Immune effector cells in patients with multiple myeloma (MM) are at the forefront of many immunotherapy treatments, and several methods have been developed to fully utilise the antitumour potential of immune cells. T and NK cell-derived immune lymphocytes both expressed activating NK receptor group 2 member D(NKG2D). This receptor can identify eight distinct NKG2D ligands (NKG2DL), including major histocompatibility complex class I (MHC) chain-related protein A and B (MICA and MICB). Their binding to NKG2D triggers effector roles in T and NK cells. NKG2DL is polymorphic in MM cells. The decreased expression of NKG2DL on the cell surface is explained by multiple mechanisms of tumour immune escape. In this review, we discuss the mechanisms by which the NKG2D/NKG2DL axis regulates immune effector cells and strategies for promoting NKG2DL expression and inhibiting its release in multiple myeloma and propose therapeutic strategies that increase the expression of NKG2DL in MM cells while enhancing the activation and killing function of NK cells.

Natural killer cells possess key regulatory function in various malignant diseases, including acute myeloid leukemia. NK cell activity is driven by signals received through ligands binding activating or inhibitory receptors. Their activity towards elimination of transformed or virally infected cells can be mediated through MICA, MICB and ULBP ligands binding the activating receptor NKG2D. Given the efficiency of NK cells, potential target cells developed multiple protecting mechanisms to overcome NK cells killing on various levels of biogenesis of NKG2D ligands. Targeted cells can degrade ligand transcripts via microRNAs or modify them at protein level to prevent their presence at cell surface via shedding, with added benefit of shed ligands to desensitize NKG2D receptor and avert the threat of destruction via NK cells. NK cells and their activity are also indispensable during hematopoietic stem cell transplantation, crucial treatment option for patients with malignant disease, including acute myeloid leukemia. Function of both NKG2D and its ligands is strongly affected by polymorphisms and particular allelic variants, as different alleles can play variable roles in ligand-receptor interaction, influencing NK cell function and HSCT outcome differently. For example, role of amino acid exchange at position 129 in MICA or at position 98 in MICB, as well as the role of other polymorphisms leading to different shedding of ligands, was described. Finally, match or mismatch between patient and donor in NKG2D ligands affect HSCT outcome. Having the information beyond standard HLA typing prior HSCT could be instrumental to find the best donor for the patient and to optimize effects of treatment by more precise patient-donor match. Here, we review recent research on the NKG2D/NKG2D ligand biology, their regulation, description of their polymorphisms across the populations of patients with AML and the influence of particular polymorphisms on HSCT outcome.

The relationships between polymorphisms of the trans-membrane(TM) region located in the major histocompatibility complex (MHC) class I chain-related gene A (MICA) and Behcet's disease (BD) have been discussed previously, however, the results were contradictory. In this study, we thoroughly assess whether MICA-TM gene variants are associated with BD by means of a systematic review and meta-analysis. Our study focused on the effects of polymorphisms of MICA-A4, A5, A5.1, A6, and A9 from the included articles. Sixteen previous original publications representing 1,555 BD patients and 2,086 unrelated healthy controls analyzed the association of BD with MICA-TM gene polymorphisms. For the five alleles, MICA-A6 showed a strongly positive correlation with BD patients and could be viewed as an increased risk factor of BD (OR = 2.34, 95%CI: 2.02-2.70). Furthermore, MICA-A4, A5, A5.1, and A9 exhibited negative associations with BD (OR = 0.71, 95%CI: 0.58-0.86; OR = 0.75, 95%CI: 0.63-0.90; OR = 0.63, 95%CI: 0.44-0.91; OR = 0.70, 95%CI: 0.58-0.84, respectively). Our meta-analysis confirmed MICA-A6 could be responsible for BD in three ethnic regions and should probably be treated as a risk factor for BD. MICA-A4, A5, A5.1, and A9 could be regarded as protective factors, especially in the Middle East and East Asia.

