Enhanced extracellular vesicles mediated uttroside B (Utt-B) delivery to Hepatocellular carcinoma cell: Pharmacokinetics based on PBPK modelling.

Our prior investigation has confirmed that the anti-hepatocellular carcinoma activity of the plant saponin, specifically Uttroside B (Utt-B), derived from the leaves of Solanum nigrum Linn. This study concentrated on formulating a novel biocompatible nanocarrier utilizing Extracellular vesicles (EVs) to enhance the delivery of plant saponin into cells. The physicochemical attributes of Extracellular Vesicles/UttrosideB (EVs/Utt-B) were comprehensively characterized through techniques such as Transmission Electron Microscopy (TEM) and Fourier-transform infrared spectroscopy (FTIR). Despite the promising therapeutic potential of this uttroside B, mechanistic know-how about its entry into cells is still in its infancy. Our research sheds light on the extracellular vesicle-mediated mechanism facilitating the entry of the saponin into cells, a phenomenon confirmed through the use of by confocal microscopy. We further analysed drug-releasing kinetics and simulated the Pharmacokinetics by PBPK modelling. The simulated pharmacokinetics revealed the bioavailability of Uttroside-B in oral administration against intravenous administration.

Pharmacological attenuation of melanoma by tryptanthrin pertains to the suppression of MITF-M through MEK/ERK signaling axis.

Melanoma is the most aggressive among all types of skin cancers. The current strategies against melanoma utilize BRAFV600E, as a focal point for targeted therapy. However, therapy resistance developed in melanoma patients against the conventional anti-melanoma drugs hinders the ultimate benefits of targeted therapies. A major mechanism by which melanoma cells attain therapy resistance is via the activation of microphthalmia-associated transcription factor-M (MITF-M), the key transcription factor and oncogene aiding the survival of melanoma cells. We demonstrate that tryptanthrin (Tpn), an indole quinazoline alkaloid, which we isolated and characterized from Wrightia tinctoria, exhibits remarkable anti-tumor activity towards human melanoma through the down-regulation of MITF-M. Microarray analysis of Tpn-treated melanoma cells followed by a STRING protein association network analysis revealed that differential expression of genes in melanoma converges at MITF-M. Furthermore, in vitro and in vivo studies conducted using melanoma cells with differential MITF-M expression status, endogenously or ectopically, demonstrated that the anti-melanoma activity of Tpn is decisively contingent on its efficacy in down-regulating MITF-M expression. Tpn potentiates the degradation of MITF-M via the modulation of MEK1/2-ERK1/2-MITF-M signaling cascades. Murine models demonstrate the efficacy of Tpn in attenuating the migration and metastasis of melanoma cells, while remaining pharmacologically safe. In addition, Tpn suppresses the expression of mutated BRAFV600E and inhibits Casein Kinase 2α, a pro-survival enzyme that regulates ERK1/2 homeostasis in many tumor types, including melanoma. Together, we point to a promising anti-melanoma drug in Tpn, by virtue of its attributes to impede melanoma invasion and metastasis by attenuating MITF-M.

The emerging role of selenium metabolic pathways in cancer: New therapeutic targets for cancer.

Selenium (Se) is incorporated into the body via the selenocysteine (Sec) biosynthesis pathway, which is critical in the synthesis of selenoproteins, such as glutathione peroxidases and thioredoxin reductases. Selenoproteins, which play a key role in several biological processes, including ferroptosis, drug resistance, endoplasmic reticulum stress, and epigenetic processes, are guided by Se uptake. In this review, we critically analyze the molecular mechanisms of Se metabolism and its potential as a therapeutic target for cancer. Sec insertion sequence binding protein 2 (SECISBP2), which is a positive regulator for the expression of selenoproteins, would be a novel prognostic predictor and an alternate target for cancer. We highlight strategies that attempt to develop a novel Se metabolism-based approach to uncover a new metabolic drug target for cancer therapy. Moreover, we expect extensive clinical use of SECISBP2 as a specific biomarker in cancer therapy in the near future. Of note, scientists face additional challenges in conducting successful research, including investigations on anticancer peptides to target SECISBP2 intracellular protein.

