Bio

Research Interests: Comparative Pathology, Retroviruses

Although officially retired, Dr. Gardner remains an active and vital member of the CIID.  Dr. Gardner's major research interest for many years has been in the natural history of retroviruses in animals and man.  As part of the Virus Cancer Program (VCP) from 1968-1981 he and his colleagues at USC School of Medicine in Los Angeles discovered and characterized oncogenic retroviruses of wild mice and domestic cats and carried out extensive studies on the epidemiology and virology of human cancer.  This led to the understanding of a new biology of retroviruses in wild mice that proved a more accurate model than inbred mice for predicting the natural history of similar retroviruses (i.e. HTLV) discovered a decade later in humans.  Study of naturally occurring retroviruses in cats led to an appreciation of the horizontal transmission of feline leukemia virus among domestic cats and contributed to the eventual development of a vaccine that now largely prevents this feline disease.  This research also led to discovery and characterization of several novel oncogenes in feline sarcomas which are now potential targets for novel cancer therapy in humans. Endogenous inherited infectious but nonpathogenic retroviruses were discovered in cats, mice and rats.  Most importantly, in respect to public health, endogenous retroviruses, despite representing ~8% of the human genome, were found to not contribute significantly to human cancer, thus ruling out a major hypothesis of the VCP.   However, the VCP established the “blue print”, technology and teamwork that led to the current molecular/genomic era and facilitated the discovery of the immunosuppressive retroviruses that cause AIDS in humans and animals.  

Since moving to UC Davis in 1981, Dr. Gardner has taken part in the discovery and description of simian AIDS caused by Type D retrovirus and also by simian immunodeficiency virus (SIV, which is closely related to human immunodeficiency virus, HIV-1).  Research on the Simian AIDS model was indispensable in leading to the development of successful antiviral drugs that have changed AIDS from an inevitable fatal disease to a long lived chronic infection.  He and his colleagues have studied the origin, natural history, pathology and pathogenesis of these retroviruses and have carried out a number of experimental vaccine trials. A potentially efficacious human AIDS vaccine based on cytomegalovirus research in the Simian AIDS model is now in phase II human trials.  His present interest is directed at better understanding of the pathogenesis of AIDS and attempts to develop more efficacious and safe vaccines against AIDS using the SIV macaque model.

Publications

'FOR half a century a small, persistent and intrepid brand of scientists insisted, often at the top of their decibel range, that cancer is caused by man's tiniest and most mysterious natural enemy: the virus. Very few scientists ever listened; if they did, they generally objected. How can a virus cause cancer when doctors don't seem to get it from their patients, or from research animals, or from tissue cultures?, the skeptics asked. Why do the old folks come down with cancer more frequently than the young? Why are there so many different agents‐1,000 chemicals alone at last count—capable of bringing about animal cancers in the laboratory? And why do Japanese males and New York Jews suffer more than other ethnic and cultural groups from cancer of the stomach, while the Chinese on Taiwan die more often from cancer of the nasopharynx?

Such thorny questions made life difficult for the virus theory proponents. But in the last few years number of leading scientists have recognized that a revolution in thinking may be brewing in their midst. Laboratory findings in molecular biology, biochemistry and immunology have begun to make scientific sense and have inspired a new wave of hope that mankind's No. 2 killer (No. is heart disease) may one day soon be effectively controlled. Meetings devoted to tumor‐producing viruses —how they work and behave—are being’ held with greater frequency than ever before and are drawing more distinguished investigators all the time.

In Bethesda, the National Cancer Institute, which controls the Government's cancer research budget, has doubled the allotment for virus research over what it was five years ago; in fact, this year's budget is twice the size of last year's, despite a general cut in research funding. The President's State of the Union message, which proposed an inspired, Apollo‐like effort to end the scourge of cancer, is likely to increase this budget even further. [Dr. Frank Joseph Rauscher II (born 1931)], who heads the institute's virus cancer task force, says that “the era of the seventies is the era of confrontation with the cancer mystery and will reveal more about the mechanism of cancer than any time since the inception of research.”

This extraordinary revival of interest in viruses can be laid in large measure to [Dr. Robert Joseph Huebner (born 1914)], 57‐year‐old head of the institute's viral carcinogenesis branch, a recent winner of the National Science Medal and one of the nation's leading disease fighters. Huebner, marshaling the findings of the last few years, has formulated a theory of cancer that not only lays the disease at the door of the virus, but actually indicts a particular type of virus known to microscopists as the C particle.

Moreover, his theory diverges from those of other viral researchers on an even more critical point: he believes that the C particle is not an infectious virus that invades the body and generates disease, but a noninfectious virus that is a normal part of all living cells—and has somehow gone haywire. The Jekyll‐and‐Hyde behind diverse cancers in man—the C particle—turns out to be, in Huebner's opinion, none other than a form of RNA, one of the two main substances that govern heredity (although the particle, as we shall see, has a different function in the cell).

[Dr. Robert Joseph Huebner (born 1914)] is an articulate man with the dominant air of one who has fathered nine children and with an Inspector Javert complex about getting his bug. He loves nothing more than to expound his ideas. “Let's be clear about one thing,” he says, “the cancer virus is definitely not the same kind of bug that causes such well‐known infections as measles, polio or the common cold. It isn't spread horizontally, that is, from person to person, or from an animal or a toilet seat. You don't catch it. The C particle, or rather the genetic material it carries, is something we all have in our bodies, and transmit vertically to our offspring, like a gene that gives you red hair or blue eyes. In fact, the cancer virus is itself a group of genes —one tiny package of inheritance units among the three‐quarters of million genes that make up the 46 corkscrew‐shaped chromosomes in the nucleus of the human cell.

“This doesn”t mean that the disease is inherited. What you do inherit is the specific group of genes that carry the potential of cancer, the oncogenes as they are known'What sets them off? That's the key question. One has to know something first about viruses and how they behave in living cells.”

Viruses are microscopic‐50,000 average‐sized ones can sit on the head of a pin—and come in a variety of shapes: some are spheres, others rods, still others come in groups of triangles. The basic composition, however, as Nobel laureate Wendell Stanley of the University of California first revealed in the nineteenthirties, is the same for all. A virus contains either deoxyribonucleic acid (DNA) molecules or ribonucleic acid (RNA) molecules, the chemical building blocks of all living things, held together by a coat of protein. Since this composition is virtually the same as that of the gene, it is not surprising that scientists have come to regard viruses as central to the study of genetics, indeed as the key to the secret of life.

They can be found anywhere, not only in the body but in the air, on the ground and in plants. According to one theory, viruses are really genes that escaped from cells. Several thousand of these “creatures” have been caught and examined in the electron microscope, and so far some 300 have been tied to disease in man and animal. In fact, a “good” virus is hard to find.

The virus exhibits the properties of a living system, but spends a good deal of its time in an inactive state. Unlike other somnolent biological units such as seeds or spores, it is not blessed with metabolic machinery; to grow and reproduce each virus must therefore latch onto host cell, with its built‐in chemical factory. Once inside the host, the virus suddenly makes up for its own deficiency by taking advantage of the cell's chemical apparatus and using it to make replicas of itself. The parasite subverts the host and reproduces at its expense, sometimes until the cell dies of exhaustion. The new viruses then stream out of the emasculated cell and invade a new cell, then another, spreading infection throughout the body—unless stopped by the body's own virus‐destroying antibodies or by a drug that prevents the virus from replicating in the host cell.

The cancer virus, the Huebner team's C particle, operates in a totally different manner. “It's been sitting quietly in the nucleus of the cell all the time, and suddenly takes command like a fifth columnist who feels the conditions are ripe,” Huebner explains. “Instead of reproducing itself, like the infectious virus does, the cancer virus programed to issue orders to the cell to grow and grow until it unhappily creates ugly cluster of cells known cancer, which invades other body structures and may cause death.”

In Huebner's thinking this virus is on hand, ironically, to serve a good purpose; it is a growth particle, perhaps one of the original growth commanders of the embryonic stage of life. “The particle really wants to go back to the good old times of its world—to grow more embryo, some arms, legs, eyes and other specialized products. In a way it wants to start again, to make the brother of the host cell. Unfortunately, in the prison of the complex, mature body structure, there is no longer an opportunity for such simplistic urges, and instead of a new and useful growth, the body grows a cancer. The particle, it seems, has the power to start us off, and the power to kill us.”

