Gregory M. Miller, PhD - US grants

The long term objectives of this proposal are to advance our understanding of the role of somatostatin gene expression in pituitary pathogenesis and in the regulation of pituitary tumor hormone hypersecretion. Because somatostatin is a potent antiproliferative agent in human tumors, the genes for somatostatin receptors which mediate its effects are hypothesized to be tumor suppressor genes. The overall goals of this proposal are: 1) to demonstrate that there are differential and pituitary tumor-specific expression of somatostatin receptor subtypes among human pituitary tumors of different phenotypes; 2) to demonstrate that the efficacy of the somatostatin analogue Octreotide to decrease growth hormone secretion and tumor size in patients with pituitary tumors is dependent upon and can be predicted by the expression of specific somatostatin receptor subtype genes; and 3) to obtain genetic evidence that somatostatin receptor subtype genes function as tumor suppressor genes in human pituitary tumors, and that pituitary tumors exhibit allelic loss and/or mutations in somatostatin receptor subtype genes. Identification of a mutant somatostatin receptor subtype gene in pituitary tumors would be the first example of a mutation in an inhibitory G-protein-linked receptor, and may provide a new pharmacological target for therapeutic intervention for patients with pituitary tumors.

[unreadable] DESCRIPTION (provided by applicant): Trace amines, distinguishable from the well-characterized biogenic amines, have been known to exist in mammalian brain for over 25 years. The recent discovery of a family of mammalian trace amine receptors (TARs) presents a significant new resource for exploring the physiological and pharmacological relevance of trace amines in brain (Borowsky et al, 2001). In conjunction with this finding is the unanticipated discovery that drugs of abuse, ranging from amphetamine, MDMA, LSD, as well as dopamine stimulate rat TAR1-mediated cAMP formation at pharmacologically relevant concentrations (Bunzow et al, 2001). Accordingly, trace amine receptors may directly or indirectly contribute to psychostimulant and hallucinogenic drug effects. As human and rodent trace amine receptors diverge in structure, subtype number and brain distribution, we postulate that non-human primates will provide a more suitable model for uncovering the physiological and pharmacological relevance of trace amine subtypes. In this regard, pilot studies indicate a 96% sequence identity for TAR1 in non-human primate and human. To establish fundamental information needed to explore TAR subtype function, we will investigate non-human primate TAR subtype structure, pharmacology, signal transduction and brain distribution. Aim 1 will determine whether human TAR subtypes (TAR1, 3, 4, 5) and structurally related orphan receptors (GPR57, GPR58 and PNR) exist in nonhuman primates. Based on these results, Aim 2 will utilize luciferase assays sensitive to different signal transduction pathways to uncover agonists as well as antagonists that block agonist-induced receptor activation. To characterize the pharmacological profile of these receptors, radioreceptor assays will be developed from lead compound(s) identified from luciferase assays. Aim 3 will map TAR subtype mRNA and protein distribution in primate brain using in situ hybridization, real-time RT-PCR and immunohistochemistry, to discern which receptor subtypes are expressed in brain for further exploration. Our pilot studies suggest that primate TARs are direct targets of psychostimulant drugs of abuse in brain. The proposed studies will form the basis for investigating the physiological and pharmacological relevance of trace amine receptors in a primate model, and may provide novel leads for developing therapeutic agents to treat addiction and possibly neuropsychiatric disorders. [unreadable] [unreadable] [unreadable]