Pyridine derivatives as anticancer lead compounds with Fatty Acid Synthase as the target: An in silico-guided in vitro study.

For the past few decades, structure-based drug discovery (SBDD) has become an inevitable technique in the drug development process for screening hit compounds against therapeutic targets. Here, we have successfully used the SBDD approach viz. virtual high-throughput screening to identify potential inhibitors against the Ketoacyl synthase (KS) domain of Fatty acid synthase (FASN). Overexpression of FASN, and subsequent enhancement of de novo lipogenesis is a key survival strategy of cancer cells. Hence, targeting lipid metabolism using FASN inhibitors has been considered as a promising method to induce metabolic stress, thereby posing a survival disadvantage to cancer cells. In the present study, we have successfully identified eight FASN inhibitors from Asinex Elite database by implementing in silico tools. Five of the hit compounds share a common ring structure, which enables characteristic binding interactions with FASN-KS. Among them, in vitro validation showed that SFA 22637550 possesses significant FASN inhibitory activity and antiproliferative effect in human cancer cells of various origins. The maximum sensitivity was exhibited towards HepG2 hepatocellular carcinoma cells (IC50 = 28 µM). The mode of cell death was found to be apoptosis with a significant increase in SubG0 population without affecting any other phases of the cell cycle. The current study puts forward an excellent core structure for the development of potent FASN inhibitors for successfully targeting cancer cell metabolism, thereby causing selective cell death.

In silico screening for identification of fatty acid synthase inhibitors and evaluation of their antiproliferative activity using human cancer cell lines.

De novo lipogenesis (DNL) by upregulation of fatty acid synthase (FASN) is an important metabolic alteration of cancer cells. FASN is over-expressed in several cancers and is often associated with a high risk of recurrence and poor prognosis. Differential expression of FASN in cancer cells and their normal counterparts leads to the impression that FASN can be an attractive druggable target in cancer therapy. Present study focuses on identification of inhibitors against FASN ketoacyl synthase (KS) domain from Asinex Biodesign compound database using in silico tools. Virtual screening resulted in the identification of two hit compounds BDD27845077 and BDD27845082 with a common core structure. Molecular Docking studies showed that BDD27845077 and BDD27845082 bind at the substrate entry channel of KS domain with GScore -12.03 kcal/mol and -12.29 kcal/mol respectively. Molecular dynamics (MD) simulation of the protein-ligand complexes shows the binding stability of ligands with FASN-KS. In vitro validation of BDD27845082 demonstrated that the compound possesses antiproliferative activity in a panel of human cancer cell lines including MDA-MB-231 (breast cancer), HCT-116 (colon cancer) and HeLa (cervical cancer) with maximum sensitivity against HCT-116 (IC 50 = 25 µM). The study put forward two lead compounds against FASN with favorable pharmacokinetic profile as indicated by virtual screening tools for the development of cancer chemotherapeutics.

Folic acid conjugated cross-linked acrylic polymer (FA-CLAP) hydrogel for site specific delivery of hydrophobic drugs to cancer cells.

BACKGROUND: The hydrogel based system is found to be rarely reported for the delivery of hydrophobic drug due to the incompatibility of hydrophilicity of the polymer network and the hydrophobicity of drug. This problem can be solved by preparing semi-interpenetrating network of cross-linked polymer for tuning the hydrophilicity so as to entrap the hydrophobic drugs. The current study is to develop a folic acid conjugated cross-linked pH sensitive, biocompatible polymeric hydrogel to achieve a site specific drug delivery. For that, we have synthesized a folic acid conjugated PEG cross-linked acrylic polymer (FA-CLAP) hydrogel and investigated its loading and release of curcumin. The formed polymer hydrogel was then conjugated with folic acid for the site specific delivery of curcumin to cancer cells and then further characterized and conducted the cell uptake and cytotoxicity studies on human cervical cancer cell lines (HeLa). RESULTS: In this study, we synthesized folic acid conjugated cross-linked acrylic hydrogel for the delivery of hydrophobic drugs to the cancer site. Poly (ethyleneglycol) (PEG) diacrylate cross-linked acrylic polymer (PAA) was prepared via inverse emulsion polymerization technique and later conjugated it with folic acid (FA-CLAP). Hydrophobic drug curcumin is entrapped into it and investigated the entrapment efficiency. Characterization of synthesized hydogel was done by using Fourier Transform-Infrared spectroscopy (FT-IR), Transmission Electron Microscopy (TEM), Differential Scanning Calorimetry (DSC). Polymerization and folate conjugation was confirmed by FT-IR spectroscopy. The release kinetics of drug from the entrapped form was studied which showed initial burst release followed by sustained release due to swelling and increased cross-linking. In vitro cytotoxicity and cell uptake studies were conducted in human cervical cancer (HeLa) cell lines. CONCLUSIONS: Results showed that curcumin entrapped folate conjugated cross-linked acrylic polymer (FA-CLAP) hydrogel showed higher cellular uptake than the non folate conjugated form. So this can be suggested as a better delivery system for site specific release of hydrophobic cancer drugs.

