M Imad Damaj - US grants

DESCRIPTION (provided by applicant): Despite evidence of strong genetic contributions to the etiology of smoking initiation (SI) and nicotine dependence (ND), we are far from identifying the specific genetic basis of individual susceptibility to ND. This project - to deepen our understanding of how genes contribute to risk for nicotine dependence (ND) - has arisen as an effort to utilize maximally the relatively unique and complementary set of skill of this group of investigators in complex human genetics, animal genetics and nicotine pharmacology. We will validate these putative risk genes using a two-step approach: replication in other human samples and the demonstration in animal models of ND that variants in these genes contribute to neurobiological pathways likely involved in ND. We will first identify promising candidate genes for smoking initiation (SI) and ND by data-mining GWA datasets and the selected candidates will be replicated. Using a mouse model of nicotine withdrawal, we will characterize behavioral QTLs relevant for ND using a recently developed expanded BXD RI mouse strain panel, focusing on strains informative for already identified areas of provisional QTLs. This approach allows us to both validate and refine our mapping of the nicotine behavioral QTL. Furthermore, we will identify candidate genes for ND by combining expression and behavioral genetics analyses in these BXD RI mouse strains. Candidate genes/pathways identified and prioritized from human and mouse studies will be validated by pharmacological or genetic manipulations to alter expression or function of candidate genes in mouse brain and determine effects on behavioral responses to in models of nicotine reward and withdrawal.

? DESCRIPTION (provided by applicant): Lung cancer is currently a leading cause of mortality. One of these serious neurotoxic side effects that develop in lung cancer patients receiving chemotherapeutic agents is chemotherapy-induced peripheral neuropathy (CIPN). Our proposal will be focused on CIPN induced by cisplatin and paclitaxel since these are two of the most effective and widely used chemotherapy drugs in the treatment of common cancers, including non- small cell lung cancer. CIPN can be a dose-limiting factor for chemotherapy or result in premature termination of treatment, thereby influencing survival and quality of life. Currently, there are no effective therapies that deal with the underlying pathogenic mechanisms such as neurodegeneration. In addition, the current symptomatic therapies that deal with painful symptoms of CIPN are generally ineffective. Therefore, the identification of alternative forms of therapy is a crucial medical need. In this application we will focus on nicotinic acetylcholine receptors (nAChRs) modulators, in particular ?7 subtypes, as potential targets for treatment of CIPN. These receptors are expressed by central and peripheral neuronal cells (dorsal root ganglia) involved in pain transmission and by macrophages and other cells involved in the inflammatory responses. In Aim 1, we will test the ability of ?7 nAChR silent agonists to prevent or ameliorate the development of peripheral neuropathy induced by cisplatin and paclitaxel, including well- defined neuropathologies that accompany CIPN. In Aim 2, we will test the ?7 nAChR silent agonists in non-small cell lung cancer cell lines and patient derived tumor cells in culture as well as in tumor bearing mice to eliminate the possibility that these agents stimulate tumor growth or compromise the potency of chemotherapy (cisplatin and paclitaxel). In addition, suppression of CIPN by the ?7 nAChR silent agonists will be confirmed in the tumor bearing mice. If effective treatment/prevention can be identified, it should be possible to treat patients or prolonged periods as dose-dependent neuropathy will not be a limiting toxicity and compliance as well as quality of life will be improved.

PROJECT SUMMARY PROJECT 3 ? ADOLESCENT NICOTINE Nicotine is one of the most commonly used drugs among adolescents. Several studies link tobacco and nicotine use in human adolescence and subsequent problems in adulthood, including later tobacco, alcohol, cocaine and other illicit drug use. Individuals who smoke cigarettes before the age of 15 are estimated to be eighty times more likely to use illegal drugs such as cocaine than those who do not. In addition, studies in animals have shown that adolescent nicotine exposure affects nicotine and cocaine reward and sensitivity in rodents; we have shown that repeated exposure to low doses of nicotine in adolescence clearly induces age- specific enhancement of the rewarding effects of nicotine and other drugs of abuse that persisted long after the termination of nicotine exposure. However, the molecular and genetic mechanisms underlining nicotine- induced enhancements to the reward effects are still unclear. This project will identify risk associated genes and gene networks using highly diverse mouse genetic populations. We will use inbred Collaborative Cross (CC)/ Diversity Outcross (DO) founder and CC strains to measure nicotine reward using the conditioned place preference (CPP) test after exposure to nicotine in early adolescence. These data will be used to conduct genetic correlations across behaviors including impulsivity, cocaine self-administration (IVSA), acute and sensitized cocaine induced locomotor activation and circadian rhythms to determine common genetic architecture shared among traits. We will also study changes in striatal dopamine (DA) signaling in CC strains that exhibit extreme behavioral responses to nicotine. Secondly, we will correlate nicotine CPP behaviors with gene expression changes in the striatum of CC mice to further identify potential candidate genes, gene networks and biological mechanisms that may mediate the effects of adolescent nicotine exposure. Finally, we will map nicotine reward in the CPP test in a large DO cohort after exposure to nicotine during early adolescence. Genotype data will be used to reveal QTLs specific to nicotine in adolescence as well as identify common QTLs shared between impulsivity, cocaine self-administration, cocaine sensitization and circadian rhythms phenotypes generated by the various projects and cores within the Center for Systems Neurogenetics of Addiction. Genetic mapping data along with expression studies will be used to identify candidate genes for validation studies in novel mouse models made through genome editing technologies. If successful, this application promises to have a significant positive impact on human health through the identification of genetically high-risk individuals who would benefit from proactive interventions following nicotine exposure in adolescence.

