Animal Studies

Humans as animals

There is the Phase 1 clinical trial [1] and a study of patients undergoing chemotherapy for liver cancer. [2] In 2004 Dr Muhammad A M El-Saadani of the University of Cairo tested 0.8 mg/kg/day cuprous nicotinic acid as an intervention in patients undergoing 5-FU (12 mg/kg/day) for hepatocellular carcinoma (HCC).  Cuprous nicotinic acid mitigated the myelosuppressive effects of 5-FU a well as a decrease in liver enzymes in the blood of the patients. [2]   Cuprous nicotinic acid decreased 5-FU induced loss of ceruloplasmin and serum albumin.  These proteins are produced by the liver as part of its normal functioning. [17] An increase in IgG titer was interpreted as an indication of an immunomodulatory role.

As an immune modulator

Some of the first indications that cuprous nicotinic acid is an immune modulator came from studies with farm fish.  The ability of 30 and 60 mg Cu(I)NA2 per kg fish food to protect farmed mullet [3] and sea bass [4] from infection by the common fish pathogen Yersinia ruckerii has been tested. Note that Cu(I)NA2 refers to a compound whose hydration state is unknown whereas “Cunermuspir” and “Copper One” refer to the anhydrous form of cuprous nicotinic acid.   The farm fish were vaccinated with inactivated Yersina ruckerii bacteria and fed regular feed or Cu(I)NA2 incorporated into the feed for four weeks.  The response of the innate immune system was measured by the phagocytic index.   The adaptive immune system response was measured by antibody titers. [3,4] The vaccinated fish were exposed to live bacteria.  Cu(I)NA2 was found to enhance the innate and adaptive immune systems as well as ability to survive an actual pathogen challenge. [3,4] Complete blood counts and liver enzymes in the blood were presented to demonstrate the safety of Cu(I)NA2 to the fish.  Cuprous nicotinic acid has also shown immunomodulatory function in chicks. [5] Indeed, the Mitosynergy clinical trial of healthy volunteers concluded a significant increase in lymphocytes, a decrease in neutrophils, and a decrease in the neutrophil to lymphocyte ratio. [1]

protection against environmental toxins

A 2011 study tested the ability of cuprous nicotinic acid to mitigate the damage caused by mercuric chloride in female catfish. [6] Oxidative damage was measured as protein carbonyls and peroxides. [6]  Enzyme activity associated with remediation of such damage, catalase and superoxide dismutase, were also measured. [6] Glycerol is used in the food, cosmetic, and drug industries and not normally considered an environmental toxin at normal doses.  Glycerol was applied topically at a dose of 3.15 ml kg-1 body weight to male Wistar rats daily for 4 weeks. [7] Cuprous nicotinic acid was given at a dose of 0.043 mg kg-1 body weight three times a week for the four-week duration of the study [7] Cuprous nicotinic acid was also applied dermally with the glycerol.  Most Cuprous nicotinic acid normalized activities of catalase and superoxide dismutase. [7] Cuprous nicotinic acid increased reduced glutathione, and decreased malondialdehyde levels in glycerol exposed rats. [7]

As a treatment for burn injuries

Female rats were scald burned on exposed skin (~10 % total area) using an established model. [8] Rats were divided into three groups to receive dermal treatments to their burns (1) Vaseline, (2) MEBO, and 3mg Cu / kg body weight / day.  Cu was applied as cuprous nicotinic acid in a Vaseline ointment. [8] MEBO is a mixture containing sesame oil, beeswax, and other edible herbs, which provide β-sitosterol, 18 natural amino acids, four essential fatty acids, vitamin E, and polysaccharides. [9]  Outcome measures were serum concentrations of total proteins, albumin and the amino acids glutamate and phenylalanine. [8] MEBO and cuprous nicotinic acid in Vaseline improved these parameters over the Vaseline control at four weeks. [9] An earlier study compared paraffin, MEBO and cuprous nicotinic acid in paraffin as a treatment of female rats subjected to full thickness scald burns over 10% of their body surface area. [10] In the first 24 hours after the burn injury a decrease in lymphocytes was seen that improved over the course of four weeks following the application of MEBO and cuprous nicotinic acid in paraffin. [10] Cuprous nicotinic acid in the paraffin ointment improved plasma super oxide dismutase activity and ceruloplasmin, a copper carrier protein, levels at three- and four-weeks post injury. [10]

