We report presence of cholinergic nerve fibers in the periphery and stroma of colon cancer tissues and their correlation with poor T cell and increased macrophage infiltration. We employed hydrogel-mediated localized delivery of an FDA-approved local anesthetic, bupivacaine (BUP), to target acetylcholine (ACh)-mediated crosstalk of cholinergic neurons with cancer and immune cells. Localized BUP-Gel therapy promotes T cell-mediated tumor inhibition and enhances the antitumor response of systemic chemotherapy and immunotherapy. Further, blockade of cancer- and immune cell-specific ACh receptors inhibits tumor growth, alters the TME, and augments the impact of chemotherapy and immunotherapy. Finally, we demonstrate that ACh receptor antagonists polarize macrophages toward an M1-like phenotype and activate T cell immunity in tumor explants of patients. Therefore, targeting cholinergic signals through localized delivery of anesthetics, as well as direct immune reprogramming via cholinergic receptor antagonists, may provide a means to modulate this tripartite crosstalk, with potential implications for therapeutic strategies.

Inflammatory bowel disease (IBD), comprising Crohn’s disease and Ulcerative colitis, is a group of multifactorial illnesses with persistent gastrointestinal inflammation and a series of undesirable consequences. IBD patients have a three times higher risk of developing colitis-associated colorectal cancer (CAC). A higher mutational burden due to persistent inflammation acts as a driver of dysplasia and even tumorigenesis. While the pathological sequence is known, the molecular mechanisms underlying the transition from chronic inflammation to CAC remain largely elusive. Post-translational modification- SUMOylation plays an integral role in shaping gut inflammation as well as several forms of cancers, including sporadic colorectal cancer. In this study, the contribution of SUMOylation to CAC pathogenesis was characterized. In the AOM-DSS CAC mice model and human IBD patient specimens, SENP5, but not other deSUMOylases, shows an altered expression. Notably, both SENP5 expression dynamics and alterations in the SUMOylome occur in chronically inflamed and neoplastic colon tissues. SENP5 interactome analysis identified Coronin1A (Coro1A), an actin-binding protein predominantly expressed in immune cells. Coro1A also shows a state-specific, distinct expression pattern in the colon. Interestingly, in contrast to wild-type mice, Coro 1A knockout mice were resistant to polyp formation, with reduced cell proliferation, oncogene expression, and epithelial-to-mesenchymal transition (EMT) gene activation, and reduced extracellular matrix (ECM) development and fibrosis, suggesting its integral role in tumorigenesis. WT mice with chronically inflamed colons and polyps have a higher number of M2-like macrophages, with increased abundance of Coro1A, suggesting a role of Coro1A in modulating the tissue microenvironment toward a pro-tumorigenic state. Mechanistically, Coro1A physically interacts with TGF-β RI and regulates TGF-β-TGF-β RI signalling endosome stability, thereby controlling TGF-β-mediated macrophage polarization. Detailed in vitro experiments revealed stabilization of Coro 1A through its interaction with SUMOylated Raftlin protein. Overall, Coro 1A is necessary and sufficient for TGF-β signalling, macrophage polarization, and tumorigenesis in CAC.

Colorectal liver metastasis is one of the major causes of poor survival in colorectal cancer patients as it causes organ dysfunction and disrupts metabolic homeostasis. Apart from the primary tumor microenvironment (TME) challenges arising from cell proliferation, immunosuppression, and angiogenesis, premetastatic niches also represent an attractive therapeutic target for preventing liver metastasis. Here, we present the engineering of a unique, highly stable sub-100 nm three-drug-loaded nanomicelles (TDC NMs) system carrying the antiproliferative drug gemcitabine, the antiangiogenic drug combretastatin A4, and the anti-inflammatory drug dexamethasone. TDC NMs mitigate tumor progression in syngeneic, xenograft, orthotopic, and metastatic tumors. In-depth quantification of the changes in immune cells revealed that TDC NMs promote T-cell-mediated antitumor immunity, limit the infiltration of protumorigenic MDSCs, and enhance the antitumorigenic M1 population. TDC NMs could also reduce tumor progression, restore abdominal circumference, and normalize ascites fluid formation in orthotopic and metastatic colon cancer models. We further demonstrated that TDC NMs inhibit the formation of premetastatic niches by targeting MDSCs and macrophages, thereby achieving metabolic homeostasis. This study provides a promising therapeutic strategy for mitigating primary tumors and premetastatic niches and can therefore be explored further in colorectal cancer patients.