N its basics, this approach to cancer isn't altogether new, as Huebner will admit. Back in the nineteen‐forties Dr. Jacob Furth of Columbia, one of the pioneers of cancer research, performed an animal experiment which suggested that leukemia potential in mice was vertically transmitted from parent to offspring, and in the nineteenfifties Ludwik Gross of the Veterans Administration proclaimed the then outrageous idea of a latent cancer gene. In later years researchers who believed that cancer must have a genetic connection put the blame for the disease on defective chromosomes, defective genes and on certain mutations of the genetic material (or some combination of these factors).

Huebner's contribution, based on considerable new data, is the conclusion that the many different forms of cancer are all traceable to morbid change in the same virus, the noninfectious C particle. He thus came to call his conception the “unitary theory” because it purports to explain the origin of all cancers, whether they seem to arise spontaneously or are induced by carcinogenic factors in the environment, such as cigarette smoke.

“Excessive cigarette smoking, chemicals, radiation and old age are simply triggers that switch on the genome [the collection of genes that make up the virus],” he says. “So are a host of other environmental and hereditary factors that make the differences in cancers between one ethnic group and another — or among geographical areas. They're all contributory causes, not the real root of the trouble, which lies in the interaction of the C particle with the genes in the host cell.”

The hypothesis is a sweeping one, and not all members of the scientific community are convinced that Huebner has provided adequate proof. RNA virus particles have been found in animal cancers, but they are not easy to identify in the tumors of man.

Huebner himself has not isolated a C particle from a human cancer, but other observers claim to have seen it, especially in some cases of breast cancer. Observations from an electron microscope are open to some interpretation, however, and there is disagreement as to whether the observers truly saw what they professed to see.

Huebner shrugs off the entire argument by saying that it is not really necessary to observe the C particle in human tumors to know that it is there. For its existence is unmistakably indicated, he maintains, by the presence of a “chemical footprint” known as an antigen.

An antigen is any substance that inspires the release of antibodies which attack a particular virus. Most proteins are good antigens, and the protein coat of an infectious virus itself is often the body's best defense against illness. (Dead or weakened viruses are introduced into the body in vaccines to combat such diseases as polio and influenza.)

The presence of an antigen can be detected by various biochemical methods, and in the nineteen fifties Huebner discovered one in animal cancers induced by DNA viruses; he called it the T (or tumor) antigen and its discovery led to the understanding of this type of cancer. Several years later, he found the antigen of the RNA tumor virus, which he named the gs (or group specific) antigen. The presence of this gs antigen is evidence of the presence of the viral RNA.

“It's a little like the devil,” Huebner says about the particle, “you know it only by its works. Medicine doesn't always have laboratory proof of the cause of a disease, but we can move forward and deal with the disease without it.”

NONETHELESS, neither Huebner nor any of his colleagues has yet performed the definitive test so dear to the virologist's heart since the days of Robert Koch, the father of bacteriology: that would require isolating the virus from a human cancer, injecting it into man and animal, and consistently obtaining the same cancer. (Obviously human volunteers for such a test would be hard to find!)

In the absence of proof of this sort, Huebner's critics argue that it may well be other genetic mechanisms that produce cancer in the cell and not the morbid expression of the RNA particle he claims is the cause. Some scientists, for instance, hold firm to the mutation hypothesis—first enunciated in 1905 and still one of the most durable theories of cancer — which maintains that a chronic irritation causes in time a genetic change that may be transferred from one cell to its progeny. This means some genetic information must have been added to, or taken away from, the genes already there or some other modification must have occurred, creating a new situation in which the cell is dominated by instructions for producing cancer.

Huebner says the mutation hypothesis is losing support even from those who believed in it for years. He points to an interesting experiment in which an investigator replaced the nucleus of a frog's egg with the nucleus taken from a cancer cell in a frog's diseased kidney. The egg was fertilized. A healthy swimming stage larva emerged from the embryo, indicating that the cancer cell had all the necessary genetic information to cause the growth of a normal organism. Evidently the cancerous state had failed to take anything away or add anything to the nucleus.

Finally, a persistent argument against‐Huebner's theory stems from those experts, mainly pathologists, who insist that cancer is not a single disease but a family of diseases. If it's just a single disease—caused by one virus —why are there so many chemically different types of cancer and why do different chemicals act specifically against different tumors?

“That's a reasonable question,” Huebner replies, “but think the many faces of cancer have to do with the type of target cell in which the virus ‘turns on.’ The diversity of cancers should not dissuade one from believing in the common cause, especially at the molecular level. After all, we all sprang from one cell, which contained the information that made us all different.”

ALTHOUGH Huebner blames a vertical, noninfectious RNA virus for all cancers, work on ‘horizontally infectious viruses was important to the development of his theory. Back in 1911 [Dr. Francis Peyton Rous (born 1879)] of the Rockefeller Institute ground up a tumor from a Plymouth Rock hen, dissolved it in a salt solution, filtered out the cancer cells and injected the resultant cell‐free “soup” into other chickens. To his intense fascination, they all developed tumors. Evidently the extract contained something that could transmit the highly malignant tumor from one hen to another.

Rous's findings, when first published, made little impact'Everyone said it was fine for the birds, but had no relevance to man. When studies of cancers in other species yielded no virus, the cancer‐virus infection theory fell into disrepute — the same kind of ignominy with which established medicine greeted the microbe theory of disease when it was first proposed. Rous himself, somewhat discouraged, turned to other fields of research. Interestingly, 55 years after Rous had made his discovery, the world of science took a second look at this feat and awarded him the Nobel Prize for Medicine.

During that half ‐ century, evidence of virus‐caused cancer slowly trickled in. The big breakthrough occurred in the nineteen‐fifties when Ludwik Gross, an Army surgeon attached to the Bronx Veterans Administration Hospital, showed by a brilliant series of experiments that leukemia, a cancer of the blood, could be transmitted in mice though only in the newborn.

Gross worked with two inbred strains of mice: the AKR strain, which shows a high incidence of inherited leukemia, and the C3H strain, which does not incur the disease naturally. He injected the leukemia fluid from the AKR mice into the low‐leukemia C3H strain and found that the latter developed the disease; in fact, the strain developed not only leukemia but a variety of other cancers. Gross was able to identify the killer virus, which is now known as the marine leukemia virus. A few years later researchers were able to see it in the electron microscope and over the years they found similar viruses in other species including — though this is in dispute—in man.

At first, Gross's monumental work was treated with the same skeptical disdain accorded early virus research. Denied working space on one occasion, he had to set up his laboratory in an unused military latrine. Nevertheless, his findings on oncogenic viruses stimulated a wave of research.

At the National Institutes of Health, for instance, Drs. Sarah Stewart and Bernice Eddy tried to duplicate Gross's experiment and found in the leukemic fluid a new virus which produced a group of cancers in newborn animals. So many cancers appeared in fact (23 in all) that the virus was christened polyoma (many tumors). More surprising, when injected into other species, such as hamsters, rats and rabbits,‐ this tiny spherical particle always yielded a display of tumorsThe virus, the first discovered which crosses species boundaries, proved to be made of DNA.

The discovery of the polyoma virus created a good deal of excitement; it meant that a single virus might be the agent in a disease that has many forms. Huebner entered the search at ‘this point, asking himself whether the polyoma could also be found in animals in their natural state and could therefore be associated with naturally occurring cancer.

He had behind him a record of having tracked down the elusive Q fever and other mysterious diseases. To find the polyoma virus, he began hunting in the places where mice were most likely to gather—crowded urban areas. In Harlem, he trapped 450 mice on the top floor of one tenement and discovered that half of them carried the virus, but no virus was found in the human beings who lived there.

He also wondered whether the Harlem mouse's wild country cousin contracted polyoma. As the owner of a, cattle farm in Maryland where he raises prize Angus bulls, he had no trouble finding natural breeding places of mice in his own hay sheds and nearby granaries. Huebner soon found that polyoma was as widespread in the country as in the city.

