Interview with researchers 4 Deployment of Cyclodextrins into the Medical Field

Successful development of a material for artificial blood from cyclodextrins

The blood donation and transfusion system in Japan is one of the safest in the world, and it would not be an exaggeration to say that it supports the healthcare system and fosters the health and well-being of the Japanese people. However, the shelf-life of red blood cell products is only about three weeks, leaving the crisis management system vulnerable when faced with disasters and other catastrophic events. What’s more, maintaining a steady supply to the blood transfusion system is also becoming more difficult due to the declining birthrate in the aging society of Japan. Due to these concerns, developing artificial blood that can successfully perform the oxygen-carrying function is of great interest. Dr. Hiroaki Kitagishi, a professor at Doshisha University, is engaged in developing such artificial blood.

The impetus for pursuing this path came when he met and studied under Professor Koji Kano (Currently a Professor Emeritus at our institution) while he was an undergraduate. Prof. Koji Kano was engaged in the research of cyclodextrins. Cyclodextrins are oligosaccharides with a cyclic structure comprising of multiple glucose subunits, and perform functions like improving the solubility of hydrophobic molecules in water through molecular inclusion and protecting materials that are reactive to water and oxygen. These properties are applied in deodorant and aromatic agents, where small molecules that become the odor source are bound to prevent their volatilization, and as food additives to stabilize materials that are otherwise unstable to heat and light. Prof. Kano focused on the characteristics of a cyclodextrin and came up with the idea of binding it to the coenzyme of hemoglobin called heme, to develop artificial hemoglobin, hemoCD, which can function as an oxygen carrier. Prof. Kano’s research was eventually passed down to Prof. Kitagishi.

It has been long accepted that when heme is exposed to water without being bound to a protein, the catalytic action of water will cause it to oxidize and lose its oxygen-binding ability. For this reason, it was assumed that it would be difficult to create artificial blood with only synthetic compounds that would have the ability to bind with oxygen in water. However, Prof. Kitagishi thought that as long as water and heme are prevented from coming into close contact through the inclusion of cyclodextrin, it would be possible to maintain the binding with oxygen. After many trials and errors, he succeeded in developing hemoCD. “The excitement I felt when I succeeded in developing a compound that functions in water which my predecessors were not able to achieve is a feeling I could never forget for the rest of my life,” he states.

Traditionally, artificial blood is created from expired blood meant for blood transfusions by purifying the hemoglobin and covering it with a synthetic compound that can act as a cell membrane. It is extremely rare to see examples of artificial blood made entirely out of synthetic compounds, such as what Prof. Kitagishi’s research group had created. “The hemoCD we developed has a small molecular size, which can be an issue as it tends to be easily filtered by the kidneys. As we advance, we are looking to solve this issue by binding the hemoCD to larger molecules,” he explained their vision for further research.

The discovery of a carbon monoxide removal method once thought impossible

Beginner’s luck is an expression used to describe how neophytes are, at times, blessed with good luck. A similar word to this is serendipity, which is a word that describes the phenomenon in which one unintentionally finds something of value while looking for something else. Occasionally, in research activities, unexpected discoveries open new paths of research.

When Prof. Kitagishi was working on developing artificial blood based on hemoCD, he observed the reaction of hemoCD during animal testing, and found that hemoCD was filtered out by the kidneys and quickly excreted. When he investigated the condition of the hemoCD in the urine, he found that hemoCD was bound to quite a bit of carbon monoxide, an unexpected discovery showing that it was selectively capturing carbon monoxide in the blood. Furthermore, he also discovered that this process triggers the decomposition of heme to generate carbon monoxide, as if to compensate for the carbon monoxide removed from the body. “It has been thought that endogenous carbon monoxide has a physiological function. However, we have proven for the first time that there is an internal mechanism within organisms that compensates for its shortage; a finding that strongly indicates that carbon monoxide is necessary for living organisms,” he reports.

In biology and genetic engineering, a technique through which genes are made inoperative in organisms (gene knockout) is used to elucidate physiological functions. However, when such a technique is used to determine carbon monoxide’s physiological function, such as stopping the activity of enzyme heme oxygenase—the source of carbon monoxide generation—it was found that heme would not degrade and began to accumulate and exhibit its toxicity. For this reason, it has not been possible to get a precise method to discover the physiological function of carbon monoxide in vivo. Prof. Kitagishi aims to elucidate the physiological function of carbon monoxide by utilizing the characteristics of hemoCD.

“HemoCD is not limited to becoming a tool to elucidate the physiological function of carbon monoxide. Since hemoCD does not remain in the body, it is proven to be an effective antidote to carbon monoxide poisoning,” says Prof. Kitagishi, with a vision to expand the use of hemoCD into the medical field as well.

The potential of hemoCD

In recent studies, carbon monoxide has been found to have anti-inflammatory and inhibitory effects on rejections during organ transplantation. HemoCD developed by Prof. Kitagishi may be able to assist in scientifically proving such functions. Furthermore, Prof. Kitagishi has begun developing a cyclodextrin that is capable of permeating the cell membranes, which will allow for hemoCD to penetrate the cells.

He speaks of his motivation for his research: “The reactivity and effect of plenty of drugs can be proven in a flask, but these drugs do not function well in practice as they fail to enter the cells. If we succeed in penetrating the cells using the cyclodextrin we just developed and have it release the bound molecules, this will likely lead to developments in a variety of research fields.”

Furthermore, Prof. Kitagishi is collaborating with other overseas researchers. “Joint research and exchanges between researchers provide important opportunities for ideas and inspirations to bubble up. In the past, when a joint researcher told me that the ability of cyclodextrin to assist molecules in penetrating cell membranes might be prove the use of cyclodextrin as an agent in assisting with impregnation during fertility treatment, and perhaps aid in resolving the social issues relating to the declining birthrate in an aging society, or with those giving birth later in life, my imagination went wild.” We can expect Prof. Kitagishi’s research to further evolve and develop through collaborations that transcend the university’s confines.