Phytochemicals as Potential Lead Molecules against Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is the most prevalent form of liver cancer, accounting for 85-90% of liver cancer cases and is a leading cause of cancer-related mortality worldwide. The major risk factors for HCC include hepatitis C and B viral infections, along with chronic liver diseases, such as cirrhosis, fibrosis, and non-alcoholic steatohepatitis associated with metabolic syndrome. Despite the advancements in modern medicine, there is a continuous rise in the annual global incidence rate of HCC, and it is estimated to reach >1 million cases by 2025. Emerging research in phytomedicine and chemotherapy has established the anti-cancer potential of phytochemicals, owing to their diverse biological activities. In this review, we report the major phytochemicals that have been explored in combating hepatocellular carcinoma and possess great potential to be used as an alternative or in conjunction with the existing HCC treatment modalities. An overview of the pre-clinical observations, mechanism of action and molecular targets of some of these phytochemicals is also incorporated.

A Novel Combinatorial Regimen Using Sorafenib and Uttroside B, A US FDA-designated 'Orphan Drug', for the Treatment of Hepatocellular Carcinoma.

INTRODUCTION: Sorafenib (Sor) is the first-line treatment option in clinics for treating advanced unresectable hepatocellular carcinoma (HCC). However, acquired chemoresistance and adverse side effects associated with Sor monotherapy limit its clinical benefits. We have previously reported the exceptional anti-HCC potential of uttroside B (Utt-B), a furostanol saponin isolated in our lab from Solanum nigrum Linn. leaves. The current study has evaluated the supremacy of a combinatorial regimen of Sor and Utt-B over Sor monotherapy. METHODS: MTT assay was used for In vitro cytotoxicity studies. A clonogenic assay was conducted to assess the anti-proliferative effect of the combination. Annexin V/PI staining, confocal microscopy, FACS cell cycle analysis, and Western blotting experiments were performed to validate the pro-apoptotic potential of the combination in HepG2 and Huh7 cell lines. Pharmacological safety evaluation was performed in Swiss albino mice. RESULTS: Our results indicate that Utt-B augments Sor-induced cytotoxicity in HepG2 and Huh7 cells. The combination inhibits the proliferation of liver cancer cells by inducing apoptosis through activation of the caspases 7 and 3, leading to PARP cleavage. Furthermore, the combination does not induce any acute toxicity in vivo, even at a dose five times that of the effective therapeutic dose. CONCLUSION: Our results highlight the potential of Utt-B as an effective chemosensitizer, which can augment the efficacy of Sor against HCC and circumvent Sor-induced toxic side effects. Moreover, this is the first and only report to date on the chemosensitizing potential of Utt-B and the only report that demonstrates the therapeutic efficacy and pharmacological safety of a novel combinatorial regimen involving Utt-B and Sor for combating HCC.

Curcuminoids as Anticancer Drugs: Pleiotropic Effects, Potential for Metabolic Reprogramming and Prospects for the Future.

