Leishmania exploits host cAMP/EPAC/calcineurin signaling to induce an IL-33-mediated anti-inflammatory environment for the establishment of infection.

Host anti-inflammatory responses are critical for the progression of visceral leishmaniasis, and the pleiotropic cytokine interleukin (IL)-33 was found to be upregulated in infection. Here, we documented that IL-33 induction is a consequence of elevated cAMP-mediated exchange protein activated by cAMP (EPAC)/calcineurin-dependent signaling and essential for the sustenance of infection. Leishmania donovani-infected macrophages showed upregulation of IL-33 and its neutralization resulted in decreased parasite survival and increased inflammatory responses. Infection-induced cAMP was involved in IL-33 production and of its downstream effectors PKA and EPAC, only the latter was responsible for elevated IL-33 level. EPAC initiated Rap-dependent phospholipase C activation, which triggered the release of intracellular calcium followed by calcium/calmodulin complex formation. Screening of calmodulin-dependent enzymes affirmed involvement of the phosphatase calcineurin in cAMP/EPAC/calcium/calmodulin signaling-induced IL-33 production and parasite survival. Activated calcineurin ensured nuclear localization of the transcription factors, nuclear factor of activated T cell 1 and hypoxia-inducible factor 1 alpha required for IL-33 transcription, and we further confirmed this by chromatin immunoprecipitation assay. Administering specific inhibitors of nuclear factor of activated T cell 1 and hypoxia-inducible factor 1 alpha in BALB/c mouse model of visceral leishmaniasis decreased liver and spleen parasite burden along with reduction in IL-33 level. Splenocyte supernatants of inhibitor-treated infected mice further documented an increase in tumor necrosis factor alpha and IL-12 level with simultaneous decrease of IL-10, thereby indicating an overall disease-escalating effect of IL-33. Thus, this study demonstrates that cAMP/EPAC/calcineurin signaling is crucial for the activation of IL-33 and in effect creates anti-inflammatory responses, essential for infection.

G-quadruplex structural dynamics at MAPK12 promoter dictates transcriptional switch to determine stemness in breast cancer.

P38γ (MAPK12) is predominantly expressed in triple negative breast cancer cells (TNBC) and induces stem cell (CSC) expansion resulting in decreased survival of the patients due to metastasis. Abundance of G-rich sequences at MAPK12 promoter implied the functional probability to reverse tumorigenesis, though the formation of G-Quadruplex (G4) structures at MAPK12 promoter is elusive. Here, we identified two evolutionary consensus adjacent G4 motifs upstream of the MAPK12 promoter, forming parallel G4 structures. They exist in an equilibria between G4 and duplex, regulated by the binding turnover of Sp1 and Nucleolin that bind to these G4 motifs and regulate MAPK12 transcriptional homeostasis. To underscore the gene-regulatory functions of G4 motifs, we employed CRISPR-Cas9 system to eliminate G4s from TNBC cells and synthesized a naphthalene diimide (NDI) derivative (TGS24) which shows high-affinity binding to MAPK12-G4 and inhibits MAPK12 transcription. Deletion of G4 motifs and NDI compound interfere with the recruitment of the transcription factors, inhibiting MAPK12 expression in cancer cells. The molecular basis of NDI-induced G4 transcriptional regulation was analysed by RNA-seq analyses, which revealed that MAPK12-G4 inhibits oncogenic RAS transformation and trans-activation of NANOG. MAPK12-G4 also reduces CD44High/CD24Low population in TNBC cells and downregulates internal stem cell markers, arresting the stemness properties of cancer cells.

A ratiometric small-molecule fluorescent probe for the selective detection of hypochlorite by an oxidative cyclization reaction: application to commercial disinfectants and live cells.

A highly selective thiophene-thioimidazole hydrazine-based ratiometric chemodosimeter (TPBN) was designed and synthesized to detect hypochlorite (ClO-). The probe showed yellow fluorescence and exhibited ultra sensitivity towards hypochlorite (detection limit 8.74 nM) through the oxidative intramolecular cyclization process to give a blue fluorescent triazole product (TPBN-P). Additionally, the as-designed sensor displayed a fast response (80 s) to hypochlorite with excellent selectivity over other competing analytes. DFT calculations, ESI-MS, and 1H NMR titration experiments supported the detection mechanism. The probe was a valuable and practical ratiometric sensor for test strips, commercial disinfectants, and water samples. The probe was successfully used in the bio-imaging of hypochlorite in human breast cancer cells due to its noteworthy photophysical characteristics and good cell permeability.