Paclitaxel is a drug commonly used for the treatment of breast, lung, and ovarian cancer. Peripheral neuropathic pain (CIPN) is one of the most common and serious adverse effects experienced by cancer patients treated with paclitaxel. CIPN can be a dose- limiting factor for chemotherapy, leading to premature termination of treatment, thereby influencing survival and quality of life. Currently, no therapies have been identified that address the underlying pathogenic mechanisms such as neurodegeneration; in fact, the current symptomatic therapies are frequently ineffective in mitigating the painful symptoms of CIPN in the majority of patients. Therefore, the identification of alternative forms of therapy is a crucial medical need. In this application we focus on agonists of peroxisome proliferator-activated receptor alpha (PPAR?) as molecules of potential therapeutic value for suppression of CIPN induced by paclitaxel. The PPAR? receptors are expressed by central and peripheral neuronal cells (dorsal root ganglia) involved in pain transmission and by macrophages and other cells immune involved in the inflammatory responses. In Aim 1, we will test whether fenofibrate, a clinically used non- selective and weak PPAR? agonist, prevents CIPN in the mouse paclitaxel model in naïve mice. In Aim 2, we will characterize the potential of more selective novel PPAR? agonists and modulators to prevent and suppress CIPN induced by paclitaxel in naïve mice. In Aim 3, we will assess the influence of these PPAR? agonists on growth and susceptibility to chemotherapy (paclitaxel) in non-small cell lung tumors in culture, in patient derived tumor cells in culture and in tumor bearing animals. If effective treatment/prevention can be identified, it should be possible to extend the treatment of patients with drugs such as paclitaxel and platinum based compounds, as dose- dependent neuropathy will no longer limit the utility of these therapies.

Paclitaxel is a cytoskeletal drug commonly used for the treatment of breast, lung, and ovarian cancer. Peripheral neuropathic pain (CIPN) is one of the most common and serious adverse effects experienced by cancer patients treated with paclitaxel. CIPN can be a dose-limiting factor for chemotherapy, leading to premature termination of treatment, thereby influencing survival and quality of life. Currently, no therapies have been identified that address the underlying pathogenic mechanisms such as neurodegeneration; in fact, the current symptomatic therapies are frequently ineffective in mitigating the painful symptoms of CIPN in the majority of patients. Therefore, the identification of alternative forms of therapy is a crucial medical need. The primary objective of this proposal is to identify novel genetic factors that contribute to paclitaxel- induced neuropathy in mice. We observed pronounced paclitaxel-induced CIPN in C57BL/6NJ strain but not in the closely related C57BL/6J substrain. Because the parental substrains are nearly genetically identical, quantitative trait locus (QTL) mapping in an experimental F2 cross (Reduced Complexity Cross; RCC) will greatly facilitate the identification of novel genetic factors that underlie differences in CIPN behaviors. In Aim 1, we will use the RCC to map genomic regions, or QTLs, that are causally associated with susceptibility versus resilience to multiple measures of CIPN. In Aim 2, we will conduct transcriptome analysis via mRNA sequencing (RNA-seq) of spinal and peripheral neuronal regions in control mice and paclitaxel-treated mice from the parental male and female C57BL/6J and C57BL/6NJ substrains. The transcriptome in control mice will aid in identifying differentially expressed, candidate CIPN susceptibility genes underlying the QTLs . Genes that are differentially expressed as a consequence of paclitaxel will reveal changes in the transcriptome relevant to central and peripheral neuronal plasticity and the behaviors/changes that support the long-term establishment of CIPN that may be important for treatment reversal. In Aim 3, we will validate candidate quantitative trait genes and functional variants that influence susceptibility to and establishment of CIPN. These studies will provide rapid genetic and neurobiological insight into CIPN. Future studies will test for translational potential in human genetics, human experimental model systems (e.g., hIPSCs), and new potential therapeutics to combat the debilitating side effects of CIPN in cancer patients.

