Corticosteroids mediate a variety of immunological actions and are commonly utilized in the treatment of a wide range of diseases. Unfortunately, therapy with this class of medications is associated with a large proportion of non-responders and significant side effects. Inhaled corticosteroids are the most commonly used asthma controller therapy. However, asthmatic response to corticosteroids also varies widely between individuals. We investigated the genetic contribution to the variation in response to inhaled corticosteroid therapy in asthma. The association of longitudinal change in lung function and single nucleotide polymorphisms from candidate genes crucial to the biologic actions of corticosteroids were evaluated in three independent asthmatic clinical trial populations utilizing inhaled corticosteroids as the primary therapy in at least one treatment arm. Variation in one gene, corticotropin-releasing hormone receptor 1 (CRHR1 ) was consistently associated with enhanced response to therapy in each of our three populations. Individuals homozygous for the variants of interest manifested a doubling to quadrupling of the lung function response to corticosteroids compared with lack of the variants (P-values ranging from 0.006 to 0.025 for our three asthmatic populations). As the primary receptor mediating the release of adrenocorticotropic hormone, which regulates endogenous cortisol levels, CRHR1 plays a pivotal, pleiotropic role in steroid biology. These data indicate that genetic variants in CRHR1 have pharmacogenetic effects influencing asthmatic response to corticosteroids, provide a rationale for predicting therapeutic response in asthma and other corticosteroid-treated diseases, and suggests this gene pathway as a potential novel therapeutic target.

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We developed a modified allele-specific PCR procedure for assaying single nucleotide polymorphisms (SNPs) and used the procedure (called SNAP for single-nucleotide amplified polymorphisms) to generate 62 Arabidopsis mapping markers. SNAP primers contain a single base pair mismatch within three nucleotides from the 3Ј end of one allele (the specific allele) and in addition have a 3Ј mismatch with the nonspecific allele. A computer program called SNAPER was used to facilitate the design of primers that generate at least a 1,000-fold difference in the quantity of the amplification products from the specific and nonspecific SNP alleles. Because SNAP markers can be readily assayed by electrophoresis on standard agarose gels and because a public database of over 25,000 SNPs is available between the Arabidopsis Columbia and Landsberg erecta ecotypes, the SNAP method greatly facilitates the map-based cloning of Arabidopsis genes defined by a mutant phenotype.

Under selective pressure from infectious microorganisms, multicellular organisms have evolved immunological defense mechanisms, broadly categorized as innate or adaptive. Recent insights into the complex mechanisms of human innate immunity suggest that genetic variability in genes encoding its components may play a role in the development of asthma and related diseases. As part of a systematic assessment of genetic variability in innate immunity genes, we have thus far have examined 16 genes by resequencing 93 unrelated subjects from three ethnic samples (European American, African American and Hispanic American) and a sample of European American asthmatics. Approaches to discovering and understanding variation and the subsequent implementation of disease association studies are described and illustrated. Although highly conserved across a wide range of species, the innate immune genes we have sequenced demonstrate substantial interindividual variability predominantly in the form of single nucleotide polymorphisms (SNPs). Genetic variation in these genes may play a role in determining susceptibility to a range of common, chronic human diseases which have an inflammatory component. Differences in population history have produced distinctive patterns of SNP allele frequencies, linkage disequilibrium and haplotypes when ethnic groups are compared. These and other factors must be taken into account in the design and analysis of disease association studies.

