Tag Archives: interferon

Julius Youngner: Slighted Polio Vaccine Pioneer

This is a tale of the hurt that a junior investigator might feel when a senior investigator takes the lion’s share of the credit for the junior investigator’s crucial breakthroughs. Jonas Salk, who conceived and oversaw the development of the first widely used polio vaccine, is the senior investigator in this anecdote. Julius Youngner, the last surviving member of the original vaccine research team that Salk assembled in the early 1950s at the University of Pittsburgh, is the slighted assistant. Youngner later had his own distinguished career. He passed away in April of this year. Here is their story.

After earning his Ph.D. in microbiology, Youngner was drafted into the World War II U.S. Army, which assigned him to the Manhattan Project, to test the toxicity of uranium salts. Youngner first learned the purpose of the Manhattan Project when the first atomic bomb was dropped on Japan.

After the war, Youngner worked as a commissioned officer for the U.S. Public Health Service. This was a significant stop in his career, since it was there that he first became interested in viruses and cell culture. But, since there was no opportunity for him to pursue that interest in Bethesda, he began to look elsewhere. Thus, it happened in 1949 that Salk recruited Youngner to join his vaccine research team in Pittsburgh, after a mutual acquaintance told Salk that Youngner was eager to work on viruses and cell culture.

Jonas Salk and Julius Youngner at the University of Pittsburgh, early 1950s

Salk hoped that Youngner might find a way to generate enough cells from monkey kidney tissue to support mass-production of the vaccine. Youngner, on his own, then developed the use of the proteolytic enzyme, trypsin, to disperse tissue fragments into individual cells, thereby generating many more cells from a given amount of tissue. Indeed, Youngner could generate enough cells to support manufacture of the vaccine. This was his first key contribution to the vaccine project. “Trypsinization” remains a mainstay of modern cell culture.

Youngner’s next major contribution to the vaccine enterprise was his development of a rapid analytical test that had two crucial applications. First, recalling that the Salk vaccine contains an inactivated virus, Youngner’s so-called “color test” made it possible to quickly screen batches of the vaccine for any live virus that might have survived the inactivation process.  Second, Youngner’s test made it possible to quickly test the vaccine’s ability to induce anti-poliovirus antibodies (1). [Youngner based his color test on an earlier observation by John Enders, Tom Weller, and Fred Robbins, that metabolic activity (as indicated by a drop in pH) was less in cultures inoculated with live virus than in control cultures (2, 3). In Youngner’s test, a color change of phenol red, resulting from a shift in pH, served as an indicator of virus activity, or of antibody activity.]

Some sources credit Youngner with having devised the process for inactivating the virus. But, that is correct in a very limited sense only. Salk selected incubation in formalin as the means to disable the virus. In truth, Salk learned of that approach a decade earlier while doing postgraduate studies under Thomas Francis at the University of Michigan. Francis was then using formaldehyde to produce his killed influenza vaccine (2).

What’s more, Salk’s choice of formalin to generate his polio vaccine was bold. Earlier, in the 1930s, Canadian scientist Maurice Brodie tested a formalin-killed polio vaccine in twelve children, with disastrous results. Several of the children developed paralytic poliomyelitis (4).

Clearly, too little exposure to formalin could leave enough live virus to cause paralytic poliomyelitis or death. On the other hand, too much exposure could so badly damage the virus’ proteins that they might no longer induce an immune response against the live virus. Brodie did not have analytical procedures to ensure that he had inactivated his vaccine to safe levels. In contrast, it was clear to Salk that getting the correct balance would be vital to his vaccine project, and Youngner’s color test was the means for doing so. Youngner used his test to determine that six days of incubation in a 1:4,000 formalin solution would result in one live virus particle in 100 million doses of the vaccine (5).

Since Youngner’s inactivation curve was based on only a few data points, and since it was likely that the slope of the curve might flatten out after a time, Salk added a margin of safety of six extra days. Thus produced, the vaccine induced antibody production in monkeys, while showing no signs of causing paralysis or other problems.