The role of major histocompatibility complex (MHC) class I chain-related gene A (MICA), a ligand of NKG2D, has been defined in human diseases by its allele associations with various autoimmune diseases, hematopoietic stem cell transplantation (HSCT) and cancer. This study describes a practical system to develop MICA genotyping by allele-specific primer extension (ASPE) on microarrays. From the results of 20 control primers, strict and reliable cut-off values of more than 30,000 mean fluorescence intensity (MFI) as positive and less than 3000 MFI as negative, were applied to select high-quality specific extension primers. Among 55 allele-specific primers, 44 primers could be initially selected as optimal primer. Through adjusting the length, six primers were improved. The other failed five primers were corrected by refractory modification. MICA genotypes by ASPE on microarrays showed the same results as those by nucleotide sequencing. On the basis of these results, ASPE on microarrays may provide high-throughput genotyping for MICA alleles for population studies, disease-gene associations and HSCT.

The NKG2D receptor is expressed on the surface of NK, T, and macrophage lineage cells and plays an important role in antiviral and antitumor immunity. To evade NKG2D recognition, herpesviruses block the expression of NKG2D ligands on the surface of infected cells using a diverse repertoire of sabotage methods. Cowpox and monkeypox viruses have taken an alternate approach by encoding a soluble NKG2D ligand, the orthopoxvirus major histocompatibility complex (MHC) class I-like protein (OMCP), which can block NKG2D-mediated cytotoxicity. This approach has the advantage of targeting a single conserved receptor instead of numerous host ligands that exhibit significant sequence diversity. Here, we show that OMCP binds the NKG2D homodimer as a monomer and competitively blocks host ligand engagement. We have also determined the 2.25-Å-resolution crystal structure of OMCP from the cowpox virus Brighton Red strain, revealing a truncated MHC class I-like platform domain consisting of a beta sheet flanked with two antiparallel alpha helices. OMCP is generally similar in structure to known host NKG2D ligands but has notable variations in regions typically used to engage NKG2D. Additionally, the determinants responsible for the 14-fold-higher affinity of OMCP for human than for murine NKG2D were mapped to a single loop in the NKG2D ligand-binding pocket.

Even when kidney allografts are well matched for human leukocyte antigen (HLA) and anti-HLA antibodies are not detected, graft rejection can still occur. There is evidence that some patients who lose their graft have antibodies specific for major histocompatibility complex (MHC) class I-related chain A (MICA) antigens. We investigated whether mismatching MICA alleles associates with MICA antibody production and graft rejection or dysfunction. MICA and HLA antibody screening in 442 recipients was performed, and specificities were confirmed in a subgroup of 227 recipients using single-antigen multiplex technology. For assignment of MICA antibody specificity, we used three independent assays. In addition, MICA alleles of 227 recipients and donors were determined by DNA sequencing. In all, 17 patients (7.5%) had MICA antibodies, and 13 patients (6%) developed MICA donor-specific antibodies (DSA). Multivariate analysis revealed MICA mismatching, as an independent significant factor associated with the presence of MICA antibodies (p = 0.009), and 14 mismatched MICA residues significantly correlated with MICA antibody production. MICA and HLA antibodies significantly associated with acute rejection (AR) and MICA DSA and HLA DSA correlated with decreased graft function by univariate and multivariate analysis. We conclude that mismatching for MICA epitopes in renal transplantation is a mechanism leading to production of MICA antibodies that associate with AR and graft dysfunction.