The first 24 hours after the burn injury were associated with necrosis and inflammatory cell infiltration of the skin.  One week later the MEBO and paraffin control groups showed persistent inflammation and tissue degeneration. [10] Burnt skin from the cuprous nicotinic acid group exhibited epidermal regeneration and angiogenesis.  [10]  The authors observed “keratin horns” in the cuprous nicotinic acid treated skin.  [10] All aspects of collagen restoration were seen in the cuprous nicotinic acid treated scalded skin. [10] Note that lysyl oxidase, an enzyme that cross-links collagen and elastin, requires a copper cofactor.

a treatment for stomach ulcers

In a 1993 study stomach ulcers were created in mice by pyloric ligation. [11]  ICuprous nicotinic acid was compared with cupric glycinate.  Both compounds were mixed with 0.25% Tween-80 in a saline solution and administered intragastrically. [11]  Both compounds increased the superoxide dismutase activity in the gastric mucosa as well as the blood plasma. [11] The changes in eicosanoids differed between the two copper compounds. [11] A second study in 2009 demonstrated that cuprous nicotinic acid alleviated several aspects of ulcer development in the rat pyloric ligation model. [12]

in cell culture models of triple negative breast cancer

All of these studies used a triple negative breast cancer (TNBC) cell line HCC1806 TNBC.  The triple negative of TNBC  refers to breast cancers negative for the estrogen receptor, progesterone receptor, and the and human epidermal growth factor receptor 2 (HER2).    

Before the advent of cuproptosis Mohamed A. Abdel-Mohsen and colleagues were looking for a therapy to overcome the resistance of a TNBC to the chemotherapy agent doxorubicin.  [13]  While cardiac toxicity is a common side effect of doxorubicin, hese authors another side effect was a process called autophagy.  They defined authophagy as the sequestration of cellular proteins and organelles in autophagosomes, the contents of which are digested when the autophagosome fuses with lysosomes. [13]  These authors used ATG8/LC3 The NBR1 gene product is the receptor for  the light chain 3 LC3, coded for by ATG8 gene, on the cell membrane of the autophagoxome. These authors induced autophage with torin-1 (TOR) autophagy inducer) and inhibited the lysosomeal digestion with chloroquine, which prevents acidification of the lysosome as well as the food digestion vacuole of the malaria parasite.

  • Doxorubicin, a chemotherapy agent that prevents DNA replication, particularly in rapidly dividing cancer cells.  In addition to cardiac toxicity, a side effect may be activation of autophagy.
  • Chloroquine inhibits acidification of the lysome, repurposed from the indication of inhibition of acidification of the digestive organelle of the malarial parasite.
  • Torin-1 is an inhibitor of mTORC1 and mTORC2, complexes which regulate mRNA translation based on input as to energy and nutrient availability and oxidative stress.
  • Cuprous nicotinic acid, the niacin being a precursor of nicotinamide adenine dinucleotide, NAD+, a ligand for sirtuin 1.  Sirt1 is a lysine deacetylase.  

Torin-1 and Doxorubicin acted in parallel inducing death by autophagy an apoptosis, but overall did not act synergistically in cell killing.  Chloroquine and cuprous nicotinic acid both promoted the accumulation of AVO (acid vesicular organelle) and promoted cell killing by apoptosis. [13]  A follow up study from this group [14 continued to elucidate the interplay between autophagy and apoptosis in doxorubicin treated TNBC cells. [14]    They speculated that doxorubicin could mediate autophagy by depletion of ATP and NAD+    and made note of the NAD+ dependent enzyme DNA repair enzyme  poly (ADP-ribose) polymerase-1 (PARP-1).  It should also be noted that ATP generation by the mitochondria involves the copper containing enzyme cytochrome C oxidase.  Cotreatment of TNBC cells with cuprous nicotinic decreased the concentration of doxorubicin needed to achieve 50% growth inhibition. [14]

Cuprous nicotinic acid was concluded to promote HCC1806 cell authophagy in the presence of other chemotherapy agent doxorubicin,       Autophagy occurs when autophagosomes fuse with lysosomes. 

The chemotherapy agent Doxorubicin exerts its anti-neoplastic effect by inserting itself between base pairs of DNA.   Cuprous nicotinic acid was tested as an adjunct treatment for triple negative breast cancer using the cell line HCC1806. [13] The small molecule drug  Chloroquine prevents acidification of the lysosome.  The mammalian target of Rapamycin (mTOR) may inhibit autophagy.     mTOR integrates signals from the environment with an output control of protein synthesis and proliferation.  Torin-1 is an inhibitor of mTORC1 and 2 complexes.  The mTORC1 complex seems to be more geared towards integrating dietary signals.   mTORC2 also responds to growth factors and may play a role in organizing the cytoskeleton.  

