Investigators: Robert K. Moyzis, Ph.D., Deborah L. Grady, Ph.D. (University of California, Irvine)
Release Date: TBA
Project 1) Clinical Trials Probands and Families
Cell line establishment funded by NIMH MH060660 (Robert Moyzis, PI) and DOE DE-FG03-97ER62485 (Robert Moyzis, PI), University of California, Irvine School of Medicine
Molecular Genetic Epidemiology Study of ADHD/DRD4 (original 2000 abstract NIMH MH60660)
Utilizing the vast number of marker loci being identified by the Human Genome Project, association studies are being applied to data on many complex disorders. Once an association is identified the issue for investigators is to determine what the association tells us about the disease process. When confirmed by numerous studies, the possibility of the association being a statistical artifact is remote. We are then left with two explanations: the associated allele plays a causative role in the disorder or the allele is in linkage disequilibrium with another locus (polymorphism) which does play a role. In order to truly understand the disorder we will have to distinguish between these, but how?
This project is designed to capitalize on our ability to rapidly and accurately sequence DNA for a locus over a large number of individuals ascertained within a rigorous research design, and apply that information to determine the reason for the association. We have chosen to examine Attention Deficit Hyperactivity Disorder (ADHD) and the dopamine receptor gene DRD4. We first reported the association of DRD4 with the D4.7 allele of this gene, which has now been confirmed by us and five other studies.
ADHD affects about 3-5% of the school-aged population and is considered the most prevalent psychiatric disorder of childhood. Family, twin, and adoption studies have established a strong genetic basis for ADHD. Initial molecular genetic investigations of candidate genes focused on the neurotransmitter dopamine because most patients are responsive to methylphenidate. The DRD4 locus is a highly variable locus where the 7-repeat form of a 48 bp segment occurs in many allelic forms. Other polymorphisms have been identified in DRD4, some of which have known functional significance, but none of which have been evaluated in ADHD studies.
Probands will be ascertained through clinical trials at the Child Development Center, UCI when they meet rigorous diagnostic criteria for ADHD. Proband, their parents and siblings will be studied and their DNA sequenced for DRD4. Other dopamine pathway loci will be genotyped. Within those families who carry D4.7 we will determine the haplotype(s) present in affected individuals for all of the polymorphisms present in the DRD4 gene. For the non-D4.7 families we will test for tight linkage with the DRD4 region. If D4.7 is merely a marker in linkage disequilibrium with a causative polymorphism, then this latter subset of families should also show evidence for linkage.
Human Telomere Mapping and Sequencing (original 1997 abstract DOE DE-FG03-97ER62485)
The Human Genome Project is undergoing a dramatic transition from an emphasis on generating physical maps to the large-scale finished sequencing of the human genome. While perhaps 80% of human DNA can be rapidly sequenced in the next 5-10 years by highly automated, high-throughput sequencing centers, a significant fraction of the human genome will not, I believe, be sequenced to completion by such “brute force” approaches. These are regions that contain: 1) a high percentage of repetitive DNA sequences; 2) internal tandem duplications including multigene families; and/or 3) are unstable in all current sequencing vectors. Such regions will likely “clog” any high throughput sequencing operation, which cannot spend the time and effort necessary to produce a quality product in such difficult DNA regions. This would be irrelevant if such regions were rare, or contained little of intrinsic informational value. Such is not the case. This includes such critical regions as centromeres and telomeres, as well as a greater abundance of low-copy repeats and multigene families than previously anticipated. Producing quality DNA sequence of these regions, which faithfully represent genomic DNA, will be a continuing challenge.
I propose that a large-scale, yet distributive, “boutique” approach to sequencing such regions is warranted, where individual laboratories specialize in genomic regions they have special expertise in investigating. Such efforts would compliment and integrate with the few truly large-scale sequencing centers that are likely to evolve during the next few years, such as the proposed DOE Joint Genome Institute sequencing center. One such “boutique” market is telomeric regions, which exhibit both high levels of repetitive DNA composition and cloning instability. Indeed, great heterogeneity exists in these regions between various individuals. Following the discovery of the human telomere nine years ago, numerous investigations have implicated genes near telomeres as likely targets for alterations during aging and cancer progression. Through the efforts of my laboratory and those of my collaborator, Dr. Harold Riethman, nearly all human telomeres have now been cloned by functional complementation in yeast. My laboratory has finished the 0.23Mb 7q telomere sequence, the first confirmed telomere region to be sequenced directly up to the terminal (TTAGGG)n repeat. Greater than 3Mb of confirmed telomeres are currently available for sequencing, with another 6-12Mb available during the next few years. Due to our extensive experience with these difficult regions, we propose to sequence all human telomeres in the next few years, at a cost competitive with the best sequencing centers world-wide ($0.30/base for finished and annotated sequence). In the process we will “cap” the sequence of all human chromosomes, and identify numerous important genes. An important QC/QA aspect of this proposal is that all sequences will be extensively confirmed against genomic DNA by PCR-sequencing.