ADRGND; AIDS; AIDS Virus; Acquired Immune Deficiency; Acquired Immune Deficiency Syndrome; Acquired Immune Deficiency Syndrome Virus; Acquired Immuno-Deficiency Syndrome; Acquired Immunodeficiency Syndrome; Acquired Immunodeficiency Syndrome Virus; Adrenal Glands; Adrenals; Affect; Agonist; Alcohols; Alleles; Allelomorphs; Area; Assay; Benzeneethanamine, N,alpha-dimethyl-, (S)-; Bioassay; Biologic Assays; Biological Assay; C-C CKR-5 Gene; C-C Chemokine Receptor Type 5 Gene; C-Fragment Endorphin; CC-CKR-5 Gene; CCCKR5 Gene; CCR-5 Gene; CCR5; CCR5 gene; CD195 Antigen Gene; CHEMR13 Gene; CKR-5 Gene; CKR5 Gene; CMKBR5 Gene; CRISP; Cell Communication and Signaling; Cell Signaling; Cells; Chemical Class, Alcohol; Chemokine (C-C) Receptor 5 Gene; Clinical; Code; Coding System; Computer Retrieval of Information on Scientific Projects Database; Crystal Meth; DNA; Deoxyephedrine; Deoxyribonucleic Acid; Dependence; Desoxyephedrine; Drugs; Endogenous Opiates; Epilepsy; Epileptic Seizures; Epileptics; Funding; Gene Expression; Gene variant; Genetic Diversity; Genetic Variation; Genotype; Grant; HIV; HIV-1 Fusion Co-Receptor Gene; HPA; HTLV-III; Host Factor; Host Factor Protein; Human; Human Immunodeficiency Viruses; Human T-Cell Leukemia Virus Type III; Human T-Cell Lymphotropic Virus Type III; Human T-Lymphotropic Virus Type III; Human, General; Hypophysis; Hypophysis Cerebri; Hypothalamic structure; Hypothalamus; Immune; Immunologic Deficiency Syndrome, Acquired; In Vitro; Infection; Institution; Integration Host Factors; Intracellular Communication and Signaling; Investigators; LAV-HTLV-III; Lipotropin 61-91; Lipotropin Fragment C; Lymphadenopathy-Associated Virus; Macaca mulatta; Man (Taxonomy); Man, Modern; Medication; Methamphetamine; Methylamphetamine; Modeling; Monkeys; N-Methylamphetamine; NIH; National Institutes of Health; National Institutes of Health (U.S.); Nervous System, Pituitary; Opiate Peptides; Opiates; Opioid Peptide; Opioid Receptor; Pharmaceutic Preparations; Pharmaceutical Preparations; Pituitary; Pituitary Gland; Polymorphism, Single Base; Psychoses; Psychotic Disorders; Receptor Gene; Receptor Protein; Receptors, Opiate; Receptors, Opioid, mu; Receptors, mu; Research; Research Personnel; Research Resources; Researchers; Resources; Rhesus; Rhesus Macaque; Rhesus Monkey; SIV; SNP; SNPs; Seizure Disorder; Signal Transduction; Signal Transduction Systems; Signaling; Simian Immunodeficiency Viruses; Single Nucleotide Polymorphism; Source; United States National Institutes of Health; Variation (Genetics); Virus-HIV; allelic variant; beta-Endorphin; beta-LPH (61-91); beta-Lipotropin C Fragment; biological signal transduction; clinical relevance; clinically relevant; drug/agent; endogenous opioid; epilepsia; epileptiform; epileptogenic; hypothalamic; hypothalamic-pituitary-adrenal (HPA) axis; hypothalamic-pituitary-adrenal axis; hypothalmus-pituitary-adrenal axis; novel; protein structure; receptor; receptor expression; response; stem; suprarenal gland

