Human Cytidine Deaminase (CDA): uncovering the physiological role through its functional genetic polymorphism K27Q

In light of the potential crucial role of CDA polymorphisms in the development of individual therapies designed to maximize response and minimize non-therapeutic side effects, we investigated first of all the CDA polymorphic distributions in healthy individuals from population of central Italy. At this purpose, we developed three polymerase chain reaction (PCR)-based methods for CDA 79AmajorC (K27Q) and CDA 208GmajorA (A70T) genotyping. Until now, these SNPs in the CDA gene have been genotyped by allelic discrimination using a TaqMan assay, or by direct sequencing or by High-resolution melting (HRM) analysis of polymerase chain reaction amplicon [...]. A RFLP (restriction fragment length polymorphism) method using RsrII as restriction enzyme was described only for A70T [...]. Our PCRbased assay [Allele Specific (AS-PCR) and Restriction Fragment Length Polymorphism (RFLP-PCR)] does not require any special equipment, and it provides rapid, reproducible, and cost effective detection of common CDA mutations. It can be carried out efficiently in a standard molecular genetics laboratory and suitable as a preliminary screen for all patients. Furthermore, we performed a new discontinuous direct HPLC assay to assess the cytidine deaminase activity from whole blood of individuals enrolled in this study, correlating it with CDA K27Q genotypes. Since genetic factors are thought to be responsible for up to 50-60% of the rheumatoid arthritis liability [...] in collaboration with Division of Rheumatology, Catholic University of the Sacred Heart (Rome, Italy), we have tested the genetic association between functional polymorphism of CDA (K27Q, rs2072671) and RA by case-control study. Finally, using mutated recombinant proteins, we performed a kinetic and molecular study on the two functional variants Q27 and K27, assessing the biochemical properties of CDA Q* and K* alleles.