Repurposing Drugs in Oncology (ReDO)—mebendazole as an anti-cancer agent...

[sukgansu]

Repurposing Drugs in Oncology (ReDO)—mebendazole as an anti-cancer agent

Pan Pantziarka^1,2, Gauthier Bouche¹, Lydie Meheus¹, Vidula Sukhatme³ and Vikas P. Sukhatme^3,4

¹ Anticancer Fund, Brussels, 1853 Strombeek-Bever, Belgium

²The George Pantziarka TP53 Trust, London KT1 2JP, UK

³GlobalCures, Inc, Newton, MA 02459, USA

⁴Beth Israel Deaconess Medical Centre and Harvard Medical School, Boston, MA 02215, USA

Correspondence to: Pan Pantziarka. Email: anticancer.org.uk@gmail.com

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Abstract

Mebendazole, a well-known anti-helminthic drug in wide clinical use, has anti-cancer properties that have been elucidated in a broad range of pre-clinical studies across a number of different cancer types. Significantly, there are also two case reports of anti-cancer activity in humans. The data are summarised and discussed in relation to suggested mechanisms of action. Based on the evidence presented, it is proposed that mebendazole would synergise with a range of other drugs, including existing chemotherapeutics, and that further exploration of the potential of mebendazole as an anti-cancer therapeutic is warranted. A number of possible combinations with other drugs are discussed in the Appendix.

Keywords: drug repurposing, anti-helminthic, metronomic chemotherapy, cancer, ReDO Project

Copyright: © the authors; licensee ecancermedicalscience. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0

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Introduction

Mebendazole (MBZ) is a broad-spectrum benzimidazole anti-helminthic drug, in the same class as albendazole, flubendazole, oxfendazole, and others. It is commonly prescribed to treat a range of parasitical worm infections, including threadworm, tapeworms, roundworms, and other nematode and trematode infections in humans and domestic animals. MBZ is available as a generic drug; common trade names have included Vermox (Janssen Pharamceutica) and Ovex (McNeil Products Ltd) in the US and Europe. It is generally available over the counter in European countries, but the last US manufacturer, Teva Pharmaceuticals, ceased production at the end of 2011, although the drug retains US Food and Drug Administration (FDA) approval. It is available in the US from compounding pharmacies, for example, Pavillion Compounding Pharmacy in Atlanta.

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Dosage

For human use, the most common formulation of MBZ is as 100 mg chewable tablets. The dosage varies according to the type of helminthic infection being treated. Pinworms are treated with a single 100 mg treatment, whereas roundworms or hookworms are treated with 100 mg twice a day for three days. MBZ, along with albendazole, is also used on a long-term basis for the treatment of human cystic and alveolar echinococcosis (also known as hydatid disease). According to the guidelines published by the World Health Organisation (http://whqlib-doc.who.int/bulletin/1996/Vol74-No3/bulletin_1996_74(3)_231-242.pdf1, 2

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Toxicity

MBZ has low toxicity, though patients may suffer from transient symptoms, such as abdominal pain and diarrhoea in cases of massive infection and excretion of parasites. Hypersensitivity reactions, such as rash, urticaria, and angioedema, have been observed on rare occasions. MBZ is contraindicated during pregnancy. Caution is also recommended in treating infants below the age of 2, primarily due to a lack of data in such cases [3

In the case of long-term administration of MBZ for echinococcosis, the evidence is that, in general, the treatment is well tolerated, but the specific treatment for some patients has to be discontinued. For example, in one open-labelled observational study, the patients treated with MBZ for alveolar echinococcosis (average: 24 months) experienced few adverse reactions, and in only three patients (of 17), the treatment was changed to albendazole due to intolerable side effects (reversible alopecia, psychological disturbance, and drop in performance) [4

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Pharmacokinetics

First-pass metabolism of MBZ ensures that only about 20% of the oral dose reaches systemic circulation, with maximum plasma concentration reached 2–4 hours post-administration. Dosing with a high-fat meal is known to modestly increase bioavailability [53, 66

The poor bioavailability has long been recognised, and strategies to improve this remain actively researched, these strategies have included alternative formulations with vegetable oils [7–9], altering the crystalline structure of MBZ [10] and investigations into PEGylation [11

