Getting a Grip on Streptococcus

If biotechnology as a science and as an industry has "grown up" over the past 20 years, the search for a vaccine against the Streptococcus A bacterium may be described as "coming of age." The bacterium presents a serious challenge. In children, it infects the mouth, nose, and throat and can cause the painful inflammation of strep throat. But it can have more dire consequences--if an infected child doesn't receive adequate antibiotic treatment, the infection could find its way into the bloodstream. The result is an antibody response that goes beyond the mouth and nose, with antibodies against Streptococcus A circulating in the bloodstream. At least some of those antibodies are cross-reactive to human tissue in the heart, kidneys, and brain, and they may cause heart damage known as acute rheumatic fever. Milan Blake, senior director of molecular biology and protein chemistry at North American Vaccines, is pictured with his wife Christa and children Richard and Susan. In the 1960s and '70s, several research groups developed vaccines that used killed or inactivated bacteria, but some troubling observations caused a backlash that continues to this day. One study performed by Benedict Massell gave a vaccine consisting of a partially purified Streptococcus A surface protein to 21 children whose siblings had developed rheumatic fever. About 18 months later, two of those children also developed rheumatic fever. Experts disagree about the validity of those findings. Siblings of children who had already had rheumatic fever may have had a genetic predisposition toward it. There was no way to be certain the rheumatic fever cases stemmed from the vaccine and not a Streptococcus A infection, says Fran Rubin, program officer for group A streptococci at the National Institute of Allergy and Infectious Diseases. "The results have had a tremendous impact," she says. The impact is still being felt. Vaccines for Streptococcus A infection went to the market anyway, but Massell's findings and others led the Food and Drug Administration to ban interstate sale of those vaccines in 1979 and call for more careful clinical evaluation of future candidates. It still isn't entirely clear how the bacterial infection leads to an autoimmune reaction, but there is some clinical and laboratory evidence to suggest that the culprit is the M protein, a surface protein on the bacteria. Most researchers believe that an epitope on the M protein induces an antibody response that is cross-reactive to an epitope on the heart, liver, kidney, and brain, where the secondary damage of rheumatic fever occurs. An epitope is a marker that triggers specific antibodies to eliminate the bacteria. "My guess is that there were interrelationships" between Massell's vaccine, the ongoing infections, and the development of rheumatic fever in two patients, speculates Rubin, "but we don't really know the mechanism of pathogenesis." Still, there appears to be good news on the horizon. Molecular biology--a neonate in 1979--grew up in the intervening years, and researchers have since picked apart the M protein, searching for the epitope that induces cross-reactive antibodies. They think they've found it, and several companies are developing subunit vaccines that eliminate the offending epitope. FDA is considering removing the restriction as several companies gear up for what promise to be more stringent vaccine clinical trials. There is plenty of incentive. In the United States, strep throat alone costs more than $2 billion to diagnose and treat, and the heart damage caused by rheumatic fever extracts a much greater toll. Recent studies have shown that Streptococcus A is also the cause of invasive fasciitis, better known as the flesh-eating disease. M Protein Ironically, though the M protein's cross-reactivity appears to have caused the defeat of those early vaccines, it has also attracted the most attention as researchers develop a new generation of vaccines. The protein itself is a highly variable surface protein that seems to divert the attention of the immune system by binding to antibodies nonspecifically and taking on a coat of antibodies. In this way, it prevents the immune system from recognizing the bacteria as foreign. James B. Dale, professor of medicine and chief of infectious diseases at the Veterans Administration Medical Center and the University of Tennessee, Memphis, has teamed with ID Biomedical Corp. of Vancouver, B.C., to develop a vaccine based on the M protein's variable regions. By linking immunogenic portions of the most common and virulent Streptococcus A strains, the company hopes to form a protein antigen that provides broad immunity. ID Biomedical has already begun Phase I trials with the vaccine formulation and hopes to begin Phase II trials some time next year. But the M protein's variable regions have a potential drawback as the basis for a vaccine. There may be more than 100 serotypes, each with variations in the M protein. It may be difficult to find a combination of epitopes that provides sufficient immunity to all the different strains of Streptococcus A bacteria. Manhattan's Siga Pharmaceuticals Inc. is working with Vincent Fischetti of Rockefeller University to develop vaccines against epitopes in the conserved