Population differences in platinum toxicity as a means to identify novel genetic susceptibility variants

Abstract

Objectives: Clinical studies show that Asians (ASN) are more susceptible to toxicities associated with platinum-containing regimens. We hypothesized that studying ASN as an 'enriched phenotype' population could enable the discovery of novel genetic determinants of platinum susceptibility.

Methods: Using well-genotyped lymphoblastoid cell lines from the HapMap, we determined cisplatin and carboplatin cytotoxicity phenotypes (IC50s) for ASN, Caucasians (CEU), and Africans (YRI). IC50s were used in genome-wide association studies.

Results: ASN were most sensitive to platinums, corroborating clinical findings. ASN genome-wide association studies produced 479 single-nucleotide polymorphisms (SNPs) associating with cisplatin susceptibility and 199 with carboplatin susceptibility (P<10). Considering only the most significant variants (P<9.99x10), backwards elimination was then used to identify reduced-model SNPs, which robustly described the drug phenotypes within ASN. These SNPs comprised highly descriptive genetic signatures of susceptibility, with 12 SNPs explaining more than 95% of the susceptibility phenotype variation for cisplatin, and eight SNPs approximately 75% for carboplatin. To determine the possible function of these variants in ASN, the SNPs were tested for association with differential expression of target genes. SNPs were highly associated with the expression of multiple target genes, and notably, the histone H3 family was implicated for both drugs, suggesting a platinum-class mechanism. Histone H3 has repeatedly been described as regulating the formation of platinum-DNA adducts, but this is the first evidence that specific genetic variants might mediate these interactions in a pharmacogenetic manner. Finally, to determine whether any ASN-identified SNPs might also be important in other human populations, we interrogated all 479/199 SNPs for association with platinum susceptibility in an independent combined CEU/YRI population. Three unique SNPs for cisplatin and 10 for carboplatin replicated in CEU/YRI.

Conclusion: Enriched 'platinum susceptible' populations can be used to discover novel genetic determinants governing interindividual platinum chemotherapy susceptibility.

Fig. 1

Comparative population cytotoxicities upon treatment of lymphoblastoid cell lines with (a) cisplatin and (b) carboplatin for Asians (ASN), Caucasians (CEU), and Africans (YRI). Each individual cell line is represented by a unique dot. Population medians are shown by horizontal bars through the population clusters. Values corresponding to these medians are shown horizontally across the bottom of the graph.

Fig. 2

Flow diagram of the single-nucleotide polymorphism (SNP)-discovery/replication methods used. Asian (ASN) cells were phenotyped by exposure to cisplatin and carboplatin, separately. Genome-wide association studies (GWAS) were performed in ASN cell lines associating genotype with IC₅₀, and SNPs of interest which associated at P<10⁻⁴ for each drug were retained. Next (analysis pathway 1), the most significant of these ASN SNPs (P<9.99 × 10⁻⁶) were included in a backwards elimination approach to identify SNPs comprising genetic signatures which robustly describe the ASN drug phenotype variation. These final reduced-model SNPs were then further validated by testing their association with the differential expression of any target genes (P<10⁻⁴), and any identified target genes were then interrogated against the original ASN platinum phenotypes to provide final, summary evidence of the importance of these genes and SNPs. In analysis pathway 2, the initial ASN GWAS SNPs were also tested for genotype–phenotype association in an independent set of combined Caucasian (CEU) and African (YRI) cell lines. SNPs replicating (as single SNPs) for each drug at P<0.05 were identified as `cross-population' platinum variants.

Fig. 3

Platinum susceptibility single-nucleotide polymorphisms (SNPs) of interest may function through regulation of a common target gene family. (a) In Asians (ASN), an example SNP of interest derived from the cisplatin analysis (left panel) and one derived from the carboplatin analysis (right panel) are each associated with the differential expression of a member of the histone H3 family of genes (HIST1H3A and HIST1H3I). Baseline histone H3 expression is plotted as a function of genotype for each SNP. (b) ASN cisplatin and carboplatin IC₅₀s are plotted separately as a function of expression for the respective target histone H3 genes. The cytotoxicity phenotypes for both drugs were correlated with histone H3-member expression levels.

Fig. 4

Representative graphical depiction of the genotype–phenotype relationships for one of the SNPs (rs7937567) identified as associated with cisplatin susceptibility (a) and a SNP (rs11120986) associated with carboplatin susceptibility (b) in Asians (ASN; top panels) and replicated in the combined population of Caucasians/Africans (CEU/YRI; bottom panels). Each individual cell line is represented by a unique dot. Genotype groups are shown on the horizontal axis. Genotype group medians are shown by horizontal bars through the respective clusters.