Introduction

Withania somnifera (W. somnifera), indigenously referred to as Ashwagandha, is a medicinal plant commonly utilized in Ayurvedic medicine known for its sedative, rejuvenative, and life-prolonging properties [1]. Extracts from various parts of W. somnifera have been linked to different biological properties, such as anti-inflammatory, anti-bacterial, and anti-cancerous effects. One of the more widely studied constituents of W. somnifera is Withaferin A (WA) [2–4]. Derived from the leaves and roots of W. somnifera, WA is a steroidal lactone, containing four cycloalkane rings, in which the lactone ring consists of five carbon atoms and a single oxygen atom. Biochemical studies revealed that the anti-cancer functionality of WA is due mainly to multiple reactive sites within the compound, including hydroxyl and ketone moieties [4, 5]. This compound has shown anti-cancer efficacy in vitro by targeting cellular pathways involved in cell proliferation, growth, survival, and angiogenesis [5]. WA therefore contains promise as a non-toxic and effective adjuvant therapy for cancer patients. We sought to investigate and review the potential efficacy of WA on brain cancer, particularly adult and pediatric high-grade gliomas (HGGs).

HGGs account for around 70 % of all malignant primary brain cancers diagnosed in American adults, with an annual incidence of approximately five new cases per 100,000 people [6, 7]. Presently, the standard of care of newly diagnosed gliomas includes maximal safe resection with concurrent administration of the alkylating cytotoxic agent temozolomide (TMZ) and fractioned 60 Gy radiation therapy (RT) [8]. Despite this treatment, nearly all malignant gliomas recur. Although disease progression or recurrence has multiple specific treatment options, these tumors are associated with high morbidity and mortality rates: median survival is around 15 months in patients with glioblastoma (GBM), and 2–5 years with anaplastic astrocytoma (AA) [9, 10]. In contrast to adult HGGs, pediatric HGGs, including anaplastic astrocytoma, glioblastoma, and diffuse intrinsic pontine gliomas, comprise approximately 8–12 % of all central nervous system (CNS) tumors diagnosed in children. These tumors are also highly aggressive, difficult to treat, and result in high morbidity and mortality rates. Furthermore, current treatment modalities for these pediatric brain tumors, including RT and chemotherapy (CT), demonstrate long-term effects on development, such as cognitive dysfunction, neuroendocrine dysregulation, and developmental delays [11, 12]. The dismal medial survival time has been attributed to the existence of the multiple drug resistance capabilities of HGGs (e.g., treatment resistant cancer stem cells, up-regulation of drug resistant genes, pro-survival responses such as autophagy, etc.) [13]. Treatment focus now relies heavily on agents with low toxicity profiles to effectively penetrate the CNS [14]. There is critical need for the emergence of innovative chemotherapeutic agents in the management of adult and pediatric HGGs that can act singularly or in synergistic combinations with other treatment modalities—a need that WA can potentially fill.

Here, we provide a brief overview of treatment resistance mechanisms adopted by HGGs, and common pathways that are altered in HGGs. Additionally, we offer a comprehensive review of WA as a potential adjunct therapy by discussing cellular responses induced within tumor tissues. By including the mechanisms of action of WA in pathways often deregulated in brain cancer, we look to provide a future perspective of WA treatment against gliomas.

Characteristics of HGGs

Mechanisms of resistance to CT and RT

The failure of the current chemoradiotherapy to eliminate all glioma cells is due to the presence of multiple resistance mechanisms. It is speculated that developed resistance to current therapy leads to 90 % of HGGs recurring at the primary site [8, 9, 15].

One such resistance mechanism is the presence of glioma stem-like cells (GSLCs)—a subset of tumor cells implicated in disease recurrence. GSLCs confer a therapeutic challenge because of their self-renewal properties and preferential activation of DNA repair machinery due to insults from CT and RT [13, 16]. RT kills tumor cells primarily through DNA damage; thus, DNA damage checkpoints play a crucial role in cellular radiosensitivity. GSLCs phosphorylate the DNA repair machinery, such as the ataxia-telangiectasia-mutated (ATM), Rad17, Chk1 and Chk2 checkpoint proteins, to a higher extent than non-stem-like cells, indicating that GSLCs have the ability to induce higher rates of DNA repair post-RT [13].