Even in his travels Huebner couldn't get away from mice with polyoma. He remembers seeing a field mouse scurrying out of sight in the vicinity of Disneyland, a rather fitting place for a mouse. Huebner promptly climbed a small fence, traced the mouse to its lair and later determined that it was carrying the virnst.

Out of these forays Huebner discovered an important, fact while the polyoma virus is present in a good many of the wild mice, the animals were singularly free of cancer. This was also true of other DNA viruses, such as the adenoviruses found in throat and adenoids of man and animals, which do not produce cancers in their natural hosts.

It seemed that these infectious DNA viruses could produce tumors in newborn laboratory rodents by injection, but did not do so when harbored by wild mice, probably, Huebner thought, be cause they entered the mouse's body later in life after some immunological and other defenses had been thrown up.

How about the other great class of viruses, the hundred or so RNA's beginning with the chicken virus—the first C particle—discovered by Peyton Rous? Here Huebner and his colleagues hit pay dirt; RNA viruses, mainly C particles, were found in large numbers of animals with naturally occurring cancer. The scientists found them in thickens, mice and cats with tumors, though they found not the virus itself but the specific antigen of the virus. The big question was: Is the virus a harmless resident in the cell or an assassin in disguise?

TO answer this question huebner had to know more about the care and feeding of the C particle. In the next few months he and his team

“‘Let's be clear about one thing,’ says Huebner. ‘The cancer virus is definitely not the same kind of bug that causes such wellknown infections as measles polio or the common cold.” hunted down antigens of the C particle in virtually every vertebrate that could be trapped and hauled into the laboratory. The particle proved to be amazingly ubiquitous. Investigators found it in rats, swine, guinea pigs, monkeys, hamsters and, to show the evolutionary continuity, in snakes. Wherever there were malignant tumors, there was evidence of C particles.

Cats were discovered to be a particularly good laboratory subject for the study of C particles. Huebner and others found that the feline leukemia virus, present in 75 per cent of cats, crosses species barriers and grows well in dog, monkey and human tissue. In fact, this finding led to a rash of public worry over catching cancer from cats. To avoid panic, Huebner and his aides, Drs. Murray Gardner and Bernard Hanes of the University of Southern California, conducted a survey in the Los Angeles area last year and established, happily, “There is no statistically significant difference in cancer incidence in the number of households with pets … and since 70 to 80 per cent of the cancer case households denied any cat ownership, it is unlikely that cats would be a causal factor in the generality of human cancer.”

Early last year several investigators created a flurry of excitement when they claimed to have seen a C particle in human cancer tissue (it looks like a miniature Chinese fortune cookie). Huebner, however, wasn't sure, though he admits he would feel like Balboa sighting the Pacific if he could identify the particle for certain. “Actually,” he says, “it wasn't that important. By this time a lot of lines of evidence from laboratories were fusing, and I became convinced that the RNA viruses are not harmless but are behind most cancers in man and animal.”

One of the major pieces of experimental evidence had been gathered the previous year when Huebner's associates, [Dr. George Joseph Todaro (born 1937)] and Stuart Aaronson, attempted to prove a closer connection between the onset of old age — especially the so ‐ called “cancer years” (beginning, by most estimates, after 50)—and an increase in the pro duction of C‐type virus.

One of the striking facts of life is that no cell lives forever. Every normal cell is exquisitely timed from birth to live its assigned lifetime and then die. Only cancer cells seem to seek immortality.

[Dr. George Joseph Todaro (born 1937)] and Aaronson grew embryonic mouse cells in a test tube until they reached the age that biologists agree amounts to dotage in tissue culture. What happened was that a number of the cells when they passed the grim boundary line of old age became cancerous — at which point most of them were starting to make the antigen of the C particle. Evidently the assembly of viral genes must have been in the cells before birth, to be released as C particles at the proper moment and with the proper trigger—in this case, some change that accompanies old age.

Numerous experiments in Huebner's own shop and elsewhere have tested cancer “triggers” other than old age. Typically, normal animal cells with and without active RNA virus have both been subjected to radiation, or have been chemically treated with known carcinogens. In one such experiment, the carcinogen produced tumor cells in a colony with RNA virus relatively quickly and in great number. A similar assault on the irusfree cells didn't transform them into cancer at all, strongly suggesting that the virus was the key factor in the cancer.

HUEBNER found another striking thing: he turned up antigens of C particles in all embryos. Perhaps nature had built the C particle into each cell for a good purpose—to produce the fast growth needed by the embryo for maturing. If the particle is associated with rapid growth, he theorized, then tissues that grow fast and thus have the highest turnover of cells should have more antigens than more slowly growing tissue. And so it proved to be. Huebner found, for instance, that the fast growing inside lining of the intestinal tract shows more footprints of the C particle than other tissues do, and indeed it shows even more antigen than can be found in most tumors. This is also true of the inside of the uterus and of the ovaries.

But some questions remained. If the cancer potential is already in the cell in the form of an RNA virus, how does one account for the transmission of cancer by the injection of the Rous sarcoma virus? Huebner worried over this for some time, but now gives this explanation:

“Rous dealt with young birds, and he forced in large amounts of the infectious virus which contained genetic information to change normal cells into cancer cells. We've shown that many chickens with C particles, unless killed by some other means, are likely to get cancer later in life, so it seems that Rous's injection of virus simply provided the disease‐causing level more quickly.”

There were even more intricate puzzlers: What, for example, causes the C particle, or indeed any other oncogenic virus, to make the cancer? And how does it actually go about its dirty work?

During the last decade, molecular biologists have successfully probed the mechanisms of the normal cell. They have shown how the DNA in the nucleus lays down chemical “orders” which are carried out by the RNA in the cytoplasm, directing it to build the living structure according to genetic plan.

At the Salk Institute in La Jolla, Renato Dulbecco has shown that a DNA virus particle can somehow incorporate itself into the genetic blueprint for a cell and cause it to behave abnormally. What happens, according to Dulbecco, is that the viral genes take up positions along the double helix of the blueprint, thus changing the genetic information passed on to the cell. (The double helix, of course, is the DNA that is known to be present in the cell; the virus particle is another form of DNA which either invades the cell or is also normally present —it is not yet clear which.)

Little was known of how the RNA virus subverts the healthy cells until last year, when Dr. Howard Temin of the University of Wisconsin and, independently, Dr. David Baltimore of the Massachusetts Institute of Technology, came up with some startling evidence. Temin and Baltimore discovered that the RNA of the virus can also replace the double helix and, by so doing reverse the transcription of information that guides the normal cell process. Here, the RNA is in charge, and, in a kind of man ‐ creates ‐ God operation, gives orders to make DNA which, in turn, creates replicas of itself. How this transfer of information goes on to cancer is still not known, but Temin's findings do indicate that the complexity of the mechanism is at last yielding to research.

HUEBNER is firm in his belief that the answer to the disease ultimately lies in somehow muzzling the viral cancer genes and forcing them to remain “silent” in the cell (until, he says, “the individual dies of something else”). This is not so fantastic as it sounds. As a result of recent work in molecular biology by Nobelists F. Jacob and J. Monod of France, each growthcontrolling gene in the cell is now known to be regulated by another gene that sits atop it and represses further cell division after the organ reaches its inherited shape and size. The tumor occurs when this “policeman” fails to do his duty, leaving the growth genes free to create the anarchy of cancer.

What Huebner envisions is that a substance can be found which will restore the repressor to its original position of control in the cell. At present he and several teams across the country are trying to find the specific natural repressor for the cancer genes of the C particle in a number of species.

Already, several virus scientists are reporting some extraordinary results in reversing malignancy in tissue culture, although they are not working directly with C particles. At Berkeley, for instance, G. S. Martin, in the famous virus laboratory of Wendell Stanley, experimented with a mutant of the Rous virus and found that he could turn a cell induced to grow abnormally by a virus into normal‐growing cell — and back again—by merely altering the temperature of the cell's environment a few degrees. At Princeton, in what may be a landmark experiment, Max Burger and Kenneth Noonan covered certain cancer cells with a plant protein called Conconavalin A and found that they suddenly grew like normal cells. He then removed the protein cover and the cancer growth took, off again!