Highly Efficient Photoswitchable Smart Polymeric Nanovehicle for Gene and Anticancer Drug Delivery in Triple-Negative Breast Cancer.

Over the past few decades, there has been significant interest in smart drug delivery systems capable of carrying multiple drugs efficiently, particularly for treating genetic diseases such as cancer. Despite the development of various drug delivery systems, a safe and effective method for delivering both anticancer drugs and therapeutic genes for cancer therapy remains elusive. In this study, we describe the synthesis of a photoswitchable smart polymeric vehicle comprising a photoswitchable spiropyran moiety and an amino-acid-based cationic monomer-based block copolymer using reversible addition-fragmentation chain transfer (RAFT) polymerization. This system aims at diagnosing triple-negative breast cancer and subsequently delivering genes and anticancer agents. Triple-negative breast cancer patients have elevated concentrations of Cu2+ ions, making them excellent targets for diagnosis. The polymer can detect Cu2+ ions with a low limit of detection value of 9.06 nM. In vitro studies on doxorubicin drug release demonstrated sustained delivery at acidic pH level similar to the tumor environment. Furthermore, the polymer exhibited excellent blood compatibility even at the concentration as high as 500 µg/mL. Additionally, it displayed a high transfection efficiency of approximately 82 ± 5% in MDA-MB-231 triple-negative breast cancer cells at an N/P ratio of 50:1. It is observed that mitochondrial membrane depolarization and intracellular reactive oxygen species generation are responsible for apoptosis and the higher number of apoptotic cells, which occurred through the arrest of the G2/M phase of the cell cycle were observed. Therefore, the synthesized light-responsive cationic polymer may be an effective system for diagnosis, with an efficient anticancer drug and gene carrier for the treatment of triple-negative breast cancer in the future.

Long non-coding intergenic RNA, LINC00273 induces cancer metastasis and stemness via miRNA sponging in triple negative breast cancer.

LncRNAs and miRNAs, being the master regulators of gene expression, are crucial functional mediators in cancer. Our study unveils the critical regulatory role of the metastatic long non-coding RNA LINC00273 as the master regulator of oncogenes involved in cancer metastasis, stemness, and chemoresistance via its miRNA sponging mechanism. M2 (a salt of bis-Schiff base) mediated G quadruplex (G4) stabilization at the LINC00273 gene promoter remarkably inhibits LINC00273 transcription. Therefore, low-level LINC00273 transcripts are unable to efficiently sponge the miRNAs, which subsequently become available to bind and downregulate their target oncogenes. We have observed significantly different global transcriptomic scenarios in LINC00273 upregulated and downregulated circumstances in MDA-MB-231 triple-negative breast cancer model. Additionally, we have found the G4 sequence in the LINC00273 RNA to play a critical role in miRNA sequestration. miRNAs (miR-6789-5p, miR200b, miR-125b-5p, miR-4268, miR3978) have base pairing complementarity within the G4 region of LINC00273 RNA and the 3'-UTR (untranslated region) of MAPK12, TGF-ß1, and SIX-1 transcripts. We have reported TGF-ß1, SIX-1, and MAPK12 to be the direct downstream targets of LINC00273. The correlation between abnormal expression of lncRNA LINC00273 and TNBC aggressiveness strongly evidenced in our study shall accelerate the development of lncRNA-based anti-metastatic therapeutics.

The miR-26a/SIRT6/HIF-1α axis regulates glycolysis and inflammatory responses in host macrophages during Mycobacterium tuberculosis infection.

Mycobacterium tuberculosis (Mtb) is the causative agent of tuberculosis. Here, a macrophage infection model was used to unravel the role of the histone deacetylase sirtuin 6 (SIRT6) in Mtb-triggered regulation of the innate immune response. Mtb infection downregulated microRNA-26a and upregulated its target SIRT6. SIRT6 suppressed glycolysis and expression of HIF-1α-dependent glycolytic genes during infection. In addition, SIRT6 regulated the levels of intracellular succinate which controls stabilization of HIF-1α, as well as the release of interleukin (IL)-1ß. Furthermore, SIRT6 inhibited inducible nitric oxide synthase (iNOS) and proinflammatory IL-6 but augmented anti-inflammatory arginase expression. The miR-26a/SIRT6/HIF-1α axis therefore regulates glycolysis and macrophage immune responses during Mtb infection. Our findings link SIRT6 to rewiring of macrophage signaling pathways facilitating dampening of the antibacterial immune response.

