Warfarin Anticoagulation Therapy in Caribbean Hispanics of Puerto Rico: A Candidate Gene Association Study

Karla Claudio-Campos, Aurora Labastida, Alga Ramos, Andrea Gaedigk, Jessicca Renta-Torres, Dariana Padilla, Giselle Rivera-Miranda, Stuart A Scott, Gualberto Ruaño, Carmen L Cadilla, Jorge Duconge-Soler, Karla Claudio-Campos, Aurora Labastida, Alga Ramos, Andrea Gaedigk, Jessicca Renta-Torres, Dariana Padilla, Giselle Rivera-Miranda, Stuart A Scott, Gualberto Ruaño, Carmen L Cadilla, Jorge Duconge-Soler

Abstract

Existing algorithms account for ~50% of observed variance in warfarin dose requirements after including common polymorphisms. However, they do not perform as well in populations other than Caucasians, in part because some ethno-specific genetic variants are overlooked. The objective of the present study was to identify genetic polymorphisms that can explain variability in warfarin dose requirements among Caribbean Hispanics of Puerto Rico. Next-Generation Sequencing of candidate genes CYP2C9 and VKORC1 and genotyping by DMET® Plus Assay of cardiovascular patients were performed. We also aimed at characterizing the genomic structure and admixture pattern of this study cohort. Our study used the Extreme Discordant Phenotype approach to perform a case-control association analysis. The CYP2C9 variant rs2860905, which was found in all the major haplotypes occurring in the Puerto Rican population, showed stronger association with warfarin sensitivity (<4 mg/day) than common variants CYP2C9*2 and CYP2C9*3. Although, CYP2C9*2 and CYP2C9*3 are separately contained within two of the haplotypes, 10 subjects with the sensitive phenotype were carriers of only the CYP2C9 rs2860905 variant. Other polymorphisms in CES2 and ABCB1 were found to be associated with warfarin resistance. Incorporation of rs2860905 in a regression model (R2 = 0.63, MSE = 0.37) that also includes additional genetics (i.e., VKORC1-1639 G>A; CYP2C9 rs1856908; ABCB1 c.IVS9-44A>G/ rs10276036; CES2 c.269-965A>G/ rs4783745) and non-genetic factors (i.e., hypertension, diabetes and age) showed better prediction of warfarin dose requirements than CYP2C9*2 and CYP2C9*3 combined (partial R2 = 0.132 vs. 0.023 and 0.007, respectively, p < 0.001). The genetic background of Puerto Ricans in the study cohort showed a tri-hybrid admixture pattern, with a slightly higher than expected contribution of Native American ancestry (25%). The genomic diversity of Puerto Ricans is highlighted by the presence of four different major haplotype blocks in the CYP2C9 locus. Although, our findings need further replication, this study contributes to the field by identifying novel genetic variants that increase predictability of stable warfarin dosing among Caribbean Hispanics.

Keywords: Caribbean Hispanics; admixture; genotyping; next-generation sequencing; pharmacogenetics; warfarin.

Figures

Figure 1
Figure 1
Histograms of them distribution of warfarin stable doses in (A) Puerto Ricans recruited from VACHS and (B) tested subcohort.
Figure 2
Figure 2
Overview of study methods and analyses. (a)See Duconge et al. PLoS ONE 2016. (b)AIMs stands for ancestral informative markers.
Figure 3
Figure 3
Single nucleotide variants (SNVs) at CYP2C9 associated with warfarin (A) sensitivity (<4 mg/day) and (B) resistance (>6 mg/day) identified with NGS. P-values correspond to case-control association test performed in Puerto Ricans of the present study but colored codes represent correlation of SNVs with rs2860905 among Hispanics from 1,000 Genomes (Puerto Ricans, Colombians, Mexican Americans and Peruvians).
Figure 4
Figure 4
Single nucleotide variants (SNVs) associated with warfarin (A) sensitivity (<4 mg/day) and (B) resistance (>6 mg/day) identified with DMET Plus array.
Figure 5
Figure 5
Representation of the most frequent haplotypes found within the region spanning CYP2C9 (chr10: 96694843 to 96750251) across Puerto Ricans in warfarin and parental populations from the 1000 Genomes Project (Caucasians, Africans and Asians). Each block represents a position within the haplotype (haplotypes 1-4, H1-4). Blue blocks indicate the presence of the variant allele for each of the positions while gray blocks indicate the presence of the wild-type allele. The green block is the presence of the variant rs2860905. Red blocks represent the variants CYP2C9*2 (rs1799853) and CYP2C9*3 (rs1057910). The data was obtained from the 1000 Genomes Project Phase 3 with phased genotypes.
Figure 6
Figure 6
Regression model for warfarin dose requirements among Puerto Ricans. The solid line depicts perfect prediction of the model. The stabilization dose refers to the dose given after three consecutive INRs values within the range (2–3 for most of the indications). The R2 value is adjusted. MAE and MSE stand for mean absolute error and mean standard error respectively.

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