In C particle investigations there have been similar promising findings. A team of scientists from the Cancer Institute and the Pasteur Institute in Paris isolated a chemical inhibitor from various types of mouse and rat tissue carrying cancer virus. They injected the material, along with cancer‐producing C‐type virus, into a colony of mice and found they developed fewer tumors than mice which were given the virus but not the inhibitor.

These and other experiments make Huebner and his colleagues more optimistic than they have been in a long time.

“I think the public will soon be able to cash the promissory note it has been holding for years from scientists working on a cancer cure,” Huebner says, adding reflectively: “You know, a few years ago that would have been enough to contemplate as a great end in itself, but now the skeptics question all values, and even ask whether it would be wise to have a cancer‐free society in view of the population explosion am old‐fashioned enough to believe that curing a disease that kills 250,000 Americans a year is a great plus for civilization — and that utopia will not be reached all at once, but step by step.”

The evolutionary conservation of genetically transmitted retrovirogenes has suggested that they may provide a critical function for normal development ( 1 ) . In the MuMTV system , specifically, it has been speculated that MuMTV expression may be required for normal functioning of the mammary gland. Bent velzen and Hilgers ( 2 ) hypothesized that MuMTV germinal provirus was associated with a mam gene, analogous with src of the avian sarcoma virus. They proposed that controlled expression of this gene was essential for normal mammary gland development, whereas uncontrolled expression ( as after exogenous infection with MuMTV ) resulted in mammary neoplasia ( 2).

COnversely, the great variation in endrogenous MuMTV restruction patterns in inbred mouse strains and individual wils mice implied a lack of selective advantage of these genes (3). This belief was supported by the finding of several apparently normal feral mice lacking detectabl eMuMTX provirus in their liver DNA (2). COhen and Varmus *3) thus favored the hypothesis that endogeneous proviruses were acquired by multiple independent infections of germ cells, and they saw no funcitonal ruole for proviral DNA in normal growth and deveopment. 

To determine whether enogeneous MuMTX provirueses are essential to normal development of the mammary gland and/or the development of mammary hyperplasia and neoplasia, we have studies these featres in a coloony of selectively bred feral mice that are totaly free of endogenous MuMTV copies in their cellular genome ( 5 ) . For ease of discussion we have labeled these endogenous virus -negative mice " ev ." In the ev mice , mammary gland development and function are compared to other feral mice homozygous for a single MuMTV provirus labeled " ev ” and to well established strains of inbred mice carrying multiple MUMTV proviruses ( 5 ) . In addition , we describe the formation in the mammary tissue of the ev and ev * mice of hyperplastic and neoplastic lesions that, although infrequently seen , are similar to those of laboratory mice ( 6 , 7 ) . We confirmed by Southern blot analysis that the ev' and ev * mouse colonies bred true for this trait and that the tumor DNA in these mice showed no alteration from the germ line MuMTV genotype. 

[...]

NEW scientific evidence has raised the possibility that acquired immune deficiency syndrome, or AIDS, may be transmissible through saliva.

The evidence, based on human and animal studies, is no more than suggestive in implicating saliva. But researchers said in interviews yesterday that they are convinced the studies raise real public health concerns.

So far, however, none of the more than 6,000 cases of AIDS reported to public health officials have been directly attributed to infection from saliva. The research has been conducted both among humans and animals.

Epidemiologic studies to date point to sexual contact as well as transfusions of blood or blood products as the major risk factors leading to AIDS.

''Right now epidemiological studies do not point to saliva as the key mode of spread of AIDS and data show that close contact is much more important,'' said Dr. Robert C. Gallo, a leading AIDS researcher.

''There is not yet clear-cut epidemiological evidence that the virus is transmitted by saliva to cause AIDS, yet this now has to be considered,'' Dr. Gallo also said. ''The question remains open whether or not saliva is a significant means of transmission. It is there and has to be studied but I don't think saliva is a major route of transmission of AIDS in humans.''

Dr. Gallo said he ''did not think that the virus all of a sudden was going all over the place'' and that he did not want to be alarmist. However, he also said, he ''did not want it underestimated and you have to deal with the facts.'' The work among human patients was carried on by Dr. Jerome E. Groopman at the New England Deaconess Hospital in Boston and Dr. Gallo at the National Cancer Institute in Bethesda, Md., and their colleagues. They found virus widely believed to cause AIDS, called HTLV-3, in the saliva of eight of 18 individuals who had so-called pre-AIDS or who had contact with individuals who had AIDS.

However, they did not find the virus in the saliva of any of 20 individuals who had AIDS.

The research report is scheduled for publication in the Oct. 26 issue of Science. But after some word of the research had come out in press reports, the work was described more fully yesterday in a telephone interview by Dr. Gallo.

The studies among animals carried on at the California Primate Research Center at the University of California at Davis found that saliva was probably the major route by which a virus similar to AIDS in humans is transmitted in monkeys who bite and scratch. Simian AIDS virus has been found in the saliva of up to 20 percent of monkeys in one cage at the California facility.

The simian AIDS virus, called SAIDS virus, belongs to the same retrovirus group as does the HTLV-3 virus that causes human AIDS. However, there are important differences in the two viruses.

Researchers have looked to the SAIDS virus in monkeys as well as the feline leukemia virus in cats as models of studying human AIDS.

Dr. Murray Gardner, a pathologist and member of the Davis research team, said in a telephone interview that ''many animals with SAIDS have the virus in saliva.''

Dr. Gardner said his colleagues have traced at least a dozen cases of SAIDS since 1976 to one apparently healthy monkey that was a carrier of the SAIDS virus.

''Wherever she went and came in direct contact with animals, the disease went with her in high incidence,'' Dr. Gardner said, referring to the movement of the monkey in the cages at the Davis facility.

Dr. Gardner also said that his team had isolated SAIDS virus from saliva taken from the carrier and injected into a healthy monkey, which then came down with SAIDS.

''It looks like the virus is not only there but also'' it causes disease, Dr. Gardner said. ''Why the carrier monkey is not sick is another unanswered question, but there are always these kinds of healthy carriers in any kind of infection.''

Dr. Gardner also said that he is ''convinced that healthy carriers can transmit the virulent SAIDS virus and that the healthy carrier may be the primary route by which the virus is spread.''

Dr. Gardner said the findings in monkeys would lead him to ''guess there's a public health danger'' in transmission of AIDS virus in saliva. Dr. Gardner also said:

''It probably takes multiple exposures to contaminated saliva. It probably will not occur overnight and probably will not result from a single drink or kiss. But with enough virus exposure you increase the chances of infection. Without a question, it is a public health matter of great concern.''

Dr. Gardner said he had presented the data about the SAIDS virus at scientific meetings in Zurich and in San Francisco and had discussed the findings with Dr. Gallo because ''it made sense to look at saliva in humans.''

Dr. Gallo called the finding of the AIDS virus in saliva ''surprising.''

SAN ANTONIO — Night has fallen here, and the monkey doctors are having a party. The scene is a dinner-dance at a Symposium on Nonhuman Primate Models for AIDS where 250 veterinarians and researchers are letting off steam after two days of intense scientific discussions.

With cowboy hats clamped to their heads, some down margaritas. Others venture onto the dance floor where a Texas trio warbles such tunes as “Your Cheatin’ Heart” and “Redneck Mother.” Mostly, though, they talk business, chattering excitedly about chimpanzees and macaques and mangabeys.

These are heady times for primatologists. AIDS has thrust them into the forefront of scientists seeking clues to one of the most confounding riddles in the history of biology. With their new role has come enhanced stature, more research grants and increased public attention--none of which means very much, to hear the monkey doctors, as they call themselves, tell it. “The driving force is to score a point or two against nature,” said Murray Gardner of UC Davis.

Moreover, any exhilaration the monkey doctors may feel is tempered by the potential dangers in their work, the terrible toll of AIDS, and by the long odds against quick success.

“I feel this great weight on my shoulders,” said Michael Murphey-Korb, who is working to create an AIDS vaccine at the Delta Regional Primate Center in Covington, La.