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Alcoholism is a pharmacogenomic disease in which multiple genes each make modest contribution, but interact to render relative protection or relative vulnerability from deleterious consequences of alcohol use. A detailed understanding of the polygenetic contributions to alcoholism can provide the basis for developing pharmacogenomics-based treatment strategies for alcohol-related problems. This grant application focuses on the development of naturally occurring, or naturalistic rhesus monkey models of the human neurogenetic variance underlying alcoholism. The concept of naturalistic modeling of genetic variances is borne out of our findings of novel functional polymorphisms in rhesus monkeys that, although consisting of different alleles than occur in humans, nevertheless share common function and phenotypic association with polymorphisms in orthologous human genes implicated in neuropsychiatric and substance abuse disorders. Accordingly, we predict that rhesus monkeys which harbor an array of functionally parallel allelic variants to those implicated in human alcoholism could be utilized to clarify the genetic interactions influencing disorder variance, and could serve as a preclinical platform for the development of individualized, pharmacogenomics-based treatment interventions. In this regard, naturalistic modeling of the human neurogenetic variance associated with alcoholism in rhesus monkeys would represent the first nonhuman primate model of a polygenetic disorder vulnerability. Our Aims are: 1) to identify novel rhesus monkey allelic variants;2) to functionally assess identified polymorphisms in vitro in comparison to human variants;and 3) to develop genotyping assays for identified functional alleles, genotype NEPRC rhesus monkeys and generate lead data on genotype/phenotype relationships in phenotypically characterized rhesus monkeys at NEPRC. Our long-term ambition is to select cohorts of rhesus monkeys that harbor overlapping yet distinct constellations of disorder-related alleles that mimic in effect human polygenetic variance underlying the phenotypes, behaviors and traits associated with alcoholism. Through the accumulation of multiple shared genotype/phenotype components, naturalistic modeling of human neurogenetic variance in rhesus monkeys can be achieved.

drug screening /evaluation

1,2-Benzenediol, 4-(2-amino-1-hydroxyethyl)-, (R)-; 3,4-Dihydroxyphenethylamine; 3-(2-Aminoethyl)-1H-indol-5-ol; 4-(2-Aminoethyl)-1,2-benzenediol; 5-HT; 5-Hydroxytryptamine; 5HT; 5HT transporter; 5HTT protein; Affinity; Amines; Assay; Autoreceptors; Benzeneethanamine, N,alpha-dimethyl-, (S)-; Binding; Binding (Molecular Function); Bioassay; Biologic Assays; Biological Assay; Brain; Brain region; CRISP; Cell Communication and Signaling; Cell Signaling; Cells; Computer Retrieval of Information on Scientific Projects Database; Corpus Striatum; Corpus striatum structure; Crystal Meth; DAT; DAT dopamine transporter; Deoxyephedrine; Desoxyephedrine; Dopamine; Drugs; Encephalon; Encephalons; Enteramine; Funding; Grant; H 89; Hippophaine; Hydroxytyramine; In Vitro; Institution; Intracellular Communication and Signaling; Investigators; Knockout Mice; Levarterenol; Levonorepinephrine; Medication; Methamphetamine; Methylamphetamine; Mice, Knock-out; Mice, Knockout; Molecular Interaction; Monkeys; N-Methylamphetamine; NET protein, neuronal; NIH; National Institutes of Health; National Institutes of Health (U.S.); Nerve Transmitter Substances; Nervous System, Brain; Neurotransmitters; Noradrenaline; Norepinephrine; Null Mouse; PKA inhibitor; Pathway interactions; Pharmaceutic Preparations; Pharmaceutical Preparations; Phosphorylation; Protein Phosphorylation; Receptor Protein; Research; Research Personnel; Research Resources; Researchers; Resources; Role; Serotonin; Signal Transduction; Signal Transduction Systems; Signaling; Source; Striate Body; Striatum; Synaptosomes; Thalamic structure; Thalamus; United States National Institutes of Health; Wild Type Mouse; biological signal transduction; brain cell; dopamine transporter; dopamine transporter proteins; drug/agent; extracellular; h89; inhibitor; inhibitor/antagonist; monoamine; noradrenaline transporter; norepinephrine transporter; norepinephrine transporter protein; pathway; psychostimulant; receptor; response; serotonin transporter; slc6a2 protein; slc6a5 protein; social role; sodium-dependent noradrenaline transporter; sodium-dependent serotonin transporter; solute carrier family 6 (neurotransmitter transporter, noradrenalin), member 2; solute carrier family 6 (neurotransmitter transporter, norepinephrine), member 5; striatal; synaptoneurosome; thalamic; uptake

