Abstract 930: A simple and inexpensive approach to allelotyping the HLA region in a case-control study of non-Hodgkin lymphoma

Abstract

Abstract Background: Non-Hodgkin lymphoma (NHL) is a heterogeneous malignancy of B- and T-cells. NHL is the fifth most common cancer in the US and NHL incidence rates are increasing, having doubled since the 1970s. As a result, finding causative factors, whether genetic or environmental, remains an important task. The human leukocyte antigen (HLA) region has a long track record of genetic association with autoimmune disease. Our group has recently published findings that indicate a genetic link between follicular lymphoma and a locus at chr6p21.33, in the HLA Class 3 region. Extended haplotypes are typical in this region, and the overall data implicate a possible genetic association between NHL and distant, yet linked, HLA alleles. Due to high genetic complexity, it is difficult to assess allelotype information in the HLA region without using genetic sequencing approaches. However, sequencing is expensive both in terms of dollars and DNA, making it unfeasible for pilot study work. To circumvent these research limitations and quickly and easily explore our hypothesis, we set out to create an assay that is high-throughput, inexpensive, uses minimal DNA, and gives meaningful HLA allelotype information. Results: Using multiplexed ligation-dependent probe amplification (MLPA) we have developed an assay that is able to obtain specific allelic information for one complex HLA gene. One to two digit, bi-allelic typing requires only 100ng of genomic DNA and can be run on 96-well plates over the course of 2 days. Samples are run through an Applied Biosystems 3730xl capillary DNA analyzer to obtain raw data. Analysis is simple relative to sequencing-based typing approaches; there is no need for expensive computer programs and translating raw data to allelotypes is trivial. Compared to sequencing, this is a very affordable assay that gives broad indications of allele associations, making it an ideal methodology for pilot studies. We are currently applying MLPA technology to screen for HLA allelotypes associated with risk of NHL. Discussion: MLPA is a useful tool that can be applied to conduct HLA allelotyping. This technology is an economically feasible approach for large-scale genotyping of closely related alleles and represents a key intermediate research tool that is more specific than genome-wide analysis and more affordable than sequencing. This assay may aid in the discovery of key HLA allelotypes associated with lymphoma, as well as help to eliminate false positive results. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 930.