Other resistance mechanisms include the up-regulation of multi-drug resistant genes (e.g., BCRP1), DNA-repair enzymes (e.g., O-6-methylguanine-DNA methyltransferase, MGMT), and antiapoptotic factors (e.g., Survivin), that increase difficulty in effective tumor management [17, 18]. MGMT works by counteracting the ability of TMZ to introduce cytotoxic methyl adducts, such as O-6-methylguanine (O-6-meG), which promote DNA strand breakage and subsequent cell death by interfering with successive cycles of DNA replication [19]. MGMT restores the structural integrity of bases with O-6-meG by transferring the added methyl group to a cysteine residue within its own active site. Naturally, its expression has been show to confer chemoresistance against TMZ in vitro and in vivo [18, 19]. Expression of Survivin, a member of the inhibitor of apoptosis (IAP) family, has been shown to correlate with treatment resistance in HGGs [20]. The importance of Survivin is illustrated in studies where Survivin disruption abolishes GSLC survival and growth [21]. Furthermore, pro-survival responses, such as autophagy, promote cell survival by allowing tumor cells to neutralize the effects of cytotoxic therapies. Lin et al. [22] showed TMZ-induced cytoprotective autophagy through a reactive oxygen species (ROS) burst and extracellular signal-regulated kinase (ERK), and, once autophagy was inhibited, malignant glioma cells underwent apoptosis. Knizhnik and colleagues [23] verified TMZ-induced autophagy is a survival mechanism, stimulating senescence rather than apoptosis.

Resistance mechanisms and deregulated pathways commonly seen in HGGs are summarized in Fig. 1.

Fig. 1
figure 1

Diagram of the various pathways deregulated in high-grade gliomas (HGG) cells, leading to overall tumor cell survival. Green arrows Upregulation of signaling pathway, red lines decreased expression or downregulation of signaling pathway (color figure online)

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Origin and characteristics of Withaferin A

Structure/anticancer properties and active sites

As previously mentioned, WA is derived from the roots and leaves of the W. somnifera plant—an erect, evergreen shrub distributed throughout India. Withanolides are a group of naturally occurring C28-steroidal lactones isolated from the roots and leaves of W. somnifera. Among them, WA is one of the most bioactive compounds, possessing anti-inflammatory, pro-apoptotic, and anti-invasive, as well as anti-angiogenic, effects. However, the chemical mechanisms by which WA accomplishes these activities are still largely unknown. Acylation or alkylation of critical macromolecules and enzymatic active sites by covalent bonds are among the several proposed explanations [24]. Chemical structural analysis of WA suggests three positions that might be involved in the alkylation reactions with nucleophilic sites. These include the C3 of the α,β-unsaturated ketone in ring A; the epoxide at position C5,6; and the C24 of the α,β-unsaturated lactone in ring E. Carbon 3 in the unsaturated A-ring has been identified by NMR spectral analysis as the main nucleophilic target site for ethyl mercaptan, thiophenol and l-cysteine ethyl ester in vitro [6].

