Cancer as Metabolic Disease and Metformin is Anticancer Drug

metformin enhance mitochondria biosynthesis via FGF21 (in brown adipose tissue)

Diabetes, Cancer and the Drug that Fights them Both - Science in the News
http://sitn.hms.harvard.edu/flash/2017/diabetes-cancer-drug-fights/

HARVARD UNIVERSITY

How does Metformin Work?

Metformin accumulates inside the mitochondria, the little energy producing organelles in our cells. Once inside , metformin inhibits complex I of the mitochondrial electron transport chain. This in turn activates AMP-Kinase(AMPK), which then inhibits the mTOR signal pathway which reduces cancer cell proliferation.(4) Left Image Electron Microscopic Image of Mitochondria.

Metformin Repurposed Anti-Cancer Drug - Jeffrey Dach MD
https://jeffreydachmd.com/2017/07/metformin-repurposed-anti-cancer-drug/

Metabolic traits of cancer stem cells
Disease Models & Mechanisms 2018 11: dmm033464 doi:
ABSTRACT
Cancer stem cells are a subpopulation of cells within a tumour believed to confer resistance to standard cancer therapies. Although many studies have addressed the specific mechanisms of tumour recurrence driven by cancer stem cells, cellular metabolism is an often-neglected attribute. The metabolic features of cancer stem cells are still poorly understood, and they thus constitute a promising field in cancer research. The findings published so far point to a distinct metabolic phenotype in cancer stem cells, which might depend on the cancer type, the model system used or even the experimental design, and several controversies still need to be tackled. This Review describes the metabolic phenotype of cancer stem cells by addressing the main metabolic traits in different tumours, including glycolysis and oxidative, glutamine, fatty acid and amino acid metabolism. In the context of these pathways, we also mention the specific alterations in metabolic enzymes and metabolite levels that have a role in the regulation of cancer stemness. Determining the role of metabolism in supporting resistance to therapy driven by cancer stem cells can raise the opportunity for novel therapeutic targets, which might not only eliminate this resistant population, but, more importantly, eradicate the whole tumour in a relapse-free scenario.

Metabolic traits of cancer stem cells | Disease Models & Mechanisms
https://dmm.biologists.org/content/11/8/dmm033464

eLife. 2014; 3: e02242.
Published online 2014 May 13. doi: 10.7554/eLife.02242

Metformin inhibits mitochondrial complex I of cancer cells to reduce tumorigenesis

Northwerstern University

There are two postulated mechanisms by which metformin reduces tumor growth. Metformin may act at the organismal level, reducing levels of circulating insulin, a known mitogen for cancer cells. Alternatively, metformin may act in a cancer cell autonomous manner. Metformin is known to inhibit mitochondrial complex I in vitro (Ota et al., 2009; El-Mir et al., 2000; Owen et al., 2000) and it is thus possible that this targeting of the electron transport chain could inhibit tumor cell growth (Birsoy et al., 2012). This latter hypothesis has been questioned as cancer cells have the ability to survive on ATP produced exclusively by glycolysis. Furthermore, cancer cells have been shown to conduct glutamine-dependent reductive carboxylation to generate the TCA cycle intermediates required for cell proliferation when the electron transport chain is inhibited (Mullen et al., 2012; Fendt et al., 2013).

Recent epidemiological and laboratory-based studies suggest that the anti-diabetic drug metformin prevents cancer progression. How metformin diminishes tumor growth is not fully understood. In this study, we report that in human cancer cells, metformin inhibits mitochondrial complex I (NADH dehydrogenase) activity and cellular respiration. Metformin inhibited cellular proliferation in the presence of glucose, but induced cell death upon glucose deprivation, indicating that cancer cells rely exclusively on glycolysis for survival in the presence of metformin. Metformin also reduced hypoxic activation of hypoxia-inducible factor 1 (HIF-1). All of these effects of metformin were reversed when the metformin-resistant Saccharomyces cerevisiae NADH dehydrogenase NDI1 was overexpressed. In vivo, the administration of metformin to mice inhibited the growth of control human cancer cells but not those expressing NDI1. Thus, we have demonstrated that metformin's inhibitory effects on cancer progression are cancer cell autonomous and depend on its ability to inhibit mitochondrial complex I.

Metformin exists as a cation at physiological pH and thus its accumulation within mitochondria is predicted to increase as a function of the mitochondrial membrane potential. The inhibition of electron transport at complex I by metformin should reduce the mitochondrial membrane potential as proton pumping is linked to electron transport.

In summary, our results indicate that metformin reversibly inhibits mitochondrial complex I within cancer cells to reduce tumorigenesis. Metformin inhibits tumorigenesis through multiple mechanisms including the induction of cancer cell death in conditions, when glucose is limited and through inhibition of mitochondrial ROS-dependent signaling pathways that promote tumorigenesis (i.e., HIF). These results indicate that metformin would be most effective in low glucose and oxygen conditions. It will be of interest to determine whether metformin treatment might provide a useful adjunct to therapies that limit glucose uptake (e.g., PI3K inhibitors) or drive tumors to low glucose and oxygen levels (e.g., anti-angiogenic inhibitors).

Metformin inhibits mitochondrial complex I of cancer cells ...
www.ncbi.nlm.nih.gov/pmc/articles/PMC4017650/

Metformin directly acts on mitochondria to alter cellular bioenergetics

Goodman Cancer Research Centre, McGill University, 1160 Pine Ave. West, Montréal, QC, H3A 1A3, Canada
Sylvia Andrzejewski, Simon-Pierre Gravel, Michael Pollak & Julie St-Pierre
Cancer & Metabolism volume 2, Article number: 12 (2014) Cite this article

Abstract
Background
Metformin is widely used in the treatment of diabetes, and there is interest in ‘repurposing’ the drug for cancer prevention or treatment. However, the mechanism underlying the metabolic effects of metformin remains poorly understood.

Methods
We performed respirometry and stable isotope tracer analyses on cells and isolated mitochondria to investigate the impact of metformin on mitochondrial functions.

Results
We show that metformin decreases mitochondrial respiration, causing an increase in the fraction of mitochondrial respiration devoted to uncoupling reactions. Thus, cells treated with metformin become energetically inefficient, and display increased aerobic glycolysis and reduced glucose metabolism through the citric acid cycle. Conflicting prior studies proposed mitochondrial complex I or various cytosolic targets for metformin action, but we show that the compound limits respiration and citric acid cycle activity in isolated mitochondria, indicating that at least for these effects, the mitochondrion is the primary target. Finally, we demonstrate that cancer cells exposed to metformin display a greater compensatory increase in aerobic glycolysis than nontransformed cells, highlighting their metabolic vulnerability. Prevention of this compensatory metabolic event in cancer cells significantly impairs survival.

Conclusions
Together, these results demonstrate that metformin directly acts on mitochondria to limit respiration and that the sensitivity of cells to metformin is dependent on their ability to cope with energetic stress.
　

Metformin directly acts on mitochondria to alter cellular bioenergetics | Cancer & Metabolism | Full Text
https://cancerandmetabolism.biomedcentral.com/articles/10.1186/2049-3002-2-12

Hexokinase II – Major Player in the Cancer Cell

Below image schematic showing Hexokinase II attached to VDAC on mitochondrial membrane, utilizing ATP to convert glucose to G6-P. Courtesy of Mathupala, S. P., YH and Ko, and P. L. Pedersen. “Hexokinase II: cancer’s double-edged sword acting as both facilitator and gatekeeper of malignancy when bound to mitochondria.” Oncogene 25.34 (2006): 4777.(58)

hexokinase-HK-II-VDACHexokinase II, the Achilles Heel of the Cancer Cell

As mentioned in my previous article, Cancer as a Metabolic Disease, the cancer cells are rapidly proliferating in uncontrolled manner. Their metabolic pathways are massively upregulated to support the rapid proliferation. These metabolic differences can be exploited to selectively kill cancer cells, leaving normal cells unharmed. The cancer cell has a voracious appetite for glucose consumption, and accomplishes this massive glucose utilization by switching to an embryonic form of the Hexokinase enzyme called Hexokinase II, not normally present in normal cells. The enzyme Hexokinase II is the first step in conversion of glucose to glucose-6-Phosphate. (58) Production of Hexokinase II in the cancer cell is over 100 times upregulated by genetic amplification.(58) Attachment of HKII to the VDAC in the cancer cell also serves to prevent mitochondrial apoptosis. Detachment of HKII from the VDAC restores mitochondrial apoptosis pathways.(58) Below Image showing apoptosis induced by release of Hexokinase II from VDAC at outer mitochondrial membrane. Fig 3 courtesy of Mathupala, S. P., YH and Ko, and P. L. Pedersen. et al. (58)

Metformin Docks in Hexokinase Two

In 2013, Barbara Salani’s group from Italy published their in vitro lung cancer cell study, showing Metformin docks in the Hexokinase II binding site, effectively blocking its function, resulting in separation Hexokinase II from the VDAC (voltage dependent anion channel) located on the outer mitochondrial membrane.(9) Dr Salani says:

This inhibition (of Hexokinase) virtually abolishes cell glucose uptake and phosphorylation as documented by the reduced entrapment of 18F-fluorodeoxyglucose.(9)

“When HK is released from VDAC …tumor cells rapidly undergo apoptosis under a variety of stimuli which were previously ineffective in inducing apoptosis.” quote from(58)

Understanding Cancer Stems Cells

Dr Patricia Sancho in her 2015 article on Pancreatic Cancer Stem Cells explains Metformin targets pancreatic cancer stem cells (CSCs), but not their differentiated non-Cancer Stem Cells.(41) Dr. Sancho’s study demonstrates that non-CSCs are highly glycolytic, while the Cancer Stem Cells (CSCs) are dependent on oxidative metabolism (OXPHOS) with “very limited metabolic plasticity”. Thus, mitochondrial inhibition by metformin creates an energy crisis and induces cancer stem cell apoptosis.(41) Dr Sanchez states that during treatment with Metformin, “resistant Cancer Stem Cell (CSC ) clones eventually emerge with intermediate glycolytic/respiratory phenotype.”(41) This is very similar to the findings of the Lisanti group who found emergence of Doxycycline resistant cancer stem cells which had acquired a purely glycolytic phenotype..(also called the Warburg Phenotype) .(59) Below image shows effect of metformin on cancer stem cells mitochondria. Ovoid Pink structures are the mitochondria. ..courtesy of Patricia Sancho, et al. MYC_PGC1a Determines Metabolic Phenotype Pancreatic Cancer Stem Cells Patricia Sancho Cell Metabolism 2015 (41) Note Warburg Phenotype = Glycolytic Phenotype present in non-cancer stem cells. Note cancer stem cells are OX-Phos dependent unless they develop resistance.

Dr Sancho found that cancer stem cells developed resistance to Metformin (see above diagram), and she states that combining Metformin with c-MYC inhibitor overcomes this resistant phenotype.

“Alternatively, combining metformin with c-MYC inhibition, prevented or reversed, respectively, resistance to metformin by enforcing their dependence on OXPHOS, suggesting a new multimodal approach for targeting the distinct metabolic features of pancreatic CSCs.”

青蒿素降解c-MYC Artesunate Degrades c-MYC

The anti-malaria drug Artesunate is now first line treatment for severe malaria in third word countries, and is commonly infused intravenously for millions of patients with virtually no adverse effects. (See my article on Artemisinin) Artesunate is also an effective anti-cancer agent which degrades the c-MYC protein. (65-66) According to Dr Lu in his 2010 article “Dihydroartemisinin accelerates c-MYC oncoprotein degradation and induces apoptosis in c-MYC-overexpressing tumor cells.” Dr Lu found Artesunate and Dihydroartemisinin (DHA) induce significant apoptosis in cancer cell lines over-expressing c-MYC protein. Dr Lu found that DHA (and Artesunate) irreversibly down-regulated the protein level of c-MYC and accelerated degradation of c-MYC protein in the cancer cells. Dr Lu concluded that Artesunate would be useful in the treatment of c-MYC-overexpressing cancer cell types, as c-Myc could serve as biomarker candidate for prediction of antitumor efficacy of Artesunate.(65-66)

Over-Expression of c-MYC Associated with Aggressive Biology and Poor Prognosis

The c-Myc gene is a transcription factor regulating proliferation, growth, and apoptosis. Overexpression or amplification of the c-Myc protein is associated with aggressive cancer cell biology with poor prognosis. (67-70) Indeed, Dr Yi studied a series of Mantle B-Cell Lymphoma patients with c-MYC overexpression in Oncotarget 2015, stating:

“Intensive chemotherapy such as HyperCVAD/MA ± R did not improve the survival of (lymphoma patients) with a c-MYC abnormality, and a new treatment strategy should be developed.”

Dr Yi found that the highly aggressive biology of the c-MYC abnormality rendered intensive chemotherapy futile, providing brief remission with no survival benefit. The combination of an OX-Phos inhibitors (such as Metformin or Doxyxyxline) targeting cancer stem cells along with the c-Myc inhibitor, Artesunate, might represent such a new treatment strategy. We await NIH funded confirmatory studies.



Metformin Targets Cancer Stem Cells

As mentioned above. cancer stems cells utilize mitochondrial OX-PHOS (oxidative phosphorylation) for their energetic migratory and metastatic capacity.(11) Indeed, Dr Diana Whitaker-Menezes in Cell Cycle 2011, reported hyperactive oxidative mitochondrial metabolism in cancer cells was blocked by Metformin.(11) Metformin treatment serves to induce purely glycolytic phenotype in surviving cancer stem cells, now rendered sensitive to glucose starvation with a second agent such as 2DG or high dose intravenous vitamin C, creating synthetic lethality. (59)(5)(9-10)

Dr Sancho found that cancer stem cells developed resistance to Metformin (see above diagram), and she states that combining Metformin with c-MYC inhibitor overcomes this resistant phenotype.

