No abstract

CTLA-4 overexpression in CD19 þ /CD5 þ cells correlates with the level of cell cycle regulators and disease progression in B-CLL patients

The results of our study indicate that CTLA-4 (A49G) exon 1 polymorphism is associated with MS progression.

Multiple myeloma (MM) is a B-lineage malignancy characterized by an accumulation of isotype-switched, immunoglobulin-producing monoclonal plasma cells. Additionaly phenotype and functional abnormalities of T cell are observed. The implication of T cells in the development of plasmocytoma has been shown in murine model. The study was undertaken to evaluate the association between polymorphisms of the genes encoding major T cell costimulatory (CD28, ICOS) and downregulatory (CTLA-4) molecules and susceptibility to MM in a Polish population. One hundred patients with MM and 202 healthy subjects were studied. Genomic DNA was isolated from whole frozen blood using the NucleoSpinR Blood kit. Allele identification was achieved by PCR amplification. The amplified product for SNP loci was purified and minisequenced using the commercial kit SNapShot (PE Applied Biosystems). The dinucleotide repeat polymorphism was studied by PCR and fluorescence based technique. The products were analyzed on the ABI PRISM 310 Genetic Analyzer (ABI PRISM 310 capillary electrophoresis system). We found an association of 49G in exon 1 and (AT)82 alleles in 3′untranslated region in the CTLA-4 gene with MM [p=0.018, OR=1.57, 95% CI=1.08–2.29 and p=0.0013, OR=1.85, 95% CI=1.27–2.72, respectively] while no association between T/C substitution in intron 3 of the CD28 gene, the dinucleotide (GT)n repeat polymorphism in intron 4 in ICOS gene and C/T base exchange in the promoter (position −318) in the CTLA-4 gene and MM was observed. Our results indicate that the A/G substitution in exon 1 and microsatelite polymorphism in the 3′untranslated region in the CTLA-4 gene might be a susceptibility locus for MM. The present study is the largest and comprehensive to date to evaluate the association between genetic polymorphisms of genes encoding T cell costimulatory and downregulatory molecules and susceptibility to MM.

There are strong evidences that altered immunological function entails an increased risk of cancer. Cytotoxic T-lymphocyte antigen 4 (CTLA-4), CD28, and Inducible co-stimulator (ICOS) are costimulatory molecules provide regulatory signals for T-cell activation. CD28 and ICOS potently enhance T-cell functions, while activation of T-cells is subsequently downregulated by CTLA-4. The abnormal expression of costimulatory molecules may be caused by polymorphisms of genes encoding these molecules. The extended study was undertaken to evaluate the association between the polymorphisms: CTLA-4+49A/G, CTLA-4-319C/T, CTLA-4(AT)n, CD28+17C/T, ICOS(GT)n and susceptibility to B-CLL in a Polish population. All together 124 B-CLL patients and 202 controls were studied. Single nucleotide polymorphisms were typed by minisequencing. The microsatellite polymorphisms were studied by PCR and fuorescence based technique. The distribution of -319T/C alleles and genotypes differed significantly between groups. The frequency of T allele and T phenotype was increased in patients with respect to controls (p=0.01, OR=1.74, 1.13∼95%CI∼2.68; p=0.01, OR =1.85, 1.13∼95%CI∼3.04, respectively). CTLA-4+49A/G and CTLA-4 (AT)n did not correlate with B-CLL. The analysis of the CD28+17C/T showed that the presence of C allele and C phenotype increased the OR of B-CLL by 1.97 and 2.07, respectively (p=0.002, 1.28∼95%CI ∼3.03; p=0.003, 1.27∼95%CI ∼3.38). For ICOS(GT)n gene polymorphism analysis we combined the alleles into 3 groups: (s)- short-alleles with 8 and 9 repeats, (m) -middle- with 10 and 11 and (l)- long- 12 and more repeats. The global distribution of alleles and genotypes was significantly different in patients and controls (p=0.00008 and p=0.00006, respectively). Individuals possessing (s) alleles were 2.75 times more prone to B-CLL than others (p=0.0000, OR=2.75, 1.67∼95%CI∼4.55), while carrying (l) alleles were protected to B-CLL (p=0.004, OR=0.47, 0.29∼95%CI∼0.79). The genotype (s)/(m) was overrepresented in patients (p=0.004, OR=2.29, 1.28∼95%CI∼2.09), but (l)/(l) individuals appeared more frequently in controls (p=0.002, OR=0.07, 0.00 ∼95%CI∼0.44). The haplotype association study of three polymorphic sites: CTLA-4-319C/T and CD28+17C/T and ICOS (s)/(m)/(l), which correlated with B-PBL in univariante analysis, showed that the distribution of haplotypes was different in cases and control, global p value was p= 3 ×108. Remarkable, the haplotypes T/C/(s) and T/T/(s) were presented only in B-CLL (p=0.0004, OR=62072, 3540∼95%CI∼1088238; p=0.00003, OR=3447, 203∼95%CI∼58334, respectively) while the haplotype C/T/(l) was significantly more frequent in control group as compared to B-CLL (p=0.000003, OR=0.3, 0.18∼95%CI∼0.51). B-CLL patients and controls polymorphic features for CTLA-4-319C/T, CD28IVS3+17C/T, ICOS (GT)n were subjected to multivariate forward stepwise logistic regression analysis. It appeared that C phenotype at CD28 T17int3C site increased twice risk of B-CLL (p=0.004, OR=2.09; 1.26∼95%CI∼3.48), while genotype (m)/(l) or (l)/(l) for ICOS gene protected from disease (p=0,0009, OR=0,41; 0,24∼95%CI∼0,69). We reported for the first time that the gene polymorphism of costimulatory molecules: CTLA-4/CD28/ICOS contribute to susceptibility to B-CLL.