Biomedicine & Pharmacotherapy

Acute myeloid leukemia (AML) remains a difficult disease to cure despite recent advances. Off-target side effects of therapy, especially prolonged cytopenia, lead to significant morbidity and mortality. There is an increased recognition that in AML, there is a potentially vulnerable dependence on OXPHOS metabolism, more so in the leukemia stem cell compartment (AML-LSC) that could be exploited. Drug re-purposing screens have suggested the possible role of artesunate (ART) in inhibiting mitochondrial respiration and arsenic trioxide (ATO) in inhibiting glycolysis. Here, we explore the potential of using a combination of ART and ATO to treat AML. Through in-vitro and in-vivo studies, we demonstrate the potential of this combination along with hypo-methylating agents and other mitocans, such as BCL-2 inhibitor venetoclax, to be used in the treatment of AML with minimal off-target effect on normal hematopoietic stem cells (HSC). These observations warrant further exploration of such novel combinations in clinical trials.

The Coronaviridae family has been implicated in several major epidemics over the past two decades, including those caused by SARS-CoV, MERS-CoV, and, most recently, SARS-CoV-2. The COVID-19 pandemic, driven by SARS-CoV-2, has led to over seven million deaths worldwide and has been associated with prolonged symptoms, chronic sequelae, and substantial socioeconomic disruptions. The limited availability of effective antiviral treatments, coupled with the ability of coronaviruses to mutate and evade immune defenses, underscores the urgent need for innovative antiviral agents. This study explores the efficacy of the nucleoside analogue DFMA as a potential antiviral agent against multiple Coronaviridae family members, including SARS-CoV-2 and two strains of murine hepatitis viruses (MHV-3 and MHV-A59). In vitro analyses demonstrated that DFMA effectively reduced the viral load in the supernatant of infected cells and enhanced cell viability for both MHV-3 and MHV-A59. Against SARS-CoV-2, DFMA showed a significant reduction in viral load, with a calculated Selectivity Index (SI) of 6.2. In vivo investigations further confirmed the antiviral potential of DFMA. In three distinct murine models--a severe COVID-19 model using MHV-3, a mild COVID-19 model employing MHV-A59, and a transgenic K18-hACE2 mouse model infected with SARS-CoV-2--DFMA administration significantly reduced viral loads in the lungs of infected mice. Additionally, DFMA mitigated inflammatory responses in all models by lowering levels of key inflammatory mediators, such as CXCL1, CCL2, and IL-6. These findings suggest that DFMA possesses broad-spectrum antiviral activity against coronaviruses and may serve as a promising therapeutic candidate for current and future coronavirus outbreaks. Further research is warranted to elucidate its mechanism of action and evaluate its efficacy in clinical settings. ImportanceCoronaviruses have caused significant outbreaks over the past two decades. Since 2020, COVID-19 has resulted in millions of deaths and lasting global impacts. The limited availability of effective antivirals and the viruss ability to mutate and evade vaccines and monoclonal antibody therapy emphasize the urgent need for new treatments. This study investigates DFMA, a promising antiviral candidate, targeting SARS-CoV-2 and two related coronaviruses.Our promising results demonstrated significant antiviral activity of DFMA, not only against SARS-CoV-2 but also against other similar coronaviruses, indicating potential future use against COVID-19 and other possible coronavirus-related diseases.

Despite progress in the treatment of non-visceral malignancies, the prognosis remains poor for malignancies of visceral organs and novel therapeutic approaches are urgently required. Here we introduce a novel therapeutic regimen by treatment with Se-methylselenocysteine (MSC) and concomitant tumor-specific induction of Kynurenine aminotransferase 1 (KYAT1) in hepatocellular carcinoma (HCC) cell lines, using either vector-based and/or lipid nanoparticle-mediated delivery of mRNA. Supplementation of MSC in KYAT1 overexpressed cells resulted in significantly increased cytotoxicity as compared to MSC alone. Furthermore, microRNA antisense targeted sites for miR122, known to be widely expressed in normal hepatocytes whilst downregulated in hepatocellular carcinoma, were added to specifically limit cytotoxicity in HCC cells, thereby limiting off-target effects. KYAT1 expression was significantly reduced in cells with high levels of miR122 supporting the concept of miR-guided induction of tumor-specific cytotoxicity. The addition of alpha-ketoacid favored the production of methylselenol, enhancing the cytotoxic efficacy of MSC in HCC cells, with no effects on primary human hepatocytes. Altogether, the proposed regimen offers great potential to safely and specifically target hepatic tumors that are currently untreatable.