Novel Sulforaphane Analog Disrupts Phosphatidylinositol-3-Kinase-Protein Kinase B Pathway and Inhibits Cancer Cell Progression via Reactive Oxygen Species-Mediated Caspase-Independent Apoptosis.

Sulforaphane, a naturally occurring isothiocyanate, has gained attention due to its tremendous anticancer potential. Thus, an array of sulforaphane analogs were synthesized and evaluated for their cytotoxic potentials on a wide range of malignant cell lines. Among these derivatives, compound 4a displayed exceptional potency in inhibiting the proliferation of cancer cell lines and a negligible effect on normal cell lines through G2/M phase arrest. The lead compound induced reactive oxygen species (ROS)-mediated mitochondrial dysfunction, leading to apoptosis. Further mechanistic studies established the interaction of the compound 4a with the insulin-like growth factor-1 receptor (IGF-R1) and blocking of the phosphatidylinositol-3-kinase (PI3K)-protein kinase B (PKB/Akt) pathway. This led to suppression of nuclear factor erythroid 2-related factor 2 (NRF-2) protein expression, thus increasing the free radicals in the tumor cells. Moreover, compound 4a induced ROS-mediated caspase-independent apoptosis. Finally, compound 4a reduced tumor progression in a 4T1 injected BALB/c syngeneic mice tumor model. In conclusion, this study summarizes the mechanism of compound 4a-mediated ROS-mediated caspase-independent apoptosis. According to the study's findings, compound 4a can be used as a powerful new anticancer agent to enhance cancer treatment.

New Tin (IV) and Organotin (IV) Complexes with a Hybrid Thiosemicarbazone/Hydrazone Ligand: Synthesis, Crystal Structure, and Antiproliferative Activity.

Nowadays, the search for new chemotherapeutic agents with low toxicity and high selectivity is a major concern. In this paper, we report the synthesis and characterization of a hybrid thiosemicarbazone/hydrazone ligand in its neutral form (L1H2) and as the chloride salt ([L1H3]Cl)-, three diorganotin (IV) complexes, and one complex with Sn (IV). The compounds have been fully characterized by IR, mass spectra, 1H, 13C, and 119Sn NMR, 119Sn CP/MAS NMR, and by single crystal X-ray diffraction. The organotin compounds have the empirical formula [SnR2L1] (R = Me, Bu, and Ph), but in the solid state, they are polymeric species with seven coordination number due to weak coordination of the pyridine nitrogen, whereas in solution, the polymeric structure is lost to afford hexacoordinate monomeric species. Reaction with SnI4 yields complex [Sn (L1)2]·EtOH, with the metal in a distorted dodecahedral arrangement. We have evaluated the antiproliferative activity of the two forms of the ligands and the four coordination compounds against MDA-MB-231, HeLa, PC3, and HepG2 cancer cell lines, and WI-38 normal cell line, and all the compounds present higher activity than cisplatin, used as the standard control. To investigate the mode of action, we have selected the most active complex, containing phenyl substituents, and used the triple negative breast cancer cell line MDA-MB-231. The results show that the complex induces apoptotic cell death promoted by generation of reactive oxygen species and by disruption of mitochondrial membrane potential.

Enhancing amyloid beta inhibition and disintegration by natural compounds: A study utilizing spectroscopy, microscopy and cell biology.

Amyloid proteins and peptides play a pivotal role in the etiology of various neurodegenerative diseases, including Alzheimer's disease (AD). Synthetically designed small molecules/ peptides/ peptidomimetics show promise towards inhibition of various kinds of amyloidosis. However, exploration of compounds isolated from natural extracts having such potential is lacking. Herein, we have investigated the repurposing of a traditional Indian medicine Lasunadya Ghrita (LG) in AD. LG is traditionally used to treat gut dysregulation and mental illnesses. Various extracts of LG were obtained, characterized, and analyzed for inhibition of Aß aggregation. Biophysical studies show that the water extract of LG (LGWE) is more potent in inhibiting Aß peptide aggregation and defibrillation of Aß40/Aß42 aggregates. NMR studies showed that LGWE binds to the central hydrophobic area and C-terminal residues of Aß40/Aß42, thereby modulating the aggregation, and reducing cell membrane damage. Additionally, LGWE rescues Aß toxicity in neuronal SH-SY5Y cells evident from decreases in ROS generation, membrane leakage, cellular apoptosis, and calcium dyshomeostasis. Notably, LGWE is non-toxic to neuronal cells and mouse models. Our study thus delves into the mechanistic insights of a repurposed drug LGWE with the potential to ameliorate Aß induced neuroinflammation.