Agonist; Amines; Assay; Bioassay; Biologic Assays; Biological Assay; Brain; CRISP; Cell Communication and Signaling; Cell Signaling; Cell/Tissue, Immunohistochemistry; Computer Retrieval of Information on Scientific Projects Database; Disease; Disorder; Encephalon; Encephalons; Funding; Grant; Human; Human, General; IHC; Immunohistochemistry; Immunohistochemistry Staining Method; Immunologic, Luciferase; Institution; Intracellular Communication and Signaling; Investigators; Luciferases; Mammals, Primates; Mammals, Rodents; Man (Taxonomy); Man, Modern; Maps; Messenger RNA; Modeling; NIH; National Institutes of Health; National Institutes of Health (U.S.); Nervous System, Brain; Numbers; Pharmacology; Physiologic; Physiological; Primates; RNA, Messenger; RT-PCR; RTPCR; Receptor Activation; Receptor Protein; Research; Research Personnel; Research Resources; Researchers; Resources; Reverse Transcriptase Polymerase Chain Reaction; Rodent; Rodentia; Rodentias; Signal Transduction; Signal Transduction Pathway; Signal Transduction Systems; Signaling; Source; Structure; System; System, LOINC Axis 4; Therapeutic Agents; Time; United States National Institutes of Health; addiction; base; biological signal transduction; disease/disorder; mRNA; neuropsychiatric; neuropsychiatry; non-human primate; nonhuman primate; novel; protein distribution; receptor; reverse transcriptase PCR

[unreadable] DESCRIPTION (provided by applicant): Serotonergic tone plays an important role in mental health, with dysfunction implicated in many prominent neuropsychiatric disorders including depression, schizophrenia, and addictive personality disorders. This grant investigates neurogenetic variation in the serotonin system by developing and exploiting naturally occurring homologous variation in rhesus monkeys. Our focus is on functional polymorphisms in tryptophan hydroxylase 2 (TPH2), serotonin transporter (SERT) and monoamine oxidase A (MAOA) genes in an effort to more accurately model serotonergic neurogenetic variance associated with human mental health disorders in rhesus monkeys. In Specific Aim 1, we will identify polymorphisms and haplotypes in rhesus monkey TPH2, SERT and MAOA genes, develop detection assays and genotype the NEPRC rhesus monkey colony. In Specific Aim 2, we will functionally assess rhesus monkey TPH2, SERT, and MAOA variants in vitro and compare these to human functional variants in the orthologous genes. Specific Aim 3 focuses on genotype/phenotype relationships in rhesus monkeys by assessing SERT function and SERT mRNA expression in platelets and immortalized B cell lines derived from SERT-genotyped rhesus monkeys, and by assessing cognitive phenotype in TPH2-, SERT-, and MAOA-genotyped rhesus monkeys. The research will develop rhesus monkeys as models of human neurogenetic variance related to phenotypes, behaviors and traits associated with neuropsychiatric and substance abuse disorders and clarify genetic interactions influencing distinct genotype/phenotype relationships influencing mental health. PUBLIC HEALTH RELEVANCE: Studies in this grant identify novel polymorphisms in serotonergic genes in rhesus monkeys. We will assess the function of TPH2, SERT, and MAOA polymorphisms and haplotypes derived from rhesus monkeys in vitro, and compare these to human functional variants in the orthologous genes which are implicated in mental health disorders. We will also assess the association between the identified functional variants in rhesus monkeys and measures of serotonin transporter function ex vivo and cognitive flexibility in vivo. [unreadable] [unreadable] [unreadable]

