In-depth characterization of myeloid cells in human milk from HIV- and HIV+ donors as potential vehicles of and defenders against vertical transmission of HIV: essential data for targeted therapeutics

Project Summary/Abstract Approximately 150,000 vertical HIV transmission events occur annually via breastfeeding; yet, only ~10-15% of infants breastfed by a mother living with HIV will become infected, suggesting components in human milk are partially protective against HIV infection [1, 2]. Human milk is comprised of cells that are >90% viable and remain functional well beyond ingestion, with infants likely to ingest ∼105–108 milk leukocytes daily [3, 4]. Functionality is often in conjunction with milk antibodies (Abs). The level and type of HIV-specific Abs in milk have been correlated with reduced milk vertical transmission (MVT), but what remains largely unexplored is the contribution of the cellular fraction [5]. Abs elicited against HIV need not be neutralizing to exhibit anti-viral effects, and numerous studies have demonstrated Fc-mediated Ab effector functions such as Ab-dependent cellular phagocytosis (ADCP) and Ab-dependent cellular cytotoxicity (ADCC) to be critical in protecting against HIV [6-10]. Our studies using milk obtained from HIV- women in the USA demonstrated milk leukocytes can perform ADCP of HIV targets when elicited by any IgG isotype, mediated by granulocytes as well as various myeloid cell types [11]. This work identified a significant population of atypical CD14- myeloid cells in milk, many of which appeared capable of ADCP. These data call for a comprehensive analysis of milk myeloid cells, especially in the context of their role in MVT of HIV – both as vehicles of and defenders against transmission. Not only must these cells be classified and compared across lactation periods, but their anti-viral functionality and HIV susceptibility determined as well [12]. Herein we intend to analyze milk obtained from donors recruited as part of the UPenn-Botswana Partnership (BUP), a subsidiary of the University of Pennsylvania, and therefore living in a geographic context relevant for MVT. We will compare milk cells from HIV- and HIV+ participants, including a subset of HIV+ participants on antiretroviral therapy (ART) with suppressed viral loads. The proposed project aims to test the hypothesis that transcriptional profiling will reclassify and reveal novel milk myeloid cell subsets unique from those expected in PBMCs, and that certain milk myeloid subsets will be susceptible to HIV infection and/or exhibit Fc-mediated function, thus identifying cellular targets to eliminate or to augment as part of a precision therapeutics strategy for lactating HIV+ women designed to eradicate MVT. As such, we aim to: 1) Conduct an in-depth analysis of human milk myeloid cells obtained from HIV- and HIV+ donors using single cell RNA sequencing (scRNAseq) in order to define cell subsets based on transcriptional profiling; 2) Determine the HIV susceptibility and HIV Envelope expression profile of each newly-defined milk myeloid subset; and 3) Assess the Fc-mediated functionality of each newly-defined milk myeloid subset. The overall goal of this proposed foundational work is to ultimately develop a targeted, therapeutic vaccine for breastfeeding women living with HIV to eliminate MVT. This work is focused on the unique needs of these women and their infants, who are doubly underserved being in a low-income country setting.