The number of published studies on curcuminoids in cancer research, including its lead molecule curcumin and synthetic analogs, has been increasing substantially during the past two decades. Insights on the diversity of inhibitory effects they have produced on a multitude of pathways involved in carcinogenesis and tumor progression have been provided. As this wealth of data was obtained in settings of various experimental and clinical data, this review first aimed at presenting a chronology of discoveries and an update on their complex in vivo effects. Secondly, there are many interesting questions linked to their pleiotropic effects. One of them, a growing research topic, relates to their ability to modulate metabolic reprogramming. This review will also cover the use of curcuminoids as chemosensitizing molecules that can be combined with several anticancer drugs to reverse the phenomenon of multidrug resistance. Finally, current investigations in these three complementary research fields raise several important questions that will be put among the prospects for the future research related to the importance of these molecules in cancer research.

Targeting receptor tyrosine kinase signaling: Avenues in the management of cutaneous squamous cell carcinoma.

Non-melanoma skin cancer (NMSC) is the most frequently diagnosed cancer worldwide. Among the various types of NMSCs, cutaneous squamous cell carcinoma (cSCC) exhibits more aggressive phenotype and is also the second-most prevalent type. Receptor tyrosine kinases (RTK) triggers key signaling events that play critical roles in the development of various cancers including cSCC. Unsurprisingly, for this reason, this family of proteins has become the cynosure of anti-cancer drug discovery pipelines and is also being considered as attractive targets against cSCC. Though inhibition of RTKs in cSCC has yielded favourable results, there is still scope for bettering the therapeutic outcome. In this review, we discuss the relevance of RTK signaling in the progression of cutaneous squamous cell carcinoma, and observations from clinical trials that used RTK inhibitors against cSCC. Backed by results from preclinical studies, including those from our lab, we also give insights into the scope of using some natural products as effective suppressors of RTK signaling and skin carcinogenesis.

Targeting BRF2: insights from in silico screening and molecular dynamic simulations.

The Transcription factor II B (TFIIB)­related factor 2 (BRF2) containing TFIIIB complex recruits RNA polymerase III multi-subunit complex to selective gene promoters that altogether are responsible for synthesizing a variety of small non-coding RNAs, including a special type of selenocysteine tRNA (tRNASec), micro-RNA (miRNA), and other regulatory RNAs. BRF2 has been identified as a potential oncogene that promotes cancer cell survival under oxidative stress through its genetic activation. The structure of the BRF2 protein was modeled using the Robetta server, refined, and validated using the Ramachandran plot. A virtual approach utilizing molecular docking was used to screen a natural compound library to determine potential compounds that can interact with the molecular pin motif of the BRF2 protein using Maestro (Schrodinger). Subsequent molecular dynamics simulation studies of the top four ligands that exhibited low glide scores were performed using GROMACS. The findings derived from the simulations, in conjunction with the exploration of hydrogen bonding patterns, evaluation of the free energy landscape, and thorough analysis of residue decomposition, collectively converged to emphasize the robust interaction characteristics exhibited by Ligand 366 (Deacetyl lanatoside C) and ligand 336 (Neogitogenin)-with the BRF2 protein. These natural compounds may be potential inhibitors of BRF2, which could modulate the regulation of selenoprotein synthesis in cancer cells. Targeting BRF2 using these promising compounds may offer a new therapeutic approach to sensitize cancer cells to ferroptosis and apoptosis.Communicated by Ramaswamy H. Sarma.

The role of AMPK in cancer metabolism and its impact on the immunomodulation of the tumor microenvironment.

