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<title>bioRxiv Subject Collection: Pharmacology And Toxicology</title>
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This feed contains articles for bioRxiv Subject Collection "Pharmacology And Toxicology"
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<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.14.718600v1?rss=1">
<title>
<![CDATA[
Kinetics of cortisol and cortisone binding to corticosteroid binding globulin and albumin in vivo 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.14.718600v1?rss=1
</link>
<description><![CDATA[
Cortisol is a major endogenous glucocorticoid that regulates numerous physiological processes. In plasma, cortisol and its inactive metabolite cortisone bind to corticosteroid-binding globulin (CBG) and albumin, leaving only the unbound fraction available for receptor activation and metabolism. Changes in ligand or protein concentrations alter unbound fractions. Existing binding equations are difficult to extend to multi-ligand, multi-protein systems and do not readily capture competitive endogenous binding interactions. The goal of this study was to develop a plasma protein binding model that quantitatively describes binding species and predicts unbound concentrations across physiological states. Total and unbound cortisol and cortisone, CBG and albumin were measured in plasma from healthy premenopausal women (n=13) at baseline and after 7 days of 30 mg hydrocortisone treatment. Reversible 1:1 binding models were implemented in COPASI and MATLAB/Simulink, and dissociation constants (Kd) were estimated by fitting binding models to observed unbound concentrations. A model describing simultaneous binding of cortisol and cortisone to CBG and albumin yielded in vivo Kd values for cortisol:CBG, cortisone:CBG, cortisol:albumin, and cortisone:albumin of 0.0130 M, 0.169 M, 172 M, and 519 M, respectively. Model predictions agreed with observed unbound cortisol and cortisone, and bootstrap resampling confirmed stable Kd estimates. This work provides a quantitative framework for predicting unbound cortisol and cortisone across physiological and disease states by accounting for both changes in ligand and protein concentrations. This enables extrapolation without reparameterization and supports exploration of conditions such as pregnancy, adrenal insufficiency, and liver disease, informing interpretation of altered cortisol concentrations in these populations.
]]></description>
<dc:creator><![CDATA[ Authement, A. K., Nath, A., Rubinow, K. B., Amory, J. K., Isoherranen, N. ]]></dc:creator>
<dc:date>2026-04-17</dc:date>
<dc:identifier>doi:10.64898/2026.04.14.718600</dc:identifier>
<dc:title><![CDATA[Kinetics of cortisol and cortisone binding to corticosteroid binding globulin and albumin in vivo]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-17</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.14.718525v1?rss=1">
<title>
<![CDATA[
A Nonsteroidal Reversal Agent Inhibits Allopregnanolone Modulation of α1β3δ GABAA Receptors 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.14.718525v1?rss=1
</link>
<description><![CDATA[
The neurosteroid allopregnanolone is a positive allosteric modulator of GABA(A) receptors, which has proved beneficial in the treatment of major depressive disorder and epilepsies. It also has a role in treating the mood swings that are associated with fluctuations in its level during the menstrual cycle. Nonetheless, a subset of women do not tolerate high levels of allopregnanolone. Iso-allopregnanolone, a negative allosteric modulator, as well as synthetic steroid antagonists are used to treat such conditions. However, steroid-based medications are difficult to deliver and their specificity of action can be unclear. Recently introduced novel nonsteroidal agents that, like iso-allopregnanolone, can reverse the action of positive allosteric modulators without changing the positive action of GABA, might provide an alternative. We surveyed a number of them on human 1{beta}3{delta} GABAARs using a [3H]muscimol binding assay. A 6-membered ring spiro-hydantoin, DKD99, allosterically reversed the positive allosteric action of allopregnanolone over a wide concentration range (6 to 1,000 nM). DKD99 shifted the modulation curve of allopregnanolone 10-fold to the right. Furthermore, it has a much lower affinity when exerting similar actions on 1{beta}3{gamma}2 receptors. Agents such as this have utility for elucidating underlying mechanisms and may offer an alternative pathway for the development of nonsteroidal therapies against the positive allosteric modulatory actions of neurosteroids.
]]></description>
<dc:creator><![CDATA[ Zhou, X., Youssef, Y., Miller, K. W. ]]></dc:creator>
<dc:date>2026-04-17</dc:date>
<dc:identifier>doi:10.64898/2026.04.14.718525</dc:identifier>
<dc:title><![CDATA[A Nonsteroidal Reversal Agent Inhibits Allopregnanolone Modulation of α1β3δ GABAA Receptors]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-17</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.14.718203v1?rss=1">
<title>
<![CDATA[
Slow Dissociation of Nitazenes from the μ-Opioid Receptor Underlies the Challenge of Overdose Reversal 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.14.718203v1?rss=1
</link>
<description><![CDATA[
Nitazenes are driving a wave of overdose deaths in the United States and Europe and often require higher doses of naloxone to reverse. To understand the molecular basis, we conducted a joint experimental and simulation study of three common nitazenes, eto-, etodes-, and protonitazene. Radioligand experiments demonstrated that all three nitazenes display higher receptor affinity and longer dissociation half-lives than fentanyl. Notably, protonitazene dissociates slower than carfentanil and its displacement requires fourfold higher antagonist concentrations. The observed trend in nitazene half-lives is recapitulated by molecular dynamics simulations, which suggest that kinetics is controlled by specific interactions with two receptor subpockets. A newly published cryo-EM structure of fluetonitazene-OR complex confirms the predicted interactions, including a -hole bond between the nitro group and Tyr1.39, a residue recently shown to modulate OR signaling bias. Our findings suggest slow receptor dissociation as a key factor challenging overdose reversal. The mechanistic insights have implications for understanding opioid toxicity and designing more effective countermeasures.
]]></description>
<dc:creator><![CDATA[ Clayton, J., Kozell, L. B., Eshleman, A. J., Bloom, S. H., Schutzer, W. E., Abbas, A. I., Stavitskaya, L., Shen, J. ]]></dc:creator>
<dc:date>2026-04-16</dc:date>
<dc:identifier>doi:10.64898/2026.04.14.718203</dc:identifier>
<dc:title><![CDATA[Slow Dissociation of Nitazenes from the μ-Opioid Receptor Underlies the Challenge of Overdose Reversal]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-16</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.13.718333v1?rss=1">
<title>
<![CDATA[
N-3 Polyunsaturated Fatty Acids Ameliorate Post-infarction Cardiac Dysfunction Through Modulation Of Adiponectin-Ceramide Metabolism 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.13.718333v1?rss=1
</link>
<description><![CDATA[
Objective:To determine whether marine-derived n-3 polyunsaturated fatty acids (PUFAs) improve post-myocardial infarction (MI) cardiac dysfunction by modulating the adiponectin-ceramide metabolic axis, and to assess potential form-specific differences between triglyceride-based (fish oil) and phospholipid-based (krill oil) n-3 PUFAs. Approach and Results: A case-control study of 100 acute MI patients and 100 healthy controls revealed significantly elevated serum C16:0-, C18:0-, C20:0-, and C24:1-ceramides in MI patients, which correlated inversely with left ventricular ejection fraction. In a murine MI model, four-week dietary intervention with fish oil or krill oil significantly reduced myocardial infarct size and improved ejection fraction compared to vehicle controls, with no significant difference between the two n-3 PUFA forms. Targeted lipidomics demonstrated that both interventions markedly reduced cardiac and serum levels of pathogenic long-chain ceramides. Mechanistically, n-3 PUFA supplementation upregulated myocardial adiponectin and AdipoR2 expression, suppressed de novo ceramide synthesis via downregulation of SPTLC2/3, and enhanced ceramide catabolism through ASAH1 activation. These changes restored mitochondrial dynamics, attenuated MFF-mediated fission, and suppressed p53- and caspase-3-dependent cardiomyocyte apoptosis. Conclusion: n-3 PUFAs exert cardioprotective effects post-MI by activating the adiponectin-ceramide axis, restoring mitochondrial homeostasis, and inhibiting apoptosis, independent of their triglyceride or phospholipid carrier form.
]]></description>
<dc:creator><![CDATA[ Liu, Y., Sun, W., Liu, J., Wu, H., Liu, P., Chen, Y., Zhang, R., Chen, W., Wang, S., Guo, X., Zhang, W., Cao, L. ]]></dc:creator>
<dc:date>2026-04-16</dc:date>
<dc:identifier>doi:10.64898/2026.04.13.718333</dc:identifier>
<dc:title><![CDATA[N-3 Polyunsaturated Fatty Acids Ameliorate Post-infarction Cardiac Dysfunction Through Modulation Of Adiponectin-Ceramide Metabolism]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-16</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.13.718206v1?rss=1">
<title>
<![CDATA[
Increased utrophin expression in healthy and DMD patient derived myoblasts in response to ERK1/2 and EZH2 inhibitor treatment 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.13.718206v1?rss=1
</link>
<description><![CDATA[
BackgroundThe X-linked muscle wasting disorder Duchenne muscular dystrophy (DMD) is a progressive and ultimately fatal disease caused by loss of function mutations in the dystrophin (DMD) gene. Upregulation of utrophin (UTRN), an embryonic homologue of dystrophin, has been proposed as a therapeutic option that could ameliorate disease. We previously generated a bioluminescent screen for utrophin-upregulating compounds using a mouse reporter of endogenous utrophin expression and discovered that inhibition of ERK1/2 and EZH2, increases utrophin expression in myoblasts.