“It is hard to remain emotionally detached from our work,” added Max Essex, director of the new AIDS Institute at Harvard. “The human suffering is so obvious.”

Essex was just one of the prominent scientists who came to the conference held by the Southwest Foundation for Biomedical Research, one of the biggest primate research centers in the country. Others included the two brightest stars in the firmament of AIDS research: Luc Montagnier and Robert Gallo, discoverers of the human immunodeficiency virus, which causes AIDS; vaccine specialist Daniel Zagury, and Jay Levy of UC San Francisco.

“This year, we’ve got a Gallo and a Montagnier at a monkey meeting,” marveled Jon Allan, a scientist at the Southwest Foundation. “Why? Because (primate research) is where the exciting breakthroughs are going to come in AIDS.”

Non-human primates are markedly similar to humans in almost all aspects of their anatomy and physiology. These similarities underlie their value as models to scientists seeking vaccines and cures to a wide range of human diseases. While mice, guinea pigs and rabbits are useful in preliminary research, monkeys and chimpanzees are usually needed for final testing of the safety and efficacy of drugs and vaccines.

Polio would never have been eradicated in this country without the use of monkeys, scientists say. Indeed, “if you look back in history at the diseases for which we have solutions, virtually all of them had animal models,” said William I. Gay, director of the animal research program of the National Institutes of Health in Bethesda, Md.

Animal models, he said, give scientists the opportunity to watch a disease unfold step by step and to conduct experiments with possible vaccines and treatments. “You do with the monkey what you cannot do with man,” said Ronald Hunt, director of the New England Primate Center in Southborough, Mass.

As a result, scientists have been searching for a primate model since shortly after the first case of AIDS was discovered in the United States in 1981. Although they have found several, none is perfect.

In AIDS, there are two requirements for the ideal animal model. First, the animal must be susceptible to infection with the human immunodeficiency virus. Second, the animal’s immune system must collapse, and the animal must develop some of the cancers and opportunistic infections that characterize AIDS. So far, scientists have found non-human primates that meet one requirement or the other, but not both.

The chimp, for example, can be readily infected with HIV. But the human’s closest relative has confounded scientists. Though a chimp can be infected with HIV, it does not get sick.

Then there is the Asian macaque, or rhesus monkey. When infected with a viral cousin of HIV called simian immunodeficiency virus (SIV), macaques develop swollen lymph nodes, enlarged spleens, diarrhea, skin rashes, tumors, brain lesions and pneumonia--in short, all the hallmarks of human AIDS. In fact, scientists call the condition SAIDS, or simian acquired immune deficiency syndrome.

“SIV-macaque is the most promising animal system we have,” said Montagnier of the Pasteur Institute of Paris.

Added Gallo: “The SIV-macaque model is reaching its maturity, and this meeting is pointing that out. In paper after paper, many groups are working on it, and data is coming in. . . . It is wonderful to have another animal system to study biology, to study mechanisms.”

Different Virus

But will the SIV-macaque model speed the search for a vaccine or therapy for AIDS? “No one can predict that because it is not the same virus” as HIV, Gallo said.

Other leading scientists say the macaque has already proved its worth. “Realistically, I don’t think we can ask for a better model,” said Patricia Fultz of the Yerkes Primate Center in Atlanta.

“It is very frustrating to read in the media, ‘There is no model for AIDS,’ when we’ve known for a year and a half what a good model we have.”

The simian AIDS virus, she said, is the same size and shape as the human AIDS virus. And while SIV has only 50% of its genes in common with HIV, Fultz said SIV shares 75% of its genes with a second human AIDS virus, known as HIV-2, which is primarily found in West Africa and causes an AIDS-like illness in some patients.

Already, researchers are testing vaccine preparations and drugs on SIV-infected macaques. But Fultz acknowledged that progress has been slowed by the biological complexity of the viruses.

“Every isolate of SIV is different,” she said. Some strains are so virulent they can kill a macaque in two weeks; others take six months to a year, or even longer.

“That kind of variation is bad. It means many of our experiments have to be repeated,” Fultz said.

Even then, experiments that work simply point up new areas for investigation. “Every answer brings five new questions,” Fultz said.

Consider the chimpanzee. Learning that chimps could be infected with HIV was easy. Now scientists want to know why infected chimps do not fall ill--or even lose their Helper-T cells, key sentinels of the immune system whose destruction by HIV is thought to lead to AIDS in humans.

“Why don’t chimps get sick?” asked Peter Nara of the National Cancer Institute. “What is the natural mechanism that keeps them healthy? More importantly, is there some way we can transfer their resistance to infected humans?”

No Protection

Chimps have also been used in the search for an AIDS vaccine. But none of the 10 preparations tried so far has protected the animals from infection with HIV, and some of them even enhanced the virus’s ability to infect.

Two African monkey species, the mangabey and green monkey, provide scientists with similar lines of inquiry as the chimp. In both species, SIV--the same virus that wreaks havoc in Asian macaques--is harmless. Indeed, more than 50% of wild African green monkeys in one recent study were infected with SIV; all were healthy.

This finding intrigues scientists. The African species “must have evolved mechanisms that kept a potentially lethal pathogen from causing disease,” Harvard’s Essex theorized.

In short, what scientists are finding is that, given enough time, species appear to develop mechanisms that allow them to coexist with AIDS-like viruses. This would explain why SIV is harmless in monkeys from Africa--where the virus presumably has been present for centuries--but devastating in newly infected species from Asia like the macaque.

Macaques are believed to have had their first experience with SIV in the 1970s when they contracted the virus from infected African green monkeys in primate research centers.

“There is a rough parallel between the differential susceptibility of green monkeys and macaques and the very different susceptibility to AIDS of chimpanzees and human beings,” Essex said.

Essex’s explanation of why some species get sick and others do not is pure Charles Darwin: Through the process Darwin called “natural selection,” the fittest African green monkeys and chimpanzees--those that could resist the immunodeficiency viruses--survived and reproduced.

“Obviously, we do not have the luxury to wait for natural selection to give humans an immune system that can resist HIV,” Essex said. So he and his colleagues in animal research centers and laboratories around the country eagerly search out every opening in the virus’s armor and every element of the immune system that might be bolstered.

At times, the work is frustrating; at others, exhilarating. Always, it is fraught with danger. “A non-human primate is a biohazardous agent, period,” said Elizabeth Muchmore of the New York University Medical Center. Scientists say the risk of being infected by lethal pathogens carried by the animals is never far from their minds.

Surprisingly, AIDS is not at the top of the list of the scientists’ worries about being infected. That honor is reserved for Herpes B--not to be confused with the well-known herpes virus that causes painful blisters on the skin and mucous membranes of humans. Herpes B is a monkey virus that coexists peacefully with macaques but is usually fatal on the rare occasions it finds its way to man, generally through monkey bites or scratches.

“It is the scariest one that we know of,” Muchmore said.

Twenty percent to 60% of the macaques in U.S. primate research centers carry the Herpes B virus. Researchers say they have become a lot more careful in the wake of an incident last year in which two handlers died after being bitten by an infected animal and now wear face shields, goggles, gowns, gloves and boots when working with macaques.

Of course, disease-causing agents are not the researchers’ only worry. They also fret that shortages of laboratory animals will delay their work.

Chimpanzees, whose susceptibility to HIV infection make them ideal for testing vaccines, are in particularly short supply. Imports of chimps were banned by the Convention on International Trade in Endangered Species, which went into effect in 1976.

With AIDS as the impetus, the government launched a $4.5-million chimp breeding program in 1986 and has managed to increase by 200 the number of research chimps in the country, to 1,400 animals. A national chimpanzee management plan allocates the scarce animals to various areas of research; already, 100 chimps have been infected with HIV.

Scientists also worry that space to house their laboratory animals is at a premium. “I just doubled my isolation facility to hold 60 monkeys,” said Hunt, director of the New England Primate Research Center. “Now, I’m expanding it again, this time to 100. I also need to hire more people, but there is no place to put them.”

