Leveraging Hitachi's renowned capillary electrophoresis technology to advance cancer diagnosis—developing a new molecular diagnostic technology capable of detecting a large number of genetic mutations in a single assay

Optical technology and biotechnology experts join forces to detect genetic mutations with high sensitivity

Ando: I studied electronic engineering in university, where I researched the application of optics to the biotechnology field. I conducted joint research with a hospital-affiliated research center on the use of lasers in gene/drug delivery and the treatment of neurological diseases, earning a doctorate degree in engineering. I chose to work at Hitachi due to the opportunity to develop a range of new technologies without being limited to one specific field. For example, if I were to develop a new optical measurement technology, I felt that Hitachi is a company where my research could be applied broadly across the organization, not only in the fields of biotechnology and healthcare, but also in other infrastructure businesses where optical technology plays a key role, such as water quality testing or railcar inspection. I chose Hitachi―a company that does not exclusively focus on healthcare―because I was interested in the possibilities of combining multiple disciplines within a single company.

After joining Hitachi in 2013, I initially worked to apply electron spin resonance technology to cellular measurement for use in regenerative medicine. Following this, I worked on dispensing technology and the development of optical sources for blood analyzers used in health checkups. Since 2021 I have been working on this project, which is focused on genetic testing.

Manri:I majored in bioengineering at university, where I completed a master's degree. During my master's studies, I researched B cells, a type of white blood cell involved in immune response. B cells produce antibodies that bind to pathogen antigens, and an interesting characteristic of B cells is that by altering the stimuli applied to them, it is possible to produce antibodies that bind more strongly to antigens. I was hoping to work in the food industry, and when my professor told me that Hitachi was establishing a Life Science Group , I decided to join the company as I believed I could put the genetic engineering skills I had acquired at university to use. When I first joined Hitachi, I was responsible for analyzing the genes expressed in people who were ill or who had taken medications, based on requests by academia and pharmaceutical companies. I really felt that my work was contributing to the treatment of diseases, such as new drugs.

In 2007, I was transferred to the R&D Group, where I was assigned to the programming group. I was a little apprehensive as I had next to no experience in programming. In fact, I was such a novice that I misheard “Core 2” as “Kouatsu (high voltage)” when buying a computer. Even so, my team trained me to the extent that today I am able to program using the C programming language, and I believe that the opportunity to grow in this way is one of the best aspects of Hitachi. I developed an algorithm that differentiates blood cells from yeast cells in a urine sediment analyzer used to test urine constituents. Although the project unfortunately ended before completion and the technology was not commercialized, it was a valuable experience for me. Following this, I transferred to my current department in 2017.

Yokoi: I completed my PhD in cell physiology, where my research topic was the growth mechanism of plant cells. Cell growth has only two mechanisms, either by elongation vertically or by expansion horizontally, and I investigated how these mechanisms are regulated at the molecular level. Following this, I spent two years at a botanical research laboratory developing new flower varieties through genetic manipulation, before joining Hitachi as a mid-career hire in 2000.

At the time, Japan was working to analyze full-length cDNA as part of the Human Genome Project that aimed to sequence all human DNA (cDNA is short for complementary DNA, which is synthesized using mRNA as a template). This was a national project conducted by the New Energy and Industrial Technology Development Organization (NEDO), with Hitachi serving as the lead organization. I had the opportunity to participate in this project, where I conducted genetic analysis for approximately three years. I was also involved in a follow-up project related to functional RNA. During this period, I gained a wealth of knowledge from discussions with researchers at national institutions and universities, and formed a strong network of personal connections. I also conducted many joint research projects with food and alcohol companies.

Following this, I was seconded to Hitachi Software Engineering (former name), where I was involved in developing software for genetic analysis and the launch of a related service business. After rejoining the R&D Group, I conducted research on sample preparation, including the extraction and labeling of genes prior to genetic testing. Since 2018, I have been working on the development of a new gene assay that utilizes this electrophoresis technology.

Engineering ways to increase the number of gene mutations that react, and adapting reagents to increase sensitivity

Yokoi: The technology we have developed detects abnormal genes in cancer patients. Cancer is caused by damage to genetic material. With the development of molecular-targeted drugs designed to target specific genetic mutations during cancer treatment, it is becoming possible to investigate the genetic mutations in the cancer of individual patients and select a molecular-targeted drug best suited to their specific needs. However, in order to test for genetic mutations, at present most patients need to have tissue samples taken, and there are limitations to the number of genetic mutations that can be screened under insurance coverage as well as the timing at when such tests can be performed. In particular, gene panel testing, which uses a next-generation sequencer1 to test a large number of genes at once, is only covered by insurance for patients with advanced or recurrent cancer who are expected to complete standard treatment. Furthermore, this test can only be taken once and is very expensive.

1 Next-generation sequencer ：A device capable of analyzing DNA sequences at high speed and in large quantities, allowing a large number of DNA bases to be sequenced in a single assay.

The concept of our development project is to utilize biological fluids such as blood at an earlier stage via an assay technology that is easy to conduct, inexpensive, and can test a large number of genes in a single run.

Hitachi markets capillary electrophoresis sequencers2, which are widely used in hospitals in Japan, Europe, and the United States. Genetic testing using the aforementioned next-generation sequencers requires time-consuming processing of specimens and other preparations. Such processes are typically outsourced, which is costly and time-consuming. By contrast, capillary electrophoresis sequencing can be performed onsite at the hospital rapidly, easily, and inexpensively.

2 Capillary electrophoresis sequencer: A device that analyzes DNA nucleotide sequences and base lengths by electrophoresis inside a capillary tube.