Viability of HCC1806 cells with IC50 values of Doxorubicin and autophagy inhibitors Torin-1 or chloroquine   was decreased when cuprous nicotinic acid was added. [13].  Another study published the same year demonstrated that cuprous nicotinic acid increased superoxide dismutase activity and promoted apoptosis. [14]  Apoptosis differs from autophagy in that the proteolytic enzymes, caspases, are secreted rather than relegated to the lysosomal contents.  The same year Dr. Abdel-Mohsen and colleagues published a report describing the actions of cuprous nicotinic acid on HCC‐1806 cells alone. [15]  The IC50 concentration needed to inhibit growth of 50% of the viable cells was about 100 μg/mL.  Growth inhibitory concentrations were used to evaluate other parameters of viability.   Concentrations of 1% and 10% of the IC50 seemed to promote early apoptosis whereas the IC50 of cuprous nicotinic acid concentration pushed cells toward necrosis. [15]  Untreated HCC-1805 and those treated with 1% of the cuprous nicotinic acid IC50  tended to be in G0 of the cell cycle.  Cuprous nicotinic acid at IC10% resulted in cells being in a sub G0 phase of the cell cycle. [15]  The increase in the programmed death protein 1, PD-1, is commonly expressed on the surface of T and B immune cells.  Binding of PD-1 to the PD-L1 receptor on tumor cells down regulates these immune cells resulting in immune tolerance.  The work of Dr. Abdel-Mohsen and colleagues and colleagues may have been the first report, unofficial, of a new form of cell death.  In March of 2022 Dr Peter Tsvetkov and colleagues published a report of  copper mediated regulated cell death that involved copper  binding to lipoic acid modified mitochondrial pyruvate dehydrogenase complex protein DHAT .   In this report the term “cuproptosis” was coined to describe another form of regulated cell death. [16]

a treatment for fatty liver disease 

Nonalcoholic fatty liver disease was induced in rats by feeding a diet high in starch and fat. [15] Unlike the fish studies in which cuprous nicotinic acid was added to the feed, 400 μg/kg cuprous nicotinic acid was given as an intragastric injection in 25% Tween-80 in saline. [15] Cuprous nicotinic acid decreased the presence of liver enzymes in the serum. [15] Anti-oxidant enzyme were also restored to control levels. [15] Fatty liver disease in another rat model was induced with a methione/choline deficient diet. [16] Rats in this study were given 0.043 cuprous nicotinic acid at mg/kg body weight, 3 times/week, for 4 weeks.  Rats on cuprous nicotinic acid and MCDD showed a decrease in hepatic transaminases in the serum at the end of the study. [16]