DESCRIPTION (provided by applicant): Rhesus monkey Trace Amine-Associated Receptor 1 (TAAR1) responds to a wide spectrum of endogenous amines (including the "trace" amines 2-phenylethylamine and tyramine, and the common biogenic amines dopamine, norepinephrine and serotonin), as well as amphetamine-like psychostimulants, including methamphetamine. In rhesus monkey brain, we have shown that TAAR1 mRNA is expressed in monoaminergic regions, and that TAAR1 is co-expressed with and modulates monoamine transporters in monoaminergic neurons. Our studies have demonstrated that TAAR1 is activated along with monoamine autoreceptors by the common biogenic amines, but that only TAAR1 is activated by methamphetamine, resulting in aberrant cAMP accumulation, triggering of cellular phosphorylation events and a consequent deregulation of monoamine transporter kinetic function. We have also recently found that TAAR1 is expressed at substantially high levels in rhesus monkey and human immune cells where it may mediate methamphetamine-induced effects on the immune system and in this regard, may play a role in methamphetamine-associated effects on human and simian immunodeficiency virus infectivity and disease progression. The emerging importance of this receptor in modulating brain monoamine systems and potentially immune cell function provides a strong rationale for determining whether polymorphic variation at the locus is functional and examining the significant similarities between human and rhesus monkeys. In this grant we apply our laboratory's significant expertise in assessing TAAR1 function, polymorphism discovery in rhesus monkey genes associated with drug addiction and neuropsychiatric disorders, and genetic variant functional assessments to initiate investigation of the TAAR1 locus. We will identify and assess functionality of the novel genetic polymorphisms in both the rhesus monkey and human TAAR1 locus to determine whether TAAR1 polymorphisms could contribute to the genetic variability that underlies susceptibility to and/or protection from neuropsychiatric and drug addiction disorders, and potentially, methamphetamine effects on the immune system. This is a completely novel area in which no investigations have been reported, and our preliminary data verifies the existence of polymorphisms in the rhesus monkey TAAR1 locus. PUBLIC HEALTH RELEVANCE: In this grant, we apply our laboratory's significant expertise in assessing the function of Trace Amine Associated receptor 1 (TAAR1) and identifying novel polymorphisms in rhesus monkey genes associated with drug addiction and neuropsychiatric disorders to initiate investigation of the rhesus monkey and human TAAR1 locus. The emerging importance of this receptor in modulating brain monoamine systems and potentially immune cell function provides a strong rationale for determining whether polymorphic variation is functional, and examining the similarities between human and rhesus monkeys.

DESCRIPTION (provided by applicant): My research focuses on the genetic variance that underlies substance abuse and neuropsychiatric disorders, and also the basic neurobiological mechanisms of addictive drugs. Under this award, I will model components of human serotonergic neurogenetic variance underlying drug addiction, alcoholism and neuropsychiatric disorders in rhesus monkeys. Though rhesus monkeys often harbor different alleles than occur in humans, genetic variants in this species have common functionality compared with those in orthologous human genes known to contribute to the variance of neuropsychiatric and addictive disease. I will systematically uncover functionally comparable polymorphisms in rhesus monkey serotonin transporter (SERT), tryptophan hydroxylase 2 (TPH2) and monoamine oxidase A (MAOA) genes, and assess genotype/phenotype relationships relevant to drug addiction, alcoholism and neuropsychiatric disorders. By selecting rhesus monkeys that naturally harbor an array of comparable functional alleles in these serotonergic genes, I will study genetic interactions influencing disorder-related phenotypes and develop a highly translational preclinical platform for the development of pharmacogenomic-informed human personalized medicine. The theme of developing novel therapeutics for addiction and psychiatric disorders extends to a complimentary research focus on the regulation of brain monoaminergic systems and the role of Trace Amine Associated Receptor 1 (TAAR1). TAAR1 is a primary target of monoamines and amphetamine like drugs. Under this award, I will study the role of TAAR1 in brain, the immune system, and the periphery. My recent work on TAAR1 strongly suggests that the receptor is a target for the development of novel therapeutics for neuropsychiatric and addiction disorders and particularly, for methamphetamine addiction. I will pursue these leads through a molecular screening approach and through assessment of novel compounds in vitro and in vivo. Both research programs join with my proposed study of the TAAR1 gene, which has polymorphisms that may underlie differences in receptor function or expression in both humans and rhesus monkeys. Under this award, I will be freed from administrative, committee, core and consortium responsibilities, so that I can devote more time (>75%) to conducting research and to activities that are directly related to the further development of my independent research career and mentoring of trainees. I will also participate in a variety of training experiences in the responsible conduct of research, and create a seminar presentation on research ethics that I will deliver to our community of researchers and students. PUBLIC HEALTH RELEVANCE: Rhesus monkeys have close genetic and physiological similarities with humans that underlie parallel behavioral, developmental, physiological, and metabolic activities. The proposed research identifies genetic variations in rhesus monkey genes that naturally mimick the function of those underlying neuropsychiatric and addictive disorder variance in humans, and models genotype/phenotype relationships relevant to basic neurobiological mechanisms and the pharmacogenomics of drugs of abuse.