By 1954, 800,000 children had been successfully immunized against polio in the first clinical trial of the vaccine. In April 1955, the outcome of the trial would be announced to a very grateful public.

By 1957, Salk’s vaccine team at Pittsburgh was no longer needed, and was dispersing. Salk was making plans to leave Pittsburgh for California, where he would found the prestigious Salk Institute. Youngner, now 34 years-old, remained at Pittsburgh, where he would begin his own distinguished career.

Although Youngner was now independent of Salk, he remained bitter over his former boss’s failure to acknowledge the underlings who had labored so diligently behind the scenes to bring the vaccine to fruition. “The first rule we learned was to call him ‘Dr Salk,’ never Jonas. He would speak to us through a wall of notes and memos…Here was a guy who could always find an hour to brief some reporter at the local Chinese restaurant, but could never find the time to sit down with his own people (6).”

Youngner was particularly appalled by events involving the paper he wrote describing his color test. “After I had what I considered to be a good draft…I gave my copy to Jonas for his comments. It should be noted this was 1954, the pre-Xerox, pre-word-processing era. I had made a working transcript of the paper for my own use and it was this copy that I handed to him. Also, it should be noted that the title page had the authors listed as ‘J.S. Youngner and E.N. Ward (6).’” Elsie Ward, who served as Youngner’s technician, was a zoologist who specialized in growing viruses.

Salk intended to read Youngner’s manuscript while away on a trip.  When Salk returned a week later, he claimed that he had lost the manuscript, but that he had jotted down some notes from which he was able to produce a draft of his own. Youngner was rather incredulous that a person as meticulous and disciplined as Salk could lose such an important manuscript. Youngner’s skepticism was further roused by the fact that Salk’s version contained all the data in Youngner’s original manuscript. Salk explained that incongruity, alleging that he found Youngner’s tables, but not the text.

In any case, Youngner was especially upset by a specific change Salk made to the title page of the manuscript: “The authors were now ‘Jonas E. Salk, J.S. Youngner, and Elsie N. Ward.’ When I (Youngner) questioned the change, Jonas said that since he had to reconstruct the whole paper it was only fair that his name go first…It was obvious to me then, and is more so now, that he considered the advance in this paper a major one and he wanted his name associated with it, even though at the time he had done nothing in the lab (no kidding!) or of an advisory nature to initiate or carry out the work (6).”

Youngner could grudgingly accept that project leaders often used their senior position to appear as co-authors, or even principal authors, on papers emanating from their labs, even if their contributions were minimal. What troubled Youngner in this instance was not that Salk pulled rank, but rather his seeming duplicity.

In yet another instance—the 1955 public announcement of the successful outcome of the clinical trial—Youngner again sensed “a pattern of deception on Salk’s part to take undue credit for the discoveries of others (6).” Salk advocated for the announcement to happen at the University of Pittsburgh. However, the National Foundation for Infantile Paralysis (better known as the “March of Dimes”), which funded the vaccine project, chose the University of Michigan in Ann Arbor as the site for the announcement. That was where Michigan professor Thomas Francis supervised the evaluation of the field trial. [Note that the NIH was not able to fund research back then the way it can today. Thus, the polio vaccine project was supported nearly entirely by private donations to the National Foundation.]

Thomas Francis spoke first. Then, when Salk spoke, he acknowledged the more prominent players in the vaccine project, including Thomas Francis, Harry Weaver (director of research at the National Foundation), Tom Rivers (chairman of the advisory committees on research and vaccines for the  National Foundation), and Basil O’Connor (law partner of Franklin Roosevelt, recruited by Roosevelt in 1928 to raise funds for polio patients at Roosevelt’s Warm Springs Foundation, and a co-founder with Roosevelt of the National Foundation in 1938; (2)). Salk then acknowledged various deans and trustees at the University of Pittsburgh. Yet, he made no mention whatsoever of his dedicated coworkers in his laboratory. They had been expecting at least some recognition from their boss.