MethodologyHere we extend this analysis to show that treatment of human myoblasts with the ERK1/2 inhibitor LY3214996 and the EZH2 inhibitor GSK503, increases UTRN expression in primary and immortalised myoblasts derived from healthy volunteers and DMD patients.

ResultsShort-term (24 hours) inhibition of ERK1/2 and EZH2 resulted in increased expression of utrophin in proliferating myoblasts. Surprisingly, in patient-derived samples, but not healthy controls, increased UTRN expression was sustained following drug removal and in vitro differentiation. Furthermore, dystrophin deficient myoblasts have altered expression of myogenic transcription factors MYOD1 and MYOG and proliferation marker Ki67, signalling an altered regenerative capacity of these cells, while ERK1/2 inhibition, alone or combined with EZH2i, reversed this transcriptional signature.

ConclusionsTreatment with ERK1/2 and EZH2 inhibitors could offer a therapeutic option for DMD by increasing UTRN and MYOD1 expression. We propose that this may compensate for DMD loss and help restore productive muscle differentiation and regeneration.
]]></description>
<dc:creator><![CDATA[ Gleneadie, H. J., Francis, T., Mo, S. P. L., Ahmed, A., Bensalah, M., Muntoni, F., Harridge, S. D. R., Merkenschlager, M., Fisher, A. G. ]]></dc:creator>
<dc:date>2026-04-15</dc:date>
<dc:identifier>doi:10.64898/2026.04.13.718206</dc:identifier>
<dc:title><![CDATA[Increased utrophin expression in healthy and DMD patient derived myoblasts in response to ERK1/2 and EZH2 inhibitor treatment]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-15</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.11.717649v1?rss=1">
<title>
<![CDATA[
Three nitrogen atoms turn old kinase inhibitors into new targetable remedy 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.11.717649v1?rss=1
</link>
<description><![CDATA[
Pathological neovascularization in the eye is a significant contributor to vision loss in diseases such as age-related macular degeneration (AMD) and diabetic retinopathy. While anti-VEGF biologics are effective, they require repeated intravitreal injections and carry procedural risks. Here, we report a novel principle for designing photoactivatable VEGFR2 inhibitors, along with two examples, EYE1090 and EYE1118, engineered from the sunitinib and vorolanib scaffolds, respectively. Azido-functionalization in these molecules enables light-triggered receptor binding while preserving potent inhibition in the dark. Both compounds exhibit significantly enhanced activity upon exposure to green light capable of reaching the retina, also in the elderly. In vitro, the compounds robustly inhibited angiogenesis and endothelial migration that was further potentiated by light. In vivo, orally administered EYE1090 and EYE1118 suppressed VEGF-induced retinal leakage and reduced lesion size in a mouse model of choroidal neovascularization at doses tenfold lower than parental compounds. Photoactivation also influenced compound biodistribution, suggesting light-guided targeting. Acute toxicity studies revealed no hepatotoxicity. This strategy exploits the natural light-focusing anatomy of the eye to locally activate systemically administered drugs, thereby reducing therapeutic doses and systemic exposure. Our findings introduce a light-targeted pharmacological approach for treating retinal diseases using photoactivable kinase inhibitors.
]]></description>
<dc:creator><![CDATA[ Besztercei, B., Antal, R., Tahtivaara, L., Lappetelainen, B., Jaaskelainen, N., Szappanos, A., Lukats, A., Pal-Kajtar, A., Budai, A., Cerrada-Gimenez, M., Kovacs, K. A. ]]></dc:creator>
<dc:date>2026-04-14</dc:date>
<dc:identifier>doi:10.64898/2026.04.11.717649</dc:identifier>
<dc:title><![CDATA[Three nitrogen atoms turn old kinase inhibitors into new targetable remedy]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-14</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.11.717944v1?rss=1">
<title>
<![CDATA[
Using Patient iPSC-derived Retinal Pigment Epithelial Cells to Evaluate Differential Susceptibility to MEK Inhibitor-Associated Retinopathy 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.11.717944v1?rss=1
</link>
<description><![CDATA[
PurposeCompare the effect of MEK inhibition on iPSC-derived retinal pigmental epithelial (RPE) cells generated from a patient who developed MEK inhibitor-Associated Retinopathy (MEKAR) versus a patient who did not develop retinopathy.

DesignCase-control

SubjectsTwo female patients with Neurofibromatosis Type 1 who were treated with MEK inhibitors. One patient developed MEKAR, the other did not.

MethodsRPE were generated from human induced pluripotent stem cells (hiPSCs) from these two patients. These hiPSC-derived RPE were treated with selumetinib for 10 days.

Main Outcome MeasuresPhagocytic activity and changes in gene expression

ResultsAs previously reported, there was a significant increase in internalized rhodopsin in phagocytosis assays, yet this was only found in hiPSC-derived RPE from the patient who developed MEKAR. Selumetinib decreased expression of genes related to fluid transport and cell volume, including aquaporins and solute transporters. At baseline, cells from the patients without MEKAR had higher expression of these genes. Interestingly, selumetinib-induced changes in gene expression only reached statistical significance in cells from the patient who did not develop MEKAR, suggesting these changes may be a compensatory protective mechanism. Patients susceptible to forming MEKAR may have increased phagocytosis without a compensatory change in expression of genes related to fluid flux, thereby inhibiting their ability to transport fluid out of the subretinal space.