“Alternatively, combining metformin with c-MYC inhibition, prevented or reversed, respectively, resistance to metformin by enforcing their dependence on OXPHOS, suggesting a new multimodal approach for targeting the distinct metabolic features of pancreatic CSCs.”

Metformin Repurposed Anti-Cancer Drug - Jeffrey Dach MD
https://jeffreydachmd.com/2017/07/metformin-repurposed-anti-cancer-drug/

voltage-dependent anion channel (VDAC)-bound HK-II contributes to the inhibition of apoptosis by suppressing the formation of mitochondrial permeability transition pores (mPTPs) [38]. On the other hand, numerous studies have indicated that inhibitors [e.g.

 Department of Otolaryngology, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, Zhejiang, China

Oncotarget | Warburg effect, hexokinase-II, and radioresistance of laryngeal carcinoma
http://www.oncotarget.com/index.php?journal=oncotarget&page=article&op=view&path%5B%5D=13044&path%5B%5D=41337

Metformin Repurposed Anti-Cancer Drug
Posted on July 27 2017
Metformin_500mg_Tablets_Jeffrey Dach MDMetformin Repurposed Anti-Cancer Drug

Metformin, FDA approved in 1994, is known as “the Good Anti-Diabetic Drug” , taken by 150 million people worldwide for control of blood sugar in Type Two Diabetes.(57) Remarkably, metformin is also an anti-cancer drug. In 2005, Dr Evans made the observation that Diabetic patients on Metformin have a 23% reduction in cancer.(4) Others have found a 30-50 per cent reduction in risk for cancer in metformin users.(1-2) (6) Since 2005, there has been considerable effort to elucidate the anti-cancer mechanism of metformin in both the laboratory and clinical setting.(3)(23) Left Image Metformin Courtesy of Wikimedia Commons.

How does Metformin Work?

Metformin accumulates inside the mitochondria, the little energy producing organelles in our cells. Once inside , metformin inhibits complex I of the mitochondrial electron transport chain. This in turn activates AMP-Kinase(AMPK), which then inhibits the mTOR signal pathway which reduces cancer cell proliferation.(4) Left Image Electron Microscopic Image of Mitochondria.

Hexokinase II – Major Player in the Cancer Cell

Below image schematic showing Hexokinase II attached to VDAC on mitochondrial membrane, utilizing ATP to convert glucose to G6-P. Courtesy of Mathupala, S. P., YH and Ko, and P. L. Pedersen. “Hexokinase II: cancer’s double-edged sword acting as both facilitator and gatekeeper of malignancy when bound to mitochondria.” Oncogene 25.34 (2006): 4777.(58)

hexokinase-HK-II-VDACHexokinase II, the Achilles Heel of the Cancer Cell

As mentioned in my previous article, Cancer as a Metabolic Disease, the cancer cells are rapidly proliferating in uncontrolled manner. Their metabolic pathways are massively upregulated to support the rapid proliferation. These metabolic differences can be exploited to selectively kill cancer cells, leaving normal cells unharmed. The cancer cell has a voracious appetite for glucose consumption, and accomplishes this massive glucose utilization by switching to an embryonic form of the Hexokinase enzyme called Hexokinase II, not normally present in normal cells. The enzyme Hexokinase II is the first step in conversion of glucose to glucose-6-Phosphate. (58) Production of Hexokinase II in the cancer cell is over 100 times upregulated by genetic amplification.(58) Attachment of HKII to the VDAC in the cancer cell also serves to prevent mitochondrial apoptosis. Detachment of HKII from the VDAC restores mitochondrial apoptosis pathways.(58) Below Image showing apoptosis induced by release of Hexokinase II from VDAC at outer mitochondrial membrane. Fig 3 courtesy of Mathupala, S. P., YH and Ko, and P. L. Pedersen. et al. (58)



Metformin Docks in Hexokinase Two

In 2013, Barbara Salani’s group from Italy published their in vitro lung cancer cell study, showing Metformin docks in the Hexokinase II binding site, effectively blocking its function, resulting in separation Hexokinase II from the VDAC (voltage dependent anion channel) located on the outer mitochondrial membrane.(9) Dr Salani says:

This inhibition (of Hexokinase) virtually abolishes cell glucose uptake and phosphorylation as documented by the reduced entrapment of 18F-fluorodeoxyglucose.(9)

“When HK is released from VDAC …tumor cells rapidly undergo apoptosis under a variety of stimuli which were previously ineffective in inducing apoptosis.” quote from(58)

Metformin, the Monkey Wrench

The Metformin molecule is a “Monkey Wrench” sabotaging the machinery of the cancer cell.

See this video of 3-D computer rendering of Metformin docking in Hexokinase II by the Salani Group (9). Click Here to view Video.

Left Image monkey wrench courtesy of wikimedia commons.

Understanding Cancer Stems Cells

Dr Patricia Sancho in her 2015 article on Pancreatic Cancer Stem Cells explains Metformin targets pancreatic cancer stem cells (CSCs), but not their differentiated non-Cancer Stem Cells.(41) Dr. Sancho’s study demonstrates that non-CSCs are highly glycolytic, while the Cancer Stem Cells (CSCs) are dependent on oxidative metabolism (OXPHOS) with “very limited metabolic plasticity”. Thus, mitochondrial inhibition by metformin creates an energy crisis and induces cancer stem cell apoptosis.(41) Dr Sanchez states that during treatment with Metformin, “resistant Cancer Stem Cell (CSC ) clones eventually emerge with intermediate glycolytic/respiratory phenotype.”(41) This is very similar to the findings of the Lisanti group who found emergence of Doxycycline resistant cancer stem cells which had acquired a purely glycolytic phenotype..(also called the Warburg Phenotype) .(59) Below image shows effect of metformin on cancer stem cells mitochondria. Ovoid Pink structures are the mitochondria. ..courtesy of Patricia Sancho, et al. MYC_PGC1a Determines Metabolic Phenotype Pancreatic Cancer Stem Cells Patricia Sancho Cell Metabolism 2015 (41) Note Warburg Phenotype = Glycolytic Phenotype present in non-cancer stem cells. Note cancer stem cells are OX-Phos dependent unless they develop resistance.

Metformin Metabolic-Phenotype-and-Plasticity-of-Pancreatic-Cancer-Stem-CellsDr Sancho found that cancer stem cells developed resistance to Metformin (see above diagram), and she states that combining Metformin with c-MYC inhibitor overcomes this resistant phenotype.

“Alternatively, combining metformin with c-MYC inhibition, prevented or reversed, respectively, resistance to metformin by enforcing their dependence on OXPHOS, suggesting a new multimodal approach for targeting the distinct metabolic features of pancreatic CSCs.”

Artesunate Degrades c-MYC

The anti-malaria drug Artesunate is now first line treatment for severe malaria in third word countries, and is commonly infused intravenously for millions of patients with virtually no adverse effects. (See my article on Artemisinin) Artesunate is also an effective anti-cancer agent which degrades the c-MYC protein. (65-66) According to Dr Lu in his 2010 article “Dihydroartemisinin accelerates c-MYC oncoprotein degradation and induces apoptosis in c-MYC-overexpressing tumor cells.” Dr Lu found Artesunate and Dihydroartemisinin (DHA) induce significant apoptosis in cancer cell lines over-expressing c-MYC protein. Dr Lu found that DHA (and Artesunate) irreversibly down-regulated the protein level of c-MYC and accelerated degradation of c-MYC protein in the cancer cells. Dr Lu concluded that Artesunate would be useful in the treatment of c-MYC-overexpressing cancer cell types, as c-Myc could serve as biomarker candidate for prediction of antitumor efficacy of Artesunate.(65-66)

Over-Expression of c-MYC Associated with Aggressive Biology and Poor Prognosis

The c-Myc gene is a transcription factor regulating proliferation, growth, and apoptosis. Overexpression or amplification of the c-Myc protein is associated with aggressive cancer cell biology with poor prognosis. (67-70) Indeed, Dr Yi studied a series of Mantle B-Cell Lymphoma patients with c-MYC overexpression in Oncotarget 2015, stating:

“Intensive chemotherapy such as HyperCVAD/MA ± R did not improve the survival of (lymphoma patients) with a c-MYC abnormality, and a new treatment strategy should be developed.”

Dr Yi found that the highly aggressive biology of the c-MYC abnormality rendered intensive chemotherapy futile, providing brief remission with no survival benefit. The combination of an OX-Phos inhibitors (such as Metformin or Doxyxyxline) targeting cancer stem cells along with the c-Myc inhibitor, Artesunate, might represent such a new treatment strategy. We await NIH funded confirmatory studies.



Metformin Targets Cancer Stem Cells

As mentioned above. cancer stems cells utilize mitochondrial OX-PHOS (oxidative phosphorylation) for their energetic migratory and metastatic capacity.(11) Indeed, Dr Diana Whitaker-Menezes in Cell Cycle 2011, reported hyperactive oxidative mitochondrial metabolism in cancer cells was blocked by Metformin.(11) Metformin treatment serves to induce purely glycolytic phenotype in surviving cancer stem cells, now rendered sensitive to glucose starvation with a second agent such as 2DG or high dose intravenous vitamin C, creating synthetic lethality. (59)(5)(9-10)

Synthetic Lethality with Glucose Starvation

My previous article discussed the combination of Doxycycline with High Dose Vitamin C as reported by the Lisanti Group’s work from Italy.(59) Dr Lisanti’s group showed that converting cancer stem cells to a purely glycolytic phenotype using repeated passages through higher doses of Doxycycline renders the cancer stem cells sensitive to synthetic lethality with a second metabolic inhibitor. One such second metabolic inhibitor is high dose IV vitamin C (Ascorbate), which serves as a potent glycolysis inhibitor, 10 times more potent than 2-DG (2-deoxy-glucose).(38-39)(59)

Similarly, by blocking mitochondrial oxidative phophorylation, Metformin converts cancer stem cells to a purely glycolytic phenotype. Since mechanisms differ, one might speculate a more robust result with combined use of both Doxycycline and Metformin. Doxycycline impairs ribosomal protein production in the mitochondria while, as mentioned above, metformin blocks the Hexokinase II enzyme, and impairs Complex One in the electron transport chain (E.T.C.) in the mitochondria.(58) Indeed, a clinical trial of the Doxycycline/ Metformin combination is underway.(22)

Metformin Inhibits Progression of B Cell Lymphocytic Leukemia

In her 2015 article in Oncotarget, Dr Silvia Bruno, Silvia reports that Metformin inhibits cell cycle progression of B-cell chronic lymphocytic leukemia cells.” (6) She reports that Metformin slowed the proliferation rate of the cancer cells, as measured by the Ki-67 iindex:

“the fraction of Ki-67 positive cells was significantly lower in metformin-treated CLL (cancer) cells than in untreated controls, in a dose-dependent way.”(6)

In addition, the stimulated cancer cells had a 10 fold increase in glucose uptake compared to quiescent cancer cells. This rise in glucose uptake was remarkably inhibited by metformin.(6)

Metformin Inhibits B Cell Lymphoma

AMPK (AMP Kinase) activity is completely lost in lymphoma cells. (8) Dr W.Y. Shi reported in Cell Death 2012 that metformin restores AMPK activity and blocks lymphoma cell growth via inhibition of the mTOR pathway.(8) Metformin remarkably blocked tumor growth in murine lymphoma xenografts at a concentration of 10 mM.(8)

Metformin Down-Regulates Inflammatory Cytokines, Enhances Immune System, Inhibits Angiogenesis

Metformin down regulates inflammatory cytokines used for cancer growth and signalling.(16) In addition, Metformin inhibits cancer cell induced angiogenesis. (44-45) Moreover, Metformin has a beneficial effect on the immune system by enhancing Killer T Cell anti-cancer activity. Dr. Kim reports in 2014 (16):

Metformin has been shown to decrease the production of inflammatory cytokines, including TNF-a, interleukin-6, and vascular endothelial growth factor (VEGF), through the inactivation of NF-KB and HIF-1a …. metformin treatment inhibits neoplastic angiogenesis, resulting in the reduction of tumor growth.(16)

Metformin Degrades Cyclin D1

Over-expression of the cell cycle regulator Cyclin D1 is a frequent feature in cancer, and predicts early metastatic spread with poor prognosis.(19) Dr Gwak reports in 2017 that Metformin degrades Cyclin D1 in ovarian cancer cell model irrespective of p53 status.(19) This is done via metformin’s ability to upregulate the AMPK/GSK3ß signaling axis . (19)

Metformin inhibits WNT pathway

In 2016 Dr Kamal reported that Metformin inhibits the WNT pathway in cancer cells at commonly used doses.(20) This is indirect inhibition via the AMPK – MTOR signalling pathway. Downstream mediators of the WNT pathway are Cyclin D1 and C-Myc.(20)

Metformin Activates Immune Response to Cancer

Dr Chae reports in 2016 , “metformin activates the T cell mediated immune response against cancer cells.” (23) In a 2015 report, Dr Eikawa’s group studied the Immune-mediated antitumor effect of metformin using a mouse xenograft model.(24) The authors state:

Metformin increased the number of CD8(+) tumor-infiltrating lymphocytes (TILs) and protected them from apoptosis and exhaustion characterized by decreased production of IL-2, TNFa, and IFN. CD8(+) TILs (tumor infiltrating lymphocytes) capable of producing multiple cytokines were mainly PD-1(-)Tim-3(+), an Effector Memory T Cell subset responsible for tumor rejection. ” (24)

Metformin for BRCA Gene Carriers

Metformin has been suggested for prevention and treatment of BRCA gene carriers.(27)

Combination of Metformin with Propranolol (Beta Blocker)

The Beta-Blocker, Propranolol has been re purposed as an anti-cancer drug. Mode of action is both directly on cancer cell metabolism as well as cancer micro-environment, disrupting catecholammine cancer signalling. (36)(60-64) The combination of metformin and propranolol has been found synergistic in Triple Negative breast cancer cell lines studied in vitro.(36)(48)

Combined with Chemotherapy or Hyperthermia

Metformin was found synergistic with conventional chemotherapy providing better results than chemo alone. This was thought to be due to metformin’s ability to target cancer stem cells. (13) Hyperthermia, or use of a sauna, increased Metformin cytotoxicity against cancer stem cells.(14)(29)

Conclusion: The evidence for Metformin as anti-cancer drug is overwhelming. It is best used in combination with other agents such as Artesunate, Doxycycline, and IV vitamin C to create Synthetic Lethality and overcome resistant cell types. There is an urgent need for NIH funding for studies confirming this combination approach to eradicating cancer stem cells.

