Biology subjects

Iyer, A. R.

Publications and source records attributed to Iyer, A. R..

5 recordsLinked to original sources

bioRxiv2026

In vivo discovery of blood-brain barrier opening small molecules with FishNAP

The blood-brain barrier (BBB) is crucial for neural homeostasis, tightly regulating molecular exchange between the circulation and brain. However, this selective protection also greatly limits drug delivery to the central nervous system, posing a major challenge for treating neurological disorders. Pharmacological strategies that transiently and safely increase BBB permeability could therefore transform brain drug delivery, yet systematic discovery of such modulators remains hampered by the limitations of current in vitro and in vivo approaches. Here we present FishNAP, a non-invasive, high-throughput zebrafish platform for real-time assessment of BBB permeability in vivo. FishNAP captures developmental changes in barrier function and detects dysfunction in genetic mutants. Using this platform, we screened 2,320 FDA-approved small molecules for compounds capable of opening an intact BBB and identified 11 that reproducibly increased permeability. Seven of these allowed entry of a 1 kDa tracer into brain tissue, and five also permitted passage of a larger 10 kDa Dextran. Barrier integrity recovered within 24 hours for all seven compounds, indicating reversible modulation. Finally, testing three representative molecules (Calcitriol, Lovastatin, and Sunitinib) in adult mice revealed increased BBB permeability and reduced Claudin-5 expression, demonstrating conserved mechanisms of BBB-regulation across vertebrates. FishNAP thus enables systematic discovery of BBB modulators with direct translational potential for brain drug delivery.

neuroscience↗

bioRxiv2025

AI-Guided Design and AKT3 Degradation Synergize to Enhance Bispecific and Trispecific CAR-T Cell Persistence and Overcome Antigen Escape

The structural design of chimeric antigen receptors (CARs) is critical for achieving robust and durable anti-tumor responses, particularly when targeting multiple antigens to prevent tumor antigen escape. However, increasing CAR complexity can introduce structural vulnerabilities, leading to antigen-independent T cell activation, activation-induced cell death, and reduced CAR-T cell persistence. To overcome these challenges, we designed 10,824 CAR molecules across diverse formats and screened 1,452 constructs in-vitro to develop an artificial intelligence model, termed CAR-Mediated Self-Destruction (CARMSeD), which predicts CAR designs susceptible to self-activation and dysfunction. Guided by CARMSeD and structural CAR-CAR interaction modeling, we identified optimized CAR architectures incorporating ICOS and 4-1BB co-stimulatory domains. Humanized bispecific CARs targeting CD20/CD19 and CD22/CD19 demonstrated superior anti-tumor efficacy and persistence both in-vitro and in various xenograft mice models. To further extend CAR-T cell persistence, we engineered bispecific CARs integrated with an AKT3-targeted PROTAC strategy. Targeted degradation of AKT3 enhanced anti-tumor potency, promoted memory T cell formation, and enabled sustained responses even under tumor rechallenge and CD19 antigen-loss conditions. Mechanistically, these effects were mediated by metabolic reprogramming involving FOXO4; notably, FOXO4-deficient CAR-T cells exhibited impaired long-term persistence. Leveraging these mechanistic insights, we developed a trispecific CAR-T cell platform incorporating a bispecific T cell engager (BiTE) targeting CD22/CD3, combined with AKT3 PROTACs. These trispecific CAR-T cells achieved potent tumor eradication, even against malignancies lacking both CD19 and CD20 expression. Collectively, this study presents a comprehensive strategy combining structure-based design, AI-guided screening, and targeted protein degradation to engineer next-generation bi and trispecific CAR-T cells with enhanced persistence, broad antigen coverage, and superior therapeutic durability.