ConclusionsMEK inhibitor-Associated Retinopathy may only affect susceptible patients whose retinal pigment epithelium cannot sufficiently regulate expression of genes related to fluid transport and cell volume, altering the ability of these cells to properly function.
]]></description>
<dc:creator><![CDATA[ Lozano, L. P., Boyce, T. M., Groves, A. P., Keen, H. L., Boldt, H. C., Mullins, R. F., Binkley, E. M., Tucker, B. A. ]]></dc:creator>
<dc:date>2026-04-14</dc:date>
<dc:identifier>doi:10.64898/2026.04.11.717944</dc:identifier>
<dc:title><![CDATA[Using Patient iPSC-derived Retinal Pigment Epithelial Cells to Evaluate Differential Susceptibility to MEK Inhibitor-Associated Retinopathy]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-14</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.10.717852v1?rss=1">
<title>
<![CDATA[
Discovery and Development of First-in-Class Cereblon-Recruiting RIPK1 Degraders 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.10.717852v1?rss=1
</link>
<description><![CDATA[
Receptor-interacting protein kinase 1 (RIPK1) is a critical regulator of programmed cell death and is implicated in various pathological conditions, particularly in mediating tumor resistance to immune checkpoint inhibitors (ICBs). In this study, we have pioneered the development of a novel cereblon (CRBN)-recruiting RIPK1 degrader, LD5095, through systematic optimization of linker and CRBN ligand portion. LD5095 demonstrates potent and selective RIPK1 degradation across cell lines, with rapid kinetics and sustained degradation over 72h post-washout. Functionally, RIPK1 degradation by LD5095 significantly sensitized Jurkat cells to TNF-induced apoptosis. Furthermore, LD5095 exhibited favorable pharmacokinetics, including metabolic stability and an extended half-life. Strikingly, in vivo, a single dose of LD5095 achieved durable RIPK1 degradation in xenograft tumors over 6 days. These findings underscore the potential of LD5095 as a chemical probe for studying RIPK1 biology and a promising candidate for cancer treatment.
]]></description>
<dc:creator><![CDATA[ Lu, D., Yu, X., Wang, J. ]]></dc:creator>
<dc:date>2026-04-14</dc:date>
<dc:identifier>doi:10.64898/2026.04.10.717852</dc:identifier>
<dc:title><![CDATA[Discovery and Development of First-in-Class Cereblon-Recruiting RIPK1 Degraders]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-14</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.13.717820v1?rss=1">
<title>
<![CDATA[
Cannabidiol (CBD) Promotes Post-TBI Astrocyte Viability and Decreases Injury-Induced Glial Stress Responses Across Zebra Finch Song Control Nuclei 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.13.717820v1?rss=1
</link>
<description><![CDATA[
The non-euphorigenic phytocannabinoid cannabidiol (CBD) has demonstrated therapeutic efficacy in childhood-onset epilepsies. Using a songbird preclinical model we have found that CBD promotes recovery of learned vocalizations following focal motor cortical injury. But questions about cellular mechanisms supporting this protection remained. Songbird vocal learning, like human speech, depends on development and maintenance of specialized neural circuits. Partial lesioning (microlesions) of the vocal pre-motor cortical-like song region HVC transiently disrupts song structure and triggers injury-associated cellular stress responses across interconnected song regions. Building on prior findings that CBD reduces neuroinflammation and synaptic loss in zebra finch song circuitry, we investigated potential astrocyte contributions. Here we report that HVC microlesions induce significant cell loss in HVC and its projection targets (vocal motor RA and striatal Area X), with a substantial fraction of apoptotic cells being astrocytes. CBD treatment reduces lesion-induced apoptosis and preserves astrocyte populations, indicating enhanced astrocyte viability as a major factor in CBD-mediated neuroprotection. Microlesions also elevate astrocyte stress, including increased lysosomal burden (LAMP1/LC3 expression) and astrocytic reactivity markers (C3, S100A10, aromatase). CBD attenuates these stress responses while enhancing neuroprotective metabolic and antioxidant mediators (glutamine synthetase [GS], glutamate-cysteine ligase modifier subunit [GCLM]), consistent with improved antioxidant and excitotoxicity resistance. Given that development-dependent sensorimotor skills (e.g. song in songbirds, language and many others in humans) depend on sensitive period establishment and ongoing post-learning maintenance of specialized neural circuits vulnerable to traumatic disruption, the zebra finch model provides a valuable preclinical platform for investigating glial-targeted interventions to promote circuit resilience and functional recovery after TBI.
]]></description>
<dc:creator><![CDATA[ Marshall, D. A., Litwa, K. A., Soderstrom, K. ]]></dc:creator>
<dc:date>2026-04-14</dc:date>
<dc:identifier>doi:10.64898/2026.04.13.717820</dc:identifier>
<dc:title><![CDATA[Cannabidiol (CBD) Promotes Post-TBI Astrocyte Viability and Decreases Injury-Induced Glial Stress Responses Across Zebra Finch Song Control Nuclei]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-14</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.09.717524v1?rss=1">
<title>
<![CDATA[
Middle-aged mice treated with GHK-Cu peptide administered intraperitoneally or intranasally show behavioral rescue but divergent hippocampal aging programs 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.09.717524v1?rss=1
</link>
<description><![CDATA[
Age-related cognitive decline (ARCD) is driven by conserved biological mechanisms of aging, yet no gerotherapeutic directly targets these processes in the brain. Glycyl-L-histidyl-L-lysine complexed with copper (GHK-Cu) is an endogenous peptide with regenerative and anti-inflammatory properties that declines with age. Whether its effects on cognitive aging depend on delivery route or exposure duration remains unclear. Aged C57BL/6J mice (20-21 months) received GHK-Cu (15 mg/kg) via short-term intraperitoneal (IP; 5 days) or longer-term intranasal (IN; 8 weeks) administration. Hippocampal-dependent escape learning was assessed using a spatial navigation task. Molecular effects were evaluated using hippocampal immunohistochemistry and bulk RNA sequencing. Differential gene expression was analyzed using DESeq2 with false discovery rate (FDR) correction, and pathway-level changes were assessed via gene set enrichment analysis (GSEA). IN GHK-Cu improved escape latency across Trials 2-4 in both sexes (P < 0.05), whereas IP dosing produced a transient improvement in males during Trial 2 (P < 0.05) without sustained effects or improvement in females. IN treatment increased synaptophysin in females (P < 0.001) and decreased GFAP in both sexes (P < 0.01), while IP treatment reduced TGF-{beta}, GFAP, and MCP-1 in males (P < 0.05) and decreased p21 in females (P < 0.0001). Transcriptomic analysis revealed distinct molecular programs. IN GHK-Cu induced coordinated suppression of oxidative phosphorylation (male NES -5.44, female NES -4.20; FDR < 0.0001) and MYC target pathways (female NES -4.31, FDR < 0.0001), with additional attenuation of PI3K-AKT-mTOR signaling in females (NES -3.15, FDR = 0.062). In contrast, IP treatment activated oxidative phosphorylation (female NES 4.97, FDR < 0.001), DNA repair (NES 5.58, FDR < 0.001), and MYC targets (NES 4.34, FDR = 0.002), indicating engagement of acute stress-response and repair pathways. GHK-Cu improves hippocampal-dependent learning in aged mice through distinct biological modes: IP exposure activates repair and stress-response pathways, whereas IN delivery induces sustained suppression of growth and mitochondrial metabolic signaling associated with aging biology. These findings demonstrate that functional cognitive improvement can arise from divergent molecular states and identify administrative route and exposure duration as key determinants of gerotherapeutic response.
]]></description>
<dc:creator><![CDATA[ Mazzola, J. M., Rosenfeld, M., Tucker, M., Wezeman, J., Ladiges, W. C., Liao, G. Y. ]]></dc:creator>
<dc:date>2026-04-13</dc:date>
<dc:identifier>doi:10.64898/2026.04.09.717524</dc:identifier>
<dc:title><![CDATA[Middle-aged mice treated with GHK-Cu peptide administered intraperitoneally or intranasally show behavioral rescue but divergent hippocampal aging programs]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-13</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.09.717469v1?rss=1">
<title>
<![CDATA[
Reversible CD28 checkpoint modulation by cyclic peptides outperforms biologic blockade under exposure-limited conditions 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.09.717469v1?rss=1
</link>
<description><![CDATA[
CD28 co-stimulatory blockade is an established therapeutic strategy in autoimmune disease, yet every clinical-stage agent shares a structural limitation: high-affinity, long-lived receptor occupancy that precludes dynamic control of immune suppression. In chronic inflammatory conditions, where prolonged immunosuppression carries infection risk and necessitates treatment interruptions, no existing agent permits rapid restoration of immune function. We report CP8, a disulfide-constrained cyclic peptide antagonist that matches the inhibitory potency of clinical-stage CD28 biologics (FR104, Acazicolcept, and Lulizumab) across primary human immune cells from healthy and ulcerative colitis donors, suppressing IL-2 and IFN-{gamma} production without agonist activity. Unlike these biologics, CP8 enables rapid and near-complete restoration of T-cell function upon compound removal, a property mechanistically inaccessible to antibody-based therapeutics and demonstrated here for the first time for any CD28-targeting agent. In a T-cell transfer colitis model, CP8 maintains efficacy under intermittent dosing and outperforms Acazicolcept, a dual CD28/ICOS inhibitor, under exposure-limited conditions, achieving superior disease suppression, tissue preservation, and cytokine reduction. These results demonstrate that potency and pharmacological persistence are decoupled properties, and reframe cyclic peptides as a superior modality for immune checkpoints where temporal control of signaling is essential to balance efficacy with the risks of chronic immune suppression.
]]></description>
<dc:creator><![CDATA[ Kuncewicz, K., Upadhyay, S., Ge, Y., Duan, H., Gabr, M. ]]></dc:creator>
<dc:date>2026-04-13</dc:date>
<dc:identifier>doi:10.64898/2026.04.09.717469</dc:identifier>
<dc:title><![CDATA[Reversible CD28 checkpoint modulation by cyclic peptides outperforms biologic blockade under exposure-limited conditions]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-13</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.08.717349v1?rss=1">
<title>
<![CDATA[
Impact of Sex on Heroin Intravenous Self-Administration by Heterogeneous Stock Rats 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.08.717349v1?rss=1
</link>
<description><![CDATA[
BackgroundIntravenous self-administration (IVSA) of opioids by rats has been shown frequently to exhibit no sex differences, in many cases a higher intake of females, and only rarely higher rates in males. A diversity of methodological parameters (opioid identity, training doses, rat strain, session duration) makes it difficult to identify consistent contributions to these outcomes.

ObjectiveTo determine if Heterogeneous Stock (HS) rats derived from 8 founder strains differ by sex in the IVSA of opioids.

MethodsMale and female Heterogeneous Stock (N=7-8 per sex) rats were permitted to self-administer heroin (20 {micro}g/kg/infusion) in 2 hour sessions under a Fixed Ratio 1 schedule of reinforcement. After acquisition, animals completed sessions in which different infusion doses of heroin (0, 15, 30, 60, 120 {micro}g/kg/infusion), oxycodone (0, 30, 60, 150, 300 {micro}g/kg/infusion) and fentanyl (0, 0.625, 1.25, 2.5, 5.0 {micro}g/kg/infusion) were assessed. Next, animals were evaluated on doses of heroin (15, 30, 60, 120 {micro}g/kg/infusion), oxycodone (30, 60, 150, 300 {micro}g/kg/infusion) and fentanyl (0.625, 1.25, 2.5, 5.0 {micro}g/kg/infusion) under a Progressive Ratio schedule. Anti-nociceptive effects of heroin (0.56-2.4 mg/kg, s.c.) were examined with a warm water tail-withdrawal assay.

ResultsFemale HS rats consistently self-administered more infusions of opioids, including heroin during acquisition, all three opioids during FR-1 dose substitution and of oxycodone and fentanyl in the PR procedure. Male rats were moderately more sensitive to the anti-nociceptive effects of heroin.