DESCRIPTION (provided by applicant): Project Summary Understanding the biological mechanisms that link HIV infection, methamphetamine dependence and consequent changes in the serotonin system that correlate to depression, stress and disruption of the hypothalamic-pituitary-adrenal axis is critical to designing new preventative and therapeutic strategies for both HIV infection and methamphetamine addiction, which have a high co-morbidity. While polymorphic variations in genes that encode the key modulators of the serotonin system, including the serotonin transporter (5-HTT), monoamine oxidase A (MAOA) and tryptophan hydroxylase 2 (TPH2) have been well-documented, epigenetic regulation of these genes is poorly understood. Epigenetics, which is defined as changes in gene expression that take place without a change in DNA sequence, is known to contribute to tissue-type and developmental stage specific gene expression, via an array of molecular modifications to both DNA and chromatin and non- coding RNAs. Borne out of a line of evidence generated in our lab, this grant explores the hypothesis that epigenetic regulation of TPH2, which codes for the rate limiting enzyme in brain serotonin synthesis, may in particular be an underlying mechanism by which HIV infection and METH dependence can cause changes in the serotonin system that lead to altered HPA axis function, neural-immune dysregulation, depression, complications for clinical treatment and ultimately, higher incidence of HIV infection, addiction and mortality. We have recently demonstrated that TPH2 5'-UTR harbors an antisense promoter, which transcribes a non- coding RNA in vitro (Chen and Miller, 2009). In Specific Aim 1, we will validate the existence of this transcript and assess its involvement in the regulation of TPH2 gene expression. We will also assess the involvement of DNA methylation and CCCTC-binding factor (CTCF) in the regulation of TPH2 gene expression. Epigenetic mechanisms are involved in spatiotemporal expression of numerous genes, as well as in environmental regulation of gene expression. Accordingly, Specific Aim 2 will investigate the role of epigenetic modification in the tissue-specific and developmental stage expression of the serotonergic genes, TPH2, 5-HTT and MAOA, as well as in the potential epigenetic regulation of those genes by specific cytokines and methamphetamine. In Specific Aim 3, we will assess the effect of SIV infection and methamphetamine on the epigenetic modification of the serotonergic genes, by comparing DNA methylation of serotonergic genes in postmortem tissues of SIV+ and SIV- rhesus monkeys. We will also perform a pilot study to explore whether methamphetamine exposure in SIV-infected rhesus monkeys exacerbates epigenetic modification of serotonergic genes. We anticipate that findings of this project will help us to better understand underlying biological mechanisms, develop new strategies to manipulate serotonin neurotransmission so as to treat HIV- and methamphetamine- associated neuropsychiatric disturbances better, reduce the spread of HIV and mortality among HIV+ patients, and enhance treatment strategies for methamphetamine addiction and HIV co-morbidity. PUBLIC HEALTH RELEVANCE: Project Narrative Understanding the link between HIV, methamphetamine abuse and changes in the serotonin system is critical to designing new preventative and therapeutic strategies for both AIDS and methamphetamine addiction, which have high co-morbidity. This project will investigate the epigenetic regulation of serotonin neurotransmission and its relevance to HIV and methamphetamine abuse. We anticipate that the research findings generated from this project will provide new insights into the regulation of the serotonin system and will lead to new strategies to treat HIV-infection and methamphetamine addiction and their co-morbidity.