Jeffrey Dach MD
7450 Griffin Road
Suite 190
Davie, Fl 33314
954 792-4663

Links to Articles with Related Interest:

Artemisinin Anti-Cancer Gift from China

IV Vitamin C as Cancer Chemotherapy

Doxycycline IV Vitamin C Anticancer Combination

Metformin the Anti-Aging Miracle Drug

Cancer as a Metabolic Disease

This article is part two. For part one, click here.

Links and References

METFORMIN as Anticancer Drug

1) Evans, Josie MM, et al. “Metformin and reduced risk of cancer in diabetic patients.” Bmj 330.7503 (2005): 1304-1305.

2) Kasznicki, Jacek, Agnieszka Sliwinska, and Józef Drzewoski. “Metformin in cancer prevention and therapy.” Annals of translational medicine 2.6 (2014). numerous meta-analyses that confirmed that metformin reduces cancer incidence by 30-50%.

3) Sacco, Francesca, et al. “The cell-autonomous mechanisms underlying the activity of metformin as an anticancer drug.” British journal of cancer 115.12 (2016): 1451.

4) Chen, Chuan-Mu, et al. “Repurposing Metformin for Lung Cancer Management.” A Global Scientific Vision-Prevention, Diagnosis, and Treatment of Lung Cancer. InTech, 2017.
In this article, we introduced the background knowledge of lung cancer management and considered repurposing old drugs to overcome therapy bottleneck. We chose metformin to prove both its antihyperglycemia and antitumor formation effects. Based on the metformin-related AMPK-dependent pathway, we tried to explore the AMPK-independent pathway in inhibition of lung tumorigenesis by metformin.

Initially, Evans et al. [2] observed that patients with type 2 diabetes mellitus (DM) under metformin treatment had a reduction of cancer incidence. It caused a 23% reduction of risk of any cancer for the metformin group.

Metformin can accumulate within the matrix of mitochondria, and it could exert the inhibition of the complex I of the mitochondrial electron transport chain.

Metformin can activate AMPK to initiate the downstream signal transduction to affect the transcription of tumor suppressor liver kinase B1 (LKB1) [14]. When metformin-related AMPK dependent pathway is affected, the inhibition of mTOR signal transduction and reduction of cancer cell proliferation are achieved [

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5) Menendez, Javier A., et al. “Metformin is synthetically lethal with glucose withdrawal in cancer cells.” Cell cycle 11.15 (2012): 2782-2792.

we recently hypothesized that stress-energy mimickers such as the AMPK agonist metformin should produce metabolic
synthetic lethality in a glucose-starved cell culture milieu imitating the adverse tumor growth conditions in vivo.

representative cell models of breast cancer heterogeneity underwent massive apoptosis (by > 90% in some cases) when glucose-starved cell cultures were supplemented with metformin.

the preferential killing of cancer stem cells (CSC) by metformin may simply expose the best-case scenario for its synthetically lethal activity because an increased dependency on Warburg-like aerobic glycolysis (hyperglycolytic phenotype) is critical to sustain CSC stemness and immortality;

6) Bruno, Silvia, et al. “Metformin inhibits cell cycle progression of B-cell chronic lymphocytic leukemia cells.” Oncotarget 6.26 (2015): 22624.

Recent studies have provided evidence that diabetic patients receiving metformin have a reduced risk of developing cancer and decreased cancer mortality [13, 14].

metformin reduces tumor growth not only indirectly (systemic effect: glucose and insulin lowering) but also by direct inhibition of energetic metabolism [18] and inhibition of pathways involved in cell proliferation [18–20], through both AMPK-dependent [21, 22] and -independent mechanisms [23–27].

the fraction of Ki-67 positive cells was significantly lower in metformin-treated CLL cells than in untreated controls, in a dose-dependent way (Figure2B).

Flow cytometric single-cell data of 2-NBDG fluorescence indicated that the average uptake of 2-NBDG after 48 hours CD40L-stimulation was almost ten fold the uptake of 2-NBDG in quiescent CLL cells (Figure ?(Figure5D).5D). The presence of metformin during CLL cell activation remarkably inhibited this rise (Figure ?(Figure5D5D).

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7) Gu, Juan J., et al. “Up-regulation of hexokinase II (HK) alters the glucose metabolism and disrupts the mitochondrial potential in aggressive b-cell lymphoma contributing to rituximab-chemotherapy resistance and is a clinically relevant target for future therapeutic development.” (2014): 1767-1767.

8) Shi, W. Y., et al. “Therapeutic metformin/AMPK activation blocked lymphoma cell growth via inhibition of mTOR pathway and induction of autophagy.” Cell death & disease 3.3 (2012): e275.
In vivo, metformin induced AMPK activation, mTOR inhibition and remarkably blocked tumor growth in murine lymphoma xenografts. Of note, metformin was equally effective when given orally.

As shown in Figure 1, the AMPK activity was completely lost in lymphoma cells. Consistent with the downregulation of AMPK expression, increased phosphorylation of mTOR, p70S6K and 4EBP1 were present in 77.3%, 66.7% and 69.7% of B-lymphoma cases

In primary lymphoma cells, metformin resulted in significant growth inhibition from the concentration of 10mM (Figure 2d). However, proliferation of CD34+ cells isolated from human cord blood, a population relatively enriched in hematopoietic progenitor cells, was not affected even at the concentrations up to 120?mM, suggesting that metformin exerted no major cytotoxic effect on normal hematopoietic precursors (Figure 2e).

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HEXOKINASE II

very important!! Use figure 2—shows molecular binding of Metformin into hexokinase 2

LUNG CANCER CELL MODEL

9) Salani, Barbara, et al. “Metformin impairs glucose consumption and survival in Calu-1 cells by direct inhibition of hexokinase-II.” Scientific reports 3 (2013).
The anti-hyperglycaemic drug metformin has important anticancer properties as shown by the direct inhibition of cancer cells proliferation. Tumor cells avidly use glucose as a source for energy production and cell building blocks. Critical to this phenotype is the production of glucose-6-phosphate (G6P), catalysed by hexokinases (HK) I and II, whose role in glucose retention and metabolism is highly advantageous for cell survival and proliferation. Here we show that metformin impairs the enzymatic function of HKI and II in Calu-1 cells. This inhibition virtually abolishes cell glucose uptake and phosphorylation as documented by the reduced entrapment of 18F-fluorodeoxyglucose.

In-silico models indicate that this action is due to metformin capability to mimic G6P features by steadily binding its pocket in HKII. The impairment of this energy source results in mitochondrial depolarization and subsequent cell death. These results could represent a starting point to open effective strategies in cancer prevention and treatment.

One of the primary metabolic changes observed in malignant transformation is an increased catabolic glucose metabolism characterized by high rates of anaerobic glycolysis regardless of oxygen concentration1. Critical to this phenotype is glucose cellular entrapment by its conversion to glucose-6-phosphate (G6P). In normal tissues, this basic process is regulated by four different hexokinase (HK) isoforms indicating that regulation of glucose phosphorylation can vary in different tissues under different condition2. In cancer cells, this reaction is mainly catalysed by HK II whose glucose affinity and mitochondrial localization are highly advantageous for cancer survival and growth3. Inhibition of HKII enzymatic activity and its mitochondrial localization, are associated with cancer cells death4,5.

Metformin effect on glucose metabolism in cancer cells

Metformin effect on cancer metabolism was evaluated by estimating Calu-1 cells capability to retain FDG. Metformin treatment decreased tracer uptake in a dose and time dependent manner up to its virtual abolition after 24 hours exposure to 10 mM drug concentration (32.7 ± 1.0% in controls vs 3.1 ± 0.4% in treated cells, p < 0.0001)

Figure 2
Molecular mechanism of HK II inhibition by metformin.
metformin is thus prefigured as an uncompetitive (Figure S1F) and allosteric inhibitor of HK II as only the enzyme-substrate complex can be bound.
reduced FDG uptake reflects a selective metformin induced impairment of glucose phosphorylation.

Figure 3. Metformin displaces HK II from Mitochondria.

In conclusion the key finding of the present study is that metformin inhibits HK II in Calu-1 cells through an allosteric modification of its molecular structure blocking the synthesis of G6P. Moreover, our results demonstrate that HK II inhibition by metformin causes release of this enzyme from the outer membrane of mitochondria, thus leading to the activation of apoptotic signals.

BREAST CANCER CELL MODEL (SAME GROUP)

10) Marini, Cecilia, et al. “Direct inhibition of hexokinase activity by metformin at least partially impairs glucose metabolism and tumor growth in experimental breast cancer.” Cell cycle 12.22 (2013): 3490-3499.

Recently, we demonstrated that metformin impairs cancer energy asset in vitro via a direct and selective enzymatic inhibition of HK isoforms I and II.19

Metformin strikingly impaired glucose consumption of MDA-MB-231 in a dose- and time-dependent manner. Maximal effect occurred with exposure to 10 mM drug concentration that progressively reduced FDG uptake down to its minimum values after 48 h (Fig. 1A).

11) Whitaker-Menezes, Diana, et al. “Hyperactivation of oxidative mitochondrial metabolism in epithelial cancer cells in situ: visualizing the therapeutic effects of metformin in tumor tissue.” Cell cycle 10.23 (2011): 4047-4064.

Similar results were obtained with NADH activity staining, which measures Complex I activity, and succinate dehydrogenase (SDH) activity staining, which measures Complex II activity. COX (Cytochrome C Oxidase) and NADH activities were blocked by electron transport inhibitors, such as Metformin. This has mechanistic and clinical implications for using Metformin as an anti-cancer drug, both for cancer therapy and chemo-prevention.

================

12) Metformin—an Adjunct Antineoplastic Therapy—Divergently Modulates Tumor Metabolism and Proliferation, Interfering with Early Response Prediction by 18F-FDG PET Imaging
Peiman Habibollahi*,1, Nynke S. van den Berg*,1, Darshini Kuruppu1, Massimo Loda2 and Umar Mahmood1
1Division of Nuclear Medicine and Molecular Imaging, Department of Radiology, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts; and 2Department of Pathology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts

MET, through activation of the AMPK pathway, produces a dose-dependent increase in tumor glucose uptake while decreasing cell proliferation in human and murine colon cancer cells.

Cancer Stem Cells

13) Bradford, Sherry A., and A. Khan. “Individualizing chemotherapy using the anti-diabetic drug, metformin, as “adjuvant”: an exploratory study.” J Cancer Sci Ther 5.6 (2013). Individualizing chemotherapy using metformin Bradford Sherry J Cancer Sci Ther 2013

when metformin was combined(with chemo) , a synergistic effect was observed resulting in high sensitivity (high cell kill);

metformin suppressed the generation of the breast cancer stem cell phenotype by regulating stem cell properties including the epithelial-mesenchymal transition status.

14) Lee, Hyemi, et al. “Response of breast cancer cells and cancer stem cells to metformin and hyperthermia alone or combined.” PloS one 9.2 (2014): e87979.

In the present study, we show that metformin is preferentially cytotoxic to Cancer Stem Cells (CSCs) relative to non-CSCs and that hyperthermia markedly increases the metformin cytotoxicity against CSCs. For the first time, we observed that hyperthermia activates AMPK, thereby suppressing mTOR. Such an activation of AMPK by hyperthermia appeared to play an important role in the hyperthermia-induced potentiation of metformin cytotoxicity against cancer cells, particularly against CSCs.

15) Song, Chang W., et al. “Metformin kills and radiosensitizes cancer cells and preferentially kills cancer stem cells.” Scientific reports 2 (2012): 362.

16) Kim, Tae Hun, et al. “Metformin against cancer stem cells through the modulation of energy metabolism: special considerations on ovarian cancer.” BioMed research international 2014 (2014).

Activation of AMPK provides a metabolic barrier to reprogramming somatic cells into stem cells [70]. The AMPK activators established a metabolic barrier to reprogramming that could not be bypassed, even through p53 deficiency, a fundamental mechanism to greatly improve the efficiency of stem cell production.

Metformin interferes with oxidative phosphorylation via interactions with respiratory complex I, resulting in reduced ATP production and metabolic stress. Metformin lowers plasma glucose levels by decreasing gluconeogenesis and glucose uptake, resulting in lower circulating insulin and IGF-1 levels.