Structure–activity relationship (SAR) studies have indicated that removal of the double bond in ring A considerably decreased the activity, indicating that the presence of the double bond in WA contributes significantly to its activity. In another example, disruption of ring A prevented the ability of WA to bind to and induce vimentin fragmentation, reversing WA’s inhibition of cancer cell invasion and apoptosis [25]. Introduction of a bulky group, such as glucose, at the C27 hydroxyl group caused a reduction in activity. This suggested that the C27 hydroxyl is important for activity. However, introduction of small groups, such as acetates, did not affect activity [2]. Inactivation of the 5(6) epoxide group with 2-mercaptoethanol resulted in the loss of activity, indicating the requirement of this group for biological activity [26]. For example, SAR studies determined that this group was important for binding and inhibiting the molecular chaperone, HSP90, resulting in HSP90 client protein degradation and pancreatic cell death [27]. Furthermore, the authors of the latter study suggested that the hydroxyl group at C4 may increase binding to HSP90 and disrupt its interaction with the key co-chaperone Cdc37. Therefore, based on SAR studies and on the chemical reactions performed, it is reasonable to conclude that biological activities of these withanolides was due mainly to an α,β-unsaturated ketone moiety in ring A, C5(6) epoxide, and hydroxyl group at C27 (Fig. 2).

Fig. 2
figure 2

Chemical structure of Withaferin A (WA) with arrows indicating pertinent reactive sites with anticancer properties. Solid arrows Essential sites, dashed arrows sites in which modification may affect biological activity. Molecular formula, C28H38O6; molecular mass, 470.6 g/mol

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Blood–brain barrier

An important limitation to current treatment for HGGs is the inability of antitumor chemotherapeutics to penetrate the blood–brain-barrier (BBB). Therefore, in order to be considered as a possible HGG treatment, WA must also be able to cross the BBB effectively. One preliminary study showed that WA treatment led to a survival advantage in a murine orthotopic GBM model, suggesting that the WA compound, or one of its active metabolites, is able to reach the brain tumor [28, 29].

Additionally, WA does not violated the Rule of Five, otherwise known as Lipinski’s Rule of Five, which assesses a compound’s properties to determine the likelihood of its efficacy as an orally absorbed drug. Lipinksi and colleagues analyzed the physicochemical properties of roughly 2000 drugs, and concluded compounds were more likely to be membrane permeable if they adhered to a set of criteria [30]. These criteria have also been applied to characterize the ability of drugs to pass through the BBB [28, 31]. WA does not display any violation of these rules, and contains a favorable blood/brain partition coefficient, which implies its efficacy in penetrating the BBB (Table 1) [32]. Additional in vivo studies are needed in mammals to determine WA’s effectiveness in crossing the BBB to truly ensure its potential for success in human trials.

Table 1 Withaferin A (WA) does not violate any condition set forth by Lipinski’s rule of five [30], which assesses the molecular likelihood of membrane permeability
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Safety considerations

The anticancerous benefits of chemotherapeutic agents must be balanced against their inherent cytotoxic properties to normal cellular tissues. Ideally, a chemotherapeutic agent targets cancer cells without affecting the integrity of surrounding tissues. WA has been shown to selectively induce cell death in cancerous cells while sparing noncancerous cells [25, 34, 35]. A study was conducted to elucidate the effects of WA on prostate adenocarcinoma cells as compared to WA effects on normal human fibroblasts. After a 2 μM treatment of WA, normal cells were found to be still viable in vitro at 96 h, whereas cancerous cells demonstrated a loss of viability at 72 h [34]. In vivo experiments also show that WA has little to no cytotoxic effects on normal cells. Thaiparambil et al. [25] discovered that, when WA was utilized to treat breast cancer in mice, histological sections of lung and liver parenchyma from WA-treated mice versus non-treated mice showed minimal necrosis at doses up to 4 mg/kg. Such experiments imply that WA spares normal tissue from the negative effects seen in cancer cells.

It important to note the dose-dependent effects of WA in human cancers remain to be determined. Further investigation is warranted to fully elucidate the effects of WA in humans; however, the compound has been shown to be safe in preliminary in vitro and in vivo studies.