Idiopathic pulmonary fibrosis (IPF) is a progressive and lethal lung disorder of unknown etiology. IPF is likely the result of complex interrelationships between environmental and host factors, although the genetic risk factors are presently uncertain. Because we have found that some MHC polymorphisms confer susceptibility to IPF, in the present study we aimed to evaluate the role of the MHC class I chain-related gene A (MICA) in the risk of developing the disease. MICA molecular typing was done by reference strand mediated conformation analysis in a cohort of 80 IPF patients and 201 controls. In addition, the lung cellular source of the protein was examined by immunohistochemistry, the expression of the MICA receptor NKG2D in lung cells by flow cytometry and soluble MICA by ELISA. A significant increase of MICA*001 was observed in the IPF cohort (OR = 2.91, 95% CI = 1.04-8.25; pC = 0.03). Likewise, the frequency of the MICA*001/*00201 genotype was significantly increased in patients with IPF compared with the healthy controls (OR = 4.72, 95% CI = 1.15-22.51; pC = 0.01). Strong immunoreactive MICA staining was localized in alveolar epithelial cells and fibroblasts from IPF lungs while control lungs were negative. Soluble MICA was detected in 35% of IPF patients compared with 12% of control subjects (P = 0.0007). The expression of NKG2D was significantly decreased in gammadelta T cells and natural killer cells obtained from IPF lungs. These findings indicate that MICA polymorphisms and abnormal expression of the MICA receptor NKG2D might contribute to IPF susceptibility.

In a study of mainly paucibacillary leprosy-affected sib-pair families from South India, in addition to the expected associations with the HLA-DRB1 locus, we have identified significant association with a functional variant of the MICA gene as well as a microsatellite in the flanking region of the MICB gene. The associations with MICA and MICB cannot be accounted for by linkage disequilibrium with the HLA class II locus indicating a role in genetic susceptibility to leprosy that is independent of HLA-DRB1. Previous studies have shown that MICA and MICB are expressed on the surface of cells in response to infection, where they are recognized by the NKG2D receptor on gammadelta T cells, CD8+ alphabeta T cells and natural killer cells, all of which contribute to defense against mycobacteria. The MICA*5A5.1 allele, associated here with leprosy susceptibility, encodes a protein lacking a cytoplasmic tail providing a possible mechanism for defective immune surveillance against mycobacteria.

During a study of MICA frequency in a healthy population and a cohort of patients suffering with inflammatory bowel disease, three DNA samples produced unusual reactivity patterns using polymerase chain reaction sequence-specific primers (PCR-SSP). These samples were subsequently characterized by sequence-based typing (SBT). Here, we report the sequence of these three novel MICA alleles.

Since the complete sequencing of a human major histocompatibility complex (MHC) haplotype, interest in non-human leucocyte antigen (HLA) genes encoded in the MHC has been growing. Non-HLA genes, which outnumber the HLA genes, may contribute to or account for HLA and disease associations. Most information on non-HLA genes has been obtained in separate studies of individual loci. To comprehensively address polymorphisms of relevant non-HLA genes in 'conserved extended haplotypes' (CEH), we investigated 101 International Histocompatibility Workshop reference cell lines and nine additional anonymous samples representing all 37 unambiguously characterized CEHs at MICA, NFKBIL1, LTA, NCR3, AIF1, HSPA1A, HSPA1B, BF, NOTCH4 and a single nucleotide polymorphism (SNP) at HLA-DQA1 as well as MICA, NOTCH4, HSPA1B and all five tumour necrosis factor short tandem repeat (STR) polymorphisms. This work (1) provides an extensive catalogue of MHC polymorphisms in all CEHs, (2) unravels interrelationships between HLA and non-HLA haplotypical lineages, (3) resolves reported typing ambiguities and (4) describes haplospecific markers for a number of CEHs. Analysis also identified a DQA1 SNP and segments containing MHC class III polymorphisms that corresponded with class II (DRB3 and DRB4) lineages. These results portray the MHC where lineages containing non-HLA and HLA variants in linkage disequilibrium may operate in concert and can guide more thorough design and interpretation of HLA-disease relationships.