Adenosine monophosphate-activated protein kinase (AMPK) is a key metabolic sensor that is pivotal for the maintenance of cellular energy homeostasis. AMPK contributes to diverse metabolic and physiological effects besides its fundamental role in glucose and lipid metabolism. Aberrancy in AMPK signaling is one of the determining factors which lead to the development of chronic diseases such as obesity, inflammation, diabetes, and cancer. The activation of AMPK and its downstream signaling cascades orchestrate dynamic changes in the tumor cellular bioenergetics. It is well documented that AMPK possesses a suppressor role in the context of tumor development and progression by modulating the inflammatory and metabolic pathways. In addition, AMPK plays a central role in potentiating the phenotypic and functional reprogramming of various classes of immune cells which reside in the tumor microenvironment (TME). Furthermore, AMPK-mediated inflammatory responses facilitate the recruitment of certain types of immune cells to the TME, which impedes the development, progression, and metastasis of cancer. Thus, AMPK appears to play an important role in the regulation of anti-tumor immune response by regulating the metabolic plasticity of various immune cells. AMPK effectuates the metabolic modulation of anti-tumor immunity via nutrient regulation in the TME and by virtue of its molecular crosstalk with major immune checkpoints. Several studies including that from our lab emphasize on the role of AMPK in regulating the anticancer effects of several phytochemicals, which are potential anticancer drug candidates. The scope of this review encompasses the significance of the AMPK signaling in cancer metabolism and its influence on the key drivers of immune responses within the TME, with a special emphasis on the potential use of phytochemicals to target AMPK and combat cancer by modulating the tumor metabolism.

Corrigendum: Targeting thymidylate synthase enhances the chemosensitivity of triple-negative breast cancer towards 5-FU-based combinatorial therapy.

[This corrects the article DOI: 10.3389/fonc.2021.656804.].

Blockade of Uttroside B-Induced Autophagic Pro-Survival Signals Augments Its Chemotherapeutic Efficacy Against Hepatocellular Carcinoma.

Our previous study has demonstrated that Uttroside B (Utt-B), a saponin isolated from the leaves of Solanum nigrum Linn induces apoptosis in hepatic cancer cells and exhibits a remarkable growth inhibition of Hepatocellular Carcinoma (HCC). Our innovation has been granted a patent from the US (US 2019/0160088A1), Canada (3,026,426.), Japan (JP2019520425) and South Korea (KR1020190008323) and the technology have been transferred commercially to Q Biomed, a leading US-based Biotech company. Recently, the compound received approval as 'Orphan Drug' against HCC from US FDA, which reveals the clinical relevance of evaluating its antitumor efficacy against HCC. In the present study, we report that Utt-B promotes pro-survival autophagy in hepatic cancer cells as evidenced by the increased expression of autophagy-related proteins, including LC3-II, Beclin1, ATG 5, and ATG 7, as well as a rise in the autophagic flux. Hence, we investigated whether Utt-B-induced autophagic response is complementing or contradicting its apoptotic program in HCC. Inhibition of autophagy using the pharmacological inhibitors, Bafilomycin A1(Baf A1), and 3-methyl adenine (3-MA), and the biological inhibitor, Beclin1 siRNA, significantly enhances the apoptosis of hepatic cancer cells and hence the cytotoxicity induced by Utt-B. We also found increased expression of autophagy markers in Utt-B-treated xenografts derived from HCC. We further analyzed whether the antimalarial drug, Chloroquine (Cqn), a well-known autophagy inhibitor, can enhance the anticancer effect of Utt-B against HCC. We found that inhibition of autophagy using Cqn significantly enhances the antitumor efficacy of Utt-B in vitro and in vivo, in NOD SCID mice bearing HCC xenografts. Taken together, our results suggest that the antitumor effect of Utt-B against HCC can be further enhanced by blocking autophagy. Furthermore, Utt-B in combination with Cqn, a clinically approved drug, if repurposed and used in a combinatorial regimen with Utt-B, can further improve the therapeutic efficacy of Utt-B against HCC.

Corrigendum: Cucurbitacin B, purified and characterized from the rhizome of Corallocarpus epigaeus exhibits anti-melanoma potential.

[This corrects the article DOI: 10.3389/fonc.2022.903832.].

Corrigendum: Blockade of uttroside B-induced autophagic pro-survival signals augments its chemotherapeutic efficacy against hepatocellular carcinoma.

[This corrects the article DOI: 10.3389/fonc.2022.812598.].

Cucurbitacin B, Purified and Characterized From the Rhizome of Corallocarpus epigaeus Exhibits Anti-Melanoma Potential.