BackgroundAcute liver failure (ALF) is associated with rapid and progressive hepatocellular injury, and severe metabolic-microbial derangements. We investigated early metabolic markers of non-survival, and a potential microbial intervention using Bacteroides intestinalis-AM1, to improve outcomes in ALF. MethodPlasma metabolomics and meta-proteomics were performed in 40 ALF patients and 5 healthy controls (training cohort). A non-survival marker panel was identified and validated in 270 ALF patients (test cohort) using high resolution mass spectrometry and machine learning. It was functionally validated in acetaminophen-induced ALF mouse model. B. intestinalis-AM1 was used to study alteration of gut bacteria and amelioration of liver injury. ResultsALF non-survivors showed a distinct metabolomic signature with elevated primary bile acids {chenodeoxycholic acid (CDCA), cholic acid (CA)}, tryptophan, tyrosine, and enrichment of pathways linked to inflammation, cell death, and stress response (p<0.01, FDR<0.01, FC>1.5). Non-survivors had higher alpha/beta diversity (p<0.05) with increase in Proteobacteria, Firmicutes, Actinobacteria (p<0.05); functionally associated with energy, amino acid and xenobiotic metabolism (p<0.05). A gut microbiota derangement in converting primary to secondary bile acids was evident as CDCA and cytotoxic metabolites (4-(2-Amino phenyl)-2,4-dioxobutanoate, L-Tyrosine) were higher. Elevated CDCA (logFC>10) levels correlated with mortality in ALF patients as well as in mouse model. In the later, administration of B. intestinalis-AM1 bacteria, (10^9) reduced CDCA and CA levels by enhancing FXR, FGF15, SLC10A1 gene expression, attenuating inflammation (IL-1beta, TLR4-signalling), necroptosis, and modulating glutathione(oxidative-repair), tryptophan(inflammation), and histidine (tissue repair) metabolism. ConclusionHigh levels of chenodeoxycholic acid (CDCA) represent a poor prognostic indicator in ALF patients. B. intestinalis-AM1, a primary-to-secondary bile acid converter, effectively reduce CDCA levels, activated FXR, reduced inflammation and protected hepatocytes, highlighting its therapeutic potential in ALF.

BackgroundDianthus superbus L. (DS) treats liver diseases, but its ALI mechanism via gut microbiota/metabolites is unclear. PurposeInvestigate DS effects on CCl-induced ALI and mechanisms. MethodsDS components (HPLC-MS/MS) were identified. ALI rats received DS. Liver/colon injury, fibrosis (biochemistry, histology, IHC, IF, WB for -SMA, Collagen I, Occludin, ZO-1), serum metabolomics, fecal 16S rRNA, and tryptophan-targeted metabolomics were assessed. Antibiotics and FMT from DS-treated donors explored microbiota roles. Results37 components, including Kaempferol and Aurantiamide, were identified. DS attenuated ALI, reducing hepatic -SMA/Collagen I while increasing AhR/Cyp1a1, and upregulating colonic Occludin/ZO-1. FMT replicated DS benefits. Untargeted metabolomics linked effects to Linoleic/Tryptophan metabolism. 16S rRNA showed increased beneficial microbiota (HT002, Lactobacillus, Romboutsia); FMT confirmed Lactobacillus enrichment. DS normalized tryptophan metabolites (4-Hydroxybenzoic acid, L-Kynurenine, Indole-3-carboxaldehyde). ConclusionDS ameliorates ALI gut-microbiota-dependently by modulating tryptophan metabolism, enhancing intestinal barrier function, activating AHR signaling, and suppressing fibrosis biomarkers, indicating its potential as a microbiota modulator for ALI. ImportanceAcute liver injury is a serious health threat with limited treatment options. This study explores how a traditional medicinal herb, Dianthus superbus L. (DS), protects the liver through an unexpected route: the gut. We discovered that DS works by rebalancing the community of gut bacteria, which in turn improves the intestinal barrier and fine-tunes the bodys tryptophan metabolism. This cascade of effects ultimately activates a key cellular pathway (AhR) that reduces liver inflammation and fibrosis. Our findings not only clarify how this traditional remedy works but also highlight the critical role of the gut-liver axis, positioning DS as a promising, microbiota-targeting therapeutic candidate for preventing and treating liver disease.