ConclusionsFemale rats drawn at random from a genetically diverse population self-administer opioids at higher rates than their male counterparts.
]]></description>
<dc:creator><![CDATA[ Taffe, M. A., Mehl, S. L., Rahman, S. R. M. U., Grant, Y. ]]></dc:creator>
<dc:date>2026-04-12</dc:date>
<dc:identifier>doi:10.64898/2026.04.08.717349</dc:identifier>
<dc:title><![CDATA[Impact of Sex on Heroin Intravenous Self-Administration by Heterogeneous Stock Rats]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-12</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.08.717353v1?rss=1">
<title>
<![CDATA[
Influence of different pharmaceuticals on the germination and early development of two leafy vegetable species 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.08.717353v1?rss=1
</link>
<description><![CDATA[
Pharmaceuticals are becoming increasingly prevalent in the environment, yet their effects on terrestrial plants, especially during early development, are poorly investigated. In this context, leafy vegetables are of particular interest because they tend to accumulate more pharmaceuticals than other crops. This study investigated the impacts of six pharmaceuticals of different classes commonly detected in soils and water on seed germination and early seedling growth of the leafy vegetables bok choy (Brassica rapa subsp. chinensis) and spinach (Spinacia oleracea) under controlled conditions. Seeds were exposed to different concentrations of the non-steroidal anti-inflammatory drugs (NSAIDs) ibuprofen, naproxen, diclofenac, or salicylic acid, the antiepileptic drug carbamazepine, or the antibiotic ciprofloxacin, and germination rates, root and shoot lengths, biomass allocation, cotyledon development, and lateral root formation (in bok choy only) measured after seven days. While germination was unaffected, early development parameters showed species-specific responses. In bok choy, high concentrations of NSAIDs and ciprofloxacin led to an increased shoot biomass and cotyledon area but a reduced primary root growth and lateral root formation, while carbamazepine had no effect. The contrasting effects on aboveground versus belowground organs of different pharmaceuticals suggest an interference with hormonal regulation, especially auxin. Spinach showed less responses than bok choy, with root length being rather increased by some NSAIDs. These results indicate that sensitivity to pharmaceuticals begins after germination and depends on both the chemical properties of the compound and the plant species. The study highlights the value of systematic comparative testing of pharmaceuticals across plant species.
]]></description>
<dc:creator><![CDATA[ Brokate, L., Mueller, C. ]]></dc:creator>
<dc:date>2026-04-12</dc:date>
<dc:identifier>doi:10.64898/2026.04.08.717353</dc:identifier>
<dc:title><![CDATA[Influence of different pharmaceuticals on the germination and early development of two leafy vegetable species]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-12</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.08.717252v1?rss=1">
<title>
<![CDATA[
Stability of Oxycodone Solutions Containing S-Ketamine or Dexmedetomidine 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.08.717252v1?rss=1
</link>
<description><![CDATA[
ObjectivesTo determine whether adding S-ketamine or dexmedetomidine to oxycodone affects the microbiological, physical, or chemical stability of patient-controlled analgesia (PCA) solutions prepared in a hospital pharmacy.

MethodsOxycodone solution (1 mg/mL) and three oxycodone-S-ketamine mixtures (0.25, 0.50, 0.75 mg/mL) and three oxycodone-dexmedetomidine mixtures (2.5, 5.0, 10 {micro}g/mL) were compounded under validated EU GMP Class A/B aseptic conditions and filled into PCA reservoirs. Reservoirs (n=42 for physicochemical studies; n=21 for sterility; n=4 for antimicrobial activity testing) were stored at 2-8{degrees}C for 28 days, then at 20-25{degrees}C for 2 days. Sterility was assessed by membrane filtration according to Ph. Eur. 2.6.1. Physical stability was evaluated by visual inspection, pH, weight, and osmolality. Chemical stability was assessed using a validated HPLC-UV method developed in accordance with FDA and ICH Q2(R1) guidelines.

ResultsAll antimicrobial activity tests showed growth of the six reference strains, indicating no inhibition by the drug mixtures. All 21 sterility-test reservoirs remained free of turbidity throughout 30 days. No visual changes, precipitation, or discolouration were observed. Weight loss was [&le;]0.3%, pH changes were between required range 4,5-7, and osmolality increased by <1.4% during the study. Measured oxycodone, S-ketamine, and dexmedetomidine concentrations remained within {+/-}5% of initial values, and no degradation products were detected.

ConclusionsOxycodone PCA solutions containing S-ketamine or dexmedetomidine remained sterile, physically stable, and chemically stable for 28 days at 2-8{degrees}C followed by 2 days at room temperature at 20-25{degrees}C. These findings support extended shelf-life and centralized batch preparation of opioid-adjuvant PCA reservoirs in hospital pharmacy practice.

Key MessagesO_ST_ABSWhat is already known on this topicC_ST_ABSOpioid-adjuvant combinations such as oxycodone with S-ketamine or dexmedetomidine are increasingly used in patient-controlled analgesia, but no commercial multi-agent formulations exist. Hospital pharmacies therefore prepare these mixtures as compounded sterile preparations, despite limited data on their chemical and microbiological stability.

What this study addsThis study demonstrates that oxycodone PCA solutions containing S-ketamine or dexmedetomidine remain chemically stable and microbiologically sterile for 28 days at 2-8{degrees}C plus 2 days at 20-25{degrees}C, when prepared under validated aseptic conditions. Concentrations of all analytes remained within {+/-}5% of initial values, and no degradation products were detected using a validated HPLC-UV method.