Albendazole and MBZ interact with cimetidine, which inhibits metabolism and has been documented to increase MBZ plasma levels, (maximum serum levels rose to 82.3 ± 41.8 ng/ml [0.28 ± 0.14 μM ] from 55.7 ± 30.2 ng/ml [0.19 ± 0.10 μM], on 1.5 g of MBZ following chronic dosing of cimetidine at 400 mg three times a day for 30 days) [1213

High intra- and inter-patient variability may be an important factor in assessing response to MBZ as a possible anti-cancer therapeutic. However, it is clear that plasma levels achieved by chronic and high-dosing schedules are in the range necessary for clinical activity based on the pre-clinical evidence assessed in the following section.

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Pre-clinical evidence in cancer—in vitro and in vivo

In 2002, Mukhopadhyay and colleagues showed that MBZ induced a dose- and time-dependant apoptotic response in a range of lung cancer cell lines [14, 15in vitro results showed that MBZ inhibited lung cancer cell growth 5-fold compared to controls. Additionally, the authors confirmed the growth inhibitory effects of MBZ against breast, ovary, colon carcinomas, and osteosarcoma, producing IC50s that varied from 0.1 to 0.8 μM.

To test the in vivo response to the MBZ treatment, nu/nu mice were inoculated with subcutaneous injections of H460 non-small cell lung cancer cells [14

Further pre-clinical evidence of MBZ anti-cancer activity was shown in adrenocortical cancer in 2008 [16], both in vitro and in vivo. H295R, SW-13 and WI-38 (normal fibroblast) cells lines were treated with different concentrations of MBZ in vitro, and the two cancer cells lines showed dose-dependent growth arrest, with IC50 of 0.23 μM for H295R and 0.27 μM for SW-13 cells, with no effect on the normal fibroblast cells. Tumour spheroid inhibition was tested against a dose of 1 μM of MBZ, which completely disaggregated the tumour spheroids and killed all cancer cells in about 20 days.

In vivo treatment of athymic nude mouse models of adrenocortical cancer showed that treatment with 1 mg and 2 mg MBZ significantly inhibited tumour growth in both implanted adrenocortical cancers. While there was little difference between the response of the primary tumours to 1 mg and 2 mg doses, the latter dose inhibited the formation of metastases from 50% of controls to 75%. No side effects were noted in the treated animals. Of note, a dose of 1 mg/day in a mouse weighing 20 gm corresponds to a human dose of approximately 500 mg daily for a 70 kg person, if extrapolated by surface area.

In 2008, the in vitro activity of MBZ against chemoresistant melanoma cell lines was assessed by Doudican et al [17

Subsequently, MBZ was shown to inhibit human melanoma xenograft growth in athymic female nude mice fed 1 mg or 2 mg oral MBZ every other day [18

MBZ activity in glioblastoma multiforme (GBM) was discovered serendipitously in 2011 by investigators, who observed that GBM xenografts were failing after mice models were fed albendazole to fight a spate of pin worm infections [19in vitro and in vivo. The in vitro IC50 of MBZ was 0.24 μM in the GL261 mouse glioma line, and 0.1 μM in the 060919 human GBM. In vivo results showed that oral MBZ treatment significantly extended mean survival up to 63% in syngeneic and xenograft orthotopic mouse glioma models.

Screening of compounds for activity against colon cancer cell lines also identified MBZ as a candidate molecule in work by Nygren and colleagues [20

Diagnosis-specific activity was assessed using the NCI 60 z score data, which showed a high level of activity against leukaemia, colon cancer, CNS and melanoma panels of cell lines, with lesser activity in breast, ovarian, renal and NSCLC lines. It should be noted that the leukaemia panel had the highest level of sensitivity to MBZ, a finding that has not been further investigated to date. In the colon cancer panel, 80% of cells lines were sensitive to MBZ. Detailed in vitro treatment against five colon cancer cell lines (HCT 116, RKO, HT29, HT-8 and SW626), showed that all displayed IC50 of <5 μM, whereas the drug was largely inactive in the non-malignant cell lines.