Some of Salk’s defenders argued that Salk had acted in the best scientific tradition by prefacing his printed remarks with the phrase, “From the Staff of the Virus Laboratory by Jonas E. Salk, M.D.” But, this was small consolation to Youngner and others of Salk’s coworkers, who expected to be individually acknowledged for their exhausting work on behalf of the life-saving vaccine. Indeed, they felt betrayed.

At any rate, the 1955 announcement of the success of the polio vaccine field trials was joyously received by the public. And while Youngner remained embittered over Salk’s slighting of his coworkers, he nonetheless understood that from the point of view of the National Foundation, “it was much easier to continue raising money when you have a hero, and they had an enormous public relations department that took up Jonas’ name as the hero, which he deserved…But in the meantime, Jonas was, how shall I say, not very generous to his colleagues and he made sure that nobody else was ever mentioned (6).”

The following excerpt is from Polio: An American Story (6). “In September 1963, Salk returned to Pittsburgh to attend the unveiling of his portrait in the auditorium of the University’s medical complex, a stone’s throw from the hospital where he had done his historic polio research. Before the ceremony, Salk told Dean George Bernier that he wished to speak privately with his former assistant, Julius Youngner, now a distinguished professor at the school of medicine. The two men hadn’t talked or crossed paths since Salk’s move to California in 1961. Salk saw the meeting as a courtesy to the only remaining member of his laboratory staff; Youngner had a different agenda. Speaking softly, he recalled, he slowly released the ‘hurt’ he had bottled up for more than thirty years. ‘Do you still have the speech you gave in Ann Arbor in1955? Have you ever reread it?’ Youngner began. ‘We were in the audience, your closest colleagues and devoted associates, who worked hard and faithfully for the same goal that you desired…Do you remember who you mentioned and who you left out? Do you realize how devastated we were at that moment and ever afterward when you persisted in making your coworkers invisible? Do you know what I’m saying,’ I asked. He answered that he did…Jonas was clearly shaken by these memories and offered little response.’…The two men engaged in some uncomfortable small talk before Dean Bernier returned to escort them to the ceremony. Speaking later to a reporter, Youngner admitted, ‘I got a lot of things off my chest. I’m beyond the point where I pull my punches with him. I think it was the first time he ever heard it so graphically.’ Asked if he had any regrets about working for Salk, Youngner replied: ‘Absolutely not. You can’t imagine what a thrill that gave me. My only regret is that he disappointed me.”’

Epilogue:

Jonas Salk is deservedly celebrated for developing the killed polio vaccine. That vaccine, together with Albert Sabin’s live attenuated vaccine, which followed soon afterwards, has nearly eradicated polio worldwide. Importantly, Sabin and other polio researchers believed that only a live vaccine could induce a level of immunity sufficient to protect against a challenge with live virulent virus. Nonetheless, Salk persevered in his conviction that a killed vaccine could protect against polio, and he was right.

Salk founded the prestigious Salk Institute in 1963. Yet he never himself made another notable contribution to science.

Youngner may be best known for his work on the Salk vaccine. Yet he had a distinguished career of his own at the University of Pittsburgh after Salk left. Youngner is especially noted for his contributions to interferon research. These include his finding that non-viral agents could trigger interferon induction in animals. And, in collaboration with colleague Samuel Salvin, he identified a second type of interferon, now known as gamma-interferon. Youngner also helped to explain the antiviral-effect of interferon, and he was the first researcher to demonstrate that some viruses express countermeasures against interferon.

Youngner also made important findings in the area of persistent virus infections. Importantly, he demonstrated that defective viral variants, including temperature-sensitive mutants, can play a role in the establishment and maintenance of viral persistence; doing so by impairing (modulating) the replication of the wild-type parental viruses. Based on that principle, Youngner sought to develop dominant-negative mutants of influenza virus as a novel means of anti-influenza therapy. In addition, Youngner and colleague Patricia Dowling developed a novel live attenuated vaccine against equine influenza virus, based on a cold-adapted influenza virus, which can replicate only at the temperatures found in the respiratory tract. That live vaccine was the first to prevent a serious respiratory disease of horses.