Acetaminophen (APAP) overdose triggers a cascade of intracellular oxidative stress events culminating in acute liver injury. The clinically used antidote, N-acetylcysteine (NAC) has a narrow therapeutic window and early treatment is essential for satisfactory therapeutic outcome. For more versatile therapies that can be effective even at late-presentation, the intricacies of APAP-induced hepatotoxicity must be better understood. Accumulation of advanced glycation end-products (AGEs) and consequent activation of the receptor for AGEs (RAGE) are considered one of the key mechanistic features of APAP toxicity. Glyoxalase-1 (Glo-1) regulates AGE formation by limiting the levels of methylglyoxal (MEG). In this study, we studied the relevance of Glo-1 in APAP mediated activation of RAGE and downstream cell-death cascades. Constitutive Glo-1 knockout mice (GKO) and a cofactor of Glo-1, {psi}-GSH, were employed as tools. Our findings show elevated oxidative stress, activation of RAGE and hepatocyte necrosis through steatosis in GKO mice treated with high-dose APAP compared to wild type controls. A unique feature of the hepatic necrosis in GKO mice is the appearance of microvesicular steatosis as a result of centrilobular necrosis, rather than inflammation seen in wild type. The GSH surrogate and general antioxidant, {psi}-GSH alleviated APAP toxicity irrespective of Glo-1 status, suggesting that oxidative stress being the primary driver of APAP toxicity. Overall, exacerbation of APAP hepatotoxicity in GKO mice suggests the importance of this enzyme system in antioxidant defense against initial stages of APAP overdose.

Omacetaxine, a semisynthetic form of Homoharringtonine (HHT), was approved for the treatment of chronic myeloid leukemia (CML). Previously, we published the synthesis of this natural alkaloid and three of its derivatives: deoxyharringtonine (DHT), deoxyhomoharringtonine (DHHT), and bis(demethyl)-deoxyharringtonine (BDHT); and reported on its refractory activity against the HL-60/RV+ cells over-expressing P-glycoprotein 1 (MDR1). In this study, we explored the extent of this resistance by first expanding the panel of established cell lines and second, using a panel of 21 leukemia patient derived primary cells. Here, we report a consistent resistance to HTT in K562 derived cells and in MES-SA/MX2 derived cells resistant to mitoxanthrone; all of them over-express MDR1, while we found U87MG-ABCG2 and H69AR cells to be very sensitive to HTT. In contrast, DHT, DHHT, and BDHT seemingly overcome this resistance due to the changes made to the acyl chain of HTT rendering the derivatives less susceptible to efflux. Surprisingly, the leukemia primary cells were very sensitive to HHT and its derivatives with low nanomolar potencies, followed by a new class of CDC7 kinase inhibitors, the anthracycline class of topoisomerase inhibitors, the DNA intercalator actinomycin-D, and the vinca alkaloid class of microtubule inhibitors. The mechanism of cell death induced by HTT and DHHT was found to be mediated via Caspase 3 cleavage leading to apoptosis. Taken together, our results confirm that HHT is a substrate for MDR1. It opens the door to a new opportunity to clinically evaluate HHT and its derivatives for the treatment of AML and other cancers.