Furthermore, LKB1-deficient cells were more sensitive to metformin-induced energy stress when cultured at low glucose concentrations and were unable to compensate for the decreased cellular ATP concentration, causing cell death [86]. These cytotoxic effects of metformin arise only in the context of a genetic defect, such as loss of p53 and/or LKB1, that is present in the cancer but not in the normal host tissue, providing opportunities for “synthetic lethality”

Metformin has been shown to decrease the production of inflammatory cytokines, including TNF-a, interleukin-6, and vascular endothelial growth factor, through the inactivation of NF-KB and HIF-1a [92–94]. Emerging results demonstrating the capacity of AMPK to inhibit the inflammatory responses suggest that metformin may also target the inflammatory component present in the tumor microenvironment [95]. In addition, several reports demonstrated that metformin treatment inhibits neoplastic angiogenesis, resulting in the reduction of tumor growth

Complex I inhibition is partially involved in metformin’s growth inhibition of EOC, possibly by increasing ROS and sensitizing cancer to additional oxidative stress.

Metformin has been demonstrated to augment the effects of various chemotherapeutic regimens by improving their efficacy as well as overcoming the chemoresistance in EOC (Table 1) [63–65, 67]. In fact, most in vitro studies used doses of metformin between 1 and 40?mM, which is well above the feasible therapeutic plasma levels (2.8–15?µM) in humans [98]. Whereas the cytotoxic effect of metformin alone was achieved at millimolar concentrations in most studies, Erices et al. observed cytotoxicity with micromolar metformin in combination with chemotherapy at concentrations where the chemotherapy alone produced no loss in viability.

Metformin Inhibits Inflammation Needed by Cancer Stem Cells 17) Hirsch, Heather A., Dimitrios Iliopoulos, and Kevin Struhl. “Metformin inhibits the inflammatory response associated with cellular transformation and cancer stem cell growth.” Proceedings of the National Academy of Sciences 110.3 (2013): 972-977. Metformin, the first-line drug for treating diabetes, inhibits cellular transformation and selectively kills cancer stem cells in breast cancer cell lines. In a Src-inducible model of cellular transformation, metformin inhibits the earliest known step in the process, activation of the inflammatory transcription factor NF-KappaBeta. Metformin strongly delays cellular transformation in a manner similar to that occurring upon a weaker inflammatory stimulus. Conversely, inhibition of transformation does not occur if metformin is added after the initial inflammatory stimulus. The antitransformation effect of metformin can be bypassed by overexpression of Lin28B or IL1ß, downstream targets of NF-KB. Metformin preferentially inhibits nuclear translocation of NF-KB and phosphorylation of STAT3 in cancer stem cells compared with non-stem cancer cells in the same population. The ability of metformin to block tumor growth and prolong remission in xenografts in combination with doxorubicin is associated with decreased function of the inflammatory feedback loop. Lastly, metformin-based combinatorial therapy is effective in xenografts involving inflammatory prostate and melanoma cell lines, whereas it is ineffective in noninflammatory cell lines from these lineages. Taken together, our observations suggest that metformin inhibits a signal transduction pathway that results in an inflammatory response. As metformin alters energy metabolism in diabetics, we speculate that metformin may block a metabolic stress response that stimulates the inflammatory pathway associated with a wide variety of cancers. IL-6 The transformed cells contain a minority population of CSCs that have an enhanced inflammatory loop that results in overproduction of IL6 (22, 24). The CSCs and non-stem cancer cells (NSCCs) within the transformed population are in a dynamic equilibrium that involves IL6 secretion Taken together, our observations suggest that metformin inhibits the inflammatory pathway necessary for transformation and CSC formation. 18) Hirsch, Heather A., et al. “Metformin selectively targets cancer stem cells, and acts together with chemotherapy to block tumor growth and prolong remission.” Cancer research 69.19 (2009): 7507-7511. Here, we show that metformin selectively kills cancer stem cells in four genetically different types of breast cancer. The combination of metformin and doxorubicin, a well-defined chemotherapeutic drug, kills both cancer stem cells and non–stem cancer cells in culture, and reduces tumor mass and prolongs remission much more effectively than either drug alone in a xenograft mouse model. These observations constitute independent support for the cancer stem cell hypothesis, and they provide a rationale for why the combination of metformin and chemotherapeutic drugs might improve treatment of patients with breast (and possibly other) cancers.’ Metformin Degrades Reduces Cyclin D1 19) Gwak, HyeRan, et al. ” Metformin induces degradation of cyclin D1 via AMPK/GSK3ß axis in ovarian cancer. ” Molecular carcinogenesis 56.2 (2017): 349-358. Metformin, which is widely used as an anti-diabetic drug, reduces cancer related morbidity and mortality. However, the role of metformin in cancer is not fully understood. Here, we first describe that the anti-cancer effect of metformin is mediated by cyclin D1 deregulation via AMPK/GSK3ß axis in ovarian cancer cells. Metformin promoted cytotoxic effects only in the cancer cells irrespective of the p53 status and not in the normal primary-cultured cells. Metformin induced the G1 cell cycle arrest, in parallel with a decrease in the protein expressions of cyclin D1 without affecting its transcriptional levels. Using a proteasomal inhibitor, we could address that metformin-induced decrease in cyclin D1 through the ubiquitin/proteasome process. Cyclin D1 degradation by metformin requires the activation of GSK3ß, as determined based on the treatment with GSK3ß inhibitors. The activation of GSK3ß correlated with the inhibitory phosphorylation by Akt as well as p70S6K through AMPK activation in response to metformin. These findings suggested that the anticancer effects of metformin was induced due to cyclin D1 degradation via AMPK/GSK3ß signaling axis that involved the ubiquitin/proteasome pathway specifically in ovarian cancer cells. Metformin Inhibits WNT pathway 20) Ahmed, Kamal, et al. “A second WNT for old drugs: Drug repositioning against WNT-dependent cancers.” Cancers 8.7 (2016): 66. A recent study revealed that anti-proliferative actions of metformin are also associated with the indirect inhibition of the WNT pathway. Surprisingly, its effects are mediated through its original target—AMPK, which then employs the MTOR signaling pathway to promote the ubiquitination and proteasomal degradation of DVL3, one of the principal WNT transducers [186]. This is very encouraging as it means that the drug can be used at its normal dose to exert its anti-WNT effects, and indeed the doses of metformin reported in the study corresponded to those found for AMPK activation in human tissues [187]. Metformin Glioblastoma Stem Cells 21) Gritti, Marta, et al. “Metformin repositioning as antitumoral agent: selective antiproliferative effects in human glioblastoma stem cells, via inhibition of CLIC1-mediated ion current.” Oncotarget 5.22 (2014): 11252. Clinical Trial 22) Metformin Hydrochloride and Doxycycline in Treating Patients With Localized Breast or Uterine Cancer . Verified May 2017 by Sidney Kimmel Cancer Center at Thomas Jefferson University 23) Chae, Young Kwang, et al. “Repurposing metformin for cancer treatment: current clinical studies.” Oncotarget 7.26 (2016): 40767. Preclinical studies have demonstrated several anticancer molecular mechanisms of metformin including mTOR inhibition, cytotoxic effects, and immunomodulation. Clinical trials in pre-surgical endometrial cancer patients exhibited a significant decrease in Ki67 with metformin monotherapy. Another interesting observation was made in patients with breast cancer, wherein a trend towards improvement in cancer proliferation markers was noted in patients without insulin resistance. metformin activates the T cell mediated immune response against cancer cells. Animal models of pancreatic cancer fed with metformin showed inhibition of insulin like growth factor-1 (IGF-1) and mTOR, along with an increase in phosphorylated AMPK In tobacco carcinogen induced lung cancer mice, the inhibition of insulin like growth factor 1 receptor/insulin receptor (IGF- 1R/IR) by metformin decreased the downstream signaling through Akt pathway. This reduced the activation of mTOR in lung tissue which corresponded to a 72% reduction in tumor burden [13]. 24) Proc Natl Acad Sci U S A. 2015 Feb 10;112(6):1809-14. Immune-mediated antitumor effect by type 2 diabetes drug, metformin. Eikawa S1, Nishida M1, Mizukami S1, Yamazaki C1, Nakayama E2, Udono H3. Metformin, a prescribed drug for type 2 diabetes, has been reported to have anti-cancer effects; however, the underlying mechanism is poorly understood. Here we show that this mechanism may be immune-mediated. Metformin enabled normal but not T-cell-deficient SCID mice to reject solid tumors. In addition, it increased the number of CD8(+) tumor-infiltrating lymphocytes (TILs) and protected them from apoptosis and exhaustion characterized by decreased production of IL-2, TNFa, and IFN?. CD8(+) TILs capable of producing multiple cytokines were mainly PD-1(-)Tim-3(+), an effector memory subset responsible for tumor rejection. Combined use of metformin and cancer vaccine improved CD8(+) TIL multifunctionality. The adoptive transfer of antigen-specific CD8(+) T cells treated with metformin concentrations as low as 10 µM showed efficient migration into tumors while maintaining multifunctionality in a manner sensitive to the AMP-activated protein kinase (AMPK) inhibitor compound C. Therefore, a direct effect of metformin on CD8(+) T cells is critical for protection against the inevitable functional exhaustion in the tumor microenvironment. ————————————————– from Targeting Cancer Stem Cells with NonToxic Therapies
　

25) Metformin Supplementation and Cancer Treatment
Feb 19, 2013 Brian D. Lawenda, M.D.

26) Bednar, Filip, and Diane M. Simeone. “Metformin and cancer stem cells: old drug, new targets.” Cancer Prevention Research 5.3 (2012): 351-354.

Metformin for BRCA GENE Carriers

27) Cell Cycle. 2017 Jun 3;16(11):1022-1028. Metformin inhibits RANKL and sensitizes cancer stem cells to denosumab. Cuyàs E1,2, Martin-Castillo B3, Bosch-Barrera J4,5, Menendez JA1,2.
The increased propensity of BRCA1 mutation carriers to develop aggressive breast tumors with stem-like properties begins to be understood in terms of osteoprotegerin (OPG)-unrestricted cross-talk between RANKL-overproducing progesterone-sensor cells and cancer-initiating RANK+ responder cells that reside within pre-malignant BRCA1mut/+ breast epithelial tissue. We recently proposed that, in the absence of hormone influence, cancer-initiating cells might remain responsive to RANKL stimulation, and hence to the therapeutic effects of the anti-RANKL antibody denosumab because genomic instability induced by BRCA1 haploinsufficiency might suffice to cell-autonomously hyperactivate RANKL gene expression. Here we report that the biguanide metformin prevents BRCA1 haploinsufficiency-driven RANKL gene overexpression, thereby disrupting an auto-regulatory feedback control of RANKL-addicted cancer stem cell-like states within BRCA1mut/- cell populations. Moreover, metformin treatment elicits a synergistic decline in the breast cancer-initiating cell population and its self-renewal capacity in BRCA1-mutated basal-like breast cancer cells with bone metastasis-initiation capacity that exhibit primary resistance to denosumab in mammosphere assays. The specific targeting of RANKL/RANK signaling with denosumab is expected to revolutionize prevention and treatment strategies currently available for BRCA1 mutation carriers. Our findings provide a rationale for new denosumab/metformin combinatorial strategies to clinically manage RANKL-related breast oncogenesis and metastatic progression.

28) Metformin suppresses triple-negative breast cancer stem cells by targeting KLF5 for degradation Cell Discovery 3, Article number: 17010 (2017) Peiguo Shi, Wenjing Liu, Tala, Haixia Wang, Fubing Li, Hailin Zhang, Yingying Wu, Yanjie Kong, Zhongmei Zhou, Chunyan Wang, Wenlin Chen, Rong Liu & Ceshi Chen

metformin significantly decreased the percentage of TNBC stem cells in two cell lines. Metformin inhibits mitochondrial complex I, which results in a decrease of ATP and the accumulation of AMP [32]. Accumulated AMP inhibits the generation of cAMP [32]. It has been established that cAMP activates PKA [32] and that activated PKA promotes mammary tumorigenesis [43]. Activated PKA also induces tamoxifen resistance in breast cancer [44]. We found that PKA has an important role in metformin-induced breast cancer stem cell suppression and that PKA is highly activated in triple-negative breast tumors. In agreement with our findings, metformin was reported to suppress breast cancer stem cells through the disruption of ATP production [45].



Synergy with Hyperthermia

29) Lee, Hyemi, et al. “Response of breast cancer cells and cancer stem cells to metformin and hyperthermia alone or combined.” PloS one 9.2 (2014): e87979.
Metformin, the most widely prescribed drug for treatment of type 2 diabetes, has been shown to exert significant anticancer effects. Hyperthermia has been known to kill cancer cells and enhance the efficacy of various anti-cancer drugs and radiotherapy. We investigated the combined effects of metformin and hyperthermia against MCF-7 and MDA-MB-231 human breast cancer cell, and MIA PaCa-2 human pancreatic cancer cells. Incubation of breast cancer cells with 0.5–10 mM metformin for 48 h caused significant clonogenic cell death. Culturing breast cancer cells with 30 µM metformin, clinically relevant plasma concentration of metformin, significantly reduced the survival of cancer cells. Importantly, metformin was preferentially cytotoxic to CD44high/CD24low cells of MCF-7 cells and, CD44high/CD24high cells of MIA PaCa-2 cells, which are known to be cancer stem cells (CSCs) of MCF-7 cells and MIA PaCa-2 cells, respectively. Heating at 42°C for 1 h was slightly toxic to both cancer cells and CSCs, and it markedly enhanced the efficacy of metformin to kill cancer cells and CSCs. Metformin has been reported to activate AMPK, thereby suppressing mTOR, which plays an important role for protein synthesis, cell cycle progression, and cell survival. For the first time, we show that hyperthermia activates AMPK and inactivates mTOR and its downstream effector S6K. Furthermore, hyperthermia potentiated the effect of metformin to activate AMPK and inactivate mTOR and S6K. Cell proliferation was markedly suppressed by metformin or combination of metformin and hyperthermia, which could be attributed to activation of AMPK leading to inactivation of mTOR. It is conclude that the effects of metformin against cancer cells including CSCs can be markedly enhanced by hyperthermia.

