Genistein for the Reduction of Adverse Cardiovascular Events (GRACE)

A. Rationale Despite the gains in life expectancy afforded by effective antiretroviral therapy (ART), people living with HIV (PLHIV) continue to exhibit persistent immune activation, immune dysfunction and chronic inflammation, all of which contribute to an elevated risk of cardiovascular disease (CVD) [1-5]. To address this excess risk, there is an ongoing need for safe adjunctive therapies capable of reducing inflammation, particularly because conventional anti-inflammatory agents can cause adverse effects or interact with ART [6-9].

. B. Background CVD in PLHIV on ART The World Health Organization estimates that there are 41 million PLHIV with 32 million (~77%) receiving ART and an additional 1.5 million more cases predicted to arise each year (https://www.who.int/teams/global-hiv-hepatitis-and-stis-programmes/hiv/strategic-information/hiv-data-and-statistics). PLHIV on ART represent a state of virologic control without full immunologic or inflammatory normalization, characterized by persistent immune activation, chronic inflammation, and multisystem dysregulation all contributing to increased risk of CVD [1-5, 10-14]. This excess risk is not fully explained by traditional CVD risk factors, such as hypertension, dyslipidemia, diabetes, smoking, obesity, sedentary lifestyle, unhealthy diet and psychosocial stress. Additionally, ARTs themselves are attributed to increased risk of hypertension, dyslipidemia and diabetes [2]. Adding to this complexity, the treatment of dyslipidemia with statins--that have the added benefits of pleiotropic anti-inflammatory off-targets--does not reduce the CVD risk to levels to those of an unaffected population [2] and may increase the incidence of diabetes [7, 15], myalgia, muscle weakness or myopathy [7].

In PLHIV, paramount among non-traditional CVD risk factors is chronic low grade systemic inflammation, characterized by persistently elevated inflammatory and immune activation markers, notably IL 6, TNF α, IL 1β, C reactive protein, D dimer, sCD14, and sCD163 [3, 16, 17]. These abnormalities are driven by multiple overlapping mechanisms, including residual HIV transcription, gut associated tissue damage with microbial translocation, chronic co infections, and incomplete immune reconstitution [18]. Gut barrier dysfunction triggered by early HIV is hypothesized to be the potential underlying cause for ongoing immune dysfunction despite effective ARTs (Fig 1). Growing evidence that gut microbiome associated metabolites regulate gene expression profiles of immune cells linked to inflammation [19]. Sustained immune activation of T cells (e.g., CD38⁺HLA DR⁺), monocytes, and innate immune pathways persists despite undetectable viral load [20]. Immune dysregulation remains, accompanied by reduced naïve T cell pools, immune senescence, and immune exhaustion. Persistent monocyte and macrophage activation has been directly linked to coronary plaque burden and vascular inflammation, reinforcing the connection between immune activation and CVD [1, 2].

These inflammatory and immune abnormalities converge on endothelial dysfunction, a central and early event in CVD pathogenesis. In PLHIV, chronic inflammation and ART exposure impair nitric oxide signaling, increase oxidative stress, and promote arterial stiffness [13]. Endothelial dysfunction is further exacerbated by comorbid exposures and conditions common in this population. Cannabis use, for example, is four times more prevalent among PLHIV [21] and has been associated with vascular inflammation, oxidative stress, and impaired endothelial dependent vasodilation [22-24]. Chronic pain states, in an estimated 25% to 80% of PLHIV on ART, similarly contribute through sustained sympathetic activation and neuroimmune signaling, which impair vascular repair mechanisms [25]. Collectively, these interrelated mechanisms drive the increased burden of CVD and other non AIDS comorbidities in PLHIV receiving ART.

Genistein as a therapeutic for PLHIV at risk of CVD Non immunosuppressive adjunctive therapeutic strategies that safely target upstream inflammation and endothelial dysfunction represent a critical unmet need to reduce CVD in PLHIV. Genistein, a naturally occurring soy derived isoflavone consumed in human diets for millennia, exhibits multi modal anti inflammatory, antioxidant, and vasoprotective properties (Fig 2) [26]. Mechanistically, genistein inhibits key pathways activated in treated HIV: NF κB-dependent transcription, pro inflammatory cytokine signaling (Fig 3), inducible nitric oxide synthase activity, and reactive oxygen species generation. Importantly, genistein does not induce global immune suppression and has demonstrated a favorable safety profile.