Ultrastructural and immunohistochemical insights on the anti-glioma effects of a dual-drug cocktail in an in vivo experimental model.

Glioma coined as 'butterfly tumor' exhibits intense heterogeneity at the molecular and cellular levels. Although, Temozolomide exerted a long-ranging and prevailing therapeutic effect against glioma, albeit it has provided modest survival outcome. Fucoidan, (marine brown algal derivative) has demonstrated potent anti-tumor effects including glioma. Nevertheless, there is paucity of studies conducted on Fucoidan to enhance the anti-glioma efficacy of Temozolomide. The present study aimed to explore the plausible synergistic anti-glioma efficacy of Fucoidan in combination with Temozolomide in an in vivo experimental model. The dual-drug combination significantly inhibited tumor growth in in vivo and prolonged the survival rate when compared with the other treatment and tumor-control groups, via down-regulation of inflammatory cascade- IL-6/T LR4 and JAK/STAT3 as per the immunohistochemistry findings. Furthermore, the ultrastructural analysis indicated that the combinatorial treatment had restored the normal neuronal architecture of glioma-induced rats. Overall, the dual-drug cocktail might enhance the therapeutic outcome in glioma patients.

Unveiling the anti-glioma potential of a marine derivative, Fucoidan: its synergistic cytotoxicity with Temozolomide-an in vitro and in silico experimental study.

Glioma coined as a "butterfly" tumor associated with a dismal prognosis. Marine algal compounds with the richest sources of bioactive components act as significant anti-tumor therapeutics. However, there is a paucity of studies conducted on Fucoidan to enhance the anti-glioma efficacy of Temozolomide. Therefore, the present study aimed to evaluate the synergistic anti-proliferative, anti-inflammatory and pro-apoptotic effects of Fucoidan with Temozolomide in in vitro and in silico experimental setup. The anti-proliferative effects of Temozolomide and Fucoidan were evaluated on C6 glioma cells by MTT and migration assay. Modulation of inflammatory markers and apoptosis induction was affirmed at the morphological and transcriptional level by dual staining and gene expression. Molecular docking (MD) and molecular dynamics simulation (MDS) studies were performed against the targets to rationalize the inhibitory effect. The dual-drug combination significantly reduced the cell viability and migration of glioma cells in a synergistic dose-dependent manner. At the molecular level, the dual-drug combination significantly down-regulated inflammatory genes with a concomitant upregulation of pro-apoptotic marker. In consensus with our in vitro findings, molecular docking and simulation studies revealed that the anti-tumor ligands: Temozolomide, Fucoidan with 5-(3-Methy1-trizeno)-imidazole-4-carboxamide (MTIC), and 4-amino-5-imidazole-carboxamide (AIC) had the potency to bind to the inflammatory proteins at their active sites, mediated by H-bonds and other non-covalent interactions. The dual-drug combinatorial treatment synergistically inhibited the proliferation, migration of glioma cells and promoted apoptosis; conversely with the down-regulation of inflammatory genes. However, pre-clinical experimental evidence is warranted for the possible translation of this combination. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-023-03814-6.

Repurposing synthetic and natural derivatives induces apoptosis in an orthotopic glioma-induced xenograft model by modulating WNT/ß-catenin signaling.

BACKGROUND: Glioblastomas arise from multistep tumorigenesis of the glial cells. Despite the current state-of-art treatment, tumor recurrence is inevitable. Among the innovations blooming up against glioblastoma, drug repurposing could provide profound premises for treatment enhancement. While considering this strategy, the efficacy of the repurposed drugs as monotherapies were not up to par; hence, the focus has now shifted to investigate the multidrug combinations. AIM: To investigate the efficacy of a quadruple-combinatorial treatment comprising temozolomide along with chloroquine, naringenin, and phloroglucinol in an orthotopic glioma-induced xenograft model. METHODS: Antiproliferative effect of the drugs was assessed by immunostaining. The expression profiles of WNT/ß-catenin and apoptotic markers were evaluated by qRT-PCR, immunoblotting, and ELISA. Patterns of mitochondrial depolarization was determined by flow cytometry. TUNEL assay was performed to affirm apoptosis induction. In vivo drug detection study was carried out by ESI-Q-TOF MS analysis. RESULTS: The quadruple-drug treatment had significantly hampered glioma proliferation and had induced apoptosis by modulating the WNT/ß-catenin signaling. Interestingly, the induction of apoptosis was associated with mitochondrial depolarization. The quadruple-drug cocktail had breached the blood-brain barrier and was detected in the brain tissue and plasma samples. CONCLUSION: The quadruple-drug combination served as a promising adjuvant therapy to combat glioblastoma lethality in vivo and can be probed for translation from bench to bedside.