The Major Histocompatibility Complex (MHC) class I chain related (MIC) A and B genes are important additional loci within the MHC. We have developed a MICA and MICB typing system using the polymerase chain reaction with sequence-specific primers (PCR-SSP), which operates under the same conditions as our routine HLA-A, -B, and -C typing method. We designed 95 primers in 84 SSP mixtures for MICA and 39 primers in 29 mixtures for MICB. This detected and differentiated all 55 MICA and 19 MICB alleles (except MICA*00701 from MICA*026, MICA*00201 from MICA*020, and three MICB alleles, which are intronic variations). A computer program confirmed the MICA amplification reactivity of each SSP mixture and evaluated the typing set for MICA allele combination ambiguities. Seventy-six "reference" DNA samples were used for validation: 50 from International Histocompatibility Workshop B lymphoblastoid cell lines (IHW BCLs) and 26 MICA-typed samples from two laboratories. The reference material identified 28 out of the 55 MICA alleles and 13 of the 19 MICB alleles, and directly validated 62 of the 84 MICA and 20 of the 29 MICB SSP mixtures. Our genotyping agreed with 283 out of the 286 (98.95%) MICA and MICB reference laboratories' allele assignments or the consensus assignments. Two of the discrepancies remain unresolved, whereas one was probably due to a reference laboratory's failure to differentiate alleles differing in exon 5 of the MICA gene. A comparison of the MICA and MICB allele assignments between laboratories identified a "disagreement rate" of 19.4% for MICA alleles and 13.1% for MICB alleles. Accordingly, we have compiled "consensus" MICA and MICB genotypes for the 50 IHW BCLs tested, which have been confirmed by our typing. We also typed 166 random blood donors. Their MICA and MICB carriage and allele frequencies and HLA-B, MICA, MICB linkage disequilibrium parameters and haplotype frequencies largely concurred with other published data on United Kingdom subjects, further supporting the validity of our typing system. This PCR-SSP system is a simple, reliable and rapid technique for typing MICA and MICB alleles. It is easily updated as new alleles are identified but clearly requires a continuing validation review until all known MICA and MICB alleles have been identified.

The human major histocompatibility complex (MHC) class I chain-related gene A (MICA) has a high degree of genetic diversity. Several methods have been used in MICA typing. Recent studies reported different results for the same reference cell lines typed by different methods. By searching the GenBank, we found an indel polymorphism in MICA intron 1 corresponding to the area where one of the sequencing-based typing primers used by others is located. We investigated this polymorphism in 43 reference samples by primer cycle sequencing. This approach revealed three haplotype-specific patterns of polymorphisms in intron 1. This study provided evidence that one of the primers commonly used in MICA typing may fail to amplify both alleles in certain heterozygous combinations. Our data showed a correlation between the three patterns in MICA intron 1 and exon 5 short tandem repeat (STR) alleles. Being neutral ones, the intron 1 and STR polymorphisms appeared to mark the ancestral lineages better than the coding region polymorphisms.

The leading model for class I-b evolution suggests non-polymorphic I-b genes evolve by gene duplication from polymorphic I-a genes. We recently found N-formyl peptide-specific orthologs of the class I-b gene H2-M3 in the rodent subfamily Sigmodontinae. To test if sigmodont M3 is a I-b gene, we sequenced M3 from wild cotton rats ( Sigmodon hispidus) diverse at the class II locus, Sihi-DQA. These haplotypes carry a single allele of M3 that closely resembles H2-M3. However, peptide-binding assays showed that cotton rat M3 bound the chemotactic N-formylpeptide fMLF better than did rat or mouse M3. The Ala116-->Lys substitution in cotton rat M3 might enhance binding of fMLF and is one of eight residues of M3 that interact with ligand residues P3 and P4 and that are positively selected, with a d(N) /d(S) ratio of 1.8. Thus, M3 is a class I-b gene in both sigmodontine and murine murids, but positive selection operates on a small subset of residues in the traditionally defined antigen recognition site.

MICB is a member of the MIC (MHC class I chain-related gene) family. Sixteen MICB alleles have been described; however, the functional relevance and population distribution of MICB alleles or their potential association to disease has not yet been evaluated. In this study, we have developed a PCR system using sequence-specific primers (PCR-SSP) that allows unambiguous amplification of all MICB alleles. This approach has been applied to type 100 healthy unrelated individuals from the Spanish population. The extent of polymorphism in this population is lower than that initially expected, and only nine alleles were detected. The alleles MICB01021 (46%), MICB0103101 (13.5%), MICB0104 (13.5%) and MICB0106 (12.5%) were found to be the most frequent alleles. HLA-B and MICA transmembrane polymorphism typing were also performed in this population. Our data showed that MICB is in linkage disequilibrium with MICA and even with HLA-B. Thus, the linkage disequilibrium with MICA and HLA-B suggests that MICB is a potential candidate for those diseases classically associated with HLA class I alleles.