How this study might affect research, practice or policyThese stability data support the assignment of extended beyond-use dates and enable centralized batch compounding of PCA reservoirs in hospital pharmacies. The findings have the potential to reduce aseptic workload, improve production efficiency and decrease medication waste while ensuring product quality.
]]></description>
<dc:creator><![CDATA[ Vaisanen, P., Makela, S., Siren, S., Pohjanoksa, K., Uusalo, P., Scheinin, M., Torniainen, K., Saari, T. I. ]]></dc:creator>
<dc:date>2026-04-12</dc:date>
<dc:identifier>doi:10.64898/2026.04.08.717252</dc:identifier>
<dc:title><![CDATA[Stability of Oxycodone Solutions Containing S-Ketamine or Dexmedetomidine]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-12</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.08.717320v1?rss=1">
<title>
<![CDATA[
Pinus sp. leaf extracts exert antileishmanial effects against Leishmania donovani by targeting trypanothione reductase 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.08.717320v1?rss=1
</link>
<description><![CDATA[
The urgent need for new antileishmanial drugs with novel mechanisms of action is driven by the limited efficacy, high toxicity, and growing resistance to currently available treatments. This study investigates the antileishmanial potential of extracts from Pinus sp., a plant widely used in the Cameroonian pharmacopoeia to treat various infections, including cough and fever conditions. The antileishmanial activity of the ethanol, methanol, hydro-ethanol, and hydro-methanol extracts of Pinus sp. leaves were evaluated against Leishmania donovani promastigotes and amastigotes using a resazurin-based assay. The most active extract was then subjected to UHPLC-LC-MS/MS metabolomic profiling. Cytotoxicity, immunomodulatory, antioxidant, and anti-inflammatory properties of Pinus sp. extracts were also investigated. Moreover, the pharmacokinetic profile and molecular interactions of the identified compounds were predicted through in silico experiments using trypanothione reductase as the target enzyme. The acute toxicity study of the most promising extract (ethanol extract) was performed at 2000 and 5000 mg/kg using the organization for economic co-operation and development (OECD) guidelines, number 423. The plant extracts inhibited the growth of L. donovani with IC50 values ranging from 6.45 to 15.78 {micro}g/mL and 16.11 to 24.44 {micro}g/mL in promastigotes and amastigotes, respectively. These extracts also showed antioxidant, immunomodulatory, and anti-inflammatory effects. The extracts indicated no cytotoxicity and high selectivity (SI >10) toward Raw264.7 and Vero CRL1586 cells. The ethanolic extract showed a lethal dose (LD) value greater than 5000 mg/kg, thus highlighting its non toxicity upon the acute toxicity test. The metabolomic profiling of the ethanol extract identified several compound classes, including flavanone O-glycosides (e.g. epiafzelechin trimethyl ether), alkaloids (e.g. harmane), and diterpenes (e.g. abietic acid). Further molecular docking studies revealed strong binding affinities of these compounds to trypanothione reductase, thus suggesting the inhibitory potential of these compounds toward the target enzyme. Overall, extracts from Pinus sp. leaves demonstrate promising antileishmanial effects. However, isolation and characterization of the antileishmanial compounds from Pinus sp. leaf, detailed pharmacokinetics, and mechanisms of action are warranted for the successful utilization of this plant in antileishmanial drug discovery.
]]></description>
<dc:creator><![CDATA[ Kemzeu, R., Tchokouaha Yamthe, L. R., Njanpa Ngansop, C. A., Madiesse Kemgne, E. A., Pone Kamdem, B., Ngouana, V., Ngoutane Mfopa, A., Donbou Djiotie, C. P., Tsouh Fokou, P. V., Tsakem Nangap, J. M., Lunga, P. K., Bruno, L. N., Fekam Boyom, F. ]]></dc:creator>
<dc:date>2026-04-12</dc:date>
<dc:identifier>doi:10.64898/2026.04.08.717320</dc:identifier>
<dc:title><![CDATA[Pinus sp. leaf extracts exert antileishmanial effects against Leishmania donovani by targeting trypanothione reductase]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-12</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.06.716751v1?rss=1">
<title>
<![CDATA[
PROTAC internalization and target degradation require clathrin-mediatedendocytosis 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.06.716751v1?rss=1
</link>
<description><![CDATA[
Proteolysis-targeting chimeras (PROTACs) are emerging as potent tools for targeted protein degradation that overcome many of the limitations of traditional small molecule inhibitors. Yet how these hetero-bifunctional therapeutics enter cells remains a mystery. While passive diffusion is conventionally assumed, the bulky structure of PROTACs suggests that active transport may be required. Recently, the fatty acid transporter CD36 was identified as a key receptor for PROTACs. However, because the uptake mechanism of CD36 is itself unknown, how PROTACs enter cells remains a mystery. Here we show that PROTAC uptake and function require clathrin-mediated endocytosis. We uncover previously unrecognized clathrin adaptor-binding motifs in the CD36 C-terminus and use live-cell imaging to visualize the recruitment of both CD36 and PROTACs to sites of clathrin-mediated endocytosis on the cellular plasma membrane. Strikingly, disruption of clathrin assembly through either genetic or pharmacological means abolishes all detectable PROTAC-induced protein degradation, demonstrating that the clathrin pathway is required for the function of PROTACs that utilize diverse E3 enzymes against multiple targets. These results elucidate the molecular mechanism of PROTAC entry into cells, providing critical information for optimizing cellular uptake and response to targeted degraders.
]]></description>
<dc:creator><![CDATA[ Liu, H.-Y., Wang, Z., Sharma, R., Perez, J., Kusaj, B., Zhou, H., Wang, M., Huibregtse, J., Li, H.-Y., Stachowiak, J. ]]></dc:creator>
<dc:date>2026-04-08</dc:date>
<dc:identifier>doi:10.64898/2026.04.06.716751</dc:identifier>
<dc:title><![CDATA[PROTAC internalization and target degradation require clathrin-mediatedendocytosis]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-08</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.06.716494v1?rss=1">
<title>
<![CDATA[
Morphological and Functional Effects of Cytoskeletal and Ion-Channel Agents on the Protoscolex of Echinococcus granulosus sensu lato 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.06.716494v1?rss=1
</link>
<description><![CDATA[
Helminthiases remain a major global health burden, and limitations of current anthelmintic therapies highlight the need for new pharmacological targets. In this study, we examined the effects of ion-channel and cytoskeletal modulators on bovine lung protoscoleces (PSCs) of Echinococcus granulosus sensu lato. Compounds acting on ion channels (praziquantel, amiloride, and amlodipine) and cytoskeletal components (albendazole and cytochalasin D) were evaluated using a semi-automated motility assay, methylene blue exclusion to assess viability, and scanning electron microscopy (SEM) to characterize structural damage. All compounds produced concentration-dependent reductions in PSCs motility. Amlodipine was the most potent inhibitor of motility, whereas praziquantel and cytochalasin D produced pronounced tegumental alterations and strong correlations between motility impairment and parasite death. In contrast, amiloride markedly reduced motility with comparatively minor effects on viability, indicating a primarily paralytic effect. Cytoskeletal disruption induced severe structural damage and parallel declines in motility and viability. SEM analysis revealed extensive tegumental collapse, loss of glycocalyx, and microtrichial damage in PSCs exposed to cytoskeletal and calcium-modulating agents. These findings highlight cytoskeletal organization and calcium-dependent ion fluxes as key physiological vulnerabilities in E. granulosus. Comparative analysis of these pharmacological targets provides mechanistic insight into how disruptions in cytoskeletal dynamics and cation homeostasis compromise parasite motility and survival.
]]></description>
<dc:creator><![CDATA[ Carabajal, M. P. A., Fernandez Salom, M. J., Martinez, L. J., Marcial, E. R., Albarracin, V. H., Cantiello, H. F. ]]></dc:creator>
<dc:date>2026-04-08</dc:date>
<dc:identifier>doi:10.64898/2026.04.06.716494</dc:identifier>
<dc:title><![CDATA[Morphological and Functional Effects of Cytoskeletal and Ion-Channel Agents on the Protoscolex of Echinococcus granulosus sensu lato]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-08</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.05.716056v1?rss=1">
<title>
<![CDATA[
Synergistic induction of a lethal Autosis-to-Apoptosis switch by phytocannabinoids and β-Caryophyllene in Triple-Negative Breast Cancer Cells 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.05.716056v1?rss=1
</link>
<description><![CDATA[
BackgroundTriple-negative breast cancer (TNBC) presents significant therapeutic limitations due to its aggressive heterogeneity and the rapid emergence of adaptive resistance to apoptosis-based regimens. Addressing these challenges requires polypharmacological strategies capable of modulating multiple signalling networks simultaneously. While the Cannabis sativa phytocomplex offers a vast chemical space for multi-target intervention, the quantitative pharmacological basis of its synergistic interactions remains largely uncharacterised.

PurposeThis study aimed to deconstruct the synergistic landscape of high-purity phytocannabinoids (CBD, CBG, CBD-A) in combination with the sesquiterpene {beta}-caryophyllene (BCP) against TNBC, using MDA-MB-231 as a primary model and Hs578T as a validation line.

MethodsGrowth Rate (GR) inhibition metrics and the SynergyFinder+ framework were used to map pharmacological interactions across four reference models. Subcellular dynamics and phenotypic transitions were characterised by high-resolution label-free holotomographic microscopy combined with live-cell kinetic imaging and single-cell fate mapping.

ResultsTwo highly potent synergistic clusters were identified for CBD-CBG-BCP combinations, with ZIP, HSA, and Bliss synergy scores exceeding 65. CBD-A exhibited minimal interaction potential and was excluded from ternary studies. GR-based quantification further revealed that these combinations produced net cytotoxicity (GR < 0) at sub-IC concentrations of each component. Single-cell fate mapping by holotomographic microscopy identified a temporally ordered death programme: an initial phase of extensive cytoplasmic vacuolisation associated with focal perinuclear space swelling and progressive nuclear compression, morphological hallmarks of autosis, which is followed by a transition to apoptotic execution. The autotic nature of the primary death phase was confirmed by pharmacological rescue with digoxin, a selective inhibitor of the Na,K-ATPase. To the best of our knowledge, this sequential engagement of autosis followed by apoptotic execution represents the first documented instance of such a two-stage death programme in any cellular model.

ConclusionThese findings provide robust evidence that specific phytocannabinoid-terpene ratios engage a Na,K-ATPase-regulated autotic programme as an upstream commitment step, followed by apoptotic execution, effectively circumventing the caspase-independent resistance mechanisms characteristic of TNBC. This study establishes a rational, quantitatively validated framework for transitioning from empirical botanical use to evidence-based, multi-target cannabinoid polypharmacology in aggressive breast cancer.
]]></description>
<dc:creator><![CDATA[ Hamann, C., Jansen, O., Jullien, K., Lhonneux, L., Ledoux, A., Frederich, M., Maquoi, E. ]]></dc:creator>
<dc:date>2026-04-07</dc:date>
<dc:identifier>doi:10.64898/2026.04.05.716056</dc:identifier>
<dc:title><![CDATA[Synergistic induction of a lethal Autosis-to-Apoptosis switch by phytocannabinoids and β-Caryophyllene in Triple-Negative Breast Cancer Cells]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-07</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.02.716001v1?rss=1">
<title>
<![CDATA[
Exofection as a Therapeutic Modality: Restoring P-gp Activity via Trophoblast-Derived EV in Neuroinflammatory Disorders 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.02.716001v1?rss=1
</link>
<description><![CDATA[
BackgroundP-glycoprotein (P-gp/ABCB1) is a key efflux transporter that maintains barrier integrity by clearing xenobiotics and toxic metabolites. At the feto-maternal interface, trophoblast-derived extracellular vesicles (CTC-EVs) naturally and transiently transfer functional P-gp to maternal decidual cells, restoring lost and or reduced P-gp function (exofection) to sustain pregnancy homeostasis. A similar loss of P-gp at the blood brain barrier (BBB) contributes to impaired amyloid-{beta} (A{beta}) clearance and neuroinflammation in Alzheimers disease. We investigated whether CTC-EV-mediated exofection could restore P-gp function in human brain endothelial cells (hBECs) and enhance A{beta} clearance under inflammatory and neurodegenerative conditions.