Anti-HGG properties of WA

WA as an effective treatment against known resistance mechanisms

GSLCs play a huge role in tumor renewal due to their self-protective properties when targeted by CT and RT, so the importance of effectively treating this subpopulation of tumor cells cannot be understated. Studies with mice bearing human ovarian tumors have shown that WA alone can preferentially target putative cancer stem-like cells (CSLCs), causing dose-related cell death [36, 37]. Furthermore, combining WA with cisplatin resulted in a synergistic effect, reducing tumor size by 70–80 % and preventing metastasis [36]. The latter is displayed in WA’s ability to inhibit epithelial-mesenchymal transition by suppressing vimentin, a key protein in mesenchymal mobility [38]. Indicators of CT resistant GSLCs, such as aldehyde dehydrogenase 1 (ALDH1) activity, and stem cell marker gene expression (CD44, CD24 and CD 117) were significantly decreased when exposed to WA [37]. Notch1 signaling, which plays an important role in cancer stem cell self-renewal, was also inhibited upon WA treatment, resulting in inhibition of Notch1 downstream signaling genes Hes1 and Hey1. These genes normally function to prevent differentiation of quiescent cells, aiding cancer progression. Whether in monotherapy or in combination effect with cisplatin, WA treatment also resulted in an increase in ROS production and DNA damage, therefore causing cell death and apoptosis in GSLCs [37]. These results reveal the ability of WA to overcome and sensitize the protective barriers exhibited by GSLCs.

Furthermore, TMZ-resistant HGGs express MGMT, which inhibits O-6-meG adducts from inducing DNA strand breakage, thereby promoting cell survival. Current research looks at combining WA and TMZ, which shows promise by allowing resistant cells to become sensitized and undergo apoptosis [39, 40]. WA treatment acts by depleting MGMT, enabling TMZ to cause DNA damage, G2/M cell cycle arrest, and death in cancerous cells [41]. The TMZ resistant cell line T98G underwent a 43 % MGMT reduction after treatment with 5 µM WA, and completely eliminated MGMT production at 10 µM WA. This study demonstrates the inverse correlation between MGMT production and TMZ effectiveness, and also provides evidence of the ability of WA to sensitize HGG cells to treatment [40].

Pro-survival proteins, such as Survivin, are also inhibited by WA, which reduces cancer cell viability. The effect of WA on Survivin was tested in in vivo studies using hamsters with induced oral carcinogenesis as well as in human breast cancer xenografted mice [41]. After 6 h of up to 5 µM of WA exposure, human breast cancer cells displayed a decrease in Survivin, XIAP, and cIAP-2 protein levels.

WA targets deregulated pathways common in HGGs

p53 signaling pathway

The p53 pathway is one of the pathways most commonly mutated in tumorigenesis with a low survival rate [42]. It acts as a tumor suppressor, modulating cell cycle arrest and apoptosis in response to cytotoxic stress and DNA damage. In addition to its tumor-suppressing functions, it acts as a transcription factor that regulates genes involved in tumor development, proliferation, and infiltration [43]. Mutations in p53 can be found in up to 80 % of adult HGG patients and approximately 50 % of pediatric HGGs [44, 45]. Mutations to p53 affect its ability to induce pro-apoptotic pathways when genetic anomalies are detected, thereby propagating the genomic instability of glioma cells [44, 46].

Panjamurthy et al. [47] found that oral administration of WA (20 mg/kg for 14 weeks) entirely prevented 7,12-dimethylbenz(a)anthracene(DMBA)-induced oral squamous cell carcinoma in golden Syrian hamsters. Furthermore, in an immunohistochemical analysis, DMBA-painted animals stained 80 % positive for p53 mutation, while WA-treated DMBA-painted animals showed a significantly weaker positive staining for p53 mutation. While this study was performed on hamsters with oral cancers, the results highlight that WA may preserve the integrity of p53 proteins during tumorigenesis.

Furthermore, an indirect mutation of p53 results in the amplification of MDM2—an E3 ubiquitin ligase that disrupts p53 activity by transcription inhibition, binds to p53 active sites, and marks it for degradation. Around 15 % of AA and GBMs display amplification of MDM2, indicating a means for tumors to deactivate p53 other than by direct mutation [26]. Increased concentrations of MDM2 within cellular structures can exert tumorigenic phenotypes. Currently, new anticancer strategies focus on inactivating MDM2 to restore p53 function. Some therapies, such as Cisplatin administration, are dependent on regular p53 function [48]. WA is able to stabilize p53 through activating ARF, which functions to prevent MDM2 binding to p53 [49].