The emergence of new SARS-CoV-2 variants, capable of escaping the humoral immunity acquired by the available vaccines, together with waning immunity and vaccine hesitancy, challenges the efficacy of the vaccination strategy in fighting COVID-19. Improved therapeutic strategies are therefore urgently needed to better intervene particularly in severe cases of the disease. They should aim at controlling the hyper-inflammatory state generated upon infection, at reducing lung tissue pathology and endothelial damages, along with viral replication. Previous research has pointed a possible role for the chaperone HSP90 in SARS-CoV-2 replication and COVID-19 pathogenesis. Pharmacological intervention through HSP90 inhibitors was shown to be beneficial in the treatment of inflammatory diseases, infections and reducing replication of diverse viruses. In this study, we analyzed the effects of the potent HSP90 inhibitor Ganetespib in vitro on alveolar epithelial cells and alveolar macrophages to characterize its effects on cell activation and viral replication. Additionally, to evaluate its efficacy in controlling systemic inflammation and the viral burden after infection in vivo, a Syrian hamster model was used. In vitro, Ganetespib reduced viral replication on AECs in a dose-dependent manner and lowered significantly the expression of pro-inflammatory genes, in both AECs and alveolar macrophages. In vivo, administration of Ganetespib led to an overall improvement of the clinical condition of infected animals, with decreased systemic inflammation, reduced edema formation and lung tissue pathology. Altogether, we show that Ganetespib could be a potential medicine to treat moderate and severe cases of COVID-19.

The combination of venetoclax (VEN) and hypomethylating agents (HMA) is the standard of care in acute myeloid leukemia (AML) for elderly patients unfit for intensive chemotherapy. Despite its clinical success, most patients eventually relapse, creating an urgent need for effective therapeutic alternatives. In this study, we aimed to evaluate the potential of romaciclib, a first-in-class CDK8/CDK19 inhibitor, in combination with VEN to overcome stroma-mediated and primary/acquired VEN-resistance. We assessed the efficacy of RVU120+VEN combination in both sensitive and resistant AML cell lines and primary patient-derived models. Our finding demonstrated that romaciclib synergizes with VEN in AML cell lines and in 8 out of 11 patient-derived cell samples. The proteomic and functional studies demonstrated that combination induced apoptosis through caspase-dependent cleavage of MCL-1. In vivo studies confirmed the efficacy of RVU120+VEN, showing eradication of leukemic cells and bone marrow recovery. Importantly, the combination effectively overcame both stroma-mediated and transcriptionally dependent VEN-resistance. Mechanistic studies, focusing on transcriptomic analyses, identified key resistance-associated pathways, including IL6/JAK/STAT3, TGF-{beta}, PI3K/AKT/MTOR, and inflammatory signaling, being suppressed by combination treatment. Furthermore, an in vivo study using a VEN-resistant patient-derived xenograft (PDX) model confirmed the efficacy of the combination, demonstrating a significant reduction in leukemia burden and a decreased proportion of leukemia initiating cells (LIC) following treatment. These findings prove the highly synergistic mechanism of action of RVU120+VEN combination and the potential to overcome primary/acquired VEN resistance in relapse/refractory AML disease. Altogether, the presented results support ongoing clinical studies evaluating romaciclib and VEN in VEN/HMA-refractory patients (NCT06191263) and provide a basis for future exploration as a frontline therapy in VEN-naive patients.

Avasimibe; a cholesterol-lowering drug with a proven safety in clinical trials, has recently been repositioned as an anticancer agent in various preclinical investigations. A study from our group reported that hypercholesterolemia promotes hepatocellular carcinoma (HCC) cell survival and hampers the anticancer effect of sorafenib, a kinase inhibitor. In the present study, we demonstrate that in HCC under hypercholesterolemic conditions the anticancer property of sorafenib is potentiated by avasimibe (AVA) co-treatment. Further, to elucidate the role of hypercholesterolemia on sorafenib efficacy, in vitro and in vivo models of HCC were used. In vitro, co-treatment of both drugs synergistically inhibited HCC cell viability and induced cell death under normal and hypercholesterolemic conditions. At the molecular level, downregulation of ERK signalling and induction of endoplasmic reticulum stress are likely to contribute to the combinatorial cytotoxic effect of sorafenib and avasimibe in vitro. In mice, fed on a high-cholesterol diet (HCD), the efficacy of sorafenib was restored by co-administration of AVA. Collectively, these findings suggest that impairment in the efficacy of sorafenib because of hypercholesterolemic phenotype could be restored by AVA co-treatment, which may have implications towards treatment strategy. HighlightsO_LICholesterol impedes sorafenib efficacy in Hepatocellular carcinoma cells. C_LIO_LIAvasimibe restores the functionality of sorafenib under hypercholesterolemic environment. C_LIO_LICombine treatment of sorafenib and avasimibe synergistically enhances cytotoxicity in hepatocellular carcinoma. C_LIO_LISorafenib and avasimibe treatment in the presence of LDLc.is associated with diminished ERK activation and increased ER stress. C_LI

