Featured image caption: Sea cucumbers are echinoderms that live on the ocean floor worldwide. Some species are also edible and serve as a rich source of nutrients. Photo credit: Anders Poulsen, Source: Wikimedia
Reference Article: Ru, R., Guo, Y., Mao, J., Yu, Z., Huang, W., Cao, X., Hu, H., Meng, M. and Yuan, L., 2022. Cancer Cell Inhibiting Sea Cucumber (Holothuria leucospilota) Protein as a Novel Anti-Cancer Drug. Nutrients, 14(4), p.786. https://doi.org/10.3390/nu14040786
Marine Life Magic
In general, marine invertebrates are a rich source of nutrients and extracts that can possess healing and even anti-tumor properties. For example, Aplidin® is an extract from a toxic sea squirt that is currently in clinical trials to treat pancreatic, stomach, and intestinal tumors. Sea cucumbers too have attracted significant attention for anti-inflammatory, anti-coagulation, and anti-bacterial effects. But first, what exactly is a sea cucumber? Despite their deceptively ordinary outward appearance, these organisms are fascinating ocean creatures. They move around the ocean floor eating algae, decaying organic matter, or small crustaceans. When threatened, sea cucumbers have a brilliant defense strategy: they forcibly eject their organs in order to surprise and confuse the predator. This shocking expulsion can serve as a temporary food source while the sea cucumber mounts a heroic escape. Thankfully, the organs then regrow a short time later. In some species, a toxic substance to harm predators can also be excreted.
Sea cucumbers have long been hallowed in Chinese medicine for their vast array of nutrients, including bioactive peptides. A bioactive peptide is a protein building block that serves a purpose beyond its nutritional value; it benefits body function or overall human health in some way. Holothuria leucospilota is an edible, tropical sea cucumber that is widely cultivated in China. Its body wall is composed of 70% protein and is as a source of rich bioactive peptides. Extracts from this particular sea cucumber have already demonstrated significant healing effects, leading Ru et al. to select this species for their study. The scientists recognized the difficulties in treating cancer, ranging from mutations to rapid cell division. Studies on the anti-cancer effects of H. leucospilota’s bioactive peptides are lacking, so the scientists set out to determine if these combined proteins possessed any benefits. Specifically, they wanted to investigate the effect of the proteins on several key process throughout a cancer cell’s life cycle, including growth, reproduction, and death.
Investigating the Inner Wall
The scientists first grew the sea cucumbers and then extracted the crude protein from the body wall of the organisms. To study anti-cancer properties, they then selected three cancer human cell lines (referred to here as HepG2, A549, and Panc02) and three “normal” human cell lines (referred to here as NIH-3T3, HaCaT, and 16HBE). The experimental treatment was the H. leucospilota body wall protein extract, while the control treatment was epirubicin hydrochloride (EPI), an established anti-tumor drug. The study is divided into smaller investigations; in each one, cultures of the six individual cell lineages received the experimental treatment, and separate cultures of each cell lineage also received the control treatment for comparison.
The effect of increasing concentrations (starting at 0.005 µg /L up to 10 µg /L) of the H. leucospilota protein on cell viability was first analyzed. Cell viability, or the proportion of healthy, living cells, can be monitored with a reagent kit that will turn viable cells a different color than inviable cells. All cell lines were treated and monitored for viability at 24 hr, 48 hr, and 72 hr. Although the results varied between the different cancer cell lines, H. leucospilota protein extract displayed cytotoxic effects for all three. In other words, the sea cucumber extracts decreased the amount of living cancer cells. The most promising results were seen in the Panc02 lines, where even the lowest concentration (0.005 µg /L) of the sea cucumber protein treatment significantly decreased cancer cell viability. H. leucospilota protein extract also decreased the viability of normal cell lines, but in all three cases, a higher concentration of protein was required to achieve the same negative effect. Therefore, the sea cucumber protein extract was more toxic to cancer cells than to normal cell lines.
Cease the Cell Cycle!
Most cells divide by going through a process called mitosis. Mitosis has four phases: G1, S, G2, and M. In G1, the cell begins to grow. During the S stage, the cell replicates its DNA. Next, in G2, the cell continues to grow and prepares for the final phase, M, or mitosis. Mitosis is cell division that results in two identical cells. Ru et al. investigated the effects of H. leucospilota on the various cell cycle stages of the selected lines. To do this, they used a procedure called flow cytometry. During this process, scientists suspend cells in a solution and pass them through a flow cytometer. This equipment beams lasers at the cells as their pass through; corresponding software provides information regarding cell characteristics and morphology based on how they fluoresce or scatter light. The EPI treatment disrupted the cell cycle in both the normal and cancer cell lines, whereas the sea cucumber protein treatment disrupted the cell cycle in only the cancer cell lines. In HepG2, the initial growth phase (G1) was reduced when cells were exposed to the protein. A549 cells witnessed a different effect: G1 was actually stimulated, but S and M stages were reduced. Decreasing the capability of cancer cells to complete cell division is a critical property of a bioactive compound intended to treat cancer.
Cease to Exist?
Ever wonder what exactly happens to old cells? They are destined for a process known as apoptosis, or programmed cell death. Cell destruction by apoptosis usually involves the shrinkage of the cell followed by disintegration. Using flow cytometry again, the effects of EPI and H. leucospilota protein on apoptosis of the cell lines were contrasted. EPI resulted in an increase in apoptosis of all the cell lines; the death rate was close to 60%. Interestingly, H. leucospilota only caused significant cell death in the cancer cell lines (again, around 60%). The normal cell lines when treated with the sea cucumber protein also experienced apoptosis, but at a lower rate (7.16%-29.65%). These rates are statistically the same as the death rate when the cells are treated with a blank control.
The proteins isolated from H. leucospilota cell walls show strong promise as an anti-cancer agent in this study. Future work will have to pinpoint specific modes of action as well as determine if there is a certain bioactive peptide responsible for the composite sample’s anti-tumor properties. Further trials in living subjects will follow, and a safe and effective drug delivery method will have to be developed. Regardless, organisms from the deep sea may hold the key to effective natural agents that will help in treating human cancers.