Ando: During the development process, we investigated whether it was possible to detect multiple gene mutations simultaneously and with high sensitivity using capillary electrophoresis, such as EGFR gene mutations, which are common in lung cancer, and KRAS gene mutations, which are common in pancreatic and colorectal cancer (see figure below). By applying a reagent labeled with a fluorescent molecule and primers set to a given base length (synthetic single-stranded DNA with a complementary base sequence that binds to the DNA template) to cancer-associated genes, and by increasing the sensitivity of the signal detectable by the capillary electrophoresis sequencer, we were able to detect multiple genetic mutations with a sensitivity of 1%.
https://link.springer.com/article/10.1007/s44211-024-00508-8

Yokoi: What distinguishes our technology is the ability to detect the presence of subtle genetic mutations in a large number of target genes with a high level of sensitivity via capillary electrophoresis. By altering the template DNA, primer length, and reagents, I believe that this technology can potentially be applied in environmental DNA testing as well as human diseases, such as infectious diseases.

We hope to use this new technology we have developed to build a new genetic testing platform together with pharmaceutical, diagnostic agent, and testing companies.

Each team member overcame hurdles at respective stages of the test development process

Yokoi: With the genetic test we are developing, the greater the number of primers types used in the single nucleotide elongation reaction, the greater the number of genes that can be analyzed. As the primers used are differentiated by their length, we believed that if we could synthesize longer primers, this would enable a greater number of genes to be tested simultaneously than in the past. However, the length of primers that can be chemically synthesized typically hits a ceiling at around 200 bases. Although we established a concept of lengthening the primers by some hypothetical means, there were no examples of such techniques available in published literature, and despite spending two to three months within the team discussing methods of adding molecular weights to the primers, we were unable to find a solution. In order to conduct electrophoretic analysis, the size of the primers must be fully aligned, which makes this difficult to accomplish with typical chemically synthesized materials. One night while I was lying in bed, I suddenly struck on a new idea for adding molecular weights to primers. I immediately put it to the test and after a couple of weeks of experimenting, I found that it appeared to work. To address the challenge of aligning primer sizes, we studied ways to improve the purity of the material and the bonding portion of the primer. These innovations made it possible to manufacture primers with a length of approximately 600 bases.

Ando: I was primarily responsible for processing the data from the fluorescence signals emitted by the capillary electrophoresis sequencer. I struggled to find a way to distinguish whether the data received was a signal that indicated genetic mutations in cancer cells compared to normal cells, or if it was simply background noise. I found that the key to analyzing this data was to compare it against baseline data that did not contain mutations. Through discussions in the team, I developed ways to improve the sensitivity by adjusting the resolution of the signal or the amount of fluorescent molecules.

Manri: I was responsible for preparing the genes and other materials used in the research and development process. Commercially-available specimens used in genetic testing contain a mixture of several genes, so I spent approximately one year working on extracting and preparing the genes that we wanted. The research experience from my university days came to good use!

Yokoi: In order to develop measurement technologies, standardized materials are essential. For example, to detect genetic mutations with a sensitivity of 1%, you first need a specimen in which you can detect genetic mutations with a sensitivity of 100%. In that regard, this research project resembled that you might find in a life science laboratory.

Ando: It is only just in recent years that the measurement capability of capillary electrophoresis sequencer instruments themselves has improved, enabling stable measurement of longer primers, which has also helped to advance our detection technology.

Yokoi: While we have successfully demonstrated the concept of the technology and that it works, next we must examine the versatility and stability of this technology to determine which targets it can be used for and how effectively it can analyze them. To achieve this, we will need to further improve the technology, including increasing its sensitivity.

A team with a flat hierarchy where problems can be brainstormed and alternative approaches proposed

Manri: I can always count on Yokoi-san and Ando-san. Yokoi-san has an extensive knowledge of genes and the skills to work with them, so when I seek his advice on a problem, he thinks through it together with me and always suggests a positive next step. Meanwhile, Ando-san offers advice from a fresh perspective from his specialty in data analysis, which often opens my eyes to alternative approaches.

Ando: I leave the ideas and the project management to Yokoi-san, while I concentrate on tasks such as data analysis. I don't think I could replicate Yokoi-san’s ability to come up with ideas and then step on the accelerator to take these ideas to the next stage. I am also impressed by the way Manri-san follows each step of the process thoroughly, even those I might skip over myself. Our research output is a true team effort.

Yokoi: I always create optimistic research plans even if there is no clear outlook. As I am a bioscience researcher and do not have experience in engineering design, I ask my colleagues to handle this part of the project. Both Ando-san and Manri-san are very open about the areas they are having problems with or aspects they think should be changed. Sharing this feedback helps us make the necessary adjustments. The best things about the R&D group’s healthcare division, including this team, is that everyone comes from different backgrounds, and that we can discuss matters with mutual respect, regardless of hierarchy.

Expanding our R&D scope to other areas such as health preservation and agriculture

Yokoi: I’d like to be able to help healthy people stay healthy for longer. For example, it would be interesting if we could use a new metric to track the progression of aging, although whether we would use genes to investigate is another story.

Manri: I previously worked in the agricultural field and am interested in genes contained in the soil. My dream is to utilize genetic analysis to find ways control the bacteria in the soil and increase agricultural productivity.

Ando: Today, people commonly use instruments such as scales, thermometers, and blood pressure monitors to monitor their health at home. I believe that when it eventually becomes possible to conduct blood tests at home, we may see a new era in which people can use genetic information to manage their own health. While the technology we introduced today uses a special compact device to detect fluorescence, I imagine that as technology advances it will become possible to conduct measurement with everyday devices such as smartphone cameras or LED lights. I anticipate that personal health care will evolve and improve as technology that is currently only available in hospitals becomes available to individuals.