Some work on in vitro efficacy against a chemoresistant breast cancer cell line (SKBr-3) was performed by Coyne and colleagues in 2013 [21

Finally, Schmit showed that a range of benzimidazoles, including MBZ, possess anti-neoplastic activity against the DS 17 canine osteosarcoma cell line in vitro [22

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Human data in cancer

No clinical trials of MBZ as a cancer treatment have been completed to date. However, there are two well-documented case reports in the literature in favour of re-purposing MBZ as an anti-cancer therapy.

In 2011, a case of long-term tumour control in metastatic adrenocortical cancer was published [23

A case of metastatic colon cancer treated with MBZ was described by Peter Nygren and Rolf Larsson in 2013 [2420

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Clinical trials

There are currently two clinical trials of MBZ in cancer, both for brain tumours.

One is a Phase I open label study, at John Hopkins Hospital, of MBZ in newly diagnosed high-grade glioma patients receiving temozolomide (http://clinicaltrials.gov/ct2/show/NCT01729260

The other clinical trial is at Cohen Children’s Medical Centre of New York in paediatric patients with low-grade gliomas (http://clinicaltrials.gov/ct2/show/NCT01837862

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Mechanism of action

The anti-parasitic action of MBZ is due to its action as a microtubule-disrupting agent acting to prevent the polymerisation of tubulin in the gut of helminths, causing the parasites to die [2526in vitro in a glioblastoma model [19] and in a melanoma model [1718

While there are rare reports of reversible alopecia, urticaria, rash, gastro-intestinal upset, leukopenia, and neutropenia in some patients treated with high-dose MBZ, all adverse effects associated with other microtubule disruption agents, there do not appear to be any reports of peripheral neuropathy, which is commonly considered a classic adverse effect of microtubule disrupting agents, including the taxanes and the vinca alkaloids [2728

MBZ appears to be effective through p53-dependent and independent pathways. For example, in lung cancer cell lines, it was found that MBZ treatment caused post-translational p53 stabilization and the downstream expression‎ of p21 and MDM2 [1417

There has been conflicting evidence regarding the effect that MBZ has on tumour neo-vascularisation, with some reports finding evidence that it has an anti-angiogenic effect and others finding none.

In the earliest work on the anti-cancer activity of MBZ, Mukhopadhyay and colleagues reported an anti-angiogenic effect on human lung cancer xenograft models [14in vivo analysis of adrenocortical cancer models failed to detect any anti-angiogenic activity compared to controls [16in silico study, which indicated that MBZ inhibits the action of VEGFR-2 by binding to it, a finding validated in vitro using a human umbilical vein endothelial cell (HUVEC) based angiogenesis functional assay [2930, 31

To date, the effect of MBZ or other benzimidazole on the immune response in cancer has not been investigated, though there is some evidence that albendazole synergised to stimulate the cellular immune response in mice treated for alveolar echinococcosis with the immunotherapeutic agent liposomal muramyl tripeptide phosphatidylethanolamine (L-MTP-PE) used in the treatment of osteosarcoma [3233–36

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Our take

Next steps

Based on the evidence summarised in Table 1

• melanoma,

• non-small cell lung cancer,

• adrenocortical cancer, and

• colon cancer.

Additional cancer types, which should be further investigated in animal studies include:

• breast cancer,

• leukaemia, and

• osteosarcoma.

As with other anti-cancer agents, it is most likely that MBZ will be more effective in combination with other drugs or treatment modalities. It should be noted that the first two clinical trials are using MBZ with current standard of care treatment in glioma, which in this case means a combination protocol with other drugs, principally temozolomide. Given the primary putative mechanism of action—microtubule disruption—there are a number of additional agents that warrant investigation for synergy with MBZ, some of which are listed in the Appendix.

Table 1. A summary of pre-clinical evidence by cancer type.