Julius Youngner, 2010

References:

  1. Salk, J.E., Youngner, J.S, Ward, E.N. (1954). Use of Color Change of Phenol Red as the Indicator in Titrating Poliomyelitis Virus or Its Antibody in a Tissue–Culture System,” American Journal of Epidemiology. 60: 214–230.
  2. Jonas Salk and Albert Sabin: One of the Great Rivalries of Medical Science, Posed on the blog March 27, 2014.
  3. John Enders: “The Father of Modern Vaccines,” Posted on the blog August 4, 2016.
  4. Vaccine Research using Children, Posted on the blog, July 7, 2016.
  5. Williams, G., Paralysed with Fear: The Story of Polio, Palgrave Macmillan, 2013.
  6. Oshinsky, D.M., Polio: An American Story, Oxford University Press, 2005.
Advertisements

Hilary Koprowski: Genesis of a Virologist

Several years before Jonas Salk and Albert Sabin developed their famous polio vaccines, Hilary Koprowski (1916-2013) in fact developed the world’s first effective, but much less well known polio vaccine (1, 2). Koprowski’s vaccine was used world-wide, but it was never licensed in the United States, ultimately losing out to Sabin’s vaccine.

Koprowski’s reputation was tarnished in 1950, when he tested his live polio vaccine on 20 children at Letchworth Village for mentally disabled children, in Rockland County, NY; an episode recounted in a recent posting Vaccine Research Using Children (1). Koprowski reported on the Letchworth Village trials at a 1951 conference of major polio researchers. Although his vaccine induced immunity in the children, and caused no ill effects, many scientists in the audience were horrified that he actually tested a live polio vaccine in human children. Afterwards, Sabin shouted at him: “Why did you do it? Why? Why?”

Although Koprowski’s polio vaccine was supplanted by the Salk and Sabin vaccines, his demonstration, that a live polio vaccine could be safe and effective, paved the way for Sabin to develop his live polio vaccine. Moreover, Sabin developed his vaccine from a sample of attenuated poliovirus that he received from Koprowski.

There is much more to tell about Koprowski. This posting relates some of the remarkable earlier events of his life, including his harrowing escape from Poland on the eve of the Second World War; a flight which inadvertently led to his career in virology. A subsequent posting will recount the now discredited, although sensational at the time, accusation that Koprowski’s polio vaccine gave rise to the HIV/AIDS epidemic.

Koprowski was born and grew up in Warsaw, where he earned a medical degree from Warsaw University in 1939. He also was an accomplished pianist, having studied piano from the age of 12 at the prestigious Warsaw Conservatory, where Chopin is said to have studied. Koprowski eventually earned a music degree from the Conservatory. He recalled, “…the first year I was the youngest and voted second best in the class (3).”

koprowski

Hilary Koprowski in Warsaw (2007)

In 1938, while Koprowski was in medical school, he married classmate Irena Grasberg who, in later years, would wonder how they had found the time for their courtship. Each had to contend with a demanding medical school program, while Hilary’s piano studies at the Conservatory was a full time program in itself (3). Irena recalled a day before both of them had an anatomy exam, and Hilary had an important recital. Hilary practiced a recital piece, while simultaneously studying a chart on the music rack showing the bones of the hand; all the while as Irena read anatomy to him.

Koprowski eventually chose a career in medicine, rather than one in music. As he explained: “…the top of the music pyramid is much narrower than that of medicine, where there is more space for successful scientists (3).” Koprowski rated himself only fourth best in his class at the Warsaw Conservatory, and he needed to excel. Yet he may have underrated himself. His piano professor at the Conservatory was “greatly disappointed” when he chose to enter medicine (3). [After the 1944 Warsaw uprising, Koprowski’s piano professor was arrested and beaten to death by German soldiers (see below and 3).] In any case, Koprowski continued to play the piano, and he even did some composing in his later years.