A combination of metformin and epigallocatechin gallate potentiates glioma chemotherapy in vivo.

Glioma is the most devastating high-grade tumor of the central nervous system, with dismal prognosis. Existing treatment modality does not provide substantial benefit to patients and demands novel strategies. One of the first-line treatments for glioma, temozolomide, provides marginal benefit to glioma patients. Repurposing of existing non-cancer drugs to treat oncology patients is gaining momentum in recent years. In this study, we investigated the therapeutic benefits of combining three repurposed drugs, namely, metformin (anti-diabetic) and epigallocatechin gallate (green tea-derived antioxidant) together with temozolomide in a glioma-induced xenograft rat model. Our triple-drug combination therapy significantly inhibited tumor growth in vivo and increased the survival rate (50%) of rats when compared with individual or dual treatments. Molecular and cellular analyses revealed that our triple-drug cocktail treatment inhibited glioma tumor growth in rat model through ROS-mediated inactivation of PI3K/AKT/mTOR pathway, arrest of the cell cycle at G1 phase and induction of molecular mechanisms of caspases-dependent apoptosis.In addition, the docking analysis and quantum mechanics studies performed here hypothesize that the effect of triple-drug combination could have been attributed by their difference in molecular interactions, that maybe due to varying electrostatic potential. Thus, repurposing metformin and epigallocatechin gallate and concurrent administration with temozolomide would serve as a prospective therapy in glioma patients.

Corrigendum: A combination of metformin and epigallocatechin gallate potentiates glioma chemotherapy in vivo.

[This corrects the article DOI: 10.3389/fphar.2023.1096614.].

Integration of synthetic and natural derivatives revives the therapeutic potential of temozolomide against glioma- an in vitro and in vivo perspective.

AIMS: Malignant gliomas constitute one of the deadly brain tumors with high degeneration rate. Though temozolomide (TMZ) is the first-line drug for glioma, its efficacy has decreased due to chemo-resistance. Repurposing synthetic and natural compounds have gained increasing interest in glioma. Hence, we combined chloroquine (CHL) a synthetic drug, naringenin (NAR) and phloroglucinol (PGL) (natural derivatives), to investigate whether the apoptotic effect of these drugs both alone and in combination, enhances the anti-tumor effects of TMZ in an in vitro and in vivo orthotopic xenograft glioma model. MAIN METHODS: The cytotoxic effect of the drugs was assessed in C6 (murine) glioma cells, U-87 MG and LN229 (human) glioblastoma cells, primary astrocytes (isolated from rat brain tissues) and HEK-293 T cells. Mitochondrial depolarization and alterations in the cell cycle was determined by confocal imaging and flow cytometry. The expression of angiogenic and apoptotic markers was evaluated using qRT-PCR and ELISA. The efficacy of the combinatorial treatment was assessed in an orthotopic xenograft model using U-87 MG cells. KEY FINDINGS: The combinatorial treatment inhibited cell proliferation, induced apoptosis and contributed to cell cycle arrest in glioma cells. The quadruple combinatorial cocktail down-regulated BCL-2 with a concomitant decrease in VEGF. As observed in vitro, the quadruple combinatorial treatment enhanced the median survival of glioma-induced rats with lower cellularity rate. SIGNIFICANCE: The combination of CHL, NAR and PGL synergistically potentiated the efficacy of TMZ on glioma in vitro and in vivo. Hence, this combination may characterize an advanced strategy for glioma treatment, thereby providing a possible translation to clinical trial.

Crosstalk between PI3K/AKT/mTOR and WNT/ß-Catenin signaling in GBM - Could combination therapy checkmate the collusion?