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Understanding the link between HIV, methamphetamine abuse and changes in the serotonin system is critical to designing new preventative and therapeutic strategies for both AIDS and methamphetamine addiction, which have high co-morbidity. This project investigates the epigenetic regulation of serotonin neurotransmission and its relevance to HIV and methamphetamine abuse. We anticipate that the research findings generated from this project will provide new insights into the regulation of the serotonin system and will lead to new strategies to treat HIV-infection and methamphetamine addiction and their co-morbidity.

DESCRIPTION (provided by applicant): Naltrexone, an opioid receptor antagonist, is an FDA-approved drug used for treating heroin overdose and decreasing alcohol intake in alcoholics. Since the mid-1980's, low dose naltrexone (LDN) has been used off- label for treating HIV infection and preventing AIDS. It has a loyal following of patients and doctors who claim remarkable benefits from LDN, yet there has been little scientific data available to validate or refute LDN efficacy. When administered to untreated HIV-infected individuals, LDN appears to stabilize CD4+ T cell counts, preserve lymphocyte responsiveness to mitogens and delay progression to AIDS. Conceivably, when taken in the early evening LDN may work by causing a normalization of a blunted endogenous circadian opioid surge in HIV-infected individuals, which in turn enhances the function of their immune system. We will take advantage of an availability of SIV-infected macaques that have not received any therapeutic confounding treatments for their SIV infection, having served as control animals for SIV vaccine development research led by various investigators who manage their research through the NEPRC. Rather than euthanize these animals, we will rigorously determine the effect of LDN in attenuating AIDS progression and enhancing immune function. Twenty-four SIV-infected rhesus macaques will be treated with 0, 0.05 or 0.3 mg/kg LDN daily (n = 8/dose). Longitudinal measures of viral RNA loads will be collected, and SIV-specific CD4+ and CD8+ T cell responses to the entire SIV proteome will be quantitated by Elispot and intracellular cytokine staining assays using overlapping SIVmac239 peptides. The profile of memory, activation, and exhaustion markers expressed by circulating memory CD4+ and CD8+ T cells will be determined by a polychromatic flow cytometry panel that includes antibodies to CD3, CD4, CD28, CD95, CCR7, CCR5, CD69, HLA-DR and PD-1, and monitored every 2 weeks. A complete blood count analysis will be performed to determine the total lymphocyte population in whole blood. The total lymphocyte population and the frequency of each subset will be used to calculate cell counts for naive, central memory and effector memory CD4+ T cell populations per ¿l of blood at each time point. Alpha interferon levels will also be measured. The mu-opioid receptor is a major target of naltrexone, and specific nonsynonymous polymorphisms in both the human (N40D) and rhesus monkey (P26R) receptor alter ligand binding and predict naltrexone sensitivity (to curtail alcohol consumption) i both humans and rhesus monkeys in a strikingly parallel manner. Accordingly, we will test the hypothesis that rhesus monkey P26R is a determinant of LDN efficacy in curtailing disease progression in the SIV/macaque model. We will assess genotype/phenotype associations and measure endorphin levels during and following treatment with LDN, revealing circadian patterns in the macaque and effects of LDN and SIV infection on these patterns. If LDN can reduce the progression of AIDS in the highly translational SIV-infected macaque model, the research could validate a low cost and safe intervention to curtail disease progression with little or no side effects. PUBLIC HEALTH RELEVANCE: Since the mid-1980's, the opioid antagonist naltrexone has been used at a low dose, off label, to attenuate AIDS progression by a relatively small group of HIV-infected individuals, and while there are strong anecdotal and testimonial reports of major beneficial effects in these patients, there has yet to be formal experimental investigation to validate its efficacy. This grant utilizes SIV-infected rhesus macaques as a translational research model to preclinically assess its efficacy, for the purpose of exploring whether this drug is beneficial in curtailing disease progression and AIDS.