Germany invaded Poland in September 1939, setting off the Second World War. As German bombs were falling on Warsaw, Koprowski answered the call for Polish men to go east, where Polish forces were organizing to resist the Germans. Irena, now pregnant, and Hilary’s mother went with him, while his father chose to remain behind. They made their way in a horse-drawn hay wagon, traveling at night to avoid German planes that were strafing the roads during the day. After a week or so on the road, they encountered refugees moving in the opposite direction. Those refugees told them that Russia had signed a pact with Germany and was now invading Poland from the east (Aside 1). So the three Koprowskis joined the flood of refugees moving to the east. When they arrived back in Warsaw, they found the city in ruins. Many of their friends and neighbors had been killed or were seriously wounded, and the city was occupied by German soldiers.

[Aside 1: The German–Soviet Non-aggression Pact was signed in Moscow in August 1939, as a guarantee of non-belligerence between Nazi Germany and the communist Soviet Union. Hitler broke the pact in June 1941 when Germany attacked Soviet positions in eastern Poland. Hitler had no intention of keeping to the pact. However, it temporarily enabled him to avoid having to fight a war on two fronts—against Britain and France in the west and the Soviet Union in the east.]

Once Germany had conquered Poland, German and Polish Jews began to be sent to concentration camps set up in Poland. The Koprowskis, who were Jewish (Salk and Sabin too were descendants of eastern European Jews), quickly made plans to leave Poland. Their first destination was to be Rome. Hilary’s father went there first to arrange living conditions for the family. To facilitate the escape of Hilary’s father from Poland, Hilary and Irena wrapped him in bandages, hoping that the authorities might gladly believe they were letting a very frail individual depart from the country.

Hilary, Irena, and Hilary’s mother then traveled by train from Warsaw to Rome. It was a harrowing trip. Irena was pregnant, and the Gestapo was roaming the trains. They feared that they might have been arrested at any time.

In Rome, the Koprowski family’s main concern was the safety of Irena and her unborn baby. Since Irena had an aunt in Paris, who would know of a good doctor there, the family thought that Paris would be a safe place for the baby to be born. Thus, Irena left for Paris, accompanied by Hilary’s father. She gave birth to Claude five days after arriving there.

Hilary did not go with Irena to France. If he had done so, he would have been impressed immediately into the Polish Army that was forming there to fight the Germans. Yet he knew that he would eventually have to leave Rome. Italy, under Mussolini’s leadership, was poised to enter the Second World War, as an Axis partner of Hitler’s Germany.

After Claude was born, Irena worked as a physician at a psychiatric hospital in Villejuif, just outside of Paris. She was the sole internist there for eight hundred patients. She kept Claude at the hospital, in a locked room, which she would slip to away every three hours to nurse him.

Back in Rome, Hilary continued to play the piano. In fact, he auditioned for, and was accepted by Rome’s L’Accademia di Santa Cecilia, which awarded him a second degree in music. Importantly, his skill at the keyboard enabled him to get visas for himself and his mother to enter Brazil, which the family hoped would be a safe haven. The best students from L’Accademia di Santa Cecilia were often in demand to play for events at the Brazilian embassy in Rome. Thus, on several occasions, Hilary played the piano at the embassy. Brazil’s consul general admired Hilary’s pianism and was pleased to arrange Brazilian entry visas for Hilary and his mother. See Aside 2.

[Aside 2: The day after Hilary arrived in Rome, he volunteered to serve as a medical examiner for a Polish draft board that was set up in the Polish embassy. The draft board’s activity at the embassy—recruiting Poles for the Polish Army—violated diplomatic protocol. In addition, Italy would soon be Germany’s Axis partner in the War. Moreover, Brazil, though neutral in the War, favored the Axis.]

Hilary and his mother had been making plans to leave Italy. Their destination was to be Spain, where they hoped they might unite with Irena, Claude, and Hilary’s father.  From Spain, the family might then go to Portugal, where they could get a boat to Brazil. But, on the very day that Hilary and his mother were to leave Italy, Mussolini issued a proclamation banning any male of military age from leaving the country. So it happened that Hilary’s escape from Italy was blocked at the boat registration. However, his mother rose to the occasion, crying and pleading with the boat registration official that she was sick, that Hilary was her sole means of support, and that she could not go on without him. “The man looked at his watch and said he must go to lunch. He looked at us and said, ‘If the boat leaves before I return, that’s my bad luck (3).’” So, Hilary and his mother boarded the boat, which left before the official returned. [Hilary’s mother was a well-educated woman, and a dentist by profession.]