Other options

Finally, improved efficacy may also be possible through improvements in the bioavailability of MBZ. As touched on previously, there is evidence that the combination of MBZ with cimetidine increases plasma levels of MBZ [1237

New protocols

Adding MBZ to the existing standard of care protocols, as the first two clinical trials have done, provides an opportunity to test whether there are incremental improvements in outcomes compared to the standard of care alone. However, we should also seek opportunities to create new protocols that combine MBZ with other repurposed drugs with similar low toxicity and potential anti-cancer activity. The intention is to create novel treatment options that are multi-targeted and which present minimal risk of toxicity. Of necessity, given our current state of knowledge, the combinations proposed in the supplementary material are speculative and informed primarily by mechanistic considerations and pre-clinical data. Additional pre-clinical studies are required, but given the urgency of unmet patient needs and the low toxicity of the proposed combinations, it may be argued that small patient trials or even off-label usage may also be warranted.

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Conclusion

The evidence for an anti-cancer effect of mebendazole treatment comes from in vitro, in vivo, in silico, and human data. Mechanistically, the microtubule action is well characterised in the laboratory and provides a similar rationale to some of the major classical chemotherapeutic drug classes, such as the taxanes and vinca alkaloids. With well-established pharmacokinetics and an excellent toxicity profile, this low-cost agent is a strong candidate for drug repurposing as an oncological treatment, both in combination with the existing standard treatments and alongside other candidate repurposing agents in a number of specific cancer types. We have outlined a number of these multi-drug combinations in the hope that clinicians can act upon this information to initiate clinical trials as a matter of some urgency.

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Acknowledgments

Peter Nygren, Gregory Riggins and Gary Gallia.

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Author contributions

Primary author: Pan Pantziarka. Contributing authors: Gauthier Bouche, Lydie Meheus, Vidula Sukhatme, Vikas P. Sukhatme. All authors read and approved the final manuscript.

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Competing interests

The authors declare that they have no competing interests. All the authors are associated with not-for-profit organisations that aim to repurpose drugs for oncology treatments.

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12. Bekhti a and Pirotte J (1987) Cimetidine increases serum mebendazole concentrations. Implications for treatment of hepatic hydatid cysts Br J Clin Pharmacol 24(3) 390–2 DOI: 10.1111/j.1365-2125.1987.tb03186.x PMID: 3663452 PMCID: 1386263

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16. Martarelli D et al (2008) Mebendazole inhibits growth of human adrenocortical carcinoma cell lines implanted in nude mice Cancer Chemother Pharmacol 61(5) 809–17 DOI: 10.1007/s00280-007-0538-0

17. Doudican N et al (2008) Mebendazole induces apoptosis via Bcl-2 inactivation in chemoresistant melanoma cells Mol Cancer Res 6(8) 1308–15 DOI: 10.1158/1541-7786.MCR-07-2159 PMID: 18667591

18. Doudican NA et al (2013) XIAP downregulation accompanies mebendazole growth inhibition in melanoma xenografts Anticancer Drugs 24(2) 181–8 DOI: 10.1097/CAD.0b013e32835a43f1

19. Bai R et al (2011) Antiparasitic mebendazole shows survival benefit in 2 preclinical models of glioblastoma multiforme Neuro Oncol 13(9) 974–82 DOI: 10.1093/neuonc/nor077 PMID: 21764822 PMCID: 3158014

20. Nygren P et al (2013) Repositioning of the anthelmintic drug mebendazole for the treatment for colon cancer J Cancer Res Clin Oncol 139(12) 2133–40 DOI: 10.1007/s00432-013-1539-5 PMID: 24135855 PMCID: 3825534

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Appendix

Introduction

The following drugs warrant further investigation in combination with mebendazole (MBZ), both in pre-clinical studies and potentially in clinical trials. These combinations listed in Table A1

Higher-priority agents

The agents listed below have a high degree of clinical evidence of efficacy and are currently either in clinical use in oncology or are currently being investigated in clinical trials. They have been selected as potential agents to be used in combination with MBZ. Note that these drugs are not listed in order of priority.