Additionally, cancer cells with mutated p53 proteins are often ROS-compromised; the ubiquitin proteasome system (UPS) is deregulated in the removal of oxidized proteins, allowing pro cell survival [49, 50]. In addition to targeting the UPS, investigators also studied the activation of TAp73, a tumor suppressor that causes chemosensitivity to Cisplatin or CT when p53 is lost or mutated [49]. TAp73 is also modulated through MDM2 concentrations in a relationship similar to the p53/MDM2 complex. WA-induced ROS activates JNK kinases to phosphorylate TAp73, thereby inhibiting the TAp73/MDM2 complex. The synergistic effect of WA-induced ROS production and the inhibition of the TAp73/MDM2 complex allowed TAp73 to induce apoptosis in cancer cells [49].

MAPK/ERK pathway

The MAPK/ERK pathway is a chain of proteins that participate in communication of signal molecules on cell surface receptors to nuclear DNA to promote cellular growth and proliferation or cell death; this pathway is frequently mutated in human cancer. Extracellular mitogens attach to a cell-surface receptor, causing the activation of Rat sarcoma protein (Ras). From here, activated Ras effects downstream activation of Raf, MEK, and, finally, ERK, to elicit cellular responses such as increased growth and cell mitosis [51, 52]. Deregulation of any point in the pathway can induce an increase in cellular proliferation, signal transduction, apoptosis, and tumorigenesis. HGGs up-regulate the MAPK/ERK pathway by producing a greater number of cell signal receptors, such as epidermal growth factor receptors (EGFR), which can be seen in 40–60 % of adult cases [53] Increased levels or increased activity of these receptors leads to amplified signal transduction and an enhancement in tumorigenesis.

WA has been shown to have many functions in the cascade, usually with p38MAPK, which functions to induce RNA interference against apoptosis [32, 35, 54]. In one case with in vitro leukemic cells from clinical patients before CT, WA phosphorylated p38MAPK, which phosphorylated ATF-2 and HSP27, ultimately leading to an increase in Bax, promoting the mitochondrial death cascade [35]. The results showed an increase in phosphorylation of p38MAPK of ~90.96 %with a 3.0 µM treatment of WA after 2.5 h. After 72 h of WA treatment, no viable leukemic cells of myeloid origin were observed.

PI3K/Akt/mTOR pathway

PI3K phosphorylation activates Akt, which then activates many downstream targets, mainly mTOR [55]. Additionally, up-regulation in Akt or its hyperactivation (due to overexpression of growth factor receptors like EGFR) has been seen to promote uncontrolled cell cycle progression in HGG cells and their subsequent protection from apoptosis. Numerous studies have found that WA produces an inhibitory effect on Akt and mTOR signaling [32, 56, 57]. Multiple oncogenic pathways, including MAPK, Akt, NF-κB, and mTOR often associate directly with Notch signaling. Overexpression of Notch-1 resulted in an increase of expression in Akt/mTOR signaling in colon cancer lines, promoting cell survival [56]. Reduction of Notch-1 reduced Akt/mTOR signaling. Western blot analysis revealed that treatment with 5 µM WA treatment reduced phosphorylated Akt (pAkt) as well as Notch-1 expression significantly over a period of 24 h. However, total Akt levels remained the same throughout the entire treatment. The WA inhibition of Notch-1 helped facilitate significant JNK-mediated apoptosis in colon cancer cells. After WA treatment, the cell percent viability of three colon cancer cell lines SW-480 (IC50: 3.56 μM), SW-620 (IC50: 5.0 μM) and HCT-116 (IC50: 5.33 μM) also showed a significant decrease.

