Woong-Ki Kim, Ph.D. - US grants

? DESCRIPTION (provided by applicant): Despite the widespread use of highly active antiretroviral therapy (HAART), HIV-associated neurocognitive disorders remain surprisingly common in both adults and children. Moreover, it is becoming clear that despite HAART, HIV and the closely-related simian immunodeficiency virus (SIV) persist in the brain 'sanctuary', where access of otherwise potent antiretrovirals is limited. However, the mechanisms that establish and maintain HIV infection in the brain during HAART are largely unknown. The overall goal of this proposal is to explore the role of long-lived brain perivascular macrophages (PVM) as a significant source of SIV infection and inflammation in the brain of SIV-infected infant macaques receiving suppressive antiretroviral therapy (ART). In our previous studies, we demonstrated that monocyte turnover predicts the onset of AIDS and is correlated with severity of SIV encephalitis in the SIV macaque model. Recently we have reported for the first time depletion of brain PVM in SIV-infected macaques with intrathecally administered liposome-encapsulated bisphosphonates. Our central hypothesis is that PVM are a significant source of persistent CNS infection and inflammation in HIV-infected newborns starting HAART with high monocyte turnover. The first aim will determine whether initiating ART after the monocyte turnover increases above pre-infection baseline levels will affect PVM infection and brain and CSF viral loads. The second aim will focus on ablating PVM in the setting of ART-treated chronic SIV infection. This study will result in better characterization of brain PVM (e.g., phenotypic markers, turnover and infection) that will help develop eradication therapy, tailored directly for the brain, and for targeting these cells selectively. The research proposed in this application is innovative because it represents an entirely novel departure from the current approach to maintaining viral suppression in HIV- infected children. Our contribution here will be significant because it is a first step toward the development of therapeutic strategies for targeting virus-infected PVM or inhibiting viral infection of PVM. Once such strategies become available, there is promise that persistent HIV reservoirs could be eradicated from brain and other tissues.

? DESCRIPTION (provided by applicant): Despite the widespread use of highly active antiretroviral therapy (HAART), HIV-associated neurocognitive disorders remain surprisingly common. Moreover, it is becoming clear that despite HAART, HIV and the closely-related simian immunodeficiency virus (SIV) persist in the brain 'sanctuary', where access of otherwise potent antiretrovirals is limited. However, the mechanisms that establish and maintain HIV infection in the brain despite HAART are largely unknown. The overall goal of this proposal is to explore the role of long-lived brain perivascular macrophages (PVM) as the target cells for SIV infection in the central nervous system during the initial peak viremia and as a major reservoir of virus during chronic infection in the presence of HAART. In our previous studies, we demonstrated that monocyte turnover predicts the onset of AIDS and is correlated with severity of SIV encephalitis in the SIV macaque model. Recently we have reported for the first time depletion of brain PVM in SIV-infected macaques with intrathecally administered liposome-encapsulated bisphosphonates. Our central hypothesis is that HIV infection of PVM contributes to persistent brain infection and inflammation despite HAART and correlates with increased turnover of blood monocytes representing systemic infection of tissue macrophages. The first aim will determine whether eliminating availability and infection of CD4 T cells in acute infection will affect de novo infection of PVM, and whether depletion of PVM in the post-peak phase of acute infection will decrease DNA proviral load in the brain. The second aim will determine whether initiating antiretroviral therapy (ART) after the monocyte turnover increases above pre- infection baseline levels will affect PVM infection, and brain and cerebrospinal fluid viral loads. The third aim will focus on ablating PVM in the setting of ART-treated chronic infection. The research proposed in this application is innovative because it represents an entirely novel departure from the current approach to maintaining viral suppression in HIV-infected patients. Our contribution here will be significant because it is a first step toward the development of therapeutic strategies for targeting virus-infected PVM or inhibiting viral infection of PVM. Once such strategies become available, there is promise that persistent HIV reservoirs could be eradicated from brain and other tissues.

? DESCRIPTION (provided by applicant): Despite the widespread use of highly active antiretroviral therapy (HAART), HIV-associated neurocognitive disorders remain surprisingly common. Moreover, it is becoming clear that despite HAART, HIV and the closely-related simian immunodeficiency virus (SIV) persist in the brain 'sanctuary', where access of otherwise potent antiretrovirals is limited. However, the mechanisms that establish and maintain HIV infection in the brain despite HAART are largely unknown. The overall goal of this proposal is to explore the role of long-lived brain perivascular macrophages (PVM) as the target cells for SIV infection in the central nervous system during the initial peak viremia and as a major reservoir of virus during chronic infection in the presence of HAART. In our previous studies, we demonstrated that monocyte turnover predicts the onset of AIDS and is correlated with severity of SIV encephalitis in the SIV macaque model. Recently we have reported for the first time depletion of brain PVM in SIV-infected macaques with intrathecally administered liposome-encapsulated bisphosphonates. Our central hypothesis is that HIV infection of PVM contributes to persistent brain infection and inflammation despite HAART and correlates with increased turnover of blood monocytes representing systemic infection of tissue macrophages. The first aim will determine whether eliminating availability and infection of CD4 T cells in acute infection will affect de novo infection of PVM, and whether depletion of PVM in the post-peak phase of acute infection will decrease DNA proviral load in the brain. The second aim will determine whether initiating antiretroviral therapy (ART) after the monocyte turnover increases above pre- infection baseline levels will affect PVM infection, and brain and cerebrospinal fluid viral loads. The third aim will focus on ablating PVM in the setting of ART-treated chronic infection. The research proposed in this application is innovative because it represents an entirely novel departure from the current approach to maintaining viral suppression in HIV-infected patients. Our contribution here will be significant because it is a first step toward the development of therapeutic strategies for targeting virus-infected PVM or inhibiting viral infection of PVM. Once such strategies become available, there is promise that persistent HIV reservoirs could be eradicated from brain and other tissues.

