Rattlesnake venom as a potential cancer treatment

 

Myotoxin-3 from the Pacific Rattlesnake Crotalus oreganus oreganus Venom Is a New Microtubule-Targeting Agent written by: María del Carmen González García Clément Radix Cédric Villard Guillaume Breuzard Philippe Mansuelle Pascal Barbier Juan Luis Valérie Nicolas Bruno O. Villoutreix Dominique Bagnard

The goal of this research was to investigate the potential of Mycotoxin-3 (Mt-3) from the rattlesnake Crotalus oreganus and its venom is a microtubule-targeting agent since microtubules are essential for the cytoskeleton playing a critical role in cell division and intracellular transport. The use of pacific rattlesnake venom is due to its high activity in an in vitro microtubule polymerization assay. They state that several microtubule targeting agents have been developed as cancer therapeutics because of their ability to disrupt microtubule dynamics and induce apoptosis. 
The pacific rattlesnake's venom has been found to hold a variety of bioactive molecules with potential therapeutic applications, one being mytoxin-3 which has been known to have anti-tumor properties in vitro. The objective of the research is to characterize the effects of MT-3 on a microtubule dynamic and investigate its potential as an anti-cancer agent which leads them to hypothesize that the MT-3 toxins found in the venom of Pacific Rattlesnakes, will affect cancer cells. 

Method: 

This article used a combination of vitro and vivo methods to study the effect of MT-3 on microtubules. They extracted MT-3 from the venom and tested its ability to affect microtubule polymerization in vitro using purified tubulin, its effect on microtubules in living cells using confocal microscopy, and used the zebrafish model to investigate the vivo toxicity and therapeutic potential in MT-3 by treating zebrafish embryos with MT-3 and monitored for developmental abnormalities and mortality. They studied the potential of MT-3 as an anti-cancer agent using in vitro assay with cancer cell lines. 

Results:

As a result, using the RP-HPLC to fractionate the venom that has been tested each fraction for its ability to alter tubulin polymerization, found that all the activity was present in fraction B4, which was further sub-fraction contained two peaks that were separated by RP-HPLC which contains a compound that increases tubulin polymerization which can then reduce the lag time before polymerization in a dose-dependent manner. 

Figure 1. shows the purification of MT-3 from Crotalus oreganus venom with the fractionated using the preparative RP-HPLC, and the activated B4 was then divided into sub-fraction with the active sub-fraction, B4-24, and its two main peaks talked about above. The graph also shows the increase in turbidity during the tubulin polymerization assay. 

During the Amino Acid Sequence Determination, the results shows that peptide peaks 1 and 2 and peptide peak 1 were sequences using in-source decay MALDI mass spectrometry, which revealed that it was MT-3, a 45-residue peptide belonging to the crotamine-like peptides that are found in crotalids. They also found that the remaining moiety of MY-3 was identified as YKRCHKKGGH, which are 10 N-terminus amino acids that were to be calculated with the monoisotopic mass of 1212.89 DA. While the second peak was subjected to N-terminal Edman sequencing which resulted in similar to MT-3 with 6/7 unidentified residues which were found to correspond to cysteines that cannot be detected due to the endangerment in disulfide bonds. and the 7th was an undefined residue due to the lysine because of the residue that was conserved in all published sequences of the crotamine family. 

Figure 2 represents the amino acid sequences of MT-3 and its isoforms with the residues in position 5 which are highlighted in bold with the blue representing the first peptide that corresponds with MT-3 while the peptides in black represent peak 2 with histidine and other leucine at position 5. The figure also shows the calculated experimental masses of each peptide.

Discussion: 

The findings of MT-3 protein toxins from the venom of the Pacific rattlesnake target the microtubules and their potential cancer treatment. Microtubules are protein filaments that play a critical role in cell division and are a common target for cancer drugs. This research was able to conclude that MT-3 binds to microtubules in vitro and distributes their assembly and then used a variety of cellular assays to show that MT-3 inhibits cell proliferation and induces cancer cell death as well as inducing drug resistance cell lines. They also stated that MT-3 was to be found in non-toxic to noncancerous cells suggesting that it could have a favorable therapeutic index. The reason for such is because of the MT-3 Mechanism of action that is possibly due to the binding of the colchicine site on the microtubules which are a target for several microtubules targeting drugs however they said that to go further in this or confirm such theory they, they would have to do further research to grasp a better understanding. 

They think that there is a potential clinical application of MT-3 noting that other snake venom proteins can also have some kind of similar effect so there is a potential in developing other drugs such as captopril, which are proteins found in the venom of the Brazilian pit viper. Overall, they concluded that MT-3 is a very promising protein agent that warrants further investigation as a potential anticancer drug.

Article:

González García, M. C., Radix, C., Villard, C., Breuzard, G., Mansuelle, P., Barbier, P., ... & Luis, J. (2022). Myotoxin-3 from the Pacific Rattlesnake Crotalus oreganus oreganus Venom Is a New Microtubule-Targeting Agent. Molecules, 27(23), 8241.