Utilization of the methodology described above will allow the establishment of a clear relation between exposure and outcome, and will also allow a clear delineation of the independent contribution of pre-HCT vs. HCT-related vs. post-HCT exposures. Having such a relationship then allows the exploration of the inter-individual variability in risk, given constant exposure. Such inter-individual variability is clearly observed in patients treated with anthracyclines, where doses exceeding 1000 mg/m2 are tolerated by some, while low doses (<150 mg/m2) result in congestive heart failure in the others. Identifying those at highest risk upfront becomes critical in determining the best therapy for the individual patient, maximizing the chance of cure, while minimizing the long-term toxicity – the basis for “personalized medicine”. In addition, understanding the pathogenetic mechanism underlying the development of an outcome of interest, given a therapeutic exposure would allow for the development of novel therapeutic interventions, allowing for early detection and potential reversal of the process.
Advances in genetics research has also been a significant contributing factor in defining the human major histocompatibility system (HLA) and high impact translational research that has guided donor selection and refined through the development of DNA-based tools for high resolution genotyping robust criteria for the optimal selection of HLA matched unrelated donors.(15
) However, despite precise matching for HLA, graft-versus-host (GVHD) disease, opportunistic infection and other complications remain significant obstacles to safety and overall success. Genetic polymorphism is responsible not only for HLA mismatching between donor and recipient, but also for the mismatching of a potentially large number of minor histocompatibility antigens encoded by genes located across the genome.(17
) Genetic variants encoding minor histocompatibility antigens include single nucleotide polymorphisms (SNPs) responsible for amino acid substitutions in cellular proteins that can function as minor histocompatibility antigens,(18
) or deletions in genes that abrogate protein production and thereby alter donor and recipient disparity for minor histocompatibility antigens as illustrated in a recent paper by McCarroll et al.(19
) In addition to genetic polymorphism causing recipient and donor disparity, certain polymorphisms can also affect gene function. Many SNPs have been identified, both intergenic and in nearby promoter regions, that modify gene function by changing expression levels, modify functional amino acid substitutions, or cause alternative splicing of gene transcripts all of which may alter gene function.(20
In order to develop a deeper understanding of the molecular underpinnings of therapy-related long-term complications, it becomes important to collect appropriate biospecimens from the patients who do and do not develop the outcome. Ideally, this should be in the form of blood, collected to allow subsequent extraction of DNA and RNA, as well as establish lymphoblastoid cell lines. In patients undergoing allogeneic HCT, the blood should be collected prior to HCT, in order to reflect the DNA of the host. Study of GvHD ideally would require the collection of paired DNA from the host and donor. There are two approaches to the study of genetic variation in disease: (1
) candidate gene studies based on the selection of a limited number of tag SNPs for analyzing specific genes and pathways; and (2
) genome-wide association studies (GWAS) using DNA arrays capable of detecting a million or more SNPs.
Both approaches have attendant strengths and limitations. Candidate gene studies are both complementary and additive to genome-wide studies. compares the characteristics of both strategies. A GWAS approach offers the ability to study complex pathways, allowing for an assessment of the action/ interaction of many genes; it also allows for new genes to be identified. GWAS has gained significant favor, partly due to the fact that several studies that have utilized a candidate gene approach have failed replication. However, a GWAS approach requires a large sample size in order to account for false discovery. In addition, there is a need for a replication cohort so that the genes identified in the discovery set can be validated in the test set.
Comparison of Candidate Gene and Genome-Wide Study Approaches
The GWAS approach does not have an a priori hypothesis, and is considered to be hypothesis-generating and more suited for complex disorders where clear etiologic lead is not established. However, this is not true for many post-HCT outcomes, where for the most part there is a clearly established etiologic association between the exposure and outcome (e.g., radiation and subsequent malignancies). In such cases, an argument could be made for a comprehensively selected (and biologically plausible) list of genes identified along the path of the action of radiation on the target organ. As mentioned earlier, GWAS is limited by the need for a large sample size due to issues related to multiple testing – this is less of an issue with a candidate gene approach. Finally, gene-gene interactions may require prohibitively large samples in a GWAS setting, but are logistically feasible when conducting a candidate gene study.