Gangliosides are sialic acid-enriched glycosphingolipids that play a vital role in regulating multiple signaling pathways during cancer progression. The diversity in their cell- and tissue-specific expression and dysregulations in cancer cells contributes to the unique pathophysiology of triple-negative breast cancer (TNBC). In this study, we follow up on our previously established hydrogel-mediated localized delivery of a combination of docetaxel (DTX) and carboplatin (CPT) (DTX-CPT-Gel therapy) that ensured effective tumor regression in multiple murine syngeneic and xenograft tumor models. Here, we demonstrate that DTX-CPT-Gel therapy downregulates GM3/GD3/GM1 gangliosides by targeting different ganglioside metabolic genes at the transcriptional and translational levels. DTX-CPT-Gel therapy-mediated alterations in ganglioside metabolism affect the activity of key growth factor receptor-mediated signaling pathways, including the epidermal growth factor receptor (EGFR) and cMET/hepatic growth factor receptor (HGFR), which positively impact tumor mitigation. Our work on DTX-CPT-Gel therapy, in continuum, highlights the potential of this therapy for TNBC treatment by intercepting multiple lipid-mediated signaling pathways and reinforces GD3 synthase/ST8SIA1 as a promising target for TNBC therapy.

The immunosuppressive tumour microenvironment (TME) is often regarded as the Achilles heel of cancer therapy, as it can limit immune cell infiltration and therapeutic efficacy. To address this, we engineered a long-lasting tetrapeptide-conjugated lithocholic acid-tamoxifen-derived injectable hydrogel (LTG4-Gel) that, upon implantation, does not exert any systemic toxicity in mice, rats and rabbits. LTG4-Gel retained its injectability and rheological flow properties even after the entrapment of multiple drugs, as confirmed by rheology, UV absorption, circular dichroism and atomic force microscopy. Doxorubicin (DOX) entrapped hydrogel (DOX-Gel) significantly induced the anti-tumour responses with enhanced survival in different syngeneic murine tumour models. We further showed that the engineered chimeric gel, upon entrapment of DOX and an immune agonist (c-di-GMP sodium salt (GMP)) (DOX-GMP-Gel), targeting the STING (stimulator of interferon gene) pathway, effectively mitigates tumour progression and increases survival across different tumour models. We demonstrated that DOX-GMP-Gel therapy activates the antitumour T cell immunity and generates a memory response to clear distant tumours. Our study provides a systemic design of long-lasting low molecular hydrogel to deliver the combination of immunostimulatory adjuvants and immunogenic cell death-inducing agents, offering a promising approach to modulate the tumour microenvironment for enhanced cancer therapy.

INTRODUCTION:Arjunolic acid, a well-known natural product with various medicinal properties, was isolated from the heartwood of Terminalia arjuna. Various amides of arjunolic acid were synthesized using different aryl and cyclic amines, characterized, and evaluated for their anti-cancer activities at the National Cancer Institute (NCI). METHODS:All the derivatives were active against all the cell lines of NCI compared to the parent molecule arjunolic acid. Eight compounds were selected for dose-dependent activity based on the preliminary results. IC50 of selected eight compounds was evaluated. Based on IC50 values against various cell lines, compound 2l was further investigated to understand the mechanism of action against HCT-116 and CT-26 colon cancer cell lines. RESULTS:Mechanistic studies of compound 2l in these two cell lines demonstrated that compound 2l arrested the colon cancer cells at the G0 /G1 phase. Compound 2l-treated cells were also found to have an increased percentage of ROS compared to untreated cells. It induced apoptosis in both these cell lines. CONCLUSION:Compound 2l was found to inhibit cancer growth in the mice model and was very effective against all the cancer cell lines. Therefore, it could be used for further development to treat colon cancer.

