Ragon Institute of MGH, MIT and Harvard

Cytokines dimerize two receptor chains to activate Janus kinases and STAT transcription factors that regulate immune cells but have therapeutic liabilities. We engineered “Trikines” to compel cis formation of three-chain cytokine receptor complexes at the cell surface that induce bespoke STAT transcriptional signaling programs optimized for therapeutic efficacy. Designed Trikines co-activated pSTAT5 and pSTAT3 signatures distinct from any natural cytokines, by assembling trimeric combinations of Interleukin-2 (IL-2), Interleukin-10 (IL-10), and Interleukin-21 (IL-21) receptors. An IL-2-based-Trikine restrained terminal differentiation of T cells, promoted stemness, and enhanced durability of tumor control without toxicity. Unexpectedly, an IL-10-based Trikine induced immune infiltration into poorly immunogenic tumors, showing striking efficacy in small cell lung cancer and pancreatic cancer models. Trikines obviate the need for cell engineering to customize STAT signatures for immunotherapy.

The HIV-1 pandemic is characterized by extensive viral genetic diversity, with strains that can be classified into distinct clades or subtypes based on sequence relatedness. Biological differences between subtypes have also been reported. HIV-1 subtype B, predominant in North America and Europe is most studied, but non-B subtypes represent the majority of infections globally and in sub-Saharan Africa, which accounts for more than two-thirds of people living with HIV. Whereas the implications of genetic and biological diversity overall for HIV prevention, treatment and cure strategies are recognized, the impact of clade differences is contested. Here we review how viral and host diversity, including subtype-specific differences may shape HIV pathogenesis, reservoir characteristics and cure strategies, highlighting data from regions where non-B subtypes circulate. Studies from African and global cohorts highlight differences in epidemiological spread and disease progression among HIV-1 subtypes that may be attributed to viral genetic sequence variations and resultant distinct properties in viral replication capacity, immune evasion, interferon resistance, co-receptor usage, latency regulation and reservoir biology. HIV-1 diversity has implications for virus biology, transmission and clinical outcomes. Clade-specific differences therefore warrant consideration in the development and design of prevention, treatment and cure strategies, including diagnostic and monitoring assays. Incorporating multi-clade research and regionally relevant cohorts will advance and accelerate efforts toward a universally applicable HIV cure.

Detecting antigen-specific lymphocytes is crucial for immune monitoring in vaccination, infection, cancer and autoimmunity. However, their low frequency and dispersed distribution make reliable detection challenging. We developed a strategy exploiting the functions of tissue-resident memory T cells (TRMs) to locally concentrate target circulating immune cells in the skin, enabling their non-invasive sampling using a microneedle (MN) skin patch. TRMs were first induced by antigen sensitization and subsequently restimulated by intradermal inoculation of the same antigen to trigger the 'alarm' and immune recruitment functions of these cells, leading to accumulation of antigen-specific T cells from the circulation over several days. In mouse vaccination models, MN patches with optimized hydrogel coatings effectively isolated thousands of live antigen-specific lymphocytes and innate immune cells. In a human participant with allergic contact dermatitis, allergen-induced TRM stimulation followed by MN application recovered diverse lymphocyte populations absent from untreated skin sites. These results establish TRM restimulation coupled with MN sampling as a non-invasive platform to monitor both local and systemic antigen-specific immune responses across disease or vaccination settings.

Broadly neutralizing antibodies (bnAbs) are evaluated as possible alternatives to standard antiretroviral treatment (ART) for maintaining control of HIV-1 replication and may enhance immune responses to reduce or control the viral reservoir. However, the immunological and virological effects of bnAbs in infants and children are unknown. We conducted a detailed analysis of proviral reservoir dynamics and antiviral immune responses in a unique group of young children from Botswana who started ART at birth and then stopped standard ART while receiving the bnAbs 10-1074 and VRC01-LS in a subsequent clinical trial. No quantitative changes in frequencies of proviral sequences were observed during bnAb treatment, but selection of genome-intact proviruses in transcriptionally repressive heterochromatin regions occurred in some study participants. Faster viral rebound following standard ART cessation was linked to elevated proportions of KIR2DL1-positive NK cells. In contrast, delayed viral rebound and more limited viral reservoir size were associated with elevated proportions of NKG2A-positive NK cells and with the HLA-B-21M signal peptide polymorphism. HIV-specific T cell responses were low in all study participants and unrelated to viral reservoir sizes or clinical outcomes following ART interruption. These results suggest that, in young children, specific NK cell subsets and KIR-HLA interactions might be linked to HIV-1 rebound kinetics after substitution of standard ART with bnAbs.