region of the M protein. Such a vaccine could theoretically provide immunity to most if not all subtypes of Streptococcus A. And unlike other approaches that use an injected vaccine, Siga's strategy uses as its carrier Streptococcus gordonii, which infects the oral cavity without causing disease. Oral administration has the added advantage of inducing a mucosal immune response and could give protection against initial infection. "In many ways I think that's what you want, because ... [children] contract the disease by respiratory infection, through droplets they inhale," says Dennis Hruby, vice president of research at Siga. The vaccine also causes a systemic response that is almost as strong as is usually seen with an injected vaccine. Siga is now completing Phase I trials using the vector alone and hopes to begin trials with the engineered bacteria by the first of the year, says Hruby. Dennis Hruby Beyond M Other companies have decided to avoid the M protein altogether. Aside from its potential immunogenicity, companies such as North American Vaccine Inc. of Columbia, Md., believe that other markers on the surface of the bacteria may confer wider protection. Although the M protein has been implicated in the autoimmune response that leads to rheumatic fever, the overall immune response is far more complicated. Most adults are immune to strep infections, yet studies show that only about 1 percent of the population expresses antibodies against the M protein at all, says Milan Blake, senior director of molecular biology and protein chemistry at North American Vaccines. And while children between ages 5 and 7 are most susceptible to infection, that vulnerability seems to wane gradually over time. All of which implies that although the M protein can apparently elicit dangerous cross-reactivity to produce rheumatic fever, it may not be the key immunogenic antigen. On a hunch, Blake and fellow University of Rockefeller researcher John Zabriskie started searching for antibodies against Streptococcus that increase over time. They found one--an inherited antibody against the polysaccharide on the surface of Streptococcus A (group A polysaccharide). The company has formulated a vaccine based on group A polysaccharide, hoping to kick-start this natural immunity, says Blake. Although it has not yet entered clinical trials, a study conducted in Mexico supports the approach. In that patient group, children with low amounts of polysaccharide antibodies tended to have Streptococcus A infections, while those with a higher count generally did not. Researchers followed those observations with in vitro tests that showed that phagocytes could destroy the bacteria in the presence of the anti-group A polysaccharide antibody. When the antibody was removed from the culture using affinity chromatography, the phagocytes had no effect. Blake reports that the company is close to entering clinical trials with the vaccine, but more preclinical work remains to be done. Peptidase A team headed by P. Patrick Cleary, a professor of microbiology at the University of Minnesota Medical School, has found another protein that, like the M protein, helps shield the bacteria from the immune system. In the mid-'80s, one of Cleary's graduate students did an experiment to see if Streptococcus A would activate the complement pathway, using the chemokine C5a as a marker. "We got a big surprise that, yes, [Streptococcus A bacteria] activated the complement pathway, but there was no C5a present. It turned out that the bacteria had des-troyed it," Cleary recalls. Soon after, they did a little fishing and discovered the culprit--a surface peptidase that selectively cleaves C5a. Experiments in mice suggest that the activity of the streptococcal C5a peptidase gives the bacteria a stay of execution long enough for it to colonize mucosal membranes and establish itself. The protein is highly conserved, even outside the group A family of bacteria. Its sequence in group A strains is 98 percent identical to the sequence of protein in Streptococcus B bacteria, which can cause meningitis in infants. That raises the "pie in the sky" possibility--as Cleary puts it--that a single vaccine could provide immunity against both groups. To produce a vaccine, Cleary's team inactivated the protein by changing an amino acid in the active site and clipping off bits of the carboxy and amino termini. Because the vaccine doesn't involve the M protein, they haven't tested it for cross-reactivity, Cleary says, but they will do so before the vaccine goes into human trials. The vaccine has been licensed by Wyeth-Lederle Vaccines (a division of American Home Products) of Rochester, N.Y. Whether it stems from the M protein or one of the other approaches, the odds seem favorable that a vaccine against Streptococcus A will eventually become available. The FDA concedes that it's probably time for the restriction on sales of a Streptococcus A vaccine to be lifted from its books, says FDA spokesperson Lenore Gelb. If it is indeed stricken from FDA's books, it could be a signature victory for the biotechnology industry. James Kling (www.nasw.org/users/jkling) is a freelance science writer in Bellingham, Wash.

Getting a Grip on Streptococcus

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