We have developed a method for major histocompatibility complex class I chain-related gene A (MICA) genotyping using multiplexed single nucleotide extension (MSNE) and flow cytometric analysis of an array of fluorescent microspheres. This technique employs a polymerase chain reaction-derived target DNA containing all the polymorphic sites of MICA, synthetic complementary primers, biotinylated dideoxynucleotide triphosphate, fluorescent reporter molecules (streptavidin-phycoerythrin), and thermophilic DNA polymerase. Genomic DNA was amplified by MICA locus-specific primers and the MSNE reactions were carried out in the presence of 30 MSNE primers used to assay polymorphisms in exons 2, 3, and 4 of the MICA genes. Thirty-two previously typed cell lines were used as reference material. The MICA gene frequencies among 201 African-American unrelated donors were determined. Of 51 previously known alleles, 18 were observed in African-Americans, compared to 16 that were found in North American Caucasians and 9 in South American Indians, suggesting a more diversified allelic distribution in African-Americans. MICA*00201 and MICA*00801 were the two most frequent alleles in African-Americans. We observed a high degree of linkage disequilibrium between certain alleles of MICA and of human leukocyte antigen-B in the African-American population. The methodology described here offers a powerful new approach to DNA typing of the MICA alleles.

Major histocompatibility complex class I chain-related genes (MIC) belong to a multicopy gene family located within the HLA class I region of chromosome 6. They encode for proteins that have a completely different organization, expression, and products from classical HLA class I gene products. One member of this family is the MICA gene, which is characterized by its high degree of polymorphism, with over 50 MICA alleles described. Moreover, MICA exon 5 presents a microsatellite polymorphism consisting of a variable number of GCT repeats that encode for 4, 5, 6, 9, or 10 alanine residues, and a variant (MICA A5.1) that includes a nucleotide insertion (GCT-->GGCT). In this study, we report a novel allele in the transmembrane region of the MICA gene consisting of seven GCT repeats found in a family based study of MICA polymorphism in celiac disease.

The ancient disease of leprosy can cause severe disability and disfigurement and is still a major health concern in many parts of the world. Only a subset of those individuals exposed to the pathogen will go on to develop clinical disease and there is a broad clinical spectrum amongst leprosy sufferers. The outcome of infection is in part due to host genes that influence control of the initial infection and the host's immune response to that infection. Identification of the host genes that influence host susceptibility/resistance will enable a greater understanding of disease pathogenesis. In turn, this should facilitate development of more effective therapeutics and vaccines. So far at least a dozen genes have been implicated in leprosy susceptibility and a genome-wide linkage study has lead to the identification of at least one positional candidate. These findings are reviewed here.

The MHC Class I related (MIC) gene family has been shown to be very polymorphic with 46 different MICA alleles being officially named by the WHO Nomenclature Committee for factors of the HLA system to date. We have identified a novel MICA allele, MICA*047, in a Coya American Indian individual from the Jujuy province of north-western Argentina. The novel MICA*047 allele differs from the MICA*030 allele by a single non-synonymous substitution in exon 2, condon 26 GTA-->GGA, Valine to Glycine1.

The human MHC class I chain-related genes (MICA and MICB) are located within the HLA class I region of chromosome 6. Their organization, expression and products differ considerably from classical HLA class I genes. MIC proteins are considered to be markers of "stress" in the epithelia, and act as ligands for cells expressing a common activatory natural killer-cell receptor (NKG2D). Molecular models are now available for the MICA protein, both bound and complexed with NKG2D. MICA molecules appear to be highly flexible and polymorphic, although the functional relevance and implications of their polymorphism have yet to be fully discerned.