Pre-Clinical Trials

Anti-Cancer Activity: Cells, Tumors and Cathepsin B

Amorphical has conducted several studies that examined the influence of ACC on cancer cell lines and animals with cancerous tumor models.

 

Cancer Cell Line Studies

The cancer cell studies focused on the influence of ACC on changes in genetic expressions, metabolic effects and Cathepsin B activity.

The studies were conducted on multiple human cancer cell lines (we tested 7 different types of cancer cells: lung, prostate, colorectal, cervical, hormonal breast, triple negative breast, and blood). The cell lines were grown in a culture medium that contained ACC, while the control group was placed in an identical medium without ACC. At the end of the culturing period, we examined and compared the expression levels of the various genes (genes whose expression was significantly increased/decreased in the ACC group compared to the control group).

For all tested cell types, we found significant changes in genetic expressions when treated with ACC compared to those grown without ACC. The genetic expressions shifted toward anti-cancer activities, enhancing the immune system’s ability to act against the tumor.

Another parameter we examined was the activity of Cathepsin B – a cysteine protease (protein degrading enzyme) active in acidic environments, which cancerous tumors use to advance and metastasize. The results of the studies showed inhibited activity of this enzyme in all the cancer cell lines treated with ACC.

In another study, performed on a mouse breast cancer cell line, the cells were grown in culture media containing ACC, calcium chloride (CaCl2) and CCC. The metabolic pathway of the cells was then tested by “Seahorse” XF Analyzer, which indicates the metabolism pathways. The results showed that the cancer cells, which had been exposed to ACC, changed their metabolic activity towards oxidative phosphorylation, while the cancer cells exposed to calcium chloride and CCC were more prone to perform a metabolic rout called glycolysis, which is characteristic of cancer cells and states of decreased oxygen supply. Increased glycolytic activity, also known as the “Warburg Effect”, usually occurs in oxygen-free environments, but in cancer cells it also appears in the presence of oxygen and is therefore one of the markers for cancer cells; in contrast, oxidative phosphorylation is more characteristic of healthy cells.

 

Animal Tumor Model Studies

Amorphical has conducted studies involving mice with a subcutaneous model of lung cancer (mouse Lewis lung carcinoma), which also measured the activity of Cathepsin B – a cysteine protease active in acidic environments, which cancerous tumors use to advance and metastasize.

The main findings of the studies were:

1. ACC decreases the growth rate of lung cancer type tumor in mice (similarly to conventional chemotherapy).

2. Administration of ACC combined with chemotherapy decreases the tumor growth rate better than either of the treatments administered separately.

3. Treatment with ACC causes statistically significant decrease in Cathepsin B activity.

Another study was performed on “nude” mice whose bodies do not reject implants due to an inhibited immune system. These mice were implanted with  a subcutaneoustumor of human non-small cell lung carcinoma cells.

Main finding:

Treatment with ACC significantly reduced the tumor growth rate in mice with human lung carcinoma.

Another study involved mice with a model of breast cancer bone metastases. In this model, breast cancer cells were injected directly into the bones. It is an extremely aggressive and lethal model of cancer. Treatment with ACC increased the survival of mice with bone metastases.

 

Anti-Inflammatory Activity

Amorphical has conducted preclinical studies in rats and mice with model of chronic inflammatory diseases including rheumatoid arthritis (RA) and multiple sclerosis (MS).

The findings were:

1. In rats with a model of RA, treatment with ACC was found to reduce the activity of Cathepsin B and Cathepsin K in the joints. Cathepsin K is an enzyme active in the bones, and particularly in the joints, whose upregulation is associated with exacerbation of RA symptoms. Cathepsin B was also found to be associated with exacerbation of RA symptoms.

2. In mice with a model of MS, treatment with ACC was found to reduce the activity of Cathepsin B in spine tissues. The general clinical condition of the mice treated with ACC was better as well. Mice that received a combination of ACC and Copaxone showed a clinical improvement in their motor abilities (same as with ACC alone) and a delay in symptom onset (same as with Copaxone alone).

 

Muscular Dystrophy

Amorphical has conducted preclinical studies in a mouse model of Duchenne muscular dystrophy (DMD)–a genetic disease that causes muscle weakness and abnormal muscle activity, increasing the risk of muscle damage.

Main findings of the study:

1. ACC helps improve muscle activity, including improved intracellular morphology and contraction.

2. ACC helps reduce Creatinine Kinase (CK) – a marker for muscle damage.

3. ACC helps improve physical ability.

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