Treating wound infections caused by Gram-positive and Gram-negative bacteria, along with associated inflammation, remains challenging due to the limited therapeutic efficacy of conventional antibiotics. Leveraging the unique properties of cholic acid derivatives, such as their cationic nature, hydrophobicity, and structural adaptability, a novel aspirin-derived facile amphiphile based on cholic acid (amphiphile 1) is designed and synthesized. The investigations reveal that the amphiphile 1 functions as a bacterial membrane disruptor through interacting with key components such as lipoteichoic acid and lipopolysaccharide of bacterial membranes. It is observed that amphiphile 1 can degrade both monomicrobial and polymicrobial preformed biofilms originating from Gram-positive and Gram-negative bacteria. Animal studies demonstrate that hydrogel-based delivery of amphiphile 1 effectively mitigates bacterial infections and resolves bacterial-associated inflammation. Collectively, these findings underscore that amphiphile 1 is a putative therapeutic agent for addressing the challenges of polymicrobial infections.

Sphingolipid and ganglioside metabolic pathways are crucial components of cell signaling, having established roles in cancer cell proliferation, invasion, and migration. However, regulatory mechanisms controlling sphingolipid and ganglioside biosynthesis in mammalian cells are less known. Here, we show that RICTOR, the regulatory subunit of mTORC2, regulates the synthesis of sphingolipids and gangliosides in human luminal breast cancer-specific MCF-7 and BT-474 cells through transcriptional and epigenetic mechanisms. We observe that RICTOR regulates glucosylceramide levels by modulating the expression of UDP-Glucose Ceramide Glucosyl transferase (UGCG). We identify Zinc Finger protein X-linked (ZFX) as a RICTOR-responsive transcription factor whose recruitment to the UGCG promoter is regulated by DNA methyltransferase 1 and histone demethylase (KDM5A), which are known AKT substrates. We further demonstrate that RICTOR regulates the synthesis of GD3 gangliosides through ZFX and UGCG, and triggers the activation of the EGFR signaling pathway, thereby promoting tumor growth. In line with our findings in human cell culture and mouse models, we observe an elevated expression of RICTOR, ZFX, and UGCG in Indian luminal breast cancer tissues and in TCGA and METABRIC datasets. Together, we establish a key regulatory circuit, RICTOR-AKT-ZFX-UGCG-Ganglioside-EGFR-AKT, and elucidate its contribution to breast cancer progression.

Arjunolic acid (AA) is a pentacyclic triterpene acid with various potent biological activities. In this work, arjunolic acid was isolated from the heartwood of Terminalia arjuna, and a series of novel arjunolic acid acetals were synthesized and characterized. The anti-cancer activity of the synthesized acetals was investigated against sixty cell lines from nine different types of cancers at the National Cancer Institute (NCI). Compounds AA-2, AA-4, AA-9, and AA-18 demonstrated significant activity against colon cancer. These compounds were selected for further studies against murine colon cancer cell line CT-26. Mechanistic studies of the most active compound AA-9 on CT-26 cells revealed cell cycle arrest in the G2/M phase, which induces ROS generation in cells, leading to cell death. Additionally, compound AA-9 showed better selectivity for tumour cells and non-tumour cells. Novel acetals of arjunolic acid were synthesized and studied for their anti-cancer activity. Among them, compound AA-9 proved to be the most potent against the CT-26 colon cancer cell line with an IC50 of 2.56 mu M.

Cancer treatment is challenged due to immunosuppressive inflammatory tumour microenvironment (TME) caused by infiltration of tumour-promoting and inhibition of tumour-inhibiting immune cells. Here, we report the engineering of chimeric nanomicelles (NMs) targeting the cell proliferation using docetaxel (DTX) and inflammation using dexamethasone (DEX) that alters the immunosuppressive TME. We show that a combination of phospholipid-DTX conjugate and PEGylated-lipid-DEX conjugate can self-assemble to form sub-100 nm chimeric NMs (DTX-DEX NMs). Anti-cancer activities against syngeneic and xenograft mouse models showed that the DTX-DEX NMs are more effective in tumour regression, enhance the survival of mice over other treatment modes, and alter the tumour stroma. DTX-DEX NMs cause a significant reduction in myeloid-derived suppressor cells, alter the polarization of macrophages, and enhance the accumulation of cytotoxic CD4+ and CD8+ T cells in tumour tissues, along with alterations in cytokine expression. We further demonstrated that these DTX-DEX NMs inhibit the synthesis of prostaglandins, especially PGE2, by targeting the cyclooxygenase 2 that is partly responsible for immunosuppressive TME. Therefore, this study presents, for the first time, the engineering of lithocholic acid-derived chimeric NMs that affect the prostaglandin pathway, alter the TME, and mitigate tumour progression with enhanced mice survival.