Another study revealed WA (2 µM) in combination with oxaliplatin (25 µM) reduced both pAkt and total Akt levels over 24 and 48 h [58]. In non-cancerous cells, the activity of PI3K is counteracted by its inhibitor, and tumor suppressor, phosphatase and tensin homolog (PTEN) [60]. However, in more than 60 % of GBMs and other HGGs, PTEN is inactivated by genetic alterations, allowing PI3K to act uninhibited [59]. Phosphorylated PTEN levels were also reduced in combination therapy with no significant change in total PTEN levels [58].

NF-κB pathway

The NF-κB family of transcription factors plays an important role in inflammatory and immune responses. It is found in almost all animal cell types and is involved in cellular responses to various stimuli such as stress, radiation, and free radicals [61, 62]. A common regulatory step in this signal cascade is the activation of IκB kinase (IKK), which prevents NF-κB function. Activation of NF-κB dimers is due to phosphorylation of IκB by IKK, enabling the active NF-κB transcription factor subunits to translocate to the nucleus and induce target gene expression [63, 64]. Deregulation of the NF-κB pathway is involved in HGGs through the up-regulation of factors that activate NF-κB, such as tumor necrosis factor-α (TNFα) [65, 66].

An in vitro and in vivo study in mice revealed that WA prevented TNF from activating downstream signaling of IκB kinase β via a thioalkylation-sensitive redox mechanism [67]. WA inhibited TNF-induced NF-κB activation in murine fibrosarcoma and human embryonic kidney cell lines [37, 67]. Another study showed that WA increased ubiquitinated-proteins in TNF-α-treated human umbilical vein endothelial cells at global levels, which suggests that WA tends to target the ubiquitin–proteasome pathway (UPP) [50, 68].

Effects on angiogenesis

HGGs are among the most angiogenic of cancers: one of the hallmarks of transformation from low-grade glioma (LGG) to HGG is the stimulation of angiogenesis and the formation of new vessels [67]. Angiogenesis supplies the cancerous microenvironment with new vasculature from pre-existing blood vessels. This process is coordinated by an increase in pro-angiogenic gene expression, including vascular endothelial growth factor (VEGF) [68]. Due to the importance of angiogenesis in HGG progression, greater attention has been paid to anti-VEGF therapies. While enthusiasm for evaluating anti-VEGF agents has been relatively dampened by safety concerns, including the risk of intracranial hemorrhage, recent trials among malignant glioma patients treated with VEGF- or VEGF receptor (VEGFR)-targeting therapeutics plus CT report an antitumor benefit, as well as acceptable safety profiles [69].

WA shows potential for inhibiting angiogenesis by binding VEGF. One study analyzed WA’s potential for molecular docking through programs such as SwissDock and Docking Server [70]. Current anti-angiogenic therapies approved by the FDA, such as Bevacizumab, have serious to life-threatening side effects, and have little effect on brain tumor invasiveness [71–73]. After treatment with Bevacizumab, glioblastomas will adapt by upregulating glycolysis production, moving towards a more anaerobic metabolism, and inducing a microenvironmental acidosis. This adaptation leads to tumor growth, invasiveness, and unconstrained proliferation. In comparison to Bevacizumab, WA showed higher cluster formation and an overall more favorable binding and affinity to VEGF using Fullfitness [70]. An in-depth study that combined Withanone, a compound found in W. somnifera (1 mg/kg) and Withaferin A (0.5 mg/kg) (WiNA), tested both in vitro and in vivo [74]. The in vivo studies consisted of human glioblastoma, osteosarcoma, fibrosarcoma, neuroblastoma, rat glioblastoma, and mouse fibroblast cell lines. WiNA was toxic to cancer cells and nontoxic on normal human cells at the optimal ratio of 20:1. Ratios such as 5:1 and 3:1 became toxic to normal cells as well. In VEGF-stimulated HUVECs, WiNA limited migration and invasiveness. In vivo studies with nude mice subcutaneous xenograft and lung metastasis models revealed nontoxicity in mice as well as tumor and VEGF reduction in small tumors. This combination effect using multiple parts of W. somnifera may be another therapy worthy of future study in HGGs.