Background/Objectives: The rapid development of safe and efficacious vaccines is often hindered by extensive, mandated non-clinical safety evaluations in animals. With the aim to provide scientific evidence supporting a "vaccine platform approach", here we present the complete non-clinical studies for two investigational vaccines, GRAd-COV2 and GRAdHIVNE1, based on GRAd, a gorilla-derived group C adenoviral vector. Methods: The biodistribution of GRAd genomes following the intramuscular administration of the vaccines was assessed in rats by a sensitive qPCR method. Local tolerance and systemic toxic effects were evaluated in single- and repeated-dose toxicity studies in rabbits. Results: GRAd-COV2 and GRAdHIVNE1 were well-tolerated. Distribution was highly confined to the injection site and draining lymph nodes, and toxicity profile consisted of transient, non-adverse inflammatory responses, while the expected immune responses to the encoded antigens were successfully induced. Notably, both vaccines demonstrated a consistent safety profile despite transgene and backbone differences, comparable to other replication-defective adenoviral vectors. Conclusions: The established non-clinical safety profile of the GRAd platform provides a robust foundation for a more efficient and streamlined regulatory pathway. By leveraging this prior knowledge, future GRAd-based vaccines can achieve accelerated clinical development while fully adhering to the ethical principles of replacement, reduction, and refinement of animal use in research.

Mouse CD8 T cell differentiation has been studied extensively in models of infections and cancer, yet no unified framework spans the full spectrum of immunological contexts. We present the CD8 immgenT framework, integrating >200,000 single-cell transcriptomes and 128-plex surface proteomes from 734 samples spanning multiple perturbations, tissues, and timepoints. Unbiased analysis identifies 21 states encompassing naive, effector, circulating memory, tissue-resident memory, progenitor-exhausted, and terminally-exhausted compartments, among others. These states re-emerge with striking molecular convergence across acute/chronic infections, cancer, autoimmunity, aging, and homeostasis, showing that near-identical transcriptional programs support protective or dysfunctional outcomes depending on developmental history and microenvironment. Classic archetypes map to discrete clusters but exhibit unappreciated heterogeneity and overlap, cautioning against rigid nomenclature. We provide validated combinatorial markers, flow cytometry gating strategies, and immgenT reference-based integration for reproducible annotation of new datasets. This universal coordinate system harmonizes fragmented CD8 T cell literature and clarifies relationships across diverse immune challenges.

Studies have demonstrated that repeated mRNA vaccination enhances the breadth of neutralization against diverse severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants. However, the development of antibodies capable of neutralizing across the Coronavirinae subfamily is poorly understood. In this study, we analyze serum samples to determine their neutralization breadth and potency and identify their antigenic targets. Using a cohort of older individuals and healthcare workers, we track correlates of broad neutralizing responses, including fusion peptide (FP) antibody elicitation. We find that although broadly neutralizing responses are often a result of receptor-binding domain (RBD)-specific antibodies, a rare subset of donors produce FP-specific broadly neutralizing responses. Interestingly, FP-specific antibodies are not observed in COVID-naive individuals, irrespective of vaccination regimen; rather, they occur following natural infection or vaccine breakthrough. This study highlights which epitope targets underpin broadly neutralizing antibody responses to coronaviruses and suggests that existing vaccines are insufficient to promote the elicitation of FP-directed broadly neutralizing coronavirus antibodies.

Prescription depressants, particularly benzodiazepines, gabapentinoids, and Z-drugs, pose overdose risk. Understanding their prevalence in overdose fatalities and co-involved substances is critical. To identify latent substance classes in prescription depressant-involved overdose deaths. Retrospective cohort study PARTICIPANTS: All individuals whose fatal overdoses involved prescription depressants from 2000-2023 in Massachusetts, US (n = 8,665). Data were obtained from the Massachusetts Registry of Vital Records and Statistics. Substances were identified using ICD-10 codes. Literal text entries were available from 2015-2023. We conducted a latent class analysis to derive substance classes and a multinomial logistic regression to examine associated factors. We assessed the proportion of deaths these classes comprised over time. Five latent classes emerged and were characterized based on the substances with highest conditional probabilities within and across classes: 1) antidepressants (21.2%); 2) fentanyl, cocaine, and benzodiazepines (34.4%); 3) antidepressants and antipsychotics (8.2%); 4) prescription opioids and benzodiazepines (19.5%); and 5) fentanyl and gabapentin or xylazine (16.7%). Over time, fentanyl-related classes came to dominate earlier prescription-only classes; deaths in Class 1 declined by 86.5%, while those in Class 5 rose from 9.6% to 45.1%. Compared to Class 2, all other classes were older, more likely female, married, educated, from rural areas, and overdosed at home. Classes 1, 3, and 5 were more likely non-White. Thousands of Massachusetts overdose deaths have involved prescribed medications, suggesting missed opportunities for intervention. Over time, the demographic patterns of deaths involving antidepressants, antipsychotics, benzodiazepines, and prescription opioids persisted, reflecting longstanding trends in polypharmacy among older women and racial disparities in prescribing. Prescription monitoring, deprescribing, screening for illicit drug use, and discussion of diversion risks are all needed. Findings underscore the need for clinical and policy interventions to reduce overdose risks among women, older individuals, and racial minorities.