Genotyping of KIR/KIRL mismatch, FCGR2A, -3A and -3B polymorphisms in patients subject to high-risk neuroblastoma immunotherapy using melting curve analysis

C Jensen; N Siebert; HN Lode

doi:10.1055/s-0033-1343654

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Klin Padiatr 2013; 225 - A37
DOI: 10.1055/s-0033-1343654

Genotyping of KIR/KIRL mismatch, FCGR2A, -3A and -3B polymorphisms in patients subject to high-risk neuroblastoma immunotherapy using melting curve analysis

C Jensen ¹, N Siebert ¹, HN Lode ¹

¹University Medicine Greifswald, Pediatric Hematology/Oncology, Greifswald, Germany

Congress Abstract

Introduction: Immunotherapy with anti-GD2 monoclonal antibody (mAB) ch14.18 prolongs survival of patients with high-risk neuroblastoma (NB). Since clinical response to some mAbs is associated with immunoglobulin-like receptor (KIR)/KIR-ligand (KIRL) mismatch and Fcγ-receptor (FCGR) polymorphisms, we established a Real-Time-PCR (qPCR) analysis of KIR, FCGR2A (H131R), -3A (V158F) and -3B (NA1/NA2) polymorphisms.

Methods and Results: Protocols of qPCR with primers for KIR, FCGR2A, -3A and -3B and subsequent melting curve analysis were established allowing for genotype determination. Importantly, all protocols included primers for an internal positive control gene. Genotyping results were confirmed by TaqMan SNP analysis, Olerup SSP Genotyping® Kit and sequencing. Due to the high allelic variation within the KIRL genes a commercially available kit (Olerup, Austria) was used to determine KIRL genotypes. With this method we analysed genomic DNA isolated from whole blood of a cohort of 41/37 patients treated with ch14.18/CHO.

Conclusion: In summary, we describe fast and reliable genotyping procedures of both KIR/KIRL-mismatch and FCGR-polymorphisms which allow correlation studies with survival following treatments with ch14.18/CHO.