Animal Rights Lobbyists

Still, many of the scientists interviewed here said worries about space and the availability of animals pale compared to the threat to research they see from the vocal and highly organized “animal rights” lobby. Gay, for example, said he had received 36,000 post cards between February and August demanding a halt to the chimp-breeding program.

“I think those animal rights people should be forced to walk through a pediatric AIDS ward to see those children dying,” one researcher said.

It is not that they are being callous about the monkeys, the scientists say.

“These are intelligent animals. They are not numbers,” acknowledged Theresa J. Smith of the Army Research Center at Camp Detrick, Md. Monkey doctors, she said, “have to have the strong belief that (what) we are doing is going to make a difference.”

That belief was reinforced at the dinner-dance here, where Maurice Hilleman, scientific director of pharmaceutical giant Merck & Co.'s research institute, told the gathered scientists that they “stand at the very center of future developments in research on the control of AIDS.”

“Not bad for a bunch of monkey doctors,” commented Lisa Krugner-Higby of the Bowman Gray School of Medicine in Winston-Salem, N.C., using the self-deprecating label often voiced by primate researchers.

And so, they persevere, urged on by the hope that they may be helping the estimated 1.5 million Americans--and countless other people around the world--infected with HIV. “That huge iceberg is going to surface” as more and more infected people develop AIDS symptoms and die, said Gardner of UC Davis.

“No one person, no one group, is going to solve it,” he added, calling for concerted national effort to conquer AIDS, akin to the Manhattan Project that developed the atom bomb during World War II.

“Sometimes in my lab, I get this great sense of history. The physicists did it 40 years ago. Now it is the biologists’ turn. The people are waiting.”

During 1991 significant headway continued to be made in simian immunodeficiency virus (SIV) vaccine research. The marked susceptibility of macaques to challenge infection with titered biologic and molecularly cloned stocks of certain strains of SIV, and the reliable induction of simian AIDS within a relatively short time (6-24 months) make this model suitable for testing the safety and immunogenicity and, most importantly, the protective efficacy of various vaccines against bloodstream infection and disease. In addition, SIV vaccines have been tested against genital and rectal routes of infection. This past year has seen a confirmation of the validity of this model for AIDS vaccine development. Protection against bloodstream infection by inactivated whole SIV vaccines has been confirmed and extended to include heterologous strains. Attenuated live virus and passive transfer of immune sera have also protected against systemic infection. The first reports of immune protection using recombinant SIV envelope vaccines have appeared. However, bearing in mind several recent failures to protect with recombinant SIV vaccines, we evidently need to learn much more about these novel vaccine strategies. Protection against rectal infection but not against vaginal mucosa! infection has been observed, raising important questions about mucosa! immunity. Solid protection against intravenous (i.v.) infection with lymphocyte associated SIV also has yet to be achieved. Finally, the recent revelation that immunization with uninfected human cells protects macaques against i.v. infection with SIV mac raises the interesting question about the role of anticellular antibodies in protective immunity. This review will summarize the SIV vaccine field, now several years old, and will highlight the most recent developments. The amount of unpublished information included in this review might render a more fitting title, '1992 - a year in preview'.

Inactivated virus

Several groups in the United States [1-5] (P. Johnson, personal communication, 1991), United Kingdom (M.P. Cranage, E.J. Stott, KA Kent, et al., submitted for publication) [6], Sweden [7], Germany (C. Stahl-Hennig, G. Voss, S. Nick, et al., submitted for publication; S. Hartung, S.G. Norley, J. Ennen, et al., submitted for publication) and the Netherlands (A Osterhaus, personal communication, 1991) have reported successful protection of macaques against experimental SIV infection and/or disease by inactivated whole SIV vaccines (Table 1). Protection of macaques against HIV- 2 challenge infection with an inactivated whole HIV- 2 vaccine [8] is covered in another chapter. Efficacy is ~90% in these trials, which now include over 100 monkeys, mostly rhesus (Macaca mulatta), cynomolgus (M fascicularis) and pig-tailed (M nemestrina) macaque species. By contrast, all controls have become persistently infected with the same challenge dose of SIV. Special laboratory conditions for obtaining successful protection include challenge with a relatively low dose, 10-200 animal infectious doses (ID) of homologous cell-free virus given intravenously (i.v.) or intramuscularly (i.m.) within 2-4 weeks after the last boost. Higher challenge doses ( > 103 ID), when tested, have not been protected against by the inactivated virus vaccines [ 1,3]. Virus for the SIV vaccine trials in the United States was grown in human Tcell lines (for example, HUT-78, CEM), whereas, to maintain maximum virulence, the live virus for challenge was usually grown in fresh human T-cells. The European Community (EC) investigators have used a common stock of formalin-inactivated SIV mac251 vaccine prepared from virus grown in a human T-cell line (C8166) and a homologous challenge stock of SIVmac25(13 2H), also grown in C8166 cells after passage through a rhesus monkey. Successful vaccines were achieved by inactivating the whole virus with formalin, ~-propiolactone, or detergent. There has been no incidence of residual live virus in any of the inactivated vaccines. Successful whole virus vaccines were made from either uncloned biologic isolates or molecular clones of SN mac or SNsm· An inactivated molecular clone of SN mac (BK28) could protect against challenge with an uncloned biologic isolate of SN mac (M. DeWilde, personal communication, 1991), and an inactivated molecular clone of SN8m (H-4) could protect against the uncloned homologous strain (SN8mE660) (P. Johnson, personal communication, 1991). Virus pelleted from culture media without further purification, and virus purified by sucrose gradient or column chromatography were used in the successful wholevirus vaccines. The total amount of virus antigen used to confer protective immunity ranged from approximately 500 μg to 3.0 mg, and the schedule of immunizations consisted of a maximum of five inoculations over a 13-month period to a minimum of three inoculations over a 2-month inteival. In a recent study comparing two total doses ( 400 μg and 2.0 mg) of the EC formalin-inactivated SN mac vaccine, one out of four monkeys receiving the lower dose and four out of four monkeys receiving the higher dose were protected against challenge infection with 33 50% infectious dose (ID50) of homologous virus (S. Hartung, S.G. Norley, J. Ennen, et al, submitted for publication). Adjuvants used include muramyl dipeptide [MOP; Syntex adjuvant formulation, monophosphyl lipid A plus 'cell wall skeletons'(RIBI)], alum, complete and incomplete Freund's adjuvant (CFA, IFA), immunostimulating complex (ISCOM) and Quil A MDP has been the most widely used adjuvant (SAF-1 Syntex, kindly provided by Dr AC. Allison, Syntex). At the German Primate Center ( C. Stahl-Hennig, G. Voss, S. Nick, et al, submitted for publication), it was shown that over 50% (four out of seven) of monkeys were protected against SN infection using a vaccine formulation (Tween ether-treated whole virus adsorbed to. alum) of the type used for many years to vaccinate humans against influenza virus. With all the vaccines tested so far antibody levels declined rapidly after each boost. In vaccine-protected monkeys, antiviral antibodies declined after challenge, reaching minimal levels by 4-6 months post-challenge. No adverse reactions have occurred in any of the vaccinated animals.

Duration of protection

Results from three groups suggest that vaccinated monkeys remain partially protected for as long as 8 months after the last boost. In one US study (M. Murphey-Corb, personal communication, 1991), two out of four SN8m-vaccinated monkeys were susceptible to challenge infection with 10 ID of the same virus 8 months after the last boost. In a second trial (M. Murphey-Corb, personal communication, 1991), using an improved whole SNsm vaccine, five out of five monkeys were still protected against the homologous virus 8 months after the last boost. However, this protection

[... much more ... ]

An experimental vaccine developed from a genetically altered virus has completely protected monkeys against an AIDS-like disease for more than four years, scientists said today. They said the protection was the strongest and longest lasting of any AIDS-related vaccine.

Federal health officials who financed the experiments hailed the finding as "a significant advance" in the search for a human vaccine to protect against H.I.V., the virus that causes acquired immune deficiency syndrome.

The new monkey vaccine contains live S.I.V., the virus that causes an AIDS-like disease in monkeys, from which a key gene was removed, the scientists are reporting Friday in the journal Science.

