Eun-Jeong Choi

Association of HLA Alleles with Autoimmune Thyroid Disease in Korean Children

Backgrounds: Data regarding genetics of Hashimoto’s disease (HD) and Graves’ disease (GD) in Korean children are lacking. Methods: 73 patients with autoimmune thyroid disease (AITD; HD 32, GD 41) were recruited. We analyzed human leukocyte antigen (HLA) class I and HLA-DRB1 by PCR-SSP, and compared them with those of 159 controls. Results: In AITD, the allele frequencies of HLA-A*02, -B*46, -Cw*01 and -DRB1*08 were higher and those of HLA-A*30, -B*07, -Cw*07 and -DRB1*01 were lower than in controls. In HD, those of HLA-B*46 and -Cw*01 were higher and those of HLA-DRB1*01 and -Cw*07 were lower than in controls. In GD, those of HLA-A*02, -B*46, -Cw*01 and -DRB1*08 were higher and those of HLA-DRB1*07 and -Cw*07 were lower than in controls. Between HD and GD, there were no significant differences in allele frequencies. The risk of AITD in the presence of both HLA-B*46 and -Cw*01 is higher than in the presence of either allele alone. Conclusion: The susceptible and protectable alleles in HD are similar to those in GD. Coexistence of HLA-B*46 and -Cw*01 may be a genetic gene marker for early-onset AITD in Koreans.

HLA-Cw polypmorphism and killer cell immunoglobulin-like receptor (KIR) gene analysis in Korean colorectal cancer patients

PURPOSE Natural killer cells (NK cells) play important roles in protecting the patient from the early development of cancers, and are activated or inhibited by killer cell immunoglobulin-like receptors (KIR), which bind to HLA class I. In the present study, we investigated the KIR genotype of Korean colorectal cancer patients. METHODS DNA samples were extracted from peripheral blood cell samples taken from Korean colorectal cancer patients and a control group. KIR genes were amplified using PCR-SSP methods, and HLA-Cw genes were characterized using PCR methods. The results were analyzed to assess the difference between colorectal cancer patients and the normal control group. RESULTS In the present study, the frequency of KIR2DS5 (33.2% vs. 20.8%, p-value < 0.007) was higher in the colorectal cancer group, and in the rectal cancer subgroup, the frequencies of KIR3DL1 (93.2%, vs. 98.1%, p-value < 0.05), KIR2DS2 (7.8% vs. 19.5%, p-value < 0.01), and KIR2DS4 (93.2% vs. 98.1%, p-value < 0.05) were lower significantly. The frequencies of HLA-Cw6 (9.1% vs. 15.7%, p-value < 0.05) and HLA-Cw7 (17.4% vs. 27.7%, p-value < 0.02) were lower in the colorectal cancer group. Of the patients with HLA-C1 homozygote, the frequency of KIR2DS2 was decreased significantly (5.8% vs. 14.5%, p-value < 0.004). CONCLUSIONS The frequency of KIR2DS5 is higher in Korean colorectal cancer patients, and in the rectal cancer subgroup, the frequencies of KIR3DL1, KIR2DS2 and KIR2DS4 are lower. Among the patients with HLA-C1 homozygote, the frequency of KIR2DS2 is decreased. Therefore, KIR2DS2 in presence of its ligand (HLA-C1 group) may have a protective effect against colorectal cancer.

Multiplex Genotyping of Cytokine Gene SNPs Using Fluorescence Bead Array

Single nucleotide polymorphisms (SNPs) of genes that affect cytokine production and function are known to influence the susceptibility and progression of immune-related conditions such as infection, autoimmune diseases, transplantation, and cancer. We established a multiplex genotyping method to analyze the SNPs of cytokine genes by combining the multiplex PCR and bead array platform. Thirteen cytokine gene regions, including 20 SNPs, were amplified, and allele-specific primer extension was performed in a single tube. High-quality allele-specific primers were selected for signals greater than 1000 median fluorescence intensity (MFI) for positive alleles, and less than 500 MFI for negative alleles. To select and improve the extension primers, modifications for the reverse direction, length or refractory were performed. 24 primers in the forward or reverse direction step and 12 primers in length or refractory modifications were selected and showed high concordance with results by nucleotide sequencing. Among the 13 candidate cytokine genes, the SNPs of 12 cytokine genes, including IL-1α, IL-1R, IL-1RA, IL-1β, IL-2, IL-4, IL-4Rα, IL-6, IL-10, IL-12, TGF-β1, and TNF-α, were successfully defined with the selected allele-specific primers in healthy Korean subjects. Our genotyping system provides a fast and accurate detection for SNPs of multiple cytokine genes to investigate their association with immune-related diseases and transplantation outcomes.

The HLA-DRB1*09:29 allele identified in a volunteer donor for hematopoietic stem cell transplant

HLA‐DRB1*09:29 differs from HLA‐DRB1*09:01:02:01 in a codon 75 in exon 2.

The HLA-C*07:478 allele identified in a volunteer donor for hematopoietic stem cell transplant

HLA‐C*07:478 differs from HLA‐C*07:02:01:01 in codon 81 in exon 2.

HLA-B*40:330 allele identified in a volunteer donor for hematopoietic stem cell transplant

HLA‐B*40:330 differs from HLA‐B*40:06:01:01 in codons 113 and 116 in exon 3.

MICB Allele Genotyping on Microarrays by Improving the Specificity of Extension Primers

Major histocompatibility complex (MHC) class I chain-related gene B (MICB) encodes a ligand for activating NKG2D that expressed in natural killer cells, γδ T cells, and αβ CD8+ T cells, which is associated with autoimmune diseases, cancer, and infectious diseases. Here, we have established a system for genotyping MICB alleles using allele-specific primer extension (ASPE) on microarrays. Thirty-six high quality, allele-specific extension primers were evaluated using strict and reliable cut-off values using mean fluorescence intensity (MFI), whereby an MFI >30,000 represented a positive signal and an MFI <10,000 represented a negative signal. Eight allele-specific extension primers were found to be false positives, five of which were improved by adjusting their length, and three of which were optimized by refractory modification. The MICB alleles (*002:01, *003, *005:02/*010, *005:03, *008, *009N, *018, and *024) present in the quality control panel could be exactly defined by 22 allele-specific extension primers. MICB genotypes that were identified by ASPE on microarrays were in full concordance with those identified by PCR-sequence-based typing. In conclusion, we have developed a method for genotyping MICB alleles using ASPE on microarrays; which can be applicable for large-scale single nucleotide polymorphism typing studies of population and disease associations.

The HLA-DQB1*04:02:03 allele identified in a volunteer donor for hematopoietic stem cell transplant

HLA‐DQB1*04:02:03 differs from HLA‐DQB1*04:02:01:01 in a codon 62 in exon 2.

The HLA-C*06:66 allele identified in a volunteer donor for hematopoietic stem cell transplant

HLA‐C*06:66 differs from HLA‐C*06:02:01:01 in codon 151 in exon 3.

The HLA-C*03:272 allele identified in a volunteer donor for hematopoietic stem cell transplant

HLA‐C*03:272 differs from HLA‐C*03:03:01:01 in exon 3, codon 163.