Modified Project Summary/Abstract Section Long-acting (LA) pre-exposure prophylactic (PrEP) strategies offer the promise of improving adherence to therapeutic regimen for maximal HIV preventive efficacy. To this end, we have developed an ultra-long acting antiretroviral (ARV) delivery implant with an unprecedented release duration of at least 2 years uninterrupted for durable and safe HIV prevention independent of user adherence. The NanoDDI is a refillable subcutaneous nanofluidic implant for constant drug release, which comprises a newly patented nanofluidic membrane, and ports for rapid, minimally invasive transcutaneous drug refilling. Refilling is performed manually via a syringe without any complex pumps or equipment to extend implant use duration beyond 2 years. The nanofluidic membrane use nanochannels to control drug release through passive diffusion without pumping mechanisms, permitting discrete, long-term user-independent dosing. Unlike injectables or other LA polymeric strategies, NanoDDI avoids burst and decay release. Zero-order release kinetics is achieved independent of physiological conditions, regardless of interindividual heterogeneity. Importantly, the NanoDDI addresses user preferences for discretion and longer dose duration. The NanoDDI development was originally planned for use with islatravir (ISL), an ARV developed by Merck. However, ISL clinical development for long-acting HIV PrEP was recently terminated by Merck based on lymphocyte depletion concerns, rendering ISL of limited clinical relevance for PrEP. Thus, we have shifted our NanoDDI focus to the potent ARVs, MK8527 (MK) and lenacapavir (LEN). Here we will test the hypothesis that constant and sustained MK8527 (MK) or lenacapavir (LEN) delivery from NanoDDI will achieve preventative drug levels for a 2-year duration and effectively prevent SHIV infection in non-human primates (NHP). This proposal outlines a comprehensive preclinical framework fundamental for developing NanoDDI as a HIV PrEP platform. We aim 1) to develop and optimize NanoDDI and MK and LEN formulations for sustained and constant release in vitro and in rats; 2) to assess pharmacokinetics (PK), tolerability, and safety of NanoDDI-MK85 and NanoDDI-LEN for 2 years in NHP and evaluate effectiveness of transcutaneous drug refilling; and 3) to comprehensively evaluate PrEP efficacy of NanoDDI-MK in NHPs using the vaginal and rectal routes of simian HIV transmission, in dose de-escalation studies. Our multidisciplinary team has a solid history of collaborative HIV PrEP studies with long-acting drug delivery implants. We will use a milestone-driven research approach to advance the NanoDDI technology towards the ultimate goal of clinical translation.