The research at the New England Regional Primate Research Center in Southborough, Mass., was led by Dr. Ronald C. Desrosiers. The findings seem certain to inject new life and controversy into AIDS-vaccine research, a field that has undergone roller-coaster waves of enthusiasm and dejection.

Many Avoid Approach

Although many successful human vaccines are derived from live viruses that have been weakened, many scientists have shunned such an approach for AIDS because of the potential dangers of injecting a highly lethal virus into uninfected individuals. Among these dangers are errors in production that would allow the vaccine virus to actually produce the illness, the possibility that the weakened virus could somehow change within humans to cause the disease and the theoretical possibility that it could lead to cancer.

Even experts in AIDS vaccines who said they were excited by the latest advance cautioned that many questions about the safety of an AIDS vaccine based on a live virus needed to be answered before one could become available for use, if it ever did.  [ Since injecting with the live virus is so dangerous ... this encourages RNA/DNA vaccination evolution ]'

The experts also said it was uncertain whether drug companies would underwrite the huge costs of developing an AIDS vaccine based on a live virus because of the liability risks if humans accidentally became infected from such a vaccine.

Nevertheless, Dr. Anthony S. Fauci, the head of the National Institute of Allergy and Infectious Diseases, a Federal agency, said, "There is nothing like the smell of potential success to lure drug companies into a field."

Dr. Fauci said scientists at his institute were "extremely enthusiastic about the possibilities" raised by Dr. Desrosiers's report.

Many Vaccines Tested

The intended use of any such vaccine would be to protect uninfected individuals against the human immunodeficiency virus. Although there is no theoretical reason to suppose that such a vaccine would benefit individuals infected with the virus, Dr. Desrosiers and Dr. Fauci said such tests would ultimately have to be carried out to determine the answer.

The new findings could lead to the first safety tests of such a vaccine on human volunteers in two years, Dr. Desrosiers said in an interview. Such initial studies are generally carried out in less than 50 volunteers.

Dr. Murray Gardner, an expert in AIDS vaccines and S.I.V. at the University of California at Davis, said his team had begun experiments to confirm Dr. Desrosiers's findings.

Most AIDS vaccines developed so far have come from the use of newer laboratory techniques in which a portion of the AIDS virus is used in an effort to stimulate the body to protect against the complete virus. But Dr. Gardner said scientists might be in danger of over-emphasizing this approach.

An Unexpected Dividend

The new vaccine came as an unexpected dividend from experiments undertaken on animals to determine the function of the nine genes in the AIDS virus on the natural course of the disease. The scientists started by deleting a gene known as nef from the simian immuodeficiency virus and then injecting the virus in the monkeys.

The function of the nef gene is unknown. When nef-deleted S.I.V. is used in test-tube experiments, the virus reproduces and damages cells. But Dr. Desrosiers said he had been surprised to discover that injection of the virus did not harm monkeys. The observation led him to wonder whether it could be used as a vaccine.

He began the experiments by injecting the altered virus into six rhesus monkeys. The animals remained healthy after two and a quarter years. An additional 12 monkeys were injected with S.I.V. as scientific controls, and all have died.

To test the effectiveness of nef-deleted S.I.V. to protect monkeys against the disease, the scientists deliberately injected the whole virus into the six monkeys as a challenge, first in small amounts and later in large amounts. None of the monkeys have shown any evidence of illness from the AIDS-like disease for up to 55 weeks after the challenge injections, the authors said.

Blood tests showed no change in the number of CD4 immune cells that play an important role in fighting off S.I.D. and that drop precipitously as the viral infection progresses to disease.

Dr. Desrosiers said his team was trying to determine the fewest number of genes that must be present in a candidate vaccine for it still to be safe and protective. He said his team had injected monkeys with S.I.V. viruses from which one to four genes had been deleted and was planning to do similar studies with gene-altered-H.I.V. in chimpanzees.

In an advance along the demanding road to development of an AIDS vaccine, researchers have shown that a vaccine derived from an AIDS-like monkey virus can prevent vaginal infection in the animals, as might occur in human heterosexual sex.

While experimental vaccines had previously been shown to give protection to monkeys, none had worked for heterosexual transmission. Experts praised the new study, which is being published today in the journal Science. But they warned that it would take years of more research to develop a human AIDS vaccine, if one can be devised.

"What makes this important is that it addresses the primary mode of H.I.V. transmission," said [Dr. Shiu-Lok Hu (born 1949)], a virologist at the University of Washington and at Bristol-Myers Squibb in Seattle.

Although H.I.V, the virus that causes human AIDS, is commonly transmitted in the United States and Europe through homosexual contact and through intravenous drug use, heterosexual transmission is the most common way it is spread in other parts of the world, especially Africa and Asia.

In the new study, scientists from New Mexico, Alabama and Georgia used new technology and strategy to immunize monkeys against the simian immunodeficiency virus, the AIDS-like virus in monkeys. They found that a vaccine consisting of chemically killed virus, followed by a booster, prevented transmission of the virus when it was introduced into the animals' vaginas.

Other experimental vaccines have protected animals against this monkey AIDS virus, and against H.I.V. in humans, when these viruses were injected into the veins of the immunized animals. Such experiments offer valuable evidence for protection of intravenous drug users.

Previously Just Theory

But because scientists know that antibodies and other factors that are produced in different areas of the body are important in immunological defenses, the evidence from previous intravenous experiments offers only theoretical reasons for believing that such vaccines would protect against heterosexual transmission.

In the new study, five of six monkeys were protected against vaginal transmission when the doses of chemically killed simian virus were injected into the muscles and then a booster was given orally or in the lungs. Oral immunization alone did not protect them. The booster vaccine was made by using a new microsphere technique -- developing microscopic-size beads less than a thousandth of an inch in diameter that contain a highly diluted, inactive form of simian virus. The beads are designed to allow slow release of the vaccine.

Dr. Preston A. Marx of the New Mexico Regional Primate Laboratory, the senior author of the study, and experts elsewhere expressed cautious optimism about the feasibility of developing a human AIDS vaccine.

Dr. Dani Bolognesi, an AIDS vaccine expert at Duke University, said that Dr. Marx's team had come up with a novel approach. Dr. Marx, Dr. Hu, Dr. Bolognesi and Dr. Murray Gardner of the University of California at Davis all said there was reason to believe that a similar vaccine could be developed to protect men. However, such experiments have yet to be carried out.

Dr. Marx said he began the experiments in July 1990 at the California Primate Center in Davis by giving the first of a series of injections of simian-virus vaccine. He took the monkeys with him to the primate center that he now heads at Hollomon Air Force Base in New Mexico.

At first the experiments seemed headed for a dead end, but one of his collaborators, Dr. John H. Eldridge at the University of Alabama in Birmingham, came across a new method that led to success, Dr. Marx said.

A year after injections of the simian vaccine, none of the monkeys had developed high levels of antibodies in the vagina. So Dr. Marx said he tried the new approach. Dr. Eldridge had learned from other experiments that when microspheres were introduced into the lungs, this significantly augmented the vaginal immune system. The researchers tested the encapsulated simian vaccine on 14 monkeys, divided into four groups.

As a scientific control, the scientists applied the vaccine to four non-vaccinated animals. Three became infected after one application to the vagina. The fourth become infected after a second application. A second group of four monkeys was given five doses of the micro-encapsulated simian vaccine in the stomach. Only small traces of antibodies were detected in blood, and no sign of immunity was found in the vagina. All four became infected within two weeks after a single application to the vagina. The two other groups received boosters of encapsulated S.I.V. in January 1992, about a year after receiving the last of three injections of the vaccine in thigh muscles.

One group, which was given boosters in the lungs, showed the strongest immunity in blood and vaginal fluids. Two of three animals were protected from vaginal transmission.

The three monkeys in the last group were given booster doses in the stomach, and all three animals resisted two vaginal exposures to the virus.

Limited Group of Animals

The experiments were limited to a small number of animals because of the high costs and a desire to use the smallest number of monkeys. Tests of experimental vaccines are intended to give live virus to immunized animals at a time when the scientists believe this might achieve the best results. In the New Mexico experiments, Dr. Marx said his team applied simian virus to the vaginas of the immunized animals at times of peak immunity, two weeks after the last booster doses.

