The vaccine can prevent millions of deaths each year and push certain diseases to the brink of survival. However, it turns out that developing effective vaccines against multiple viruses is extremely challenging, if not impossible.
Despite decades of research, there are no approved vaccines that can prevent influenza virus, Epstein Barr virus (EBV) or respiratory syncytial virus (RSV) for long periods of time. For example, for RSV, a pathogen that causes respiratory infections, high-risk infants receive a monthly synthetic antibody injection to protect these infants in the short term.
In recent years, several groups of researchers have explored a different approach: using the immune system to produce antibody-producing B cells, and using CRISPR to modify them, in order to express large amounts of antibodies against these difficult-to-treat viruses as required in the case of infection.
In a new study, researchers from the Fred Hutchinson Cancer Research Center and the University of Washington in the United States succeeded in genetically modifying human and mouse B cells to express targeted HIV, EBV, and influenza virus and RSV antibodies. They also confirmed that these genetically engineered B cells blocked RSV infection in a mouse infection model. The results of the study were published in the journal of Science Immunology.
Branden Moriarity, a genomics engineer at the University of Minnesota, who did not participate in the new study, said, " This is a concept that people have been thinking for a long time."
In order to modify B cells, the immunologist Justin Taylor of the Fred Hutchinson Cancer Research Center and his team explored antibodies that have been shown to be effective against each of these four viruses in previous studies. They used a conventional CRISPR strategy to insert the antibody-encoding DNA into a small incision made on the antibody-coding gene of primary human B cells.
These researchers designed this insertion so that expression of the new antibody protein is regulated by the promoters of these cells, allowing these cells to produce them as normal after being subjected to viral antigen triggering. They will, as much as possible, differentiate these modified B cells into antibody-secreting plasma B cells that express these antibodies on their cell surface and secrete them.
Once these researchers confirmed that a reasonable proportion of these modified B cells did express these new antibodies, they investigated whether these cells were resistant to viral infection in mice. They repeated this process in mouse B cells, inducing them to express antibodies targeting RSV, perfusing successfully modified B cell populations into normal rodents, and then waiting for a period of time.
Six days later, RSV-specific antibodies appeared in their serum but did not appear in control mice. Some of these mice received an intranasal dose of RSV injection and these modified B cells successfully resisted the virus. Five days later, these researchers detected almost no level of RSV in the lungs of mice with these modified B cells in their blood. However, they found high levels of this virus in control mice that did not contain these B cells.
Taylor said he believes these modified B cells may be useful for patients who have undergone hematopoietic stem cell transplantation. These patients usually take immunosuppressive agents and are very susceptible to viral infections. RSV infections suffered by these patients can be prevented by injections of synthetic antibodies such as palivizumab, but such injections must be received on a regular basis. On the other hand, modified B cells theoretically only need to be injected once.
To investigate whether engineered B cells might provide protection in this setting, the researchers studied mutant mice lacking T cells and B cells. After they injected these cells into these mutant mice, they were surprised to find that RSV-specific antibodies in these mutant mice appeared to stay in the blood for a longer period of time than wild-type mice. In fact, immunodeficient mice were able to prevent RSV infection 82 days after receiving these engineered B cell transplants. Taylor explains why this is something they need to explore more.
In general, the approach taken by the Taylor team is similar to that used in other recent studies, such as a study published in the eLife journal (eLife, 2019, doi:10.7554/eLife.42995) and published in the Journal of Experimental Medicine. A study in the journal (Journal of Experimental Medicine, 2019, doi: 10.1084/jem. 20190287), both of which use CRISPR to construct B cells expressing HIV neutralizing antibodies. But the difference is that the current new study addresses some of the key challenges in the process of transforming antibody-encoding genes.
This is a tricky thing: First, antibodies are synthesized by two genes—one for the heavy chain encoding the antibody and one for the light chain of the antibody. Once the heavy and light chains are translated into proteins, respectively, they fuse together to form the final antibody.
The challenge in editing these genes is that mismatches can sometimes occur, resulting in hybrid antibodies consisting of the engineered heavy chain paired with the B cell's own light chain. In some cases, this can lead to an autoimmune response. Taylor explained, “This risk is very low...but, given that millions of B cells have millions of unique light chains, this risk is not worth the risk.”
The Taylor team solved this problem by physically joining together the DNA encoding the heavy and light chains and inserting the ligated DNA into a single CRISPR cleavage site. (Other research groups have devised different strategies to solve this problem, such as knocking out the light chain-encoding genes of B cells themselves.)
Richard James, an immunologist at the Seattle Children's Institute in the United States (not participating in the new study) commented, "I think the genetic modification strategy they use to fuse light and heavy chains is a good idea."
However, he added, “In practice, this would be an expensive treatment,” and noted that personalized CAR-T cell therapy has been thought to cost close to $500,000. For the time being, B-cell therapies must be personalized: Based on the expression of individual cell surface antigens in these cells, cells from one donor will suffer from immune rejection in different recipients. He added, "Unless we can find a way to build allogeneic B cells, it will only become more cost effective."
Moriarity agrees. He said, "Only when you have a patient with a disease you will die with and there is no other treatment," prophylactic use of engineered B cells may be just a viable treatment option. However, he said, the new study offers other possibilities for future research and is one of the first studies to modify antibodies in mouse B cells. For scientists studying B-cell differentiation and cell- and immune-related processes, "I think this will make a huge contribution to the field of basic biology."
Michael Goldberg, a former immunologist at Harvard University and now CEO of STIMIT, an immunotherapeutics startup, believes the new study is "a good extension of previous research." For him, B cell antibody modification has other advantages besides preventing viral infections: for example, antibody levels that respond to antigen concentrations in the blood may be used to treat diseases such as rheumatoid arthritis, among which Rheumatoid arthritis is usually treated with antibodies that neutralize the cytokine TNFα.
However, "the duration of antibody expression needs to be increased. In this regard, it will be important to strengthen our understanding of the transplantation process and B cell culture conditions. However, given that these are the first to involve the retrofitting of engineered B cells. The preliminary results of the experiment on sexual transfer are encouraging."
Taylor agrees that there is still more research work to be done. He said, "One thing we completely ignore in this research is to really assess the safety of what we are doing and to ensure that no off-target effects occur. I think this has always been a concern for CRISPR-Cas9. Before using it for human body treatment, we must absolutely determine that it is safe. ”