TBX21 encodes for the transcription factor T-bet (T-box expressed in T cells), which influences naïve T lymphocyte development and has been implicated in asthma pathogenesis. Specifically, the T-bet knockout mouse spontaneously develops airway hyperresponsiveness and other changes consistent with asthma. Because airway responsiveness is moderated by the use of inhaled corticosteroids in asthma, it is conceivable that genetic variation in TBX21 may alter asthma phenotypes in a treatment-specific fashion. Here we demonstrate that the nonsynonymous variation in TBX21 coding for replacement of histidine 33 with glutamine is associated with significant improvement in the PC 20 (a measure of airway responsiveness) of asthmatic children in a large clinical trial spanning 4 years. We note that this increase occurs only in the children randomized to inhaled corticosteroids and that it dramatically enhances the overall improvement in PC 20 associated with inhaled corticosteroid usage. The average PC20 at trial end for subjects on inhaled corticosteroids possessing a variant allele was in the normal range for nonasthmatics. In cellular models, we show that the TBX21 variant increases T helper 1 and decreases T helper 2 cytokine expression comparably with wild type. TBX21 may thus be an important determinant pharmacogenetic response to the therapy of asthma with inhaled corticosteroids. PC20 ͉ pharmacogenetics ͉ T-bet ͉ interaction C orticosteroids mediate a variety of immunological actions and are commonly used in the treatment of a diverse number of diseases. Inhaled corticosteroids are the most effective and commonly used therapy in the management of asthma (1, 2) but may be associated with serious adverse reactions (3). In evaluating asthma therapy response, measures of lung function, such as forced expiratory volume at 1 second (FEV 1 ), and of airway responsiveness, as measured by the provocative concentration of methacholine causing a 20% decrement in FEV 1 (PC 20 ) are commonly used. However, there is large interindividual variation in the FEV 1 and PC 20 responses to inhaled corticosteroids (4, 5). Thus, identifying those patients most likely to benefit from this treatment would be valuable. Because the intraindividual response to inhaled corticosteroid treatment in patients with asthma is highly repeatable (6) and because both FEV 1 and PC 20 are heritable traits (7, 8), a genetic basis for the heterogeneity of this therapeutic response is plausible.The gene TBX21 (GenBank accession no. NM 013351) encodes for transcription factor T-bet (T-box expressed in T cells), which is responsible for the induction of T helper (Th)1 cells and the repression of Th2 cells from naïve T lymphocytes (9). T-bet has been implicated in the pathogenesis of asthma (10, 11). Because the T-bet knockout mouse develops spontaneous airway hyperresponsiveness (10), a phenotype that is modulated by corticosteroids, we assessed the relationship of TBX21 with PC 20 outcomes in asthma. Only one common (estimated heterozygosity Ն 5%) nonsynonymo...

Background-Defensins are antimicrobial peptides that may take part in airway inflammation and hyperresponsiveness.

During acute cytomegaloviral (CMV) mononucleosis, mononuclear leukocytes displayed diminished responsiveness to certain mitogens. Responses to phytohemagglutinin were normal in patients with community-acquired CMV illness but were reduced in patients with transfusion-associated infection. Both groups were hyporesponsive to pokeweek mitogen and concanavalin A. Cells from convalescent CMV mononucleosis patients responded as well as cells from normal donors to the three mitogens. Preculture of cells from patients with acute CMV mononucleosis for up to seven days before addition of concanavalin A greatly enhanced the blastogenic response to that mitogen. The effect was increased further by depletion of adherent cells after preculture for seven days and could be markedly reduced by the addition of fresh, autologous adherent cells to the precultured nonadherent cells. Suppressor activity was not observed in the serum of patients with acute CMV mononucleosis. These studies suggest that mitogen hyporesponsiveness in CMV mononucleosis may be mediated by suppressor cells included among the plastic-adherent mononuclear leukocyte population.

The IL12B gene on chromosome 5q31-33 encodes the p40 subunit of interleukin 12, an immunomodulatory cytokine. To test the hypothesis that the IL12B gene contains polymorphisms associated with asthma, we genotyped six haplotype-tagging polymorphisms in the IL12B gene, both in 708 children enrolled in the Childhood Asthma Management Program (CAMP) and in their parents. Using the family-based association test (FBAT) program and its haplotype (HBAT) and phenotype (PBAT) options, we tested each polymorphism and haplotype for association with asthma and asthma-related phenotypes. We tested positive associations for replication in a case-control study comparing 177 adult moderate-to-severe asthmatics with 177 nonasthmatic controls. In whites in the CAMP cohort, the A allele of the IL12B G4237A polymorphism was undertransmitted to asthmatic children (P=.0008, recessive model), the global test for haplotypes for affection status was positive (P=.009, multiallelic chi (2)), and two polymorphisms were associated with different atopy phenotypes. In addition, we found a strong association between the IL12B_4237 and IL12B_6402 polymorphisms and an asthma-severity phenotype in whites, which we also found in the independent population of white adult asthmatics. IL12B may be an important asthma gene.