30) Metformin targets multiple signaling pathways in cancer
Yong Lei†, Yanhua Yi†, Yang Liu, Xia Liu, Evan T. Keller, Chao-Nan Qian, Jian Zhang Chinese Journal of Cancer 2017 36:17

31) A phase II clinical trial of metformin as a cancer stem cell targeting agent in stage IIc/III/IV ovarian, fallopian tube, and primary peritoneal cancer. Meeting:2017 ASCO Annual Meeting Abstract No:5556
Poster Board Number:Poster Session (Board #378)
Citation:J Clin Oncol 35, 2017 (suppl; abstr 5556)

Author(s): Ronald J. Buckanovich, Jason Brown, Jessica Shank, Kent A. Griffith, R. Kevin Reynolds, Carolyn Johnston, Karen McLean, Shitanshu Uppal, J. Rebecca Liu, Lourdes Cabrera, Geeta Mehta; Department of Internal Medicine, University of Michigan, Ann Arbor, MI; Department of Obstetrics and Gynecology, Naval Medical Center San Diego, San Diego, CA; Department of Biostatistics, University of Michigan, Ann Arbor, MI; Department of Obstetrics and Gynecology, University of Michigan, Ann Arbor, MI; Department of Bioengineering, University of Michigan, Ann Arbor, MI

Background: Epidemiologic and preclinical studies suggest that Metformin has antitumor effects which may be due to an impact on cancer stem-like cells (CSC). We present a phase II trial of metformin administered in combination with chemotherapy for patients with advanced stage epithelial ovarian cancer (EOC). Primary endpoints were 18 month progression free survival (PFS) and CSC number in Metformin treated tumors. Methods: Thirty-eight patients with confirmed stage IIC(n=1)/III(n=25)/IV(n=12) EOC were treated with either neoadjuvant metformin followed primary debulking surgery and adjuvant Metformin+chemotherapy, or neo-adjuvant metformin+chemotherapy, followed by interval debulking and adjuvant chemotherapy+Metformin. Patients were evaluated for side effects, PFS and overall survival (OS). Metformin treated tumors were evaluated for the presence of CSC via FACS and sphere assays. Results: Thirty-two patients (84%) completed at least six cycles of metformin+chemotherapy. Metformin was well tolerated with only one grade III/IV treatment-related adverse event (3%) noted. Common adverse effects were diarrhea (18%) and nausea (16%). Eighteen month PFS was 65.4% (95% confidence interval 47.9-78.3), Median PFS was 21.7 months (CI-17-26.7). Estimated three year OS was 73.5% (CI-54.7-84.3) with median OS not reached after a media follow-up of 33 months. Finally, tumors treated with metformin were noted to have a 3-fold decrease in ALDH+ CSC at baseline, increased sensitivity to Cisplatin in vitro, and a reduced ability to amplify ALDH+ CSC with passage in vitro. Conclusions: This is the first prospective study of Metformin in EOC patients. Translational studies confirm an impact of metformin on CSC. Metformin was well tolerated and outcome results were favorable, supporting the use of Metformin in phase-III studies. Clinical trial information: NCT01579812

32) Leão, Ricardo, et al. “Cancer Stem Cells in Prostate Cancer: Implications for Targeted Therapy.” Urologia Internationalis (2017).

33) DORAN, Elena, and Andrew P. HALESTRAP. “Evidence that metformin exerts its anti-diabetic effects through inhibition of complex 1 of the mitochondrial respiratory chain.” Biochemical Journal 348.3 (2000): 607-614. Metformin exerts effects through inhibition of complex 1 of the mitochondrial respiratory chain DORAN Elena Biochemical Journal 2000

34) Ward, N. P., et al. “Complex I inhibition augments dichloroacetate cytotoxicity through enhancing oxidative stress in VM-M3 glioblastoma cells.” PloS one 12.6 (2017): e0180061.

The robust glycolytic metabolism of glioblastoma multiforme (GBM) has proven them susceptible to increases in oxidative metabolism induced by the pyruvate mimetic dichloroacetate (DCA). Recent reports demonstrate that the anti-diabetic drug metformin enhances the damaging oxidative stress associated with DCA treatment in cancer cells. We sought to elucidate the role of metformin’s reported activity as a mitochondrial complex I inhibitor in the enhancement of DCA cytotoxicity in VM-M3 GBM cells. Metformin potentiated DCA-induced superoxide production, which was required for enhanced cytotoxicity towards VM-M3 cells observed with the combination. Similarly, rotenone enhanced oxidative stress resultant from DCA treatment and this too was required for the noted augmentation of cytotoxicity. Adenosine monophosphate kinase (AMPK) activation was not observed with the concentration of metformin required to enhance DCA activity. Moreover, addition of an activator of AMPK did not enhance DCA cytotoxicity, whereas an inhibitor of AMPK heightened the cytotoxicity of the combination. Our data indicate that metformin enhancement of DCA cytotoxicity is dependent on complex I inhibition. Particularly, that complex I inhibition cooperates with DCA-induction of glucose oxidation to enhance cytotoxic oxidative stress in VM-M3 GBM cells.

These data suggest that complex I inhibition cooperates with DCA activation of oxidative glucose metabolism to promote catastrophic oxidative stress in VM-M3 glioblastoma cells.

136) Wheaton, William W., et al. “Metformin inhibits mitochondrial complex I of cancer cells to reduce tumorigenesis.” Elife 3 (2014): e02242.

35) Griss, Takla, et al. “Metformin antagonizes cancer cell proliferation by suppressing mitochondrial-dependent biosynthesis.” PLoS biology 13.12 (2015): e1002309.

Metformin Propranolol Combination

36) also see (48) Rico, María, et al. “Metformin and propranolol combination prevents cancer progression and metastasis in different breast cancer models.” Oncotarget 8.2 (2017): 2874. Metformin and propranolol combination prevents cancer progression and metastasis in different breast cancer models.

37) Saengboonmee, Charupong, et al. “Metformin Exerts Antiproliferative and Anti-metastatic Effects Against Cholangiocarcinoma Cells by Targeting STAT3 and NF-KB.” Anticancer research 37.1 (2017): 115-123.

38) Zhu, Jie, et al. “Targeting cancer cell metabolism: The combination of metformin and 2-Deoxyglucose regulates apoptosis in ovarian cancer cells via p38 MAPK/JNK signaling pathway.” American journal of translational research 8.11 (2016): 4812.

Targeting cancer cell metabolism is a new promising strategy to fight cancer. Metformin, a first-line treatment for type 2 diabetes mellitus, exerts anti-cancer and anti-proliferative action. 2-deoxyglucose (2-DG), a glucose analog, works as a competitive inhibitor of glycolysis. In this study, we show for the first time that metformin in combination with 2-DG inhibited growth, migration, invasion and induced cell cycle arrest of ovarian cancer cells in vitro. Moreover, metformin and 2-DG could efficiently induce apoptosis in ovarian cancer cells, which was achieved by activating p38 MAPK and JNK pathways. Our study reinforces the growing interest of metabolic interference in cancer therapy and highlights the potential use of the combination of metformin and 2-DG as an anti-tumor treatment in ovarian cancer.

39) Ben, Sahra I., et al. “Targeting cancer cell metabolism: the combination of metformin and 2-deoxyglucose induces p53-dependent apoptosis in prostate cancer cells.” Cancer research 70.6 (2010): 2465.

Metformin targets cancer stem cells

40) Bost, F., et al. “Energy disruptors: rising stars in anticancer therapy?.” Oncogenesis 5.1 (2016): e188. Energy disruptors: rising stars in anticancer therapy?

Biguanides Metformin target cancer stem cells

Cancer stem cells (CSCs) are localized in tumors, resistant to chemotherapy, and capable of self-renewal and differentiation. Importantly, CSCs are the cause of disease relapse. Biguanides appear to target this cancer cell population. The combination of metformin with chemotherapy has been shown to be more efficient than either drug alone in xenograft models using several cancer cell lines, and this treatment specifically targets CSCs. Furthermore, treatment with both drugs significantly prolongs the remission following xenograft implantation.33, 34 This specific effect was confirmed in several other cancer models, including pancreas, breast and ovary.35, 36, 37 Interestingly, Sancho et al. have shown that CSCs rely mainly on OXPHOS and are unable to effectively induce glycolysis to compensate for reduced ATP production upon mitochondrial inhibition. The level of MYC expression controls this metabolic characteristic of CSCs; low MYC expression allows high PGC1-a expression, which results in enhanced mitochondrial biogenesis. Consequently, the observation that metformin specifically affects the viability of CSCs to a greater extent than non-CSCs is not surprising.38

Metformin and Cancer Stem cells

41) Patricia Sancho, et al. “MYC/PGC-1a Balance Determines the Metabolic Phenotype and Plasticity of Pancreatic Cancer Stem Cells.” Cell Metabolism 22 (2015): 1-16. MYC_PGC1a Determines Metabolic Phenotype Pancreatic Cancer Stem Cells Patricia Sancho Cell Metabolism 2015

The anti-diabetic drug metformin targets pancreatic cancer stem cells (CSCs), but not their differentiated progenies (non-CSCs), which may be related to distinct metabolic phenotypes. Here we conclusively demonstrate that while non-CSCs were highly glycolytic, CSCs were dependent on oxidative metabolism (OXPHOS) with very limited metabolic plasticity. Thus, mitochondrial inhibition, e.g., by metformin, translated into energy crisis and apoptosis. However, resistant CSC clones eventually emerged during treatment with metformin due to their intermediate glycolytic/respiratory phenotype. Mechanistically, suppression of MYC and subsequent increase of PGC-1a were identified as key determinants for the OXPHOS dependency of CSCs, which was abolished in resistant CSC clones. Intriguingly, no resistance was observed for the mitochondrial ROS inducer menadione and resistance could also be prevented/reversed for metformin by genetic/pharmacological inhibition of MYC. Thus, the specific metabolic features of pancreatic CSCs are amendable to therapeutic intervention and could provide the basis for developing more effective therapies to combat this lethal cancer.

Metformin Enhances Rituxan in B-cell Lymphoma

42) http://ascopubs.org/doi/abs/10.1200/jco.2015.33.15_suppl.e19513
Metformin enhances the activity of rituximab in B-cell lymphoma pre-clinical models. Priyank P. Patel, Juan J Gu, Cory Mavis, Myron Stefan Czuczman, Francisco J. Hernandez-Ilizaliturri

Background: The Warburg effect is primarily observed in rapidly growing tumors including aggressive B-cell lymphomas and is thought to be a consequence of the progression to cancer rather than the cause of it and altering the glucose metabolism in cancer cells appears to be an attractive strategy in cancer medicine. Retrospective studies have shown survival benefit in solid tumors and diffuse large B-cell lymphoma cohorts who were on metformin for type-2 diabetes. In an attempt to characterize the mechanism by which metformin affects the biology of B-cell lymphoma, we studied its effect on rituximab activity. Methods: A panel of B-cell lymphoma cells was exposed to metformin +/- rituximab or isotype control, changes in cell cycle distribution or induction of apoptosis was determined by flow cytometry. Antibody-dependent cellular cytotoxicity (ADCC) and complement mediated cytotoxicity (CMC) assays were performed to demonstrate changes in sensitivity to rituximab following metformin exposure. For in vivo studies, SCID mice were inoculated via tail vein injection (iv) with Raji cells (day 0) and assigned to observation, rituximab (at 10mg/kg/dose on days +3,7,10 and 14), metformin (at 2mg/ml in drinking water) or metformin and rituximab. Differences in survival (measured at the time for limb paralysis development) were evaluated by log-rank test between treatment arms. Results: In vitro exposure to metformin resulted in S/G1 cell cycle arrest and induction of apoptosis in a dose-dependent manner. Metformin enhanced the anti-proliferative effects of mAbs targeting CD20. Moreover, pre-incubation of lymphoma cells enhanced rituximab-mediated CMC. In vivo, significant improvement in survival was observed in metformin + rituximab arm (mean survival not reached at 69+/- 5.3 days) compared to rituximab (mean survival 57.1 +/- 4.2 days) (p = 0.05). Conclusions: Our data suggests that metformin inhibits the proliferation of B-cell lymphoma cell lines and enhances the anti-tumor activity of rituximab. Our finding highlights a potential role for metformin in the treatment of B-cell malignancies.

43) Rodríguez-Lirio, A., et al. “Metformin induces cell cycle arrest and apoptosis in drug-resistant leukemia cells.” Leukemia research and treatment 2015 (2015).

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Anti Angiogenesis

44) https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4792577/
Wang, Jichang, et al. “Suppression of tumor angiogenesis by metformin treatment via a mechanism linked to targeting of HER2/HIF-1a/VEGF secretion axis.” Oncotarget 6.42 (2015): 44579.

45) https://www.ncbi.nlm.nih.gov/pubmed/25196138/
Int J Cancer. 2015 Mar 15;136(6):E534-44. doi: 10.1002/ijc.29193. Epub 2014 Sep 18. The biguanides metformin and phenformin inhibit angiogenesis, local and metastatic growth of breast cancer by targeting both neoplastic and microenvironment cells. Orecchioni S1, Reggiani F, Talarico G, Mancuso P, Calleri A, Gregato G, Labanca V, Noonan DM, Dallaglio K, Albini A, Bertolini F.