In Spain, Hilary and his mother stayed at a hotel in Barcelona. Despite the wartime conditions, they were able to communicate, if only sporadically, with Irena and Hilary’s father, who were still in France. Then, after Germany invaded France in 1940, Irena, Claude, and Hilary’s father reunited with Hilary and his mother in Barcelona. [The escape of Irena, Claude, and Hilary’s father from France was far more harrowing than the escape of Hilary and his mother from Italy (See 3 for details).]

The family now needed to get to Portugal, where they could then get a boat to Brazil. Irena had already obtained Portuguese visas for herself and for Claude. But Hilary and his mother only had visas for Brazil. Hilary’s applications for visas at the Portuguese embassy were repeatedly denied, until a fellow Pole at Hilary’s Barcelona hotel advised him of the obligatory bribe that must accompany visa applications. The advice was right-on, and the family (minus Hilary’s father, who chose to go to England) sailed for Brazil without further incident.

In Brazil, Irena found work in Rio de Janeiro as a nurse. But she soon managed to secure a position as a pathologist at the largest hospital in the city. Hilary, on the other hand, could not find a job in medicine and, so, he turned to teaching piano. After six months of teaching unenthusiastic piano students, Hilary by chance recognized a man on the street in Rio who happened to be a former schoolmate from Warsaw. The man also happened to be working at the Rockefeller Foundation’s outpost in Rio. He told Hilary that the Foundation was looking for people, and he also told Hilary who he should contact there. Hilary interviewed at the Foundation the next day, and was told to report for work the day after that.

The Foundation assigned Hilary to research how well, and for how long the attenuated yellow fever vaccine—developed by Nobel laureate Max Theiler in 1935 (4) —might protect against yellow fever. The disease was endemic in Brazil, and it was actually the Rockefeller Foundation’s first priority.

Hilary’s supervisor at the Foundation was Edwin Lennette; a staff member of the International Health Division of the Rockefeller Foundation, assigned to its Brazilian outpost, specifically because of his interest in yellow fever. In 1944, Lennette would be reassigned to the Rockefeller Foundation laboratory in Berkeley, California, where he would establish the first diagnostic virology laboratory in the United States. Indeed, Lennette is known as one of the founders of diagnostic virology. But, in Brazil, he introduced Hilary Koprowski to virology.

Hilary’s apprenticeship under Lennette was going very well. It would result in nine papers—published between 1944 and 1946— that Hilary would co-author with Lennette. Moreover, Lennette was interested in other viruses, in addition to yellow fever. Thus, their co-authored papers included studies of Venezuelan equine encephalitis virus, Japanese encephalitis virus, St. Louis encephalitis virus, and West Nile virus, as well as yellow fever.

Most importantly, Koprowski’s work under Lennette introduced him to Max Theiler’s methods and approach to viral attenuation. In brief, Theiler found that propagating yellow fever virus in an unnatural host—chick embryos—caused the virus to adapt to that host, thereby reducing its capacity to cause disease in humans.  Koprowski would later acknowledge that Theiler provided him with a “most encouraging model” for attenuating poliovirus. [Koprowski attenuated poliovirus by propagating it first in mice and then in rats. Recall that Sabin developed his live polio vaccine from attenuated poliovirus that he received from Koprowski (1).] See Asides 3 and 4.

[Aside 3: The rabies vaccine, which Louis Pasteur developed in 1885, is often referred to as the first attenuated virus vaccine. Nevertheless, while Pasteur did passage his vaccine virus in rabbit spinal cords, the virus may have been killed when the spinal cords were later dried for up to fourteen days. Also, in Pasteur’s day, nothing was known about immunity or mutation, and viruses had not yet been identified as microbes distinct from bacteria. The yellow fever vaccine developed by Max Theiler at the Rockefeller Institute (now University) in New York may have been the first deliberately attenuated viral vaccine.]