cancer biology↗

bioRxiv2025

Synaptic Targets of Circadian Clock Neurons Influence Core Clock Parameters

Neuronal connectivity in the circadian clock network is essential for robust endogenous timekeeping. In the Drosophila circadian clock network, four pairs of small ventral lateral neurons (sLNvs) serve as critical pacemakers. Peptidergic communication via sLNv release of the key output neuropeptide Pigment Dispersing Factor (PDF) has been well characterized. In contrast, little is known about the role of the synaptic connections that sLNvs form with downstream neurons. Connectomic analyses revealed that the sLNvs form strong synaptic connections with a group of previously uncharacterized neurons, SLP316. Here, we show that silencing synaptic output in the SLP316 neurons via tetanus toxin (TNT) expression shortens the free-running period, whereas hyper-exciting them by expressing the Na[+] channel NaChBac results in period lengthening. Under light-dark cycles, silencing SLP316 neurons also causes lower daytime activity and higher daytime sleep. Our results revealed that the main postsynaptic partners of the Drosophila pacemaker neurons are a non-clock neuronal cell type that regulates the timing of sleep and activity.

neuroscience↗

bioRxiv2024

The Circadian Neuropeptide PDF has Sexually Dimorphic Effects on Activity Rhythms

The circadian system regulates the timing of multiple molecular, physiological, metabolic, and behavioral phenomena. In Drosophila, as in other species, most of the research on how the timekeeping system in the brain controls the timing of behavioral outputs has been conducted in males, or sex has not been included as a biological variable. A critical set of circadian pacemaker neurons in Drosophila release the neuropeptide pigment-dispersing factor (PDF), which functions as a key output factor in the network with complex effects on other clock neurons. Lack of Pdf or its receptor, PdfR, results in most flies displaying arrhythmicity in activity-rest cycles under constant conditions. However, our results show that female circadian rhythms are less affected by mutations in both Pdf and PdfR. Crispr-Cas9-mediated mutagenesis of Pdf, specifically in ventral lateral neurons (LNvs), also has a greater effect on male rhythms. We tested the influence of M-cells on the circadian network and showed that speeding up the molecular clock specifically in M-cells led to sexually dimorphic phenotypes, with a more pronounced effect on male rhythmic behavior. Our results suggest that the female circadian system is more resilient to manipulations of M-cells and the PDF pathway, suggesting that circadian timekeeping is more distributed across the clock neuron network in females.

neuroscience↗

bioRxiv2023

Selective Enhancer Dependencies in MYC-Intact and MYC-Rearranged Germinal Center B-cell Diffuse Large B-cell Lymphoma

High expression of MYC and its target genes define a subset of germinal center B-cell diffuse large B-cell lymphoma (GCB-DLBCL) associated with poor outcomes. Half of these high-grade cases show chromosomal rearrangements between the MYC locus and heterologous enhancer-bearing loci, while focal deletions of the adjacent non-coding gene PVT1 are enriched in MYC-intact cases. To identify genomic drivers of MYC activation, we used high-throughput CRISPR-interference (CRISPRi) profiling of candidate enhancers in the MYC locus and rearrangement partner loci in GCB-DLBCL cell lines and mantle cell lymphoma (MCL) comparators that lacked common rearrangements between MYC and immunoglobulin (Ig) loci. Rearrangements between MYC and non-Ig loci were associated with unique dependencies on specific enhancer subunits within those partner loci. Notably, fitness dependency on enhancer modules within the BCL6 super-enhancer (BCL6-SE) cluster regulated by a transcription factor complex of MEF2B, POU2F2, and POU2AF1 was higher in cell lines bearing a recurrent MYC::BCL6-SE rearrangement. In contrast, GCB-DLBCL cell lines without MYC rearrangement were highly dependent on a previously uncharacterized 3 enhancer within the MYC locus itself (GCBME-1), that is regulated in part by the same triad of factors. GCBME-1 is evolutionarily conserved and active in normal germinal center B cells in humans and mice, suggesting a key role in normal germinal center B cell biology. Finally, we show that the PVT1 promoter limits MYC activation by either native or heterologous enhancers and demonstrate that this limitation is bypassed by 3 rearrangements that remove PVT1 from its position in cis with the rearranged MYC gene. Key pointsO_LICRISPR-interference screens identify a conserved germinal center B cell MYC enhancer that is essential for GCB-DLBCL lacking MYC rearrangements. C_LIO_LIFunctional profiling of MYC partner loci reveals principles of MYC enhancer-hijacking activation by non-immunoglobulin rearrangements. C_LI

cancer biology↗