The researchers used a complex test known as polymerase chain reaction to show that the immunized monkeys challenged with the virus have remained free of infection for more than a year and a half after the challenge.

Dr. Marx said he had failed in previous efforts with a different vaccine to protect against vaginal challenge with the siminan virus and that others had failed with still other vaccines.

He said he did not know the immunological mechanism that allowed the encapsulated vaccine to work and that his team has found no satisfactory explanation for the failure in one monkey.

Dr. Marx said his team did not test rectal immunity because the study was intended to explore the feasibility of a vaccine against heterosexual transmission. An earlier study of animals in Britain has shown that another vaccine has given partial protection against rectal transmission.

Dr. Marx and others said that the success of the New Mexico study leaves a number of questions that need to be answered in future tests, such as:

• *How long does immunity last?

• *If it is not lifelong, how many boosters will be needed?

• *Will the vaccine protect if the virus is given weeks or months after peak immunity?

• *Will such a vaccine protect against strains of virus that are different from the ones used to make the vaccine?

• *Can a recombinant vaccine be made from a fragment of H.I.V.? If so, which component is the critical fragment?

Dr. Gardner, the expert from the University of California at Davis, said the new study "is meaningful, promising, and points us in a new direction that we will have to follow until we understand how the vaccine works or we come to a dead end. In this line of work, any step forward is welcome."

TEN years ago Margaret M. Heckler, who was then Secretary of Health and Human Services, predicted an AIDS vaccine by 1986. Although the deadline passed, the effort continued.

But this month work on the two experimental vaccines furthest along in development received a serious setback. A Government-appointed panel of 30 experts rejected plans for full-scale tests. They said they had little confidence that the vaccines would protect a sufficiently large proportion of recipients.

The decision on June 17 was surprising because in late April another panel, made up of some of the same people, recommended such a large-scale study. Both the April and June recommendations were nearly unanimous.

The June panel did recommend continuing smaller studies to test the safety of the two vaccines as well as that of more than 20 others that have been injected into about 2,500 uninfected humans throughout the world. Work on immune response also continues.

Ever since H.I.V., the virus that causes AIDS, was identified in the early 1980's, development of an AIDS vaccine has been the highest public health priority. Since 1984, the United States Public Health Service has spent at least $639 million on developing an AIDS vaccine.

Mrs. Heckler's optimism reflected the enormous confidence then prevalent that state-of-the-art genetic engineering techniques would allow this to be done quickly. But the AIDS virus turned out to be arguably the wiliest virus known. H.I.V. constantly mutates, evading the body's defenses and eventually destroying the immune system. Thus making an AIDS vaccine is a far more difficult effort than had been thought.

How big a setback for the development of an AIDS vaccine was the panel's decision? The answer depends on whom you talk to. The quest for a vaccine in the midst of a raging epidemic almost inevitably touches off a clash between empirical and highly scientific approaches.

Empiricists say they must rely at least in part on intelligent guesswork. Scientists favoring a more cautious approach say the deadliness of AIDS means that as much as possible should be learned through laboratory and animal experiments before subjecting volunteers to the crucial test -- finding out whether the vaccine prevents infection.

Both the April and June panels said that too many pieces of the scientific puzzle were missing. A particular concern was the inability to identify the immune response that correlates with protection against H.I.V.

The two vaccines are now in the second of the three stages of testing that the Food and Drug Administration requires before considering a product for licensing. One is made by Biocine, a joint venture of Chiron and Ciba-Geigy in Emeryville, Calif. The other is made by Genentech of South San Francisco.

Although the members of the April panel favored expanded studies, they were not overly impressed with the vaccines. Their recommendation reflected a belief that little more could be learned from continuing studies and the larger ones might show that the vaccines worked.

But the June panel, reviewing fresher data, learned that 13 participants in AIDS vaccine trials, including five in the Biocine and Genentech studies, had become infected with H.I.V. The infections came from risky behavior, not from the vaccine. Some infections are expected in vaccine trials. Because none of the newly infected participants had completed a full course of immunization (up to four shots over several months), their cases probably are not likely to be helpful in assessing the efficacy of the vaccines.

And because a full-scale study would involve up to 9,000 more participants, many panelists wanted to wait for a thorough analysis of data concerning infected participants before proceeding to a study that could cost more than $50 million.

The June panelists said they hoped that more promising potential vaccines would emerge from among the second and third generation vaccines now in their earliest stages of testing. Were they wise in waiting, or too timid? Is it possible there will never be an AIDS vaccine?

Dr. Anthony S. Fauci, head of the National Institute of Allergy and Infectious Diseases, a Federal agency in Bethesda, Md., that convened the panels, denied that the latest action was a setback. But he conceded that it would be at least three years before a full-scale effectiveness study could be carried out on any experimental AIDS vaccine.

To an empiricist like Dr. Sten H. Vermund, who served on the April panel and recently moved from the institute to the University of Alabama in Birmingham, the decision was a mistake: "It is wrong to halt progress in one vaccine concept because of a pipe dream that another vaccine concept will be the panacea."

Scientists are generally more cautious in testing a prevention method than they are concerning new therapies for a disease. But Dr. Vermund argued that the risk-to-benefit ratio should tilt toward taking more risks in dealing with prevention for a major public health problem. In his opinion, the June panelists should have recommended forging ahead in testing the efficacy of the two experimental AIDS vaccines. "In the face of a raging epidemic in which existing preventions have limited efficacy, why aren't we compelled to take more risks in prevention research?" he asked.

Dr. Vermund also criticized the lack of explicit criteria to determine what is needed to move an experimental AIDS vaccine into full-scale efficacy studies. "At what height is the high jump bar set?" he asked.

Dr. Vermund said it was doubtful that an experimental vaccine that is fundamentally different from the existing ones would be on the horizon for several years. "If we have a partially effective vaccine and do not know it because we are not testing it, that would be a public health tragedy," he said.

Scientists boast that they have learned more quicker about the AIDS virus than other other infectious agent. Most of that knowledge has come from research on the fundamental structure of viruses and cells. And many in it have been eager to apply their newest techniques, believing they hold the key to a successful AIDS vaccine.

Have Government officials put too much emphasis on molecular biology in developing an AIDS vaccine?

Dr. Murray Gardner, an AIDS vaccine expert at the University of California at Davis who did not serve on either panel, said the area was very specialized and, by its nature, often inflexible. He likened such scientists to football place kickers who do only what they know how to do best: place kick.

"AIDS has been wonderful for molecular biologists," Dr. Gardner said. "But have molecular biologists done anything for AIDS yet?"

High technology techniques may not be the answer, he said. "We may be losing sight of the real world where we should be trying simpler preventions such as a vaginal microbicide or an acid douche," he said. "Something that simple that could be distributed for pennies might do more to contain this pandemic than all of this high-powered technology."

Dr. Dani P. Bolognesi, an expert on AIDS vaccines from Duke University who voted with the June panel, said any number of developments could turn the situation around. "There may be lots of surprises in the woodpile," he said.

To Dr. Ruth Berkelman, an AIDS official at the Centers for Disease Control and Prevention in Atlanta who served on the June panel, the decision was not so much a setback as a lesson in overreliance on technology. "It shows we cannot always count on technology to solve a problem," she said in interview.

If genetic engineering techniques fail to deliver an AIDS vaccine, Mrs. Heckler's prediction will turn out to be one of the biggest overpromises in the history of science.

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Dr. Murray Gardner, a virologist at the University of California at Davis, said Dr. Huebner ''was just what we needed to get people moving, a visionary who foresaw the importance of viruses, in particular retroviruses.''

Retroviruses, little sacs of genetic material, put out a kind of reverse code or defective blueprint causing cell overgrowth. Dr. Huebner put together a team of scientists to study them, Dr. Gardner said. ''And within 15 years it paid off,'' he said.

By then the existence of the oncogene was confirmed by others and it was clear that vaccines could be developed to prevent tumors. As a result of such vaccines, including one for hepatitis B, rates of some viral cancers, like that of the liver, have been sharply reduced.

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