MethodsCTC-EVs were isolated and characterized by nanoparticle tracking analysis and western blotting for P-gp and EV markers. Transcriptomic profiling of CTC-EVs identified enrichment of transporter-related genes, including solute carriers and ABC transporters, along with inflammatory mediators. Network analysis revealed coordinated modules linking EV cargo to transporter regulation, endocytosis/trafficking pathways, and inflammatory remodeling processes converging on BBB efflux activity. hBECs were exposed to LPS (500 ng/mL, 48 h) with or without CTC-EVs. P-gp expression was assessed by immunofluorescence (mean fluorescence intensity, MFI) and western blotting, while functional efflux was measured using Calcein-AM assays. A{beta} oligomer transport was evaluated using a transwell hBEC model. In vivo, 3xTg-AD mice received intravenous CTC-EVs (1x10L/day for 5 days), followed by assessment of P-gp expression, A{beta} burden, and neuroinflammatory markers. Pharmacokinetic studies in P-gp knockout mice were conducted to confirm functional transporter recovery.

ResultsLPS exposure significantly reduced P-gp expression in hBECs (41.3% decrease in MFI, p=0.0084), which was restored by CTC-EVs (46.7% increase vs. LPS, p=0.0121). Exofection increased P-gp by a 2.1-fold following EV treatment as determined by western blot. Functional assays demonstrated enhanced efflux, with a 38.5% reduction in intracellular Calcein fluorescence (p<0.001). Network-informed mechanisms supported coordinated regulation of transporter and trafficking pathways. CTC-EVs improved A{beta} transport across inflamed hBEC monolayers. In vivo, EV-treated 3xTg-AD mice exhibited increased P-gp expression in the frontal cortex (38.6%) and hippocampus (42.1%), reduced A{beta} plaque burden (27.9%), and decreased inflammatory markers (IL-1{beta} and TNF-, p<0.05). In P-gp knockout mice, EVs reduced brain drug accumulation by 22.4% (p=0.032), confirming restoration of transporter function.

ConclusionCTC derived EVs are natural carriers of functional transporter proteins and restore efflux capacity in compromised endothelial barriers. Integration of transcriptomic and network analyses highlights coordinated regulation of transporter, trafficking, and inflammatory pathways underlying exofection. This reproductive biology inspired strategy offers a promising therapeutic approach for enhancing A{beta} clearance and mitigating neuroinflammation in Alzheimers disease.
]]></description>
<dc:creator><![CDATA[ Kammala, A. K., Tatiparthy, M., Sreenivasa Murthy, S. G. S., Garza, K., Budhwani, S., Richardson, L. S., Menon, R., Krishnan, B. ]]></dc:creator>
<dc:date>2026-04-06</dc:date>
<dc:identifier>doi:10.64898/2026.04.02.716001</dc:identifier>
<dc:title><![CDATA[Exofection as a Therapeutic Modality: Restoring P-gp Activity via Trophoblast-Derived EV in Neuroinflammatory Disorders]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-06</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.04.03.716347v1?rss=1">
<title>
<![CDATA[
Simian Immunodeficiency Virus and Antiretroviral Therapy Impact Rhesus Macaque Brain Lipid Distribution 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.04.03.716347v1?rss=1
</link>
<description><![CDATA[
Human immunodeficiency virus (HIV) infection promotes considerable bioenergetic, spatially heterogenous strain to the brain that is incompletely ameliorated through viral suppression afforded by antiretroviral therapy (ART). Disrupted homeostasis of brain lipids after HIV in humans or simian immunodeficiency virus (SIV) infection in rhesus macaques occurs due to elevated energetic demands, neuroinflammation, reactive oxygen species, and barrier leakiness. Brain lipids are particularly vulnerable to HIV-associated dysregulation due to their high abundance, unique composition, and specialized functional roles. Using rhesus macaques exposed to SIV and ART (tenofovir disoproxil fumarate (TDF), emtricitabine (FTC), and dolutegravir (DTG), we investigated the spatial distribution and abundance of lipids across brain regions and metabolically relevant peripheral tissues using mass spectrometry imaging. When comparing lipid abundance, individual lipids representing a multitude of species were more varied across tissues than by treatment condition.

Further, we discerned either solely SIV infection or ART outweighed one another in altering phospholipids in different tissues Presence of ART had a greater influence on phospholipid homeostasis in the temporal cortex and hippocampus than in the midbrain, possibly due to differences in penetrance and turnover of ART across brain regions.