Project 2) MTA Probands and Families
Cell line establishment funded by NIH 5R01NS043740-03 (James Swanson, PI, Robert Moyzis, Co-PI) and DOE DE-FG03-97ER62485 (Robert Moyzis, PI), University of California, Irvine School of Medicine
Molecular Genetic of ADHD in the MTA Sample (original 2004 abstract NIH 5R01NS043740-03)
This revised application focuses on extending our prior investigations of genotype-phenotype relationships in Attention Deficit Hyperactivity Disorder (ADHD), the most commonly recognized, diagnosed and treated psychiatric disorder of childhood in the USA (NIH Consensus Conference, 2000).
The Multimodality Treatment of ADHD (MTA) study (see MTA Group, 1999) provides a valuable sample for the proposed research. The MTA sample was defined by 579 ADHD cases and 288 classmate control cases, who were recruited from 7 North American sites and diagnosed by a rigorous evaluation of ADHD symptoms and comorbid conditions. The MTA was designed as a randomized clinical trial of long-term (14 month) treatment with pharmacological and psychosocial interventions. The initial findings (MTA Group, 1999) and ongoing follow-up (MTA Group, 2002) provide extraordinary detail about response to treatment and long-term outcome. Since the MTA follow-up is still in progress, we have easy access to most (about 85%) of the MTA cases and families still in the study. Given these special characteristics, we consider the MTA sample to be a valuable national resource that could be used to increase our understanding of the biological bases of ADHD. So far, genetic factors have not been investigated in this extraordinary sample.
Molecular genetic studies have been performed to evaluate the association of ADHD with candidate genes for the dopamine type 4 receptor (DRD4) and for the dopamine transporter (DAT), based on variable number of tandem repeat (VNTR) polymorphisms. Multiple replications as well as non-replications have been reported and have identified risk alleles: the DRD4 7-repeat (7R) of a 48 bp VNTR and the DAT 10-repeat (10R) of a 40 bp VNTR. Functional significance of these risk alleles has been proposed.
We have four specific aims to investigate molecular genetic bases of ADHD in the MTA sample:
1. To collect DNA from the MTA sample. We propose to obtain DNA from the four treatment groups of children with ADHD and local normative comparison group children (and their parents) who are subjects in the ongoing MTA follow-up study.
2. To replicate and extend association studies of candidate genes. Many investigations have already replicated the DRD4-7R allele association with the categorical diagnosis of ADHD, thus clearly confirming the association, but the association of the DAT-10R allele with ADHD is less clear. We will determine whether the reported associations are replicated in the large MTA sample. In addition to the associations with qualitative diagnosis, in this sample we will extend the findings by using two quantitative traits (severity of behavioral symptoms and reaction time on neuropsychological tests) to evaluate the association patterns with DRD4 and DAT genotypes.
3. To evaluate functional significance of candidate genotype. The MTA follow-up has shown that the most sensitive measure of response to treatment is provided by a “source by domain” assessment based on the SNAP rating scale completed by parents and teachers. SNAP ratings of ADHD and ODD symptoms from the follow-up assessments will be used to evaluate the impact of the specific DRD4 and DAT genotypes on response to the pharmacological and psychosocial treatments of the MTA study and to the long-term outcome of this sample. We will also determine DRD4 haplotypes based on sequence analysis, and tests the hypothesis derived from our recent preliminary studies that subgroups with “old” and “new” haplotypes differ in variability of psychometric measures of ability.
4. To investigate multiple genes in dopamine pathways. In addition to the 48 bp and 40 bp VNTRs of the DRD4 and DAT genes, associations with additional polymorphisms in these and other genes have been suggested by several studies. In this application, we propose to extend our work by (a) evaluating multiple loci for association with the categorical diagnosis or quantitative traits of ADHD and for prediction of responses to treatments, and (b) creating “genomic haplotypes” across combinations of the loci [haplo-specific genotypes at more than one locus simultaneously] to investigate possible interactive roles for multiple polymorphisms in neuroanatomical and neurochemical pathways on manifestation of ADHD symptoms and responses to treatments.
Additional information on the MULTIMODAL TREATMENT STUDY OF ADHD (MTA):
The MTA study was supported by cooperative agreement grants and contracts from NIMH and the National Institute on Drug Abuse (NIDA) to the following: University of California–Berkeley: U01 MH50461, N01MH12009, and HHSN271200800005-C; DA-8-5550; Duke University: U01 MH50477, N01MH12012, and HHSN271200800009-C; DA-8-5554; University of California– Irvine: U01 MH50440, N01MH 12011, and HHSN271200800006-C; DA-8-5551; Research Foundation for Mental Hygiene (New York State Psychiatric Institute/Columbia University): U01 MH50467, N01 MH12007, and HHSN271200800007-C; DA-8-5552; Long Island–Jewish Medical Center U01 MH50453; New York University: N01MH12004, and HHSN271200800004-C; DA-8-5549; University of Pittsburgh: U01 MH50467, N01MH 12010, and HHSN271200800008-C; DA-8-5553; and McGill University N01MH12008, and HHSN271200800003-C; DA-8-5548.
No instrument available