  1. Safety and Efficacy of Cunermuspir on Energy, Strength, and Fatigue in Patients With Nerve or Muscle Pain.  URL https://clinicaltrials.gov/ct2/show/NCT04737278
  2. El-Saadani MA.  (2004) A combination therapy with copper nicotinate complex reduces the adverse effects of 5-fluorouracil on patients with hepatocellular carcinoma. J Exp Ther Oncol. 2004 Apr;4(1):19-24. PMID: 15255289.
  3. Abo-Ghanema I, Saad TT , El-Bahrc SM, Fayza AIE.  Some Studies on the Effect of Copper (I) Nicotinate Complex on the Immune Response and Some Biochemical Parameters of Mugil cephalus Fish Vaccinated With Yersinia Ruckeri Bacterin J Arabian Aquaculture Soc. 2007 2(2)105-130  [Google Scholar] free paper
  4. Saad TT, El-Meadawy Z.Kh.Ali MA, Fayza AIE.  Some Studies on the Effect of Copper (I) Nicotinate Complex on the Immune Response and Some Biochemical Parameters of Sea Bass Fish Vaccinated With Yersinia Ruckeri Bacterin World’s Vet. J. 2013 3(4): 74-81 [Google Scholar]
  5. Musa S.A., Hafez A.H., Nassar A.Y., Gohar M.A. (1987) Copper(I)-Nicotinic Acid Complex: An Immunopotentiator in Chickens Vaccinated Against Newcastle Disease. In: Sorenson J.R.J. (eds) Biology of Copper Complexes. Experimental Biology and Medicine, vol 16. Humana Press. 1987 [Google Scholar]
  1. Al-Salahy MB. (2011) Physiological studies on the effect of copper nicotinate (Cu-N complex) on the fish, Clarias gariepinus, exposed to mercuric chloride. Fish Physiol Biochem. 2011 Sep;37(3):373-85.PMID 20927584  DOI: 10.1007/s10695-010-9437-0
  2. Medhat Hegazy A, Hafez AS, Eid RM. (2020) Protective and antioxidant effects of copper-nicotinate complex against glycerol-induced nephrotoxicity in rats. Drug Chem Toxicol. 2020 May;43(3):234-239. PMID 29944001 DOI: 10.1080/01480545.2018.1481084
  3. Nassar MA, Ali AM, El Din H, Omar M. (2018) Comparative plasma phenylalanine and glutamate profiles in scalded rats treated with Mebo or Copper (I) Nicotinate Complex. Ann Burns Fire Disasters. 2018 Dec 31;31(4):329-334. PMID  30983934
  4. http://en.mebo.com/Detail/engoods/id/296/aid/7616.html
  5. Nassar MA, Eldien HM, Tawab HS, Saleem TH, Omar HM, Nassar AY, Hussein MR. (2012) Time-dependent morphological and biochemical changes following cutaneous thermal burn injury and their modulation by copper nicotinate complex: an animal model. Ultrastruct Pathol. 2012 Oct;36(5):343-55.  PMID   23025652   DOI: 10.3109/01913123.2012.685687
  6. el-Saadani MA, Nassar AY, Abou el-Ela SH, Metwally TH, Nafady AM. (1993) The protective effect of copper complexes against gastric mucosal ulcer in rats. Biochem Pharmacol. 1993 Sep 14;46(6):1011-8. -j. PMID 8216343 doi: 10.1016/0006-2952(93)90665
  7. Tuorkey MJ, Abdul-Aziz KK. (2009) A pioneer study on the anti-ulcer activities of copper nicotinate complex [CuCl (HNA)2] in experimental gastric ulcer induced by aspirin-pylorus [corrected] ligation model (Shay model). Biomed Pharmacother. 2009 Mar;63(3):194-201PMID: 18343628DOI: 10.1016/j.biopha.2008.01.015
  8. Abdel-Mohsen MA, Abdel Malak CA, Abou Yossef MA, El-Shafey ES. Antitumor Activity of Copper (I)-Nicotinate Complex and Autophagy Modulation in HCC1806 Breast Cancer Cells. Anticancer Agents Med Chem. 2017 Nov 24;17(11):1526-1536. https://pubmed.ncbi.nlm.nih.gov/28356017/
  9. Abdel-Mohsen MA, Abdel Malak CA, El-Shafey ES. (2019) Influence of copper (I) nicotinate complex and autophagy modulation on doxorubicin-induced cytotoxicity in HCC1806 breast cancer cells. Adv Med Sci. 2019 Mar;64(1):202-209. PMID 30798072  DOI: 10.1016/j.advms.2018.08.014
  1. Abdel-Mohsen MA, Toson EA, Helal MA. (2019) Oncostatic treatment effect of triple negative breast cancer cell line with copper (I)-nicotinate complex. J Cell Biochem. 2019 Mar;120(3):4278-4290 PMID 30362603   DOI: 10.1002/jcb.27713
  2. Cohen L, Humeidi R, Spangler RD, Eaton JK, Frenkel E, Kocak M, Corsello SM, Lutsenko S, Kanarek N, Santagata S, Golub TR. Copper induces cell death by targeting lipoylated TCA cycle proteins. Science. 2022 Mar 18;375(6586):1254-1261. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9273333/
  3. Kahlson MA, Dixon SJ. Copper-induced cell death. Science. 2022 Mar 18;375(6586):1231-1232.
  4. Salama RH, Nassar AY, Nafady AA, Mohamed HH. (2007) A novel therapeutic drug (copper nicotinic acid complex) for non-alcoholic fatty liver. Liver Int. 2007 May;27(4):454-64.   PMID 17403185   DOI: 10.1111/j.1478-3231.2007.01460.x
  5. Hegazy AM, Farid AS, Hafez AS, Eid RM, Nasr SM. (2019) Hepatoprotective and immunomodulatory effects of copper-nicotinate complex against fatty liver in rat model. Vet World. 2019 Dec;12(12):1903-1910. PMID  32095039 PMCID: PMC6989322