365 SNPs were genotyped using Illumina Human-1 BeadChip in 168 cancer patients treated with irinotecan chemotherapy. We identified the SNP rs9351963 in potassium voltage-gated channel subfamily KQT member 5 (<i>KCNQ5</i>) as a candidate factor related to incidence of irinotecan-induced diarrhea. The <i>p</i> value for rs9351963 was 3.31×10⁻⁵ in Fisher's exact test and 0.0289 in the permutation test (when multiple testing problems were corrected). Additionally, rs9351963 was clearly superior to the clinical parameters and the model involving rs9351963 showed sensitivity of 77.8% and specificity of 57.6% in the evaluation by means of logistic regression. Recent studies showed that <i>KCNQ4</i> and <i>KCNQ5</i> genes encode members of the M channel expressed in gastrointestinal smooth muscle and suggested that these genes are associated with irritable bowel syndrome and similar peristalsis diseases. These results suggest that rs9351963 in <i>KCNQ5</i> is a possible predictive factor of incidence of diarrhea in cancer patients treated with irinotecan chemotherapy and for selecting chemotherapy regimens, such as irinotecan alone or a combination of irinotecan with a KCNQ5 opener. Nonetheless, clinical importance of rs9351963 should be further elucidated.</p></div

Drug metabolic pathways and the drug response of irinotecan.

<p>Drug metabolic pathways and the drug response of irinotecan.</p

Irinotecan-treated cancer patients with SNP information, genetic factor, and clinical parameters for incidence of diarrhea.

<p>“<i>UGT1A1</i>*<i>6</i> or *<i>28</i>” is a genetic factor constructed from 2 polymorphisms (UGT1A1*<i>6</i> and *<i>28</i>); “2” indicates *<i>6</i>/*<i>6</i>, *<i>28</i>/*<i>28</i> or *<i>6</i>/*<i>28</i>, “1” indicates *<i>6</i> or *<i>28</i>, and “0” indicates “other than 2 and 1.” Area: body surface area (m²), PS: performance status, Cr: grade of creatinine, Hg: grade of hemoglobin, Alb: grade of albumin, ALP: grade of alkaline phosphatase, and GOT: grade of glutamic oxaloacetic transaminase. Each laboratory test value (Alb, Hg, GOT, ALP, and Cr) was recorded before the irinotecan therapy. For each type of clinical tests the grade and aberrant values were defined according to the National Cancer Institute - Common Toxicity Criteria (NCI-CTC, Version 2.0). C_max/dose: SN38 C_max/dose [10⁻³×m²/L]. AUC: area under the concentration-time curve. AUC ratio: Ratio of AUC_SN38/AUC_CPT-11. 5-FU: 5-fluorouracil, CDDP: cisplatin, MMC: mitomycin C, VP16: etoposide. * and † indicate <i>p</i><0.05 and 0.05≤<i>p</i><0.10, respectively. For each concomitant drug, 0 means “not administered,” 1 indicates administered.</p

Extracted 7 SNPs with <i>q</i><1 for the second dataset.

<p>RS number: reference SNP identification number in dbSNP, MAF: minor allele frequency, Chr: chromosome number, i.e., a position in human genome GRCh37.p10 build 104, <i>p</i>_F indicates a <i>p</i> value calculated using Fisher's exact test, <i>q</i>_BH indicates adjusted <i>p</i>_F value by the Benjamini-Hochberg method, <i>p</i>_per indicates <i>p</i> values adjusted using a permutation test for multiple testing problems, * indicates <i>p</i>_per<0.05. NearGene-5 indicates that the SNP is within 2 kb upstream of a gene.</p

Contingency tables for rs9351963 in <i>KCNQ5</i> for each model using each dataset.

<p>(A) irinotecan monotherapy (first dataset), (B) any irinotecan chemotherapy (including irinotecan monotherapy; second dataset), and (C) irinotecan combination chemotherapy (excluding irinotecan monotherapy). OR: odds ratio. The <i>p</i> values were calculated using Fisher's exact test. CI: confidence interval.</p

An outline of chemotherapeutic response analysis in irinotecan-treated cancer patients using a combined method consisting of a knowledge-based algorithm for identifying SNPs (KB-SNP), 2 stages of screening, and the permutation test.

<p>(A) The KB-SNP algorithm. (B) Extraction of SNPs using KB-SNP for statistical analysis. (C) Two-stage screening of irinotecan-treated cancer patients. (D) Calculation of the <i>p</i> value in the permutation test based on the 2 stages of screening. The second dataset (includes first dataset) was permutated. The permutated first dataset was extracted from the permutated second dataset. By using Fisher's exact test, SNPs with <i>p</i><0.005 for the first dataset were selected from among 5,242 SNPs. Among the selected SNPs, those with the lowest <i>p</i> value in Fisher's exact test for the second dataset were selected. This procedure was repeated 100,000 times and empirical null distribution was constructed. Using the distribution, the actual <i>p</i> value obtained from the second stage of screening was converted to the adjusted <i>p</i> value (based on correction of multiple testing problems). At these screening steps, allele models were used for each SNP.</p

Comparison of AIC, AUC, and ROC curves for logistic regression models.

<p>(A) Parameters of each model. (B) The ROC curve of a model consisting of rs9351963+MMC+ Amrubicin. ROC: receiver operating characteristic, AUC: area under the ROC curve, NULL indicates the model without parameters. Each genetic factor conforms to the proportional odds model, AIC: Akaike's information criterion, AUC: area under the ROC curve, Sens.: Sensitivity (%), Spec.: Specificity (%).</p