Project Summary Despite evidence of strong genetic contributions to the etiology of nicotine dependence (ND), we are far from identifying the specific genetic bases of individual susceptibility to ND. The primary objective of this proposal is to identify novel genetic factors that contribute to nicotine withdrawal, an important aspect of ND that contribute relapse, in mice. We observed pronounced nicotine withdrawal traits differences in C57BL/6NJ strain but not in the closely related C57BL/6J substrain. Because the parental substrains are nearly genetically identical, quantitative trait locus (QTL) mapping in an experimental F2 cross (Reduced Complexity Cross; RCC) will greatly facilitate the identification of novel genetic factors that underlie differences in withdrawal behaviors. In Aim 1, we will use the RCC to map genomic regions, or QTLs, that are causally associated with susceptibility versus resilience to multiple measures of nicotine withdrawal. In Aim 2, we will conduct transcriptome analysis via mRNA sequencing (RNA-seq) of four brain regions regions in control mice and chronic nicotine-treated mice from the parental male and female C57BL/6J and C57BL/6NJ substrains. The transcriptome in control mice will serve as a useful tool both in identifying candidate genes for future genome editing that are differentially expressed and underlie the behavioral QTLs as well as providing genomic insight into the neuronal context that influences susceptibility versus resilience to nicotine withdrawal. Genes that are differentially expressed as a consequence of nicotine will reveal changes in the transcriptome relevant to central neuronal plasticity and the behaviors/changes that support ND. In Aim 3, we will validate candidate quantitative trait genes and functional variants identified and ranked by Aims 1-2.

Evidence of significant co-occurrence between pain and alcohol consumption are emerging. There is also indication that alcohol can induce acute analgesia with cross- sectional evidence that some individuals may be motivated to use alcohol to cope with pain. However, potential moderators and mechanisms of action remain largely uncharacterized and poorly understood. This proposal will focus on examining potential pharmacological and genetic mechanisms mediating the alcohol-pain connection using the mouse BXD recombinant inbred (RI) panel. The primary objective of this proposal is to identify novel genetic factors that contribute to alcohol acute analgesic effects and development of tolerance in mice. We observed for the first time strain differences between C57BL6/J and DBA/2J for alcohol-induced antinociceptive effects in the hot- plate test after oral administration. In Aim 1, we will examine and characterize alcohol?s analgesic properties in acute pain models after oral dosing in the mouse. In Aim 2, we will use BXD RI lines to map genomic regions, or QTLs, that are causally associated with susceptibility versus resilience to alcohol effects and the development of tolerance in the hot-plate test. In Aim 3, we will identify changes in the transcriptome associated with acute analgesic phenotype and tolerance of alcohol. We will measure changes in gene expression in relevant neuronal tissues (amygdala, periaqueductal grey and prefrontal cortex) associated with alcohol initial sensitivity and tolerance in extreme RI strains. In Aim 4, we will validate candidate quantitative trait genes and functional variants identified and ranked by Aims 2-3.

Project Summary Virginia Commonwealth University (VCU) seeks to renew the Health Educational Research Opportunities for Teachers (HERO-T) program, a two -year research and professional development opportunity for science teachers, and expand the Health Educational Research Opportunity and Exploration for Students (HEROES) enrichment program for secondary students in the Richmond, Virginia Metropolitan Statistical Area. These programs, grounded in theory, are designed based upon a combination of data derived from current literature related to the impact of research experiences for teachers and literature on student engagement in science. The wealth of experience of the investigators, steering committee, and advisory board members will enable the successful implementation of this program. The goals of the HERO-T and HEROES programs are to 1. cultivate in teachers the skills needed to conduct cutting edge biomedical research and translate that research experience into curricula for their students and 2. increase student awareness of and exposure to research in the biomedical sciences and foster in them scientific investigation skills. These goals will be accomplished by: 1. Selecting three new secondary science teachers annually (15 teachers total), HERO-T fellows, to participate in a two year experience consisting of two eight week summer research experiences and a two year professional learning community (PLC) during the school year. 2. Providing teachers with research training, professional development, and coaching. 3. Engaging twenty secondary students annually in the HEROES Academy (up to 100 students total), a monthly enrichment program to build their awareness of careers in biomedicine/ biomedical research and cultivate their scientific investigation skills. 4. Selecting ten talented students annually (50 students total), HEROES scholars, to participate in the HEROES Externship, a one week summer research program, and the Center on Health Disparities Summer Research Symposium. In the first four years of the grant, 12 teachers were trained, 83% from high-need schools and 67% from underrepresented groups. The HERO-T fellows have developed more than 12 lesson plans from their research to be used in an average of 2 classes per fellow per year impacting over 150 students in the first year to now over 1,200 secondary students annually. More than $10,000 has been used to purchase laboratory supplies for the teachers? classes thus amplifying the long lasting impact of their research experience. Thirty-nine students have participated in the HEROES program. Approximately 75% of the students have come from high need schools and 80% are from underrepresented groups. This grant request is for another five years of support.