The FLT3 protein is a well-established therapeutic target in the treatment of Acute Myeloid Leukemia (AML), with its inhibition playing a crucial role in disease management. In this study, we identify and propose a novel FLT3 inhibitor that demonstrates superior binding affinity, stability, and pharmacokinetic properties compared to currently available inhibitors. We initially characterized the binding interactions of known FLT3 inhibitors through molecular docking and then strategically modified functional groups to enhance binding affinity, optimize drug-likeness, and minimize toxicity. The resulting analogue exhibits improved metabolic stability, lower toxicity, higher intestinal absorption, and superior permeability. Molecular dynamics simulations further confirm that the novel inhibitor forms stable and persistent interactions with FLT3, as evidenced by reduced conformational fluctuations and compact structural integrity. Free energy calculations reveal stronger ligand stabilization, while dynamic correlation analysis suggests enhanced engagement with critical residues, reinforcing its potential as an effective therapeutic agent. These findings highlight a promising candidate for further experimental validation and potential development in AML treatment.

BackgroundPalmitoyl-protein thioesterase-1 (PPT1) is an exciting druggable target for inhibiting autophagy in cancer. MethodsIn this study, we aimed to evaluate the effects of ezurpimtrostat-targeting PPT1 in combination with an anti-PD-1 antibody in liver cancer using a transgenic immunocompetent mouse model. ResultsHerein, we revealed that inhibition of PPT1 using ezurpimtrostat, a safe anticancer drug in humans, decreased the liver tumor burden by inducing the penetration of lymphocytes within tumors when combined with anti-programmed death-1 (PD-1). Inhibition of PPT1 potentiates the effects of anti-PD-1 immunotherapy by increasing the expression of major histocompatibility complex (MHC)-I at the surface of liver cancer cells and modulates immunity through recolonization and activation of cytotoxic CD8+ lymphocytes. ConclusionsEzurpimtrostat turns cold into hot tumors and, thus, constitutes a powerful strategy to improve T cell-mediated immunotherapies in liver cancer. Summary boxWe reported that inhibiting palmitoyl-protein thioesterase-1 enzyme (PPT1) enhances the antitumor activity of anti-programmed death-1 (PD-1) in liver cancer in preclinical models. This study provides the rational for this combination in cancer clinical trials. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=143 SRC="FIGDIR/small/524541v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@1f44bf3org.highwire.dtl.DTLVardef@1a66ca8org.highwire.dtl.DTLVardef@127a51corg.highwire.dtl.DTLVardef@1c8f2af_HPS_FORMAT_FIGEXP M_FIG C_FIG Ezurpimtrostat activities in cancerThe absence of immune effectors especially cytotoxic cells in the microenvironment of cold tumor is associated with a lack of response to ICI. This condition is mainly due to an increase in the autophagy process responsible for the sequestration and destruction of an antigen-presenting molecule, MHC-I. The inhibition of PPT1 using ezurpimtrostat treatment led to (1) the inhibition of PPT1 and consequently the autophagy process, (2) the increase of MHC-I surface expression, and (3) the recruitment and the activation of CD8+ T cells at tumor site leading to (4) the improvement of CD8+ T cell cytotoxic activity. Thus, ezurpimtrostat-treated tumors become eligible for anti-PD-1 immunotherapy as the combination of both led to decreased macronodules, micronodules, and tumor growth.