The ethnomedicinal plant from the Cucurbitaceae family, Corallocarpus epigaeus, or its bioactive derivatives have been widely utilized in traditional medicine owing to their distinct applications against various human ailments and have lured the interest of ethnobotanists and biochemists. Here, we report for the first time, the anti-cancer potential of a bio-active fraction isolated from the dried rhizome of C. epigaeus, and the bioactive principle identified as cucurbitacin B (Cu-B). The purification processes involving the utilization of multiple organic extracts of C. epigaeus rhizome powder, yielded Cu-B from the Ethyl acetate Cytotoxic Fraction (ECF), obtained by the chromatographic separation of the ethyl acetate extract. Amongst the various cancer lines tested, melanoma cells exhibit maximal sensitivity towards the Cu-B-containing ECF fraction. Cu-B induces an apoptotic mode of cell death initiated intrinsically as well as extrinsically in A375 melanoma cells whilst remaining comparatively less toxic to normal skin fibroblasts. In vivo studies involving a NOD-SCID murine model of human melanoma demonstrate the ability of Cu-B to attenuate tumor growth, while being pharmacologically safe in vivo, as assessed in Swiss albino mice. Furthermore, Cu-B inhibits MEK 1/2 as well as the constitutive and EGF-induced ERK 1/2 activation, indicating a definitive involvement of MAPK signal transducers in regulating Cu-B-mediated anti-melanoma activity. Together, our study demonstrates the anti-melanoma potential of C. epigaeus-derived Cu-B, which indicates the Cucurbitaceae succulent as a prospective source for deriving potent and pharmacologically safe anti-cancer compounds.

Augmented Efficacy of Uttroside B over Sorafenib in a Murine Model of Human Hepatocellular Carcinoma.

We previously reported the remarkable potency of uttroside B (Utt-B), saponin-isolated and characterized in our lab from Solanum nigrum Linn, against HCC. Recently, the U.S. FDA approved Utt-B as an 'orphan drug' against HCC. The current study validates the superior anti-HCC efficacy of Utt-B over sorafenib, the first-line treatment option against HCC. The therapeutic efficacies of Utt-B vs. sorafenib against HCC were compared in vitro, using various liver cancer cell lines and in vivo, utilizing NOD.CB17-Prkdcscid/J mice bearing human HCC xenografts. Our data indicate that Utt-B holds an augmented anti-HCC efficacy over sorafenib. Our previous report demonstrated the pharmacological safety of Utt-B in Chang Liver, the normal immortalized hepatocytes, and in the acute and chronic toxicity murine models even at elevated Utt-B concentrations. Here, we show that higher concentrations of sorafenib induce severe toxicity, in Chang Liver, as well as in acute and chronic in vivo models, indicating that, apart from the superior therapeutic benefit over sorafenib, Utt-B is a pharmacologically safer molecule, and the drug-induced undesirable effects can, thus, be substantially alleviated in the context of HCC chemotherapy. Clinical studies in HCC patients utilizing Utt-B, is a contiguous key step to promote this drug to the clinic.

Virtual screening-based identification of novel fatty acid synthase inhibitor and evaluation of its antiproliferative activity in breast cancer cells.

Cancer cells activate de novo lipogenesis by overexpressing the lipogenic enzymes ACLY, ACC and FASN to support rapid cell division. FASN, previously known as oncogenic antigen-519 (OA-519) catalyzes seven sequential reactions to synthesize palmitic acid (C16) from substrates acetyl CoA, and malonyl CoA. The dependence of cancer cells on FASN-derived lipids and the differential expression of FASN in cancer cells compared to their normal counterparts make it an attractive metabolic drug target in cancer therapy. In the present study, an attempt has been made to identify potent FASN inhibitors from Asinex-Synergy compound database using structure-based virtual screening. The serial docking protocols of increasing precisions identified LEG-17649942, with glide score -10.34 kcal/mol as a promising compound which can directly interact with active site residues H293 and H331. LEG-17649942 possesses drug-like pharmacokinetic properties as predicted by Qikprop. LEG-17649942 exhibited cytotoxicity in breast cancer cell lines SK-BR-3, MCF-7 and MDA-MB-231 with maximum activity against MDA-MB-231 cells with IC50 of 50 µM. The study put forward LEG-17649942 as a novel drug-lead compound against triple negative breast cancer with an exquisite binding pattern to FASN-KS domain.