Tumour cells secrete various proangiogenic factors like VEGF, PDGF, and EGF that result in the formation of highly vascularized tumours with an immunosuppressive tumour microenvironment. As tumour growth and metastasis...

The mechanistic target of rapamycin kinase (MTOR) is pivotal for cell growth, metabolism, and survival. It functions through two distinct complexes, mechanistic TORC1 and mechanistic TORC2 (mTORC1 and mTORC2). These complexes function in the development and progression of cancer by regulating different cellular processes, such as protein synthesis, lipid metabolism, and glucose homeostasis. The mTORC1 complex senses nutrients and initiates proliferative signals, and mTORC2 is crucial for cell survival and cytoskeletal rearrangements. mTORC1 and mTORC2 have therefore emerged as potential targets for cancer treatment. Several mTOR inhibitors, including rapamycin and its analogs (rapalogs), primarily target mTORC1 and are effective for specific cancer types. However, these inhibitors often lead to resistance and limited long-term advantages due to the activation of survival pathways through feedback mechanisms. Researchers have created next-generation inhibitors targeting mTORC1 and mTORC2 and dual PI3K/mTOR inhibitors to address these difficulties. These inhibitors demonstrate enhanced anti-tumor effects by simultaneously disrupting multiple signaling pathways and show promise for improved and long-lasting therapies. However, development of resistance and adverse side effects remain a significant obstacle. Recent additions known as RapaLinks have emerged as a boon to counter drug-resistant cancer cells, as they are more potent and provide a more comprehensive blockade of mTOR signaling pathways. This Review combines current research findings and clinical insights to enhance our understanding of the crucial role of mTOR signaling in cancer biology and highlights the evolution of mTOR inhibitors as promising therapeutic approaches.

Synthesis, characterization of pyrazole acetals of andrographolide and their in vitro anticancer activity.

Treatment of triple-negative breast cancer (TNBC) is challenging because of its “COLD” tumor immunosuppressive microenvironment (TIME). Here, we present a hydrogel-mediated localized delivery of a combination of docetaxel (DTX) and carboplatin (CPT) (called DTX-CPT-Gel therapy) that ensured enhanced anticancer effect and tumor regression on multiple murine syngeneic and xenograft tumor models. DTX-CPT-Gel therapy modulated the TIME by an increase of antitumorigenic M1 macrophages, attenuation of myeloid-derived suppressor cells, and increase of granzyme B + CD8 + T cells. DTX-CPT-Gel therapy elevated ceramide levels in tumor tissues that activated the protein kinase R (PKR)–like endoplasmic reticulum kinase (PERK)–mediated unfolded protein response (UPR). This UPR-mediated activation of apoptotic cell death led to release of damage-associated molecular patterns, thereby activating the immunogenic cell death that could even clear the metastatic tumors. This study provides a promising hydrogel-mediated platform for DTX-CPT therapy that induces tumor regression and effective immune modulation and, therefore, can be explored further for treatment of TNBC.

Emergence of vancomycin resistance in Gram-positive bacteria and the prevalence of vancomycin-resistant Enterococci (VRE) infections are highly alarming as very limited antibiotic options are available against VRE infections. Here, we present the synthesis of cholic acid-derived dimeric amphiphiles where two cholic acid moieties are tethered through carboxyl terminals using different alkylene spacers. Our investigations revealed that dimer 5 possessing a propylene spacer and glycine-valine peptides tethered on hydroxyl groups is the most effective antimicrobial against VRE. Dimer 5 can permeabilize bacterial membranes, generate reactive oxygen species, and clear preformed biofilms. We further demonstrate that dimer 5 downregulates vancomycin-mediated transcriptional activation of the vanHAX gene cluster and does not allow VSE to develop vancomycin resistance until 100 generations. Therefore, this study, for the first time, presents a bacterial membrane-targeting amphiphile that can mitigate VRE infections and inhibit the emergence of vancomycin resistance.