Despite known adverse effects of hydroxychloroquine (HCQ) and azithromycin (AZM) on cardiac function, HCQ and AZM have been used as combination therapy in the treatment of COVID-19 patients. Recent clinical data indicate higher complication rates with HCQ/AZM combination treatment in comparison to monotherapy. Here, we used human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) to systematically investigate the effects of HCQ and AZM individually and in combination. The clinically observed QT prolongation caused by treatment with HCQ could be recapitulated in iPSC-CMs based on prolonged field potential duration (FPDc). Interestingly, HCQ-induced FPDc prolongation was strongly enhanced by combined treatment with AZM, although AZM alone slightly shortened FPDc in iPSC-CMs. Furthermore, combined treatment with AZM and HCQ leads to higher cardiotoxicity, more severe structural disarrangement, and more pronounced contractile and electrophysiological dysfunctions, compared to respective mono-treatments. First mechanistic insights underlying the synergistic effects of AZM and HCQ on iPSC-CM functionality are provided based on increased Cx43- and Nav1.5-protein levels. Taken together, our results highlight that combined treatment with HCQ and AZM strongly enhances the adverse effects on cardiomyocytes, providing mechanistic evidence for the high mortality in patients receiving HCQ/AZM combination treatment.

Background and aimsAlcohol-related liver disease (ALD) has high mortality due to systemic inflammation. We analysed liver and monocyte kinome profiles in a chronic ethanol-fed pre-clinical rat model to identify therapeutic targets that could mitigate inflammation in ALD. MethodKinome profile was performed in liver and circulating monocytes at baseline, and after 8,12,16,20 and 24-weeks of 40% ethanol administration. Pathway-specific inhibitors, including PS1145 (IKK-phosphorylation inhibitor), PH-797804 (MAPK14 inhibitor), resveratrol and prednisolone were tested for their anti-inflammatory effects in ALD-rats, PBMC from patients and NIAAA mouse model. ResultsKinome profiling identified 497 liver and 345 monocyte kinases in ALD rats (FDR<0.01). A time-dependent increase in MAPK14-associated kinases was observed in both tissues (FC>1.5, p<0.05). By 24 weeks, 172 liver and 48 monocyte kinases were significantly upregulated, particularly those linked to MyD88-TLR4, PI3K-Akt, TNF, TGF{beta}, and cellular senescence pathways. Key contributors included TGF{beta}R1, ROS-generating kinases, and IL-1-driven MAPK14 and IKK phosphorylation. Targeting this axis, PS1145 (an IKK inhibitor) suppressed NF{kappa}B activation and inflammation in THP1 and HepG2 cells, as well as PBMCs from healthy and SAH patients, outperforming PH797804, resveratrol, and prednisolone (p<0.05, FC>1.5). PS1145 significantly reduced IL-6, TNF, and NF{kappa}B, while increasing IL-10. In vivo, PS1145 treatment in the NIAAA mouse model markedly reduced hepatic steatosis, cellular stress and inflammatory pathways (cytokine signalling, IL-36 signalling and others) more effectively than standard therapies, Notably, it also downregulated IL36R, impairing TLR receptor dimerization, suggesting a dual mechanism of action (p<0.05) highlighting its therapeutic potential in ameliorating systemic inflammation in ALD. ConclusionOur study highlights the key role of MAPK14 and MYD88-TLR4 pathway kinases in driving systemic inflammation in SAH. The IKK inhibitor PS1145, by blocking both IKK phosphorylation and TLR dimerization, effectively suppresses inflammatory signalling and improves liver pathology, positioning it as a promising targeted therapy for ALD.

Intra-tumoral heterogeneity has been shaping the field of precision medicine for cancer patients ever since its emergence. Prodrugs, which require activation by tumor associated enzymes (TAEs), are a rapidly emerging approach for targeted therapeutics. SULT1A1, a sulfotransferase enzyme and TAE, is over-expressed in about 5-15% of cancer patients including breast, prostate and renal cell carcinoma (RCC); however, it is either not expressed or expressed at low level in most normal tissue. Bioinformatic RNA analyses revealed that SULT1A1 over-expression in tumors is correlated with worse patient prognosis. We have identified a new compound, FIS103, which is a small molecule anti-cancer prodrug that is activated by SULT1A1 once internalized. This class of compounds, N-benzyl indole carbinols (N-BICs), cause rapid cell death by inducing widespread non-specific covalent alkylation of proteins in the cancer cell. We report that FIS103 displays potent antitumor activity in SULT1A1 over-expressing RCC cell lines (A498 and Caki-1). Contrarily, low SULT1A1 expressing RCC cells (786-O and ACHN) did not show any antitumor effects, which suggests low FIS103 toxicity in the absence of SULT1A1. In silico modeling validated the predicted SULT1A1-FIS103 interaction. Furthermore, FIS103 demonstrates potent SULT1A1-dependent antitumor activity in NU/J mouse xenografts injected with A498 cells. Remarkably, the flank tumors in mice regressed to non-measurable 14 days post-FIS103 treatment and did not regrow through the study conclusion. Additionally, SD rats treated with FIS103 once daily for 14 days demonstrated a promising liver toxicity profile with serum liver enzymes falling within the normal range and histopathology analysis indicated no difference between FIS103 or vehicle treated rats. We hereby demonstrate that FIS103 may have the potential to improve survival as well as quality of life of RCC patients and its application could be extended to other SULT1A1 expressing cancers.

