Association of dopamine-related genetic loci to dopamine D3 receptor antagonist ABT-925 clinical response.

ABT-925, a selective dopamine D3 receptor (DRD3) antagonist, was tested in schizophrenia. A DRD3 gene polymorphism results in an S9G amino-acid change that has been associated with lower risk of schizophrenia, higher affinity for dopamine and some antipsychotics, and differential response to some antipsychotics. The effect of S9G genotype on response to ABT-925 was examined. DNA samples (N=117) were collected in a proof-of-concept, double-blind, randomized, placebo-controlled study of ABT-925 (50 or 150 mg QD) in acute exacerbation of schizophrenia. A pre-specified analysis assessed impact of genotype (SS versus SG+GG) on change from baseline to final evaluation for the Positive and Negative Syndrome Scale (PANSS) total score using analysis of covariance with genotype, treatment and genotype-by-treatment interaction as factors, and baseline score as covariate. Significant genotype-by-treatment interaction (P=0.015) was observed for change from baseline to final evaluation for the PANSS total score. Within subgroup analyses showed significant improvement from placebo in the SG+GG group treated with ABT-925 150 mg. More favorable clinical outcomes were observed in patients treated with ABT-925 150 mg who carried the DRD3 G allele than in those who carried the DRD3 SS genotype.

Construction of internal cDNA competitors for measuring IL-10 and IL-12 cytokine gene expression in swine.

A competitive PCR assay (cPCR) was used to quantify swine cytokine responses to parasite infection. Internal standards (deleted cDNA competitor molecules [DcDNA mimics]) were produced and tested for swine interleukin-12 (IL-12), interleukin-10 (IL-10) and hypoxanthine phosphoribosyltransferase (HPRT) from PCR generated cDNA cloned in plasmid vectors. Deletion clones for the cDNA competitor molecules (DcDNA mimics) were generated for IL-10, IL-12 and HPRT by PCR in a single step and verified by (1) amplification of the expected smaller PCR product with the original primers, (2) appropriate fragment size released by restriction digestion of the deleted clone, and (3) correct sequence of the new DcDNA insert. DcDNA mimics were used to quantitate cytokine gene mRNA production during experimental and natural infections of swine with the gastrointestinal nematode parasite Trichuris suis. Mesenteric lymph node cells were collected from control and infected pigs at the time of maximal pathogenicity (35 days after infection) and snap frozen. After RNA extraction, samples were reverse transcribed (RT) to cDNA. cPCR was performed using the housekeeping gene HPRT DcDNA mimic and HPRT specific primers to insure RNA integrity and concentration. Cytokine cDNA content in these samples was then quantitated using cytokine mimics and gene specific primers. IL-10 gene expression in MLN draining the colon of pigs experimentally infected with T. suis increased 10-20 fold at day 35 compared to control pigs. IL-12 gene expression was not detectable in MLN of these pigs, but was detectable in MLN of pigs exposed naturally to T. suis on a contaminated dirt lot that also exhibited signs of secondary bacterial invasion. Swine IL-10 and IL-12 gene expression can be quantitated in local mesenteric tissues. This cPCR assay will enable scientists to quantitate cytokine gene expression in swine and determine the nature of immune responses to important infectious diseases.