• Metformin: There is pre-clinical evidence that metformin potentiates the action of existing microtubule disrupting drugs in a range of cancer types, including endometrial cancers and paediatric sarcomas [1–3

• Metronomic chemotherapy: While there is intense interest in the area of metronomic chemotherapy using taxanes or vinca alkaloids, progress has been restricted because of a lack of oral formulations of many of these drugs, with the exception of oral vinorelbine. Where existing microtubule targeting drugs without oral formulations are used in metronomic settings, it is normally as a weekly infusion in combination with daily dosing of oral cyclophosphamide or capecitabine. A number of clinical trials using oral vinorelbine have reported both low toxicity and evidence of clinical benefit in advanced cancers [4, 5

• Taxanes or Vinca Alkaloids: Combinations of microtubule targeting agents, for example, paclitaxel or docetaxel and vinorelbine, act synergistically, and there are numerous trials exploring multiple combinations of different microtubule agents [6in vitro and in vivo [7

• Albendazole or other benzimidazole: There is evidence that the different benzimidazoles vary in their molecular targets and that combining them may improve efficacy and reduce the risks of acquired resistance. While this approach has not been explored in a cancer setting, there is pre-clinical and clinical evidence that the combination of MBZ and albendazole is a more effective treatment in certain hard to treat parasitic conditions [8, 9in vitro and in vivo evidence where albendazole exerts an anti-angiogenic action by down-regulating vascular endothelial growth factor (VEGF), an effect mediated through inhibition of tumoural hypoxia inducible factor (HIF-1α) [10

• Itraconazole: The anti-fungal drug itraconazole has shown some evidence of having anti-cancer activity, possibly through an anti-angiogenic action and inhibition of Hedgehog signalling pathway [11, 12www.clinicaltrials.gov/ct2/show/NCT008874581314] and prostatic cancer [15in vitro, at doses achievable in humans [16

• Cimetidine: The H2 receptor antagonist cimetidine, primarily used to treat peptic ulcers and heartburn, has shown in vitro and in vivo anticancer activity in a range of cell types and animal models, with a number of possible methods of action [171819

• Diclofenac: The non-steroidal anti-inflammatory drug (NSAID) is a commonly used anti-inflammatory analgesic with known activity as a COX-2 inhibitor, and is available both in topical and oral form. While there is evidence that perioperative or intraoperative diclofenac may be associated with lower risk of cancer recurrence or metastases following surgery [20in vivo evidence in melanoma [21] and ovarian cancer [222324

• Chloroquine/Hydroxychloroquine: The anti-malarial drugs chloroquine and hydroxychloroquine are under active investigation in a range of clinical trials for cancer in combination with radiotherapy and/or alongside existing chemotherapy regimens. The putative mechanism of action of chloroquine is that it acts as an inhibitor of autophagy, acting therefore to restrict the ability of cancer cells to move to an autophagic state such that they move into apoptosis in response to cellular stresses initiated by chemotherapy or radiotherapy [2526

• Clarithromycin: A well-established macrolide antibiotic, clarithromycin has been used in an oncological setting for the eradication of Helicobacter Pylori infection or as a treatment for treatment-associated mycobacterial infection. It has also been used in combination therapy with lenalidomide, and dexamethasone for the treatment of multiple myeloma [27] or as a monotherapy for B cell lymphoma [28293026

Other agents

The drugs listed below may also be suitable for combination treatments with MBZ and other agents, however, the evidence is not as strong and therefore this list must be viewed as more speculative.

• 2-Methoxyestradiol (2Me): A natural metabolite of estradiol, 2Me has shown promising anti-cancer activity in a number of clinical trials and is currently being developed as a drug under the trade name of Panzem (EntreMed Inc), with trials on-going in a range of solid tumours. Proposed methods of action include anti-angiogenesis, suppression of microtubule dynamics and inhibition of proliferation [31in vitro and in vivo studies assessing the synergistic action of 2Me with other microtubule-targeting drugs, including a recent study that assessed the combination of 2Me and albendazole in a xenograft colorectal cancer model and reported a significantly improved survival time [32

• Losartan: The angiotensin II receptor antagonist losartan, used mainly to treat hypertension, is currently being investigated as a possible anticancer therapeutic, primarily for its role in counter-acting the reduced vascular perfusion caused by physical stresses within the tumour mass [3334

• Omega 3 PUFAs: There is pre-clinical evidence that omega 3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) can have chemosensitising effects in a range of cancer cell types and for a range of standard chemotherapeutic drugs [3536, 3738

Table A1. Proposed drug combinations with MBZ for specific indications.

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