The cannabinoid CB2 receptor (CB2R) is a potential therapeutic target for distinct forms of tissue injury and inflammatory diseases. To thoroughly investigate the role of CB2R in pathophysiological conditions and for target validation in vivo, optimal pharmacological tool compounds are essential. Despite the sizable progress in the generation of potent and selective CB2R ligands, pharmacokinetic parameters are often neglected for in vivo studies. Here, we report the generation and characterization of a tetra-substituted pyrazole CB2R full agonist named RNB-61 with high potency (Ki 0.13-1.81 nM, depending on species) and a peripherally restricted action due to P-glycoprotein mediated efflux from the brain. 3H and 14C labelled RNB-61 showed apparent Kd values < 4 nM towards human CB2R in both cell and tissue experiments. The >6000-fold selectivity over CB1 receptors and negligible off-targets in vitro, combined with high oral bioavailability and suitable systemic pharmacokinetic (PK) properties, prompted the assessment of RNB-61 in a mouse ischemia-reperfusion model of acute kidney injury (AKI) and in a rat model of chronic kidney injury/inflammation and fibrosis (CKI) induced by unilateral ureteral obstruction. RNB-61 exerted dose-dependent nephroprotective and/or antifibrotic effects in the AKI/CKI models. Thus, RNB-61 is an optimal CB2R tool compound for preclinical in vivo studies with superior biophysical and PK properties over generally used CB2R ligands.

The combination of direct-acting and host-directed antivirals targeting SARS-CoV-2 represents an attractive treatment strategy to combat COVID-19. In our previous work, we showed that the FDA-approved anti-arthritis drug Auranofin restricts SARS-CoV-2 replication and pathology in an animal model. Here, we report that Auranofin inhibits SARS-CoV-2 by targeting viral entry and main protease (Mpro) activity without affecting viral transcription. Time-of-addition studies combined with functional assays of viral entry, protease activity, and cell-cell fusion delineated its inhibitory effects at both early and late stages of the viral life cycle. Molecular docking and isothermal titration calorimetry analyses of Auranofin and the known Mpro inhibitor Nirmatrelvir indicated competitive binding within the Mpro active-site pocket. In addition, Auranofin attenuated NF-{kappa}B-dependent signalling and suppressed proinflammatory cytokine production. Combination studies with SARS-CoV-2 targeting nucleoside analogues, revealed the strongest synergistic antiviral activity with remdesivir in vitro. Comparable synergy was observed between auranofin and GS-621763, the orally bioavailable derivative of remdesivir, and was further validated in a preclinical animal model. Collectively, these findings provide a rationale for the further development of auranofin-nucleoside analog combinations targeting SARS-CoV-2. FundingThis research has been supported by ICMR (IIRPIG-2023-0000978) and BIRAC grant (BT/CS0070/06/22) to ST. We acknowledge the infrastructure and research support provided to IISc by the Crypto Relief Fund, L&T Trust, DST-FIST program, Institute of Eminence Fund, Ministry of Education, and the DBT-IISc partnership program (Phase II). RN acknowledges DBT-RA fellowship, SK acknowledges PMRF fellowship, and RS acknowledges FICCI-PMRF fellowship. Research in contextO_ST_ABSEvidence before this studyC_ST_ABSMultiple previous studies have provided evidence showing that auranofin is an antiviral agent targeting SARS-CoV-2 through a host-targeting mechanism, involving the inhibition of thioredoxin reductase and redox homeostasis. Mixed results have been reported regarding the effect of this drug on inhibiting virus-induced syncytia. A prior study from our lab demonstrated the drugs effectiveness in reducing SARS-CoV-2 viral loads in both cell and animal models. Added value of the studyOur study firmly establishes the synergistic use of Auranofin and Remdesivir GS 621763 in the treatment of SARS-CoV-2 infection.