Glioblastoma multiforme is one of the calamitous primary glial brain tumors with extensive heterogeneity at cellular and molecular levels. While maximal surgical resection trailed by radio and chemotherapy employing temozolomide remains the gold-standard treatment for malignant glioma patients, the overall prognosis remains dismal and there exists an unmet need for effective therapeutic strategies. In this context, we hypothesize that proper understanding of signaling pathways responsible for glioblastoma multiforme proliferation would be the first trump card while searching for novel targeted therapies. Among the pathways aberrantly activated, PI3K/AKT/mTOR is the most significant pathway, that is clinically implicated in malignancies such as high-grade glioma. Further, the WNT/ß-Catenin cascade is well-implicated in several malignancies, while its role in regulating glioma pathogenesis has only emerged recently. Nevertheless, oncogenic activation of both these pathways is a frequent event in malignant glioma that facilitates tumor proliferation, stemness and chemo-resistance. Recently, it has been reported that the cross-talk of PI3K/AKT/mTOR pathway with multiple signaling pathways could promote glioma progression and reduce the sensitivity of glioma cells to the standard therapy. However, very few studies had focused on the relationship between PI3K/AKT/mTOR and WNT/ß-Catenin pathways in glioblastoma multiforme. Interestingly, in homeostatic and pathologic circumstances, both these pathways depict fine modulation and are connected at multiple levels by upstream and downstream effectors. Thus, gaining deep insights on the collusion between these pathways would help in discovering unique therapeutic targets for glioblastoma multiforme management. Hence, the current review aims to address, "the importance of inter-play between PI3K/AKT/mTOR and WNT/ß-Catenin pathways", and put forward, "the possibility of combinatorially targeting them", for glioblastoma multiforme treatment enhancement.

Wnt/ß-catenin Antagonists: Exploring New Avenues to Trigger Old Drugs in Alleviating Glioblastoma Multiforme.

BACKGROUND: Glioblastoma Multiforme (GBM) is one of the most heterogeneous primary brain tumors with high mortality. In spite of the current therapeutic approaches, the survival rate remains poor, with death occurring within 12 to 15 months after the preliminary diagnosis. This warrants the need for an effective treatment modality. The Wnt/ß-catenin pathway is presumably the most noteworthy pathway upregulated in almost 80% of GBM cases, contributing to tumor initiation, progression, and survival. Therefore, therapeutic strategies targeting key components of the Wnt/ß-catenin cascade using established genotoxic agents like temozolomide and pharmacological inhibitors would be an effective approach to modulate the Wnt/ß-catenin pathway. Recently, drug repurposing by means of effective combination therapy has gained importance in various solid tumors, including GBM, by targeting two or more proteins in a single pathway, thereby possessing the ability to overcome the hurdle implicated by chemoresistance in GBM. OBJECTIVE: In this context, by employing computational tools, an attempt has been made to find out the novel combinations against the Wnt/ß-catenin signalling pathway. METHODS: We have explored the binding interactions of three conventional drugs - namely temozolomide, metformin and chloroquine - along with three natural compounds, viz. epigallocatechin gallate, naringenin and phloroglucinol, on the major receptors of Wnt/ß-catenin signalling. RESULTS: It was noted that all the experimental compounds showed profound interaction with two major receptors of the Wnt/ß-catenin pathway. CONCLUSION: To the best of our knowledge, this study is the first of its kind to characterize the combined interactions of the aforementioned drugs with the Wnt/ß-catenin signalling in silico, and this will putatively open up new avenues for combination therapies in GBM treatment.

Disentangling the therapeutic tactics in GBM: From bench to bedside and beyond.

Glioblastoma multiforme (GBM) is one of the most common and malignant form of adult brain tumor with a high mortality rate and dismal prognosis. The present standard treatment comprising surgical resection followed by radiation and chemotherapy using temozolomide can broaden patient's survival to some extent. However, the advantages are not palliative due to the development of resistance to the drug and tumor recurrence following the multimodal treatment approaches due to both intra- and intertumoral heterogeneity of GBM. One of the major contributors to temozolomide resistance is O6 -methylguanine-DNA methyltransferase. Furthermore, deficiency of mismatch repair, base excision repair, and cytoprotective autophagy adds to temozolomide obstruction. Rising proof additionally showed that a small population of cells displaying certain stem cell markers, known as glioma stem cells, adds on to the resistance and tumor progression. Collectively, these findings necessitate the discovery of novel therapeutic avenues for treating glioblastoma. As of late, after understanding the pathophysiology and biology of GBM, some novel therapeutic discoveries, such as drug repurposing, targeted molecules, immunotherapies, antimitotic therapies, and microRNAs, have been developed as new potential treatments for glioblastoma. To help illustrate, "what are the mechanisms of resistance to temozolomide" and "what kind of alternative therapeutics can be suggested" with this fatal disease, a detailed history of these has been discussed in this review article, all with a hope to develop an effective treatment strategy for GBM.