Overall, these data demonstrate ART robustly increased phospholipids across brain regions while SIV infection had a varied impact depending on the brain region. These findings inform the need to further evaluate the neurologic consequences that may result in the brain due to disrupted lipid homeostasis across ART regimens.
]]></description>
<dc:creator><![CDATA[ White, C. J., Vanderschoot, K. A., Brown, D. R., Espley, A. F., Neumann, E. K., Tressler, C. M., Williams, D. W. ]]></dc:creator>
<dc:date>2026-04-04</dc:date>
<dc:identifier>doi:10.64898/2026.04.03.716347</dc:identifier>
<dc:title><![CDATA[Simian Immunodeficiency Virus and Antiretroviral Therapy Impact Rhesus Macaque Brain Lipid Distribution]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-04</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.03.31.715705v1?rss=1">
<title>
<![CDATA[
Changes in dysbiosis and gene expression in the gut of wharf roach (Ligia Spp.) fed with expanded polystyrene 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.03.31.715705v1?rss=1
</link>
<description><![CDATA[
The East Asian region, known for its high levels of human and fishery activities, experiences serious plastic pollution in the marine environment, especially in seawater and along coastlines. Wharf roaches (Ligia spp.) collected from the coast of western Japan frequently ingest expanded polystyrene (EPS), which is then excreted as microplastic through their feces. However, the impact of EPS exposure and ingestion on the gut microbiome of wharf roaches remains unclear. Thus, this study aimed to investigate the effects of EPS ingestion on the gut microbiota of wharf roaches by examining their gut microbiota and gene expression. The expression levels of more than 400 genes, including those associated with xenobiotic metabolism, and the abundance of gut microbial community were altered. Microbial analysis revealed that at least five archaeal types, two to four bacterial types, three to seven eukaryotic types, and three viral types were involved in a correlation network composed of strong associations. Among them, Haloquadratum, Halalkalicoccus, and Methanospirillum (archaea); Volvox (eukaryote); and Varicellovirus and T4-like viruses showed significantly increased abundance. Furthermore, covariance structure analysis indicated that the viruses and methanogens played key causal roles as characteristic factors related to EPS administration. In conclusion, EPS disrupts the intestinal environment of wharf roaches and serves as a potential material for viral activation and methane production. Building on our previous field study that identified wharf roaches as potential indicators of coastal EPS pollution, this study provides novel insights into the ecological impacts of EPS ingestion and consequences of plastic pollution.
]]></description>
<dc:creator><![CDATA[ LEE, S., Miyamoto, H., Takai, Y., Suda, W., Ohno, H., Simasaki, Y., Oshima, Y. ]]></dc:creator>
<dc:date>2026-04-03</dc:date>
<dc:identifier>doi:10.64898/2026.03.31.715705</dc:identifier>
<dc:title><![CDATA[Changes in dysbiosis and gene expression in the gut of wharf roach (Ligia Spp.) fed with expanded polystyrene]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-03</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.03.31.715675v1?rss=1">
<title>
<![CDATA[
Low cadmium concentrations alter B and T cell responses in Jamaican fruit bats (Artibeus jamaicensis) 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.03.31.715675v1?rss=1
</link>
<description><![CDATA[
Bats are exposed to a variety of pollutants, including cadmium (Cd), that can impair immune function and potentially increase viral shedding and burden. Despite this, little is known about the impacts of heavy metals on bats. This study aimed to determine the impacts of Cd exposure on bat T and B cell immune responses in naive and coronavirus infected bats and determine the impact of Cd on viral replication in Jamaican fruit bat (JFB; Artibeus jamaicensis) cells. To determine the impact of Cd exposure on adaptive immune responses, splenocyte cultures from naive and BANAL-52 coronavirus infected JFB were treated with 0, 1, and 10 {micro}M Cd and stimulated overnight with concanavalin A. RNA was extracted, a SYBR Green qPCR was used to assess gene expression. To determine if Cd exposure increased viral replication, two JFB kidney cell clones were treated with 0, 1, 10, and 50 {micro}M of CdCl2 overnight and then infected with Cedar virus (CedV). Supernatants were collected and viral titers determined. Several transcripts were upregulated in both naive and virus infected JFB splenocytes treated with Cd. B cell transcripts were significantly upregulated in a dose-dependent manner and T cell transcripts were also increased in Cd treated splenocytes. Assessment of transcripts associated with T cell subsets suggest a predominant Th2 response in Cd treated splenocytes. Viral replication was not significantly different in Cd treated kidney clones compared to the non-treated cells. These studies provide evidence that JFB adaptive immune responses are altered when exposed to low Cd concentrations.
]]></description>
<dc:creator><![CDATA[ Pulscher, L. A., Charley, P. A., Zhan, S., Reasoner, C., Burke, B., Schountz, T. ]]></dc:creator>
<dc:date>2026-04-03</dc:date>
<dc:identifier>doi:10.64898/2026.03.31.715675</dc:identifier>
<dc:title><![CDATA[Low cadmium concentrations alter B and T cell responses in Jamaican fruit bats (Artibeus jamaicensis)]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-03</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.03.31.715632v1?rss=1">
<title>
<![CDATA[
Nicotine self-administration increases impulsive action: differential effects of nAChR modulators in a Go/No-Go task 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.03.31.715632v1?rss=1
</link>
<description><![CDATA[
Tobacco use disorder is a chronic condition characterized by compulsive nicotine use, withdrawal, and relapse following abstinence. Impulsivity contributes to persistent nicotine use and poor cessation outcomes. This study examined whether nicotinic acetylcholine receptor (nAChR) modulators alter impulsive action in a nicotine self-administration Go/No-Go task in male and female rats. Rats acquired intravenous nicotine self-administration and were then trained in a Go/No-Go procedure in which active lever presses were reinforced during Go periods but not during No-Go periods. We then assessed the effects of varenicline (0.1-3 mg/kg), nicotine (0.1-0.6 mg/kg), and the nAChR antagonist mecamylamine (0.5-2 mg/kg) in the Go/No-Go procedure. Varenicline and nicotine pretreatment reduced active responding during both Go and No-Go periods, whereas mecamylamine selectively reduced responding during No-Go periods. Mecamylamine decreased the percentage of active responses during No-Go trials, indicating reduced bias toward the nicotine-associated lever. In contrast, nicotine and varenicline did not alter response allocation, suggesting that their effects reflected nonspecific reductions in responding rather than changes in impulsive action. No sex differences were observed. Substituting saline for nicotine during self-administration did not alter active responding during Go periods, but rats in the saline group had fewer active responses during No-Go periods than rats in the nicotine group. These results show that chronic nicotine self-administration increases impulsive action and that nAChR antagonism, but not agonism or partial agonism, reduces nicotine-related impulsive action. This work supports the utility of the Go/No-Go self-administration task for investigating nAChR-dependent mechanisms underlying nicotine-induced impulsivity.
]]></description>
<dc:creator><![CDATA[ Chellian, R., Huisman, G., Bruijnzeel, A. ]]></dc:creator>
<dc:date>2026-04-02</dc:date>
<dc:identifier>doi:10.64898/2026.03.31.715632</dc:identifier>
<dc:title><![CDATA[Nicotine self-administration increases impulsive action: differential effects of nAChR modulators in a Go/No-Go task]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.03.30.715362v1?rss=1">
<title>
<![CDATA[
Decoding the Structural and Functional Impact of the Leukaemia-Associated A338V Mutation in GPR183 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.03.30.715362v1?rss=1
</link>
<description><![CDATA[
G protein-coupled receptors rely on dynamic conformational changes to coordinate G protein activation and recruitment of regulatory transducers such as G protein-coupled receptor kinases and {beta}-arrestins. The chemotactic receptor GPR183 has been implicated in a context-dependent role in hematological malignancies. Here, we investigated the impact of A338V mutation located within the C-terminal tail of GPR183. This mutation is associated with acute myeloid leukaemia. Using bioluminescence resonance energy transfer-based assays in HEK293A cells, we assessed receptor-proximal signaling events. The A338V variant displayed preserved agonist potency and comparable agonist-induced Gi activation relative to wild type, although constitutive activity towards Gi was modestly reduced. In contrast, recruitment of GRK2 and {beta}-arrestin2 was consistently impaired across multiple assay configurations. These differences were not attributable to altered receptor abundance, as the C-tail untagged mutant exhibited increased plasma membrane expression despite reduced regulatory transducer engagement. While intramolecular conformational biosensor measurements revealed subtle differences in global receptor conformation between WT and A338V, extensive molecular dynamics simulations supported the altered conformational sampling of the C-terminal tail in the A338V variant. Together, these data support a model in which the A338V substitution selectively alters C-terminal structural dynamics, impairing GRK2 and {beta}-arrestin2 recruitment while preserving G protein activation.
]]></description>
<dc:creator><![CDATA[ Andersson, L., Wesolowski, P. A., Jahrstorfer, L., De Rosa, A., Heger, T., Neuman, V., Sieradzan, A. K., Wales, D. J., Kozielewicz, P. ]]></dc:creator>
<dc:date>2026-04-01</dc:date>
<dc:identifier>doi:10.64898/2026.03.30.715362</dc:identifier>
<dc:title><![CDATA[Decoding the Structural and Functional Impact of the Leukaemia-Associated A338V Mutation in GPR183]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-01</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.03.30.715248v1?rss=1">
<title>
<![CDATA[
Electroconvulsive seizures for alcohol use disorder: a preclinical study 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.03.30.715248v1?rss=1
</link>
<description><![CDATA[
The use of neuromodulation techniques for the treatment of alcohol use disorder is receiving increasing attention, especially non-invasive approaches, such as repetitive transcranial magnetic stimulation or transcranial direct current stimulation, while the hypothetical use of electroconvulsive therapy remains unexplored. Given our experience inducing electroconvulsive seizures (ECS) for therapeutic purposes in psychopathology rodent models, we evaluated the role of ECS on reducing the increased voluntary ethanol consumption caused by adolescent ethanol exposure in our validated preclinical model. Rats were treated in adolescence with a binge paradigm of ethanol (2 g/kg, i.p.; 3 rounds of 2 days at 48-h intervals; post-natal day, PND 29-30, PND 33-34 and PND 37-38) or saline. Following persistent withdrawal until adulthood, rats were allowed to: voluntarily drink ethanol (20%) by a two-bottle choice test, for 3 days (PND 80-82); treated with ECS (95 mA for 0.6 s, 100 Hz, pulse width 0.6 ms; ear-clip electrodes) or SHAM for 5 days (PND 86-90); re-exposed to voluntarily ethanol exposure (PND 94-96). Brains were collected on PND 97 to evaluate hippocampal markers of ethanol toxicity and/or treatment response (e.g., NeuroD, NF-L, BDNF and NF-L/BDNF ratio). Our results reproduced the increased voluntary ethanol consumption in adult rats induced by adolescent ethanol exposure and demonstrated that ECS could improve this abuse-prone response. Moreover, we suggested a possible role for BDNF in the beneficial effects induced by ECS, especially reducing the neurotoxic ratio NF-L/BDNF. Overall, we provide preclinical evidence for the potential use of ECS as an efficacious treatment for alcohol use disorder.
]]></description>
<dc:creator><![CDATA[ Garcia-Cabrerizo, R., Bergas-Cladera, P., Colom-Rocha, C., Garcia-Fuster, M. J. ]]></dc:creator>
<dc:date>2026-04-01</dc:date>
<dc:identifier>doi:10.64898/2026.03.30.715248</dc:identifier>
<dc:title><![CDATA[Electroconvulsive seizures for alcohol use disorder: a preclinical study]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-01</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.03.26.714567v1?rss=1">
<title>
<![CDATA[
Steroid-based Tide Quencher 1 probes enable real-time mapping of novel non-canonical cholesterol sites on the M1 muscarinic receptor 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.03.26.714567v1?rss=1
</link>
<description><![CDATA[
Steroid-based fluorescent-quencher probes now enable real-time, residue-level mapping of previously inaccessible cholesterol-binding sites on G-protein-coupled receptors. We designed Tide Quencher 1 (TQ1) conjugated steroids that target two distinct peripheral sites on the M1 muscarinic receptor. One near the extracellular N-terminus and another adjacent to the intracellular C-terminus. Using pregnanolone glutamate as a versatile scaffold, we synthesised a library of probes varying in C-3 linker length ({gamma}-aminobutyric acid vs. L-glutamic acid) and C-3/C-5 stereochemistry (3/3{beta}/5/5{beta}). Fluorescence-quenching assays with CFP-tagged receptors revealed that TQ1 probes consistently outperformed Dabcyl, delivering up to 40 % quenching within minutes and sub-micromolar EC50 values. The most potent N-terminal probe (35-PRG-Glu-TQ1 (5)) achieved 300 nM potency, while the best C-terminal probe (35{beta}-PRG-Glu-TQ1 (3)) reached 1 {micro}M potency with rapid association. Molecular docking and MD simulations identified key residues (K20, Q24, W405 at the N-site; K57, Y62, W150 at the C-site) mediating binding, a prediction confirmed by alanine-scan mutagenesis that markedly reduced quenching at the N-terminus and only modestly affected the C-terminus. Competition experiments with non-quenching analogues further validated probe specificity. Crucially, the pregnane core proved essential; alternative steroid backbones failed to generate robust quenching. This fluorescence-quenching platform overcomes the limitations of traditional radioligand assays, providing kinetic insight, high-throughput compatibility, and the ability to dissect lipid-GPCR interactions in native membranes. The approach is readily extensible to other GPCR families, opening new avenues for structure-guided drug discovery targeting allosteric cholesterol sites.
]]></description>
<dc:creator><![CDATA[ Chetverikov, N., Szanti-Pinter, E., Jurica, J., Vodolazhenko, M., Budesinsky, M., Zima, V., Svoboda, M., Dolejsi, E., Janouskova-Randakova, A., Urbankova, A., Jakubik, J., Kudova, E. ]]></dc:creator>
<dc:date>2026-04-01</dc:date>
<dc:identifier>doi:10.64898/2026.03.26.714567</dc:identifier>
<dc:title><![CDATA[Steroid-based Tide Quencher 1 probes enable real-time mapping of novel non-canonical cholesterol sites on the M1 muscarinic receptor]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-01</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.03.30.715187v1?rss=1">
<title>
<![CDATA[
In vivo pharmacokinetics and tissue distribution profile of a Wnt/β-catenin pathway-targeting anticancer cassane diterpene isolated from Caesalpinia pulcherrima 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.03.30.715187v1?rss=1
</link>
<description><![CDATA[
A cassane diterpene, 6{beta}-cinnamoyl-7-hydroxyvouacapen-5-ol (6{beta}CHV), isolated from Caesalpinia pulcherrima, has emerged as a promising anticancer drug lead with reported Wnt/{beta}-catenin pathway inhibitory activity and in vivo safety. The present study reports the in vivo pharmacokinetics and tissue distribution of 6{beta}CHV in Wistar rats following a single oral dose of 200 mg/kg. A reproducible RP-HPLC-UV method was developed and validated for quantifying 6{beta}CHV in rat plasma and tissues. Chromatographic separation was achieved using a gradient elution of methanol and water. The method was subsequently applied to investigate the pharmacokinetics and tissue distribution of 6{beta}CHV. Plasma pharmacokinetic analysis revealed delayed and moderate absorption, with a Tmax of 4 h and a Cmax of 1314.12 ng/mL. Following absorption, 6{beta}CHV is distributed widely across peripheral tissues, including the liver, heart, lungs, spleen, and kidneys, as well as pharmacological sanctuary sites such as the brain and testes. The highest concentrations were observed in the stomach, small intestine, and liver, with detectable levels persisting up to 24 h, reflecting extensive tissue partitioning and retention. Overall, these findings demonstrate that oral administration of 6{beta}CHV is feasible. However, the delayed absorption suggests that further optimization of formulation or alternative administration routes may enhance systemic exposure. This study provides the first comprehensive pharmacokinetic and tissue distribution profile of 6{beta}CHV, supporting its continued preclinical development as a potential anticancer therapeutic.