The last century has witnessed the establishment of neoplastic disease as the second cause of death in the world. Nonetheless, the road towards desirable success rates of cancer treatments is still long and paved with uncertainty. With this work, we aim to select natural products that act via endoplasmic reticulum (ER) stress, since the latter is known to be a vulnerability of malignant cells, and display selective toxicity against cancer cell lines. Starting from a chemical library of over 90 natural products, nontoxic molecules towards non-cancer cells were selected to be assessed for their toxicity towards cancer cells, namely the human gastric adenocarcinoma cell line AGS and the lung adenocarcinoma cell line A549. The active molecules towards at least one of these cell lines were studied in a battery of ensuing assays designed to show the involvement of ER stress in the observed cytotoxic effect, for instance, by evaluating ER stress-related gene expression or caspase activation. We show that several natural products are selectively cytotoxic against malignant cell lines, with their cytotoxicity relying on ER stress and activation of the unfolded protein response (UPR). Berberine and emodin are proposed as potential leads for the development of more potent ER stressors to be used as selective anticancer agents. Berberine was effective against the two cell models we worked with, disrupting Ca2+ homeostasis, inducing UPR target gene expression and ER-resident caspase-4 activation, standing out as the most promising candidate to aid in the development of novel ER stress-based strategies against neoplastic disease.

Malaria tropica, caused by Plasmodium falciparum (P. falciparum), remains a global health challenge with limited therapeutic options. In mammalian cells, the small-molecule compound RSL3 induces ferroptosis via lipid peroxidation. In this study, we demonstrate that RSL3 synergizes with Pyrimethamine, an inhibitor of P. falciparum dihydrofolate reductase (DHFR), to suppress parasite proliferation in red blood cells (RBCs). A similar synergistic effect was observed with Cycloguanil, a structural analogue of Pyrimethamine, but not with other DHFR inhibitors or alternative ferroptosis inducers. Notably, Ferrostatin-1, an antagonist of lipid peroxidation, largely failed to rescue parasite growth in the presence of RSL3, suggesting a mechanism distinct from canonical ferroptosis. These findings suggest that the synergy may involve unidentified targets of RSL3 and Pyrimethamine in P. falciparum, divergent from those described in mammalian systems. Moreover, RSL3 and related compounds could serve as promising adjuvants to enhance the antimalarial efficacy of Pyrimethamine and potentially overcome drug resistance. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=173 HEIGHT=200 SRC="FIGDIR/small/643256v1_ufig1.gif" ALT="Figure 1"> View larger version (21K): org.highwire.dtl.DTLVardef@1282707org.highwire.dtl.DTLVardef@11a6631org.highwire.dtl.DTLVardef@2b2cf8org.highwire.dtl.DTLVardef@955b34_HPS_FORMAT_FIGEXP M_FIG C_FIG

Liver regeneration is impaired in patients suffering from alcohol-associated liver (ALD) diseases. Wnt ligands and their FZD receptors are dysregulated in diseased livers. R-spondin and their receptors are known to regulate Wnt activity via the stabilization of FZD receptors. Here, we investigated the components of the Wnt and R-Spondin-signaling pathways and their activity in patients with ALD. We found that while hepatocytes retained high levels of differentiation markers such as ASGR1 and ASGR2, the expression of two R-spondin co-receptors, LGR4 and LGR5, and of Wnt target genes, CYP1A2 and others, were strongly reduced. SZN-043, a hepatocyte-targeted R-Spondin mimetic, is a new investigational drug that stimulates the physiological Wnt repair pathway and proliferation of hepatocytes. Here, we show that SZN-043 induced hepatocyte proliferation in all models tested, including humanized mouse livers, a chronic-binge alcohol-induced liver injury, and a CCl4-induced fibrosis mouse model. Altogether, SZN-043 could be beneficial for the treatment of ALD.