The human white adipose tissue (WAT) contains progenitors with cooperative roles in breast cancer (BC) angiogenesis, local and metastatic progression. The biguanide Metformin (Met), commonly used for Type 2 diabetes, might have activity against BC and was found to inhibit angiogenesis in vivo. We studied Met and another biguanide, phenformin (Phe), in vitro and in vivo in BC models. In vitro, biguanides activated AMPK, inhibited Complex 1 of the respiratory chain and induced apoptosis of BC and WAT endothelial cells. In coculture, biguanides inhibited the production of several angiogenic proteins. In vivo, biguanides inhibited local and metastatic growth of triple negative and HER2+ BC in immune-competent and immune-deficient mice orthotopically injected with BC. Biguanides inhibited local and metastatic BC growth in a genetically engineered murine model model of HER2+ BC. In vivo, biguanides increased pimonidazole binding (but not HIF-1 expression) of WAT progenitors, reduced tumor microvessel density and altered the vascular pericyte/endothelial cell ratio, so that cancer vessels displayed a dysplastic phenotype. Phe was significantly more active than Met both in vitro and in vivo. Considering their safety profile, biguanides deserve to be further investigated for BC prevention in high-risk subjects, in combination with chemo and/or targeted therapy and/or as post-therapy consolidation or maintenance therapy for the prevention of BC recurrence.

VEGF Overexpressed in Mantle Cell Lymphoma

46) Anticancer Res. 2002 Sep-Oct;22(5):2899-901.
Immunohistochemical detection of C-kit (CD117) and vascular endothelial growth factor (VEGF) overexpression in mantle cell lymphoma. Potti A1, Ganti AK, Kargas S, Koch M.

Mantle cell lymphoma (MCL) is a low-grade lymphoproliferative malignancy that is extremely refractory to chemotherapy. Commonly used treatments have yielded unfavorable response rates (30% complete remission). We evaluated the incidence of c-kit (CD117) and vascular endothelial growth factor (VEGF) overexpression in patients with MCL in an effort to identify possible targets for therapeutic

Patients with a diagnosis of MCL based on CD5 positivity associated with cyclin D1 positivity and CD23 negativity on the lymph node/bone marrow specimen were included in our retrospective study. CD117 overexpression was performed using immunohistochemistry on archival specimens. VEGF expression was detected by the avidin-biotin-complex method.
RESULTS:Between 1997 and 2001, we identified 17 patients with MCL (9 males, 8 females) with a mean age of 57 years (age range: 42-66 years). The mean overall survival was 34 months (range: 11-60 months). VEGF expression was identified in 7 out of 17 (41.18%) patients with MCL. Among the VEGF-positive patients (n = 7, 41.1%), the mean survival was 24 months (range: 11-42 months), while patients without VEGF expression (n = 10, 58.9%) had a mean survival of 44 months (range: 21-60 months). CD117 expression was identified in only 2 out of 17 (1.17%) patients in our study.
CONCLUSION:Our study evaluated the role of c-kit and VEGF overexpression in MCL. Although CD117 may not be of therapeutic significance, target-directed signal transduction inhibition therapy using VEGF-inhibitors may be a distinct possibility in a select group of patients with MCL. Future larger studies are urgently needed to elaborate the role of VEGF in MCL.

metformin with 2DG

47) http://www.fasebj.org/content/31/1_Supplement/824.5
Anti-angiogenic Effects of Metformin in 2-Deoxyglucose Treated Microvascular Endothelial Cells: Role of Thrombospondin-1
Samson Mathews Samuel, Suparna Ghosh, Yasser Majeed, Hong Ding and Chris R Triggle. Pharmacology, Weill Cornell Medicine-Qatar, Doha, Qatar

Background & objective The biguanide metformin, which is widely used in the management of type 2 diabetes, has received considerable interest as a potential anti-cancer agent in many forms of cancer. However, the effect of metformin in tumor endothelial cells (TECs) has not been studied. TECs play a key role in tumor angiogenesis thereby supporting tumor growth, cancer cell survival and metastasis and hence targeting TECs in order to inhibit angiogenesis could prove to be a potential anti-angiogenic cancer therapy in a wide range of cancers. Reports show that metformin increases the levels of anti-angiogenic thrombospondin-1 (TSP1) in the serum of women with polycystic ovarian syndrome (1). Data from our preliminary studies in Mouse Microvascular Endothelial Cells (MMECs) that overexpress angiogenic VEGF (CRL-2460; ATCC; sarcoma cell line; cell type: endothelial) has revealed that metformin (2mM) significantly increased TSP1 in glucose-starved cells when compared to metformin treated glucose exposed cells. We therefore investigated the effects of metformin on angiogenesis, in MMECs in combination with the glycolytic inhibitor, 2-deoxyglucose (2DG).

Materials & methods MMECs were treated with 2DG (5mM) for 48h in the presence or absence of metformin (2mM) & western blot analysis was performed to assess the status of angiogenic and anti-angiogenic marker proteins. Alternatively, the putative AMPK activator, A769662 (150mM), was also used instead of metformin. Cell proliferation assay, cell migration assays and wound healing assays were also performed.

Results We observed a significant up-regulation of TSP1, while the levels of pVEGFR2 (Y1175) were markedly decreased; in 2DG-exposed cells treated with metformin (2mM) when compared to cells maintained in normal glucose exposed cells that were treated with metformin. A769662, did not show any effect on TSP-1 levels in normal glucose or 2DG-exposed cells. Furthermore, treatment with metformin (2mM) in 2DG exposed cells significantly increased the levels of pRap (S792), which in turn should have caused the inhibition of the mTOR pathway as evidenced by the significant decrease in the levels of pmTOR (S2448), p4E-BP1 (T36/47), pS6 (S235/236) and pS6 (240/244) when compared to metformin treated normal glucose exposed cells. Levels of cell cycle related proteins such as pCycB1 (S147), CycD1 and CycD2 significantly decreased in cells treated with a combination of 2DG and metformin when compared to cells that were treated with either 2DG or metformin alone. The rate of cell proliferation and endothelial cell migration also significantly decreased in cells treated with a combination of 2DG and metformin when compared to cells that were treated with either 2DG or metformin alone.

Conclusion Our findings show that using metformin in combination with 2DG has an anti-angiogenic activity associated with a significant up-regulation of thrombospondin-1 and could prove to be therapeutic strategy in a wide range of cancers.

Metformin and Propranolol

48) Rico, María, et al. “Metformin and propranolol combination prevents cancer progression and metastasis in different breast cancer models.” Oncotarget 8.2 (2017): 2874.

Taken together our results suggest that metformin plus propranolol combined treatment might be beneficial for triple negative breast cancer control, with no symptoms of toxicity.

The indirect anticancer effect of Met involves insulin dependent actions, associated with a reduction of insulin circulating levels that lead to a decrease in the mitogenic and antiapoptotic potential of insulin. In this way, Met may diminish the pro-stimulatory effect of insulin on cancer cells. Met direct effects are linked to inhibition of mitochondrial complex I [16]. This inhibition interrupts mitochondrial respiration, decreasing proton-driven synthesis of ATP, causing cellular energetic stress and elevation of the AMP:ATP ratio which, in turn, activates AMP-activated protein kinase (AMPK), a key cellular energy sensor kinase [10]. AMPK activation leads to a reduction in mammalian target of rapamycin (mTOR) signaling, protein synthesis and proliferation [16–18].

Propranolol (Prop) is a noncardioselective ß-adrenergic receptor blocker with reported antioxidant and anti-inflammatory properties, used traditionally for hypertension, angina pectoris, myocardial infarction, migraines, anxiety disorders, and tremor [19]. It was previously shown that Prop reduces intracellular calcium levels, Bax-mediated cytochrome C release and inhibits protein kinase C (PKC) activity in a ß-adrenoreceptor independent manner [19–21], and it induces cell cycle arrest and apoptosis via Akt/MAPK pathway in melanoma cells [22]. Many studies in humans have demonstrated its efficacy for the treatment of infantile haemangioma. In this regard, it seems that Prop exerts its suppressive effects acting through the HIF-1a-VEGF-A angiogenesis axis, with effects mediated through the PI3K/Akt and p38/MAPK pathways [23]. In relation to breast cancer, retrospective studies reported an improved survival with reduction in the risk of recurrence in woman receiving this ß-blocker therapy [24].”’

49) https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3248132/
Oncotarget. 2010 Nov; 1(7): 466–469. Beta-adrenergic signaling, a novel target for cancer therapy? Hildegard M. Schuller

itraconazole

50) https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4703001/
Head, Sarah A., et al. “Antifungal drug itraconazole targets VDAC1 to modulate the AMPK/mTOR signaling axis in endothelial cells.” Proceedings of the National Academy of Sciences 112.52 (2015): E7276-E7285.

free pdf
51) Tsubamoto, Hiroshi, et al. “Repurposing itraconazole as an anticancer agent.” Oncology Letters 14.2 (2017): 1240-1246.
Itraconazole may be a promising agent for targeting CSCs in relapsed disease of multiple types of cancer; therefore, further preclinical studies on CSCs and the surrounding stroma cells are warranted.

52) https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4406527/
Pantziarka, Pan, et al. “Repurposing Drugs in Oncology (ReDO)—itraconazole as an anti-cancer agent.” ecancermedicalscience 9 (2015).

========================================

53) https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4096030/
Pantziarka, Pan, et al. “The repurposing drugs in oncology (ReDO) project.” ecancermedicalscience 8 (2014).

Mebendazole Anthelminthic Threadworm infections Generic
Nitroglycerin Vasodilator Angina Generic
Cimetidine H2-receptor antagonist Peptic ulcer Generic
Clarithromycin Antibiotic Respiratory tract infection Generic
Diclofenac NSAID Pain relief Generic
Itraconazole Antifungal Broad spectrum antifungal Generic

pdf
55) Pantziarka, Pan, et al. “Repurposing drugs in your medicine cabinet: untapped opportunities for cancer therapy?.” Future oncology 11.2 (2015): 181-184.

56) https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4488459/
McCabe, Bronagh, Fabio Liberante, and Ken I. Mills. “Repurposing medicinal compounds for blood cancer treatment.” Annals of hematology 94.8 (2015): 1267-1276.

57) Cao, Jia, et al. “Low concentrations of metformin suppress glucose production in hepatocytes through AMP-activated protein kinase (AMPK).” Journal of Biological Chemistry 289.30 (2014): 20435-20446.

58) Mathupala, S. P., YH and Ko, and P. L. Pedersen. “Hexokinase II: cancer’s double-edged sword acting as both facilitator and gatekeeper of malignancy when bound to mitochondria.” Oncogene 25.34 (2006): 4777.

59) De Francesco, E. M., Michael Lisanti et al. “Vitamin C and Doxycycline: a synthetic lethal combination therapy targeting metabolic flexibility in cancer stem cells (CSCs).” Oncotarget (2017). Vitamin C and Doxycycline: A synthetic lethal combination therapy targeting metabolic flexibility in cancer stem cells (CSCs).

60) Sci Rep. 2016; 6: 18673. Aspirin and atenolol enhance metformin activity against breast cancer by targeting both neoplastic and microenvironment cells Giovanna Talarico,1,* Stefania Orecchioni,1,* Katiuscia Dallaglio,2 Francesca Reggiani,1 Patrizia Mancuso,1 Angelica Calleri,1 Giuliana Gregato,1 Valentina Labanca,1 Teresa Rossi,2 Douglas M. Noonan,3,4 Adriana Albini,3,* and Francesco Bertolinia,1,*
61) http://jnm.snmjournals.org/content/55/3/439.full
Propranolol Inhibits Glucose Metabolism and 18F-FDG Uptake of Breast Cancer Through Post-transcriptional Downregulation of Hexokinase-2. Fei Kang1, Wenhui Ma1, Xiaowei Ma1, Yahui Shao1, Weidong Yang1, Xiaoyuan Chen2, Liwen Li1,3 and Jing Wang1 Author Affiliations. 1Department of Nuclear Medicine, Xijing Hospital, Fourth Military Medical University, Xi’an, China

62) β-adrenoceptor can influence the 18F-FDG PET imaging of breast cancer through its regulation to the posttranscriptional level of hexokinase-2. Fei Kang1, Xiaowei Ma1, Wenhui Ma1, Yahui Shao1, Liwen Li1, Weidong Yang1 and Jing Wang1

63) https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5102691/

Ecancer medical science. 2016; 10: 680. Repurposing Drugs in Oncology (ReDO)—Propranolol as an anti-cancer agent. Pan Pantziarka,1,2 Gauthier Bouche,1 Vidula Sukhatme,3 Lydie Meheus,1 Ilse Rooman,1,4 and Vikas P Sukhatme3,5

Dosage

The PRO dose varies by indication. The anti-hypertensive dose is in the range 160 – 320 mg/day, starting at 80 mg and increasing as required to a maintenance dose that is generally 160 mg – 240 mg, in divided doses or as once a day use of extended release tablets. For angina the dose is 120 – 240 mg/day. Migraine prophylaxis is in the range 80 – 240 mg/day [4].

64) http://www.tandfonline.com/doi/full/10.3109/13880209.2014.892513

İşeri, Özlem Darcansoy, et al. “beta-Adrenoreceptor antagonists reduce cancer cell proliferation, invasion, and migration.” Pharmaceutical biology 52.11 (2014): 1374-1381.