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=125 SRC="FIGDIR/small/715187v1_ufig1.gif" ALT="Figure 1">
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]]></description>
<dc:creator><![CDATA[ De Vass Gunawardane, S., Epitawala Arachchige, O. V., Wijerathne, S. K., Punyasiri, P. A. N., Murugananthan, A., Samarakoon, S. R., Senathilake, K. S. ]]></dc:creator>
<dc:date>2026-04-01</dc:date>
<dc:identifier>doi:10.64898/2026.03.30.715187</dc:identifier>
<dc:title><![CDATA[In vivo pharmacokinetics and tissue distribution profile of a Wnt/β-catenin pathway-targeting anticancer cassane diterpene isolated from Caesalpinia pulcherrima]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-04-01</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.03.29.713781v1?rss=1">
<title>
<![CDATA[
Cirrina: LLM-driven pharmacological reasoning agent enables preclinical CNS drug evaluation 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.03.29.713781v1?rss=1
</link>
<description><![CDATA[
Assessing whether a preclinical drug candidate will work is not a prediction problem but a reasoning problem. The same numerical output warrants different interpretations depending on the target and therapeutic context. CNS drug development presents the most demanding instance of this reasoning problem. For example, a compound must cross the blood-brain barrier, resist efflux transport, and achieve adequate receptor occupancy at a dose that clears safety margins. The constraints interact with each other in a web that needs careful interpretation. Here, we show that Cirrina, an LLM agent coupled to eight mechanistic pharmacology tools, can reason across the input data to provide better decisions and a well documented reasoning trace. The LLM agent reasons across multiple data tiers from SMILES to animal PK/PD measurements adjusting thresholds based on target-specific requirements. Validated against 181 CNS compounds, it achieved a 68% accuracy compared to a rule-based deterministic pipeline of 31% accuracy. In 103 discordant cases, the agents reasoning was correct in 75% of instances compared to only 10% for deterministic pipelines. Cirrina provides a scalable, documented framework for preclinical decision-making, effectively identifying failure-prone candidates that generic thresholds overlook, and thereby reducing the chances of failure along the clinical development cycle.
]]></description>
<dc:creator><![CDATA[ Rajbanshi, B., Iqbal, K., Guruacharya, A. ]]></dc:creator>
<dc:date>2026-03-31</dc:date>
<dc:identifier>doi:10.64898/2026.03.29.713781</dc:identifier>
<dc:title><![CDATA[Cirrina: LLM-driven pharmacological reasoning agent enables preclinical CNS drug evaluation]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-03-31</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.03.27.714742v1?rss=1">
<title>
<![CDATA[
Study comparing characteristics of ademetionine-containing tablets from different countries 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.03.27.714742v1?rss=1
</link>
<description><![CDATA[
Background/ObjectivesThe quality and characteristics of approved medicines can vary substantially depending on manufacturing processes and standards within a given country. The aim of the study was to compare the available marketed brands of ademetionine tablets derived from various countries in order to identify potential differences between the different formulations.

MethodsWe performed comprehensive analyses of the physical, chemical, and dissolution characteristics of different formulations of ademetionine tablets marketed in China, India, Russia, Ukraine, and Uzbekistan, using the originator formulation of Heptral(R) as the reference standard. The formulations were evaluated at initial analysis and after 3 months at 40{degrees}C/75% relative humidity. Clinical parameters such as ademetionine content, degradation products, S,S-isomer, and water content were assessed using HPLC, and a dissolution profile analysis performed in 2 hours of acid solution followed by 90 minutes in a buffer solution.

ResultsThe Nusam (India) and Ximeixin (China) products were the two products most comparable to the Heptral products. Adenomak (Ukraine), the only food-grade product and only one with the tosylate salt showed the most significant quality variations compared to Heptral including dissolution failure as well as considerable variability between batches.

ConclusionsThe study highlights the importance of using pharmaceutical-grade ademetionine products to maintain clinical efficacy and ensuring standards are maintained across global markets.
]]></description>
<dc:creator><![CDATA[ Mato, J. M., Wong, G. L., Gooijer, Y., Safaei, A. ]]></dc:creator>
<dc:date>2026-03-30</dc:date>
<dc:identifier>doi:10.64898/2026.03.27.714742</dc:identifier>
<dc:title><![CDATA[Study comparing characteristics of ademetionine-containing tablets from different countries]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-03-30</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.03.27.714572v1?rss=1">
<title>
<![CDATA[
Targeting Protease-activated Receptor 4 (PAR4) Protects Against Acute Kidney Injury (AKI) in Ischemia Reperfusion Injury 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.03.27.714572v1?rss=1
</link>
<description><![CDATA[
Acute kidney injury (AKI) is a serious and common clinical syndrome that currently has no effective treatment. Emerging evidence links coagulation pathways to kidney injury, particularly through coagulation proteases. Protease-activated receptors (PARs) are a family of G-protein coupled receptors (GPCRs) that are activated by proteolytic cleavage of their N termini, exposing a tethered ligand that initiates receptor signaling. PARs have been shown to play a major role in inflammation, vascular regulation, and tissue injury. PARs play key roles in inflammation, vascular regulation, and tissue injury. Previous work from the Hamm laboratory demonstrated that PAR4 contributes to AKI progression, as PAR4 knockout mice were protected in both unilateral ureteral obstruction and ischemia-reperfusion-based models of kidney disease.

In this study, we investigated the potential of a PAR4 antagonist, VU6073819, at mitigating AKI progression in an ischemia-reperfusion injury (IRI) mouse model. PAR4 antagonism not only alleviated kidney injury and inflammatory response, but it significantly improved the survival. These findings identify PAR4 as a promising therapeutic target for AKI.
]]></description>
<dc:creator><![CDATA[ Webb, E. M., Cao, S., Pan, Y., Zhang, M.-Z., Harris, R., Boutaud, O., Bouchard, J. L., Jones, C. K., Lindsley, C. W., Hamm, H. E. ]]></dc:creator>
<dc:date>2026-03-30</dc:date>
<dc:identifier>doi:10.64898/2026.03.27.714572</dc:identifier>
<dc:title><![CDATA[Targeting Protease-activated Receptor 4 (PAR4) Protects Against Acute Kidney Injury (AKI) in Ischemia Reperfusion Injury]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-03-30</prism:publicationDate>
<prism:section></prism:section>
</item>
</rdf:RDF>