Psoriasis is a systemic, relapsing, and chronic autoimmune inflammatory disease of the skin. Topical use of betamethasone, a glucocorticoid, in the form of creams is a common treatment for psoriasis. However, topical use of these creams is challenging due to the ineffective entrapment of steroids, burst release of the entrapped drugs, poor skin permeability, and high toxicity. Herein, we present the engineering of a betamethasone-loaded topical hydrogel (B-Gel) that can efficiently entrap steroids with high spreadability, and can also maintain the sustained release of drugs. We used an imiquimod (IMQ) induced ear psoriasis model, and demonstrated that topical application of B-Gel can mitigate the autoimmune inflammation reactions, and leads to a reduction in erythema, induration, scaling, and ear thickness. As interleukin 17 (IL-17) secreting T helper 17 (Th17) cells and γδ+ T cells are responsible for psoriasis, B-Gel treatment witnessed a reduction in the infiltration of leukocytes, CD4+ T cells, Th17 T cells, and dermal γδ+ T cells. We further demonstrated that B-Gel mediated reduction of IL-1β, IL-17, and K16 (marker for keratinocyte proliferation) is responsible for alleviation of psoriasis. Therefore, the non-greasy nature of the hydrogel with a cooling effect provides an alternative for topical application of steroids.

Psoriasis is a systemic, relapsing, and chronic autoimmune inflammatory disease of the skin. Topical use of betamethasone, a glucocorticoid, in the form of creams is a common treatment for psoriasis. However, topical use of these creams is challenging due to the ineffective entrapment of steroids, burst release of the entrapped drugs, poor skin permeability, and high toxicity. Herein, we present the engineering of a betamethasone-loaded topical hydrogel (B-Gel) that can efficiently entrap steroids with high spreadability, and can also maintain the sustained release of drugs. We used an imiquimod (IMQ) induced ear psoriasis model, and demonstrated that topical application of B-Gel can mitigate the autoimmune inflammation reactions, and leads to a reduction in erythema, induration, scaling, and ear thickness. As interleukin 17 (IL-17) secreting T helper 17 (Th17) cells and γδ+ T cells are responsible for psoriasis, B-Gel treatment witnessed a reduction in the infiltration of leukocytes, CD4+ T cells, Th17 T cells, and dermal γδ+ T cells. We further demonstrated that B-Gel mediated reduction of IL-1β, IL-17, and K16 (marker for keratinocyte proliferation) is responsible for alleviation of psoriasis. Therefore, the non-greasy nature of the hydrogel with a cooling effect provides an alternative for topical application of steroids.

We present a non-immunogenic, injectable, low molecular weight, amphiphilic hydrogel-based drug delivery system (TB-Gel) that can entrap a cocktail of four front-line antitubercular drugs, isoniazid, rifampicin, pyrazinamide, and ethambutol. We showed that TB-Gel is more effective than oral delivery of the combination of four drugs in reducing the mycobacterial infection in mice. Results show that half the dose of chemotherapeutic drugs is sufficient to achieve a comparable therapeutic effect to that of oral delivery.

Treatment of chronic wound infections caused by Gram-positive bacteria such as Staphylococcus aureus is highly challenging due to the low efficacy of existing formulations, thereby leading to drug resistance. Herein, we present the synthesis of a nonimmunogenic cholic acid-glycine-glycine conjugate (A6) that self-assembles into a supramolecular viscoelastic hydrogel (A6 gel) suitable for topical applications. The A6 hydrogel can entrap different antibiotics with high efficacy without compromising its viscoelastic behavior. Activities against different bacterial species using a disc diffusion assay demonstrated the antimicrobial effect of the ciprofloxacin-loaded A6 hydrogel (CPF-Gel). Immune profiling and gene expression studies after the application of the A6 gel to mice confirmed its nonimmunogenic nature to host tissues. We further demonstrated that topical application of CPF-Gel clears S. aureus-mediated wound infections more effectively than clinically used formulations. Therefore, cholic acid-derived hydrogels are an efficacious matrix for topical delivery of antibiotics and should be explored further.