Artesunate Degrades c-MYC and Inhibits WNT

65) Lu, Jin-Jian, et al. “Dihydroartemisinin accelerates c-MYC oncoprotein degradation and induces apoptosis in c-MYC-overexpressing tumor cells.” Biochemical pharmacology 80.1 (2010): 22-30.

Artemisinin and its derivatives (ARTs) are effective antimalarial drugs and also possess profound anticancer activity. However, the mechanism accounted for its distinctive activity in tumor cells remains unelucidated. We computed Pair wise Pearson correlation coefficients to identify genes that show significant correlation with ARTs activity in NCI-55 cell lines using data obtained from studies with HG-U133A Affymetrix chip. We found c-myc is one of the genes that showed the highest positive correlation coefficients among the probe sets analyzed (r=0.585, P<0.001). Dihydroartemisinin (DHA), the main active metabolite of ARTs, induced significant apoptosis in HL-60 and HCT116 cells that express high levels of c-MYC. Stable knockdown of c-myc abrogated DHA-induced apoptosis in HCT116 cells. Conversely, forced expression of c-myc in NIH3T3 cells sensitized these cells to DHA-induced apoptosis. Interestingly, DHA irreversibly down-regulated the protein level of c-MYC in DHA-sensitive HCT116 cells, which is consistent to persistent G1 phase arrest induced by DHA. Further studies demonstrated that DHA accelerated the degradation of c-MYC protein and this process was blocked by pretreatment with the proteasome inhibitor MG-132 or GSK 3beta inhibitor LiCl in HCT116 cells. Taken together, ARTs might be useful in the treatment of c-MYC-overexpressing tumors. We also suggest that c-MYC may potentially be a biomarker candidate for prediction of the antitumor efficacies of ARTs.

66) Oncotarget. 2015 Jun 30;6(18):15857-70.
Structurally diverse c-Myc inhibitors share a common mechanism of action involving ATP depletion. Wang H1, Sharma L1, Lu J1, Finch P1, Fletcher S2,3, Prochownik EV1,4,5.

The c-Myc (Myc) oncoprotein is deregulated in a large proportion of diverse human cancers. Considerable effort has therefore been directed at identifying pharmacologic inhibitors as potential anti-neoplastic agents. Three such groups of small molecule inhibitors have been described. The first is comprised of so-called “direct” inhibitors, which perturb Myc’s ability to form productive DNA-binding heterodimers in association with its partner, Max. The second group is comprised of indirect inhibitors, which largely function by targeting the BET-domain protein BRD4 to prevent the proper formation of transcriptional complexes that assemble in response to Myc-Max DNA binding. Thirdly, synthetic lethal inhibitors cause the selective apoptosis of Myc over-expressing either by promoting mitotic catastrophe or altering Myc protein stability. We report here a common mechanism by which all Myc inhibitors, irrespective of class, lead to eventual cellular demise. This involves the depletion of ATP stores due to mitochondrial dysfunction and the eventual down-regulation of Myc protein. The accompanying metabolic de-regulation causes neutral lipid accumulation, cell cycle arrest, and an attempt to rectify the ATP deficit by up-regulating AMP-activated protein kinase (AMPK). These responses are ultimately futile due to the lack of functional Myc to support the requisite anabolic response. Finally, the effects of Myc depletion on ATP levels, cell cycle arrest, differentiation and AMPK activation can be mimicked by pharmacologic inhibition of the mitochondrial electron transport chain without affecting Myc levels. Thus, all Myc inhibitors promote a global energy collapse that appears to underlie many of their phenotypic consequences.

67) Patricia Sancho, et al. MYC_PGC1a Determines Metabolic Phenotype Pancreatic Cancer Stem Cells Patricia Sancho Cell Metabolism 2015

However, eventually tumors relapsed under metformin due to emergence of resistant CSCs with an intermediate metabolic phenotype with increased c-MYC expression.

Mechanistically, we found that low MYC expression in human CSCs allowed high PGC1A expression levels, which resulted in enhanced mitochondrial biogenesis, strong mitochondrial activity and antioxidant properties, and subsequently low mitochondrial ROS levels, as a prerequisite for their stemness functions. Intriguingly, sustained suppression of MYC was required for maintaining stemness but rendered CSCs unable to substantially activate glycolysis and thus highly susceptible to mitochondrial targeting, e.g., by metformin or menadione.

MYC promotes a Warburg-like glycolytic phenotype, probably via dual mechanism: (1) upregulation of key glycolytic enzymes and (2) suppression of PGC1A . The latter is a transcriptional co-activator for nuclear receptor PPARs , which allows the protein to interact with multiple transcription factors, e.g., cAMP response element-binding protein (CREB) and nuclear respiratory factors (NRFs) (Fernandez-Marcos and Auwerx, 2011 ). While it is known that proteasomal degradation inhibits PGC-1 a(Ahuja et al.,2010), we here show for the first time a direct inhibitory effect of MYC on PGC-1 a at the transcriptional level, upon MYC binding to the PGC1A promoter.
Interestingly, while MYC has been linked to stemness properties in other tumors, e.g., hepatocellular carcinoma, breast, and lung cancer (
Akita et al., 2014; Zhao et al., 2015 ), we found in PDAC that MYC was associated with a more differentiated phenotype and MYC overexpression actually reduced stemness in CSCs (Figure 7). Notably, while suppression of MYC was essential for maintaining CSC phenotypes, the mere inhibition of MYC in non-CSCs did not equip them with stemness features.

68) Yi, Shuhua, et al. “High incidence of MYC and BCL2 abnormalities in mantle cell lymphoma, although only MYC abnormality predicts poor survival.” Oncotarget 6.39 (2015): 42362. In multivariate analysis, the MYC abnormality was the independent adverse factor for both PFS and OS, and intensive chemotherapy did not improve the outcome of these patients.

In the literatures, patients with a MYC abnormity always had extensive bone marrow involvement or leukemic presentation. So, cases of MCL with a MYC abnormality represent a relatively unique group with highly aggressive clinical and biological behavior.

Then, we determined if the HyperCVAD/MA ± R regimen could improve the survival of patients with a MYC abnormality. Unfortunately, this regimen had no impact on the survival of these patients.

“Intensive chemotherapy such as HyperCVAD/MA ± R did not improve the survival of patients with a MYC abnormality, and a new treatment strategy should be developed.”

69) Histopathology. 2016 Feb;68(3):442-9. MYC overexpression correlates with MYC amplification or translocation, and is associated with poor prognosis in mantle cell lymphoma.
Choe JY1,2, Yun JY1,2, Na HY2,3, Huh J4, Shin SJ4, Kim HJ5, Paik JH1, Kim YA6, Nam SJ3, Jeon YK3, Park G7, Kim JE2,6.

We aimed to investigate MYC expression and chromosomal aberration in mantle cell lymphoma (MCL), and the clinical significance of these factors.
METHODS AND RESULTS: Sixty-five patients with MCL, including 54 classic, nine blastoid and two pleomorphic variants, were enrolled. Expression of MYC, Ki67 and p53 was assessed by immunohistochemistry. MYC amplification or translocation was examined by fluorescence in-situ hybridization. MYC expression was higher in blastoid/pleomorphic MCL variants (mean, 19.0%) than in classic MCL (mean, 1.9%; P < 0.001). Expression of p53 and Ki67 was also significantly higher in these variants. MYC amplification was found in two of 53 cases tested, both of which were blastoid variants with high MYC expression (29.7% and 20.4%). MYC translocation was found in two of 52 cases tested, both of which were pleomorphic variants with remarkably high MYC expression (68.5% and 71.0%). High MYC or p53 expression was significantly associated with shortened overall survival and progression-free survival in univariable and multivariable analyses (all P < 0.05).
CONCLUSIONS: MYC overexpression is a negative predictor of MCL patient outcomes. MYC gene amplification or translocation might be related to the pathogenesis of MCL, particularly in blastoid/pleomorphic variants.

70) Nguyen, Lynh, Peter Papenhausen, and Haipeng Shao. “The Role of c-MYC in B-Cell Lymphomas: Diagnostic and Molecular Aspects.” Genes 8.4 (2017): 116.

c-MYC is one of the most essential transcriptional factors, regulating a diverse array of cellular functions, including proliferation, growth, and apoptosis. Dysregulation of c-MYC is essential in the pathogenesis of a number of B-cell lymphomas, but is rarely reported in T-cell lymphomas. c-MYC dysregulation induces lymphomagenesis by loss of the tight control of c-MYC expression, leading to overexpression of intact c-MYC protein, in contrast to the somatic mutations or fusion proteins seen in many other oncogenes. Dysregulation of c-MYC in B-cell lymphomas occurs either as a primary event in Burkitt lymphoma, or secondarily in aggressive lymphomas such as diffuse large B-cell lymphoma, plasmablastic lymphoma, mantle cell lymphoma, or double-hit lymphoma. Secondary c-MYC changes include gene translocation and gene amplification, occurring against a background of complex karyotype, and most often confer aggressive clinical behavior, as evidenced in the double-hit lymphomas. In low-grade B-cell lymphomas, acquisition of c-MYC rearrangement usually results in transformation into highly aggressive lymphomas, with some exceptions. In this review, we discuss the role that c-MYC plays in the pathogenesis of B-cell lymphomas, the molecular alterations that lead to c-MYC dysregulation, and their effect on prognosis and diagnosis in specific types of B-cell lymphoma.

Double Hit Lymphomas Venetoclax

71) Friedberg, Jonathan W. “How I treat” Double Hit” lymphoma.” Blood (2017): blood-2017. How I treat Double Hit Lymphoma c-MYC Friedberg Jonathan W Blood 2017

The 2016 revision of the WHO classification for lymphoma classification has included a new category of lymphoma, separate from diffuse large B-cell lymphoma termed High grade B-cell lymphoma with translocations involving myc and bcl-2 or bcl-6. These lymphomas, which occur
in less than 10% of cases of diffuse large B-cell lymphoma, have been referred to as “double hit” lymphomas (or triple hit lymphomas if all three rearrangements are present). It is important to differentiate these lymphomas from the larger group of “double expressor” lymphomas which have increased expression of MYC and BCL-2 and/or BCL-6 by immunohistochemistry, using variable cut-off percentages to define positivity. Double hit lymphomas have a poor prognosis when treated with standard chemoimmunotherapy, and have increased risk of central nervous system involvement and progression.

Novel agents with particular promise in patients with double hit DLBCL may include small molecule inhibitors of BCL-2 such as venetoclax, which has demonstrated in vivo efficacy against aggressive myc-driven mouse lymphomas 64 and has been studied in patients with
relapsed lymphoma with limited activity in aggressive histologies.65

72) Rattan, Ramandeep, et al. “Metformin suppresses ovarian cancer growth and metastasis with enhancement of cisplatin cytotoxicity in vivo.” Neoplasia 13.5 (2011): 483IN26-491IN28.
Ovarian cancer is the most lethal gynecologic cancer in women. Its high mortality rate (68%) reflects the fact that 75% of patients have extensive (>stage III) disease at diagnosis and also the limited efficacy of currently available therapies. Consequently, there is clearly a great need to develop improved upfront and salvage therapies for ovarian cancer. Here, we investigated the efficacy of metformin alone and in combination with cisplatin in vivo. A2780 ovarian cancer cells were injected intraperitoneally in nude mice; A2780-induced tumors in nude mice, when treated with metformin in drinking water, resulted in a significant reduction of tumor growth, accompanied by inhibition of tumor cell proliferation (as assessed by immunohistochemical staining of Ki-67, Cyclin D1) as well as decreased live tumor size and mitotic cell count. Metformin-induced activation of AMPK/mTOR pathway was accompanied by decreased microvessel density and vascular endothelial growth factor expression. More importantly, metformin treatment inhibited the growth of metastatic nodules in the lung and significantly potentiated cisplatin-induced cytotoxicity resulting in approximately 90% reduction in tumor growth compared with treatment by either of the drugs alone. Collectively, our data show for the first time that, in addition to inhibiting tumor cell proliferation, metformin treatment inhibits both angiogenesis and metastatic spread of ovarian cancer. Overall, our study provides a strong rationale for use of metformin in ovarian cancer treatment.

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Diabetes, Cancer and the Drug that Fights them Both

by Megan L Norris
figures by Bradley Wierbowski

The emerging link between cancer and diabetes
In the early 2000s, observations that diabetics are more likely to get cancer than non-diabetics began piling up. Was this because diabetes and cancer share general risk factors such as diet, aging and obesity? Or was there a direct link between them, with cancer benefiting from the sugar-rich and inflamed environment brought on by diabetes? Making bad news worse, it became apparent that cancer thrives in the presence of excess insulin, like that injected by many diabetics as therapy. Thus, one of the ways to treat diabetes could be making the cancer risk even worse.

Almost as soon as this dark cloud began to loom, rays of light broke through from an unexpected source. Research on a popular type II diabetes treatment called metformin revealed that metformin actually seemed to lower the risk for colorectal and other cancers in diabetics. Though it may seem paradoxical that metformin and insulin injections, two treatments for the same disease, could have such opposite effects on cancer, years of research in both the clinic and the laboratory has begun to pull back the curtain on this mystery. Broadly speaking, metformin makes the body more sensitive to the insulin it already is. For type II diabetics, not only does this increased insulin sensitivity treat diabetes, but it drains the fuel on which some cancers may thrive.