At low doses, WA exerts potent anti-angiogenic activity in vivo [66, 75]. Treating human umbilical vein endothelial cells (HUVEC) with WA resulted in induced cytostatic cell cycle G1 arrest. WA also inhibited vessel formation in vitro in endothelial cells and in vivo in mice-induced tumors [66, 76]. WA also has the ability to bind to vimentin and covalently modify its cysteine residue, which is present in the highly conserved α-helical coiled-coil 2B domain. This binding causes an aggregation of vimentin, ultimately leading to apoptosis [76]. WA suppressed neovascularization in wild type mice at 73 % inhibition and only marginally inhibited (29 %) neovascularization in vimentin-null mice. This revealed that inhibition of capillary growth from WA treatment relies on vimentin expression.

A summary of cellular responses induced by WA can be found in Fig. 3.

Fig. 3
figure 3

A diagram of the effects of WA on various pathways in HGG cells, leading to tumor cell death and inhibition. Green arrows Increased protein expression, red arrows decreased protein expression, red bars inhibition of downstream signaling targets (e.g., PTEN inhibits PI3K) (color figure online)

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Future perspectives

Recent laboratory investigations have begun to elucidate the multitude of effects WA exerts on cancer cells while simultaneously having minimal consequences on normal tissue. WA in various cancer types primes it to be efficacious in HGGs because many deregulated signaling pathways commonly identified in HGGs are also characteristically preserved in other cancer types [77]. Unfortunately, there is little information available on WA as a treatment against HGGs. In the literature, there are a few papers investigating the use of WA directly against HGGs (Table 2). Along with these other authors, we believe that WA can be applied to treating HGGs, and that WA therapy has the potential to yield remarkable results.

Table 2 Studies in the literature that include WA as a treatment against high-grade gliomas (HGGs)
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Interestingly, WA has been found to have promising effects in multiple other disease entities. In many cases, safety has already been shown and studies are now investigating efficacy. Table 3 summarizes these trials and each of their focuses.

Table 3 Clinical trials that include WA as a treatment/drug for various conditions
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The future of utilizing WA for HGG treatment involves determining its ability to cross the BBB in humans. WA adheres to Lipinski’s Rule of Five; therefore, pharmacokinetic study could confirm the ability of WA to be BBB permeable. Additionally, more in vivo studies are needed to allow scientists to gain additional evidence of WA safety in relevant biological contexts. Overall, the future perspectives behind the use of WA in HGGs are positive. The innocuous nature and efficacy of WA in completed clinical and pre-clinical trials make it a compound highly worthy of consideration for the treatment of pediatric HGGs. A great deal of preliminary work has emerged to solidify WA’s place as an efficient anti-cancer treatment. Its success across various tumor types justifies its use in brain cancer settings.

Conclusions

Targeting HGGs requires a novel approach in response to tumor resistance to current treatment modalities, and the multitude of deregulated signaling pathways characteristic of malignant brain cancers. Metabolites from the indigenous Indian plant W. somnifera have garnered attention from the scientific community for their novel cancer preventative and anti-tumor properties. Specifically, WA derived from the root of W. somnifera has shown its anti-cancer efficacy in humans across multiple cancer types, and its effects should warrant its further study in HGGs. WA is hypothesized to be BBB permeable, allowing for infiltration into the brain, which further strengthens its potential for use in the setting of HGGs.

WA effectively targets and prevents over-proliferation of GSLCs, while downregulating survival proteins expressed in HGGs, such as Survivin. In preclinical studies, WA was able to prevent alterations to p53 in a dose-dependent manner, as well as inactivate MDM2 through activation of ARF and ROS production. Additionally, WA promotes a mitochondrial death cascade by upregulating Bax, inhibits mutated Akt, mTOR, and NF-κB pathways, and inhibits angiogenesis in tumors. However, while its effects are promising in preclinical settings, further investigation is warranted to fully unearth the ability of WA to act against HGGs.