Project Summary Despite the advent of highly active antiretroviral therapy (HAART), HIV-associated neurocognitive disorders remain surprisingly common. HIV and the closely-related simian immunodeficiency virus (SIV) may persist in the brain ?sanctuary?, where access of otherwise potent antiretrovirals is limited. It is now becoming clear that myeloid cells support HIV/SIV infection independently of CD4 T cells and can be the source of rebound virus in tissues including brain upon cessation of suppressive antiretroviral therapy (ART). To date, however, therapeutic strategies for targeting HIV in the myeloid cells and in the central nervous system (CNS) have not yet been developed. The overall goal of this proposal is to lead collaborations to exploit the known pharmacological colony-stimulating factor 1 receptor (CSF1R) inhibition for macrophage targeting to target long-lived infected CSF1Rhigh myeloid cells in the CNS. Recently, we found, for the first time, overexpression and activation of the CSF1R in CNS myeloid cells including perivascular macrophages (PVM) and activated microglia in SIV-infected macaques with encephalitis, as well as in the brain of virally suppressed HIV patients. We also found that CSF1R blockade in vitro selectively ablated rhesus monocyte-derived CSF1Rhigh macrophages. Our central hypothesis is that resident CSF1Rhigh myeloid cells in the brain contribute to persistent HIV brain infection and neuroinflammation despite HAART. Consequently, selective targeting of infected myeloid cells by CSF1R signaling blockade will eliminate the persistent viral reservoir from the CNS. The first aim will determine whether ablation of CSF1Rhigh myeloid cells in the CNS in during acute infection will decrease DNA proviral load in the brain. The second aim will focus on ablating the CSF1Rhigh brain myeloid cells in the setting of ART-treated chronic infection. The research proposed in this application is innovative because it represents an entirely novel departure from the current approach to maintaining viral suppression in HIV-infected patients. Our contribution here will be significant because it is a first step toward the development of therapeutic strategies for targeting virus-infected CNS myeloid cells or inhibiting viral infection of myeloid cells in the CNS. Once such strategies become available, there is promise that persistent myeloid HIV reservoirs could be eradicated from brain and other tissues.

Abstract Although people living with HIV (PLWHIV) have comparable life-expectancy as the HIV-negative population does, their life-quality is still deeply compromised due to the prevalence of neuropsychiatric disorders including depression and anxiety. The main pathological changes in the brains of those people include aberrant microglial (Mg) activation and neuronal injuries. Drug abuse is a high comorbidity of HIV infection and abused drugs could exaggerate the existing neurologic complications in PLWHIV. The detailed mechanisms underlying such phenomenon remain much elusive. Several contributing factors have been suggested for such neurological disorders including the continued expression HIV proteins such as trans-activator of transcription (TAT), the long-term use of antiretrovirals (ARVs), and drugs of abuse. Our preliminary data demonstrated that :1) NLRP3 inflammasome signaling was involved in HIV-TAT-mediated Mg activation; 2) cocaine activated Mg through dysregulating miR-124/TLR4 axis, which could be reversed by inhibition of NLRP3 inflammasome; 3) combination ARVs used in clinical practice (tenofovir:TFV, emtricitabine:FTC, and dolutegravir:DTG) could activate Mg via lysosomal dysfunction; 4) co-exposure of Mg to three agents (TAT, cocaine, and ARVs) intriguingly resulted in increased activity of NLRP3 inflammasome in vitro; 5) IL1?, the final executor of NLRP3 inflammasome activation, decreased the spine density and increase the glutamate receptor ionotropic NMDA subunits (Grins) in vitro; and 6) increased NLRP3 inflammasome activity was shown in the brains of SIV-infected macaque. Based on these findings and two distinct steps of NLRP3 inflammasome activation, we hypothesize that exacerbated NLRP3 inflammasome activation in the context of HIV-TAT/HIV, cocaine, ARVs will lead to exaggerated Mg activation and neuronal injuries, which are responsible for the high incidence of neuropsychiatric disorders in PLWHIV with cocaine use. We will test this hypothesis in the following two specific aims (SA) using complimentary in vitro and in vivo approaches. SA1: Investigate the role of NLRP3 inflammasome in Mg activation and neuronal injuries in the context of HIV-TAT/HIV, cocaine, & ARVs. We will split this SA into three sub aims. SA1A will explore the detailed mechanisms responsible for exaggerated NLRP3 inflammasome activation in vitro; SA1B will explore the mechanisms underlying NLRP3 inflammasome-mediated neuronal injuries; and SA1C will investigate the status of NLRP3 inflammasome and neuronal injuries in archived SIV- infected macaque brains and HIV-infected individuas with or without cocaine use. SA2: Explore the potential therapeutic effects of NLRP3 inflammasome inhibition on neuropsychiatric behaviors in HIV iTat mice in vivo.