Figure 1: An overview of diabetes. In non-diabetics, sugar (glucose) from food exits the bloodstream in response to insulin and is made into cellular energy. In type I diabetics, the pancreatic beta-cells in charge of making insulin fail, causing a toxic build-up of glucose in the bloodstream called hyperglycemia. Type I diabetes can be treated with injection of insulin, which is significantly less precise than what a healthy pancreas can do. In type II diabetes, insulin is created but not sensed properly causing hyperglycemia. In response, the pancreas creates even more insulin resulting in high blood insulin levels called hyperinsulinemia.
Figure 1: An overview of diabetes. In non-diabetics, sugar (glucose) from food exits the bloodstream in response to insulin and is made into cellular energy. In type I diabetics, the pancreatic beta-cells in charge of making insulin fail, causing a toxic build-up of glucose in the bloodstream called hyperglycemia. Type I diabetes can be treated with injection of insulin, which is significantly less precise than what a healthy pancreas can do. In type II diabetes, insulin is created but not sensed properly causing hyperglycemia. In response, the pancreas creates even more insulin resulting in high blood insulin levels called hyperinsulinemia.
Diabetes is a pervasive disease
Diabetes is the 7th leading cause of death in the US and is on the rise. More than 10% of Americans over 20 years old have diabetes, with type II diabetes accounting for 95% of those cases. Untreated, diabetes leads to excessive amounts of glucose in the blood, a state called hyperglycemia, which can damage organs such as the heart, kidney and nerves.

In a non-diabetic individual following a meal, sugars and starches break down into glucose, which circulates in the blood. Beta-cells in the pancreas release insulin to trigger the movement of glucose out of the bloodstream and into cells where it can be used or stored. As shown in Figure 1, when no insulin is produced (type I diabetes) or is produced but not sensed correctly (type II diabetes), glucose amounts remain elevated in the blood causing dangerous hyperglycemia.

In addition to high blood glucose, diabetics also have a condition called hyperinsulinemia, which refers to excessively high amounts of insulin in their bloodstream. Since type I diabetes is marked by the body’s inability to produce insulin, a common treatment is insulin injection. Fine-tuning both the concentration and timing of these injections, and properly mimicking what a non-diabetic body can do, has proven a great challenge. As a result, these injections lead to significantly higher insulin levels than a non-diabetic would ever experience, resulting in hyperinsulinemia. Conversely, type II diabetics can make their own insulin, but their body isn’t sensitive to it. As a result, their bodies often pump out more and more insulin, resulting in a nearly constant state of hyperinsulinemia.

Additionally, this constant demand on the beta-cells can cause them to wear out and lose their ability to make insulin at all; patients who experience this are often given the insulin injections more common to type I diabetics, perpetuating their hyperinsulinemia through a different means. These elevated insulin levels are much less toxic to organs as compared to excess glucose, so the fact that treating hyperglycemia can lead to hyperinsulinemia, though not ideal, has been historically viewed as a considerably lesser evil.

Figure 2: The relationship between cancer and diabetes. Scenario I: Diabetics may have an increased occurrence of cancer because traits that make them susceptible to diabetes also make them susceptible to cancer. Scenario II: Side effects of diabetes, like hyperglycemia and hyperinsulinemia, may directly increase the ability of cancer to grow in a diabetic’s body.

Cancer runs on insulin
How could diabetes be affecting cancer? The simplest possibility is that what makes a person predisposed for diabetes is the same as what makes them predisposed for certain cancers, such as obesity and age. In this scenario, shown in Figure 2, one disease isn’t causing the other. Rather, both diseases simply capitalize on the same health situations. Though shared predispositions and risk factors may certainly explain some of the co-occurrence between diabetes and cancer, doctors and researchers are collecting mounting evidence that implicates diabetes as having a more direct role in increasing the risk of cancer. For one, cancer cells require a lot of energy to grow, divide, and spread. Importantly, one of cancer’s most vital energy sources is glucose, meaning that the high blood glucose levels in many diabetics could greatly benefit the growth and progression of cancer. More striking, however, is that insulin has been found to promote tumor growth and metastasis. Though this happens in diabetics and non-diabetics alike, people with diabetes have especially high levels of insulin in their blood, which could make for extra fertile grounds for cancer progression (Figure 2). As such, the insulin injections aimed at treating diabetes may also be increasing cancer risk.

Metformin treats diabetes without giving cancer a leg up
With the excess glucose and insulin present in diabetics and cancer potentially thriving off of both, must diabetics just live with an increased cancer risk? Enter metformin: the first-choice drug to treat type II diabetes. The exact way that metformin works is still being explored, but in the last two decades we have learned that metformin seems to have two important, and likely overlapping, methods of action. First, it sensitizes the body to insulin and, second, it coerces cells to make more energy (see Figure 3).

Figure 3: Metformin affects insulin signaling and energy production. Metformin sensitize the body to insulin already present in the bloodstream. As a result of better insulin signaling, cells begin to remove glucose from the blood and produce cellular energy.
Figure 3: Metformin affects insulin signaling and energy production. Metformin sensitize the body to insulin already present in the bloodstream. As a result of better insulin signaling, cells begin to remove glucose from the blood and produce cellular energy.
By increasing sensitivity to insulin, metformin helps type II diabetics start using the insulin already present in their bodies. This results in decreased insulin levels, which many researchers believe is how metformin is affecting cancer. This also means that metformin requires some insulin to be present in order to work best, so it alone may not be able to treat type I diabetics. However, it has recently been proposed to treat type I diabetes with a mix of insulin therapy and metformin, with the hope that lower concentrations of insulin would be required. In either case, after cells are treated with metformin and begin responding better to insulin, the cells can remove glucose from the blood to make cellular energy, thereby decreasing hyperglycemia and treating diabetes.

Cancer as a metabolic disease
Insulin and glucose are both metabolites, meaning that they are produced as byproducts of metabolism — the processes that all cells perform to obtain energy. The realization that cancer is tightly linked to metabolism only came to light in the early 2000s from unexpected, and seemingly unrelated, sources. While one group of researchers in Scotland was studying the basics of metabolism in yeast, separate groups across the world were studying cancer in rats, humans and frogs. Unexpectedly, the groups realized that they were all studying the same genes. Nearly simultaneously, one of the same genes was predicted to be the target of metformin. When all the pieces were put together, the idea that metformin could treat diabetes and cancer began to make sense.

Today, the data demonstrating the efficacy of metformin protecting against cancer is clear. A 2015 paper in the journal Cancer revealed metformin treatment decreased the chance of colorectal cancer in diabetics by 12%. Other studies demonstrated metformin selectively blocks tumor growth, can suppress cancer-promoting genes and provides a reduced risk of cancer in general. Intriguingly, some data suggests that, for certain cancers, this protective effect is only seen in diabetic settings. That is, cancers that are thriving from the hyperinsulinemic environment were treatable with metformin, while other cancers were not affected. Research into which cancers are likely to be susceptible to metformin is ongoing and relies heavily on understanding with more detail just how metformin is working. Ideally, we will soon know exactly how metformin is impacting insulin, metabolism and cancer.

Basic science makes big discoveries
The case of metformin is not unique. The more scientists follow the rabbit hole of basic research, which is research targeted at fundamental biology as opposed to medicine or disease, the more we equip our researchers in the clinic to make important, perhaps startling, medical advancements. Though possibly a pervasive idea, it is inaccurate to think we would be better off if we focused all our research efforts on disease. We have certainly come a long way, but with each decade of basic research, we uncover things cells do that we could have never predicted the decade before. Though aiming research directly at a disease is of course useful, persevering in our understanding of the foundations of biology is both crucial and expedient to making meaningful medical discoveries.

Megan L Norris is a 5th year PhD student in the department of Molecules, Cells and Organisms at Harvard University.

For more information:
Drug discovery: metformin and the control of diabetes

LKB1 and AMPK and the cancer-metabolism link – ten years after

Cover image modified from Sugar Cubes by David Pacey [CC BY 2.0], via Wikimedia Commons and Metastatic Melanoma Cells by Julio C. Valencia [CC BY-NC 2.0] via NIH Image Gallery flickr

Diabetes, Cancer and the Drug that Fights them Both - Science in the News
http://sitn.hms.harvard.edu/flash/2017/diabetes-cancer-drug-fights/

Pleiotropic Benefits of Metformin: Macrophage Targeting Its Anti-inflammatory Mechanisms | Diabetes
https://diabetes.diabetesjournals.org/content/64/6/1907

二甲双胍通过内质网相关降解PD-L1促进抗肿瘤免疫。
Metformin Promotes Antitumor Immunity via Endoplasmic-Reticulum-Associated Degradation of PD-L1

University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA


强调
•二甲双胍通过阻断PD-L1 / PD-1轴增强抗肿瘤CTL免疫力

•二甲双胍激活的AMPK在S195直接与PD-L1结合并磷酸化

•pS195诱导的PD-L1糖基化异常导致ERAD降解PD-L1

•二甲双胍和抗CTLA4联合治疗具有协同抗肿瘤作用

摘要
据报道，二甲双胍具有抗肿瘤活性并维持细胞毒性T淋巴细胞（CTL）高度免疫监测。但是，尚未完全了解二甲双胍在癌症免疫中的功能和详细机制。在这里，我们显示二甲双胍通过降低程序性死亡配体1（PD-L1）的稳定性和膜定位来增加CTL活性。此外，我们发现被二甲双胍激活的AMP激活的蛋白激酶（AMPK）直接磷酸化PD-L1的S195。 S195磷酸化诱导异常的PD-L1糖基化，导致其ER积累和与ER相关的蛋白质降解（ERAD）。一致地，二甲双胍治疗的乳腺癌患者的肿瘤组织在AMPK激活下表现出降低的PD-L1水平。通过二甲双胍阻断PD-L1的抑制信号可增强针对癌细胞的CTL活性。我们的发现确定了通过ERAD途径表达PD-L1的新调节机制，并表明二甲双胍-CTLA4阻断剂组合具有提高免疫疗法疗效的潜力。

Metformin Promotes Antitumor Immunity via Endoplasmic-Reticulum-Associated Degradation of PD-L1 - ScienceDirect
https://www.sciencedirect.com/science/article/pii/S1097276518305999#!

Biomed Pharmacother. 2019 Mar;111:1156-1165. doi: 10.1016/j.biopha.2019.01.021. Epub 2019 Jan 12.
Metformin enhances mitochondrial biogenesis and thermogenesis in brown adipocytes of mice.
Karise I1, Bargut TC2, Del Sol M3, Aguila MB4, Mandarim-de-Lacerda CA5.
Author information
1
Laboratory of Morphometry, Metabolism and Cardiovascular Disease, Biomedical Center, Institute of Biology, The University of the State of Rio de Janeiro, Rio de Janeiro, Brazil. Electronic address: iarakarise@hotmail.com.
2
Laboratory of Morphometry, Metabolism and Cardiovascular Disease, Biomedical Center, Institute of Biology, The University of the State of Rio de Janeiro, Rio de Janeiro, Brazil. Electronic address: therezabargut@gmail.com.
3
Doctoral Program in Morphological Sciences, Universidad de La Frontera, Temuco, Chile. Electronic address: mariano.delsol@ufrontera.cl.
4
Laboratory of Morphometry, Metabolism and Cardiovascular Disease, Biomedical Center, Institute of Biology, The University of the State of Rio de Janeiro, Rio de Janeiro, Brazil. Electronic address: mbaguila@uerj.br.
5
Laboratory of Morphometry, Metabolism and Cardiovascular Disease, Biomedical Center, Institute of Biology, The University of the State of Rio de Janeiro, Rio de Janeiro, Brazil. Electronic address: mandarim@uerj.br.
Abstract
AIMS:
We studied the effect of metformin on the brown adipose tissue (BAT) in a fructose-rich-fed model, focusing on BAT proliferation, differentiation, and thermogenic markers.

MAIN METHODS:
C57Bl/6 mice received isoenergetic diets for ten weeks: control (C) or high-fructose (F). For additional eight weeks, animals received metformin hydrochloride (M, 250 mg/kg/day) or saline. After sacrifice, BAT and white fat pads were prepared for light microscopy and molecular analyses.

KEY FINDINGS:
Body mass gain, white fat pads, and adiposity index were not different among the groups. There was a reduction in energy intake in the F group and energy expenditure in the F and FM groups. Metformin led to a more massive BAT in both groups CM and FM, associated with a higher adipocyte proliferation (β1-adrenergic receptor, proliferating cell nuclear antigen, and vascular endothelial growth factor), and differentiation (PR domain containing 16, bone morphogenetic protein 7), in part by activating 5' adenosine monophosphate-activated protein kinase. Metformin also enhanced thermogenic markers in the BAT (uncoupling protein type 1, peroxisome proliferator-activated receptor gamma coactivator-1 alpha) through adrenergic stimuli and fibroblast growth factor 21. Metformin might improve mitochondrial biogenesis in the BAT (nuclear respiratory factor 1, mitochondrial transcription factor A), lipolysis (perilipin, adipose triglyceride lipase, hormone-sensitive lipase), and fatty acid uptake (lipoprotein lipase, cluster of differentiation 36, adipocyte protein 2).

SIGNIFICANCE:
Metformin effects are not linked to body mass changes, but affect BAT thermogenesis, mitochondrial biogenesis, and fatty acid uptake. Therefore, BAT may be a metformin adjuvant target for the treatment of metabolic disorders.

Copyright © 2019 Elsevier Masson SAS. All rights reserved.

KEYWORDS:
Brown adipose tissue; Fructose; Metformin; Mouse; Thermogenesis

Metformin enhances mitochondrial biogenesis and thermogenesis in brown adipocytes of mice. - PubMed - NCBI
https://www.ncbi.nlm.nih.gov/pubmed/30841429