[Aside 4: Koprowski and Lennette were among the first researchers to observe that infection by one virus (yellow fever, in this instance) might inhibit the growth of another unrelated virus (West Nile virus, in this instance). That is, they had inadvertently detected what later would be known as interferon. Yet while they looked for an anti-viral substance in their tissue culture media, and while their results suggest that it actually was there, they stated in their summary that nonspecific anti-viral factors were not present (5). Koprowski and Lennette collaborated again in the 1970s; this time to investigate subacute sclerosing panencephalitis, a rare late complication of measles infection that results in neurodegeneration.]

Hilary continued to give piano recitals in Brazil, regretting only that he did not have time to practice the piano as much as he would have liked. Nonetheless, his piano playing expanded his circle of friends to include musicians, artists and writers, in addition to his fellow scientists. Moreover, Irena was satisfied with her medical practice, and with the many friends and rich social life that she and Hilary had in Brazil.

Earlier, in 1940, while Hilary was still in Rome, and expecting that the family would soon have to leave Europe, he believed that the United States would likely be the best destination for them. Thus, he applied to the United States for visas. He had nearly forgotten those applications when, in 1944, their numbers came up.

The Koprowski family now faced somewhat of a dilemma. It was happily settled in Brazil, and had no prospects in the United States. On the other hand, the Rockefeller Foundation’s yellow fever project was drawing to a close, and the Foundation was planning to leave Rio. Importantly, coming to America was now a “dream come true (3)”.  So, in December 1944, the Koprowskis boarded an aging steamer in Brazil, and sailed under wartime blackout conditions, through German submarine-infested waters, for New York City.

During Hilary’s his first days in America, he used the Rockefeller Institute library in Manhattan to work on manuscripts reporting his research in Brazil. During one of his visits to the Rockefeller, he happened to meet Peter Olitzky (Aside 5), an early polio researcher there, who arranged for Hilary to meet Harold Cox, the director of the virology department at Lederle Laboratories, in Pearl River, New York.  Hilary interviewed with Cox, who offered him a research position at Lederle, which Hilary accepted. Meanwhile, Irena was appointed an assistant pathologist at Cornell Medical College in Manhattan.

[Aside 5: In 1936, Olitzky and Sabin collaborated on a study at the Rockefeller Institute, which, although carefully done, wrongly concluded that poliovirus could attack nerve cells only; a result that did not bode well for the development of an attenuated polio vaccine.]

At Lederle, Hilary began the experiments that led to the world’s first successful polio vaccine. In 1950 he tested the live vaccine in eighteen mentally disabled children at Letchworth Village (1). None of these children had antibodies against poliovirus before he vaccinated them, but each of them was producing poliovirus antibodies after receiving the vaccine. Importantly, none of the children suffered ill effects. What’s more, Koprowski did not initiate the test. Rather, a Letchworth Village physician, fearing an outbreak of polio at the facility, came to Koprowski’s office at Lederle, requesting that Koprowski vaccinate the Letchworth children (1).

References:

   

  1. Vaccine Research Using Children, Posted on the blog July 7, 2016.
  2. Jonas Salk and Albert Sabin: One of the Great Rivalries of Medical Science, Posed on the blog March 27, 2014.
  3. Roger Vaughan, Listen to the Music: The Life of Hilary Koprowski. Springer-Verlag, 2000.
  4. The Struggle Against Yellow Fever: Featuring Walter Reed and Max Theiler, Posted on the blog May 13, 4014.
  5. Lennette EH, Koprowski H., 1946. Interference between viruses in tissue culture, Journal of Experimental Medicine, 83:195–219.

 

 

 

 

 

Maurice Hilleman: Unsung Giant of Vaccinology

In January 2005, more than 100 of the world’s most renowned biomedical researchers got together to pay tribute to the 85-year-old Maurice Hilleman. When it was Hilleman’s turn to address the gathering, he alluded to them as his “peers in the world of science.” Referring to Hilleman’s gracious comment, science journalist Alan Dove wrote: “By any objective measure, a gathering of Maurice Hilleman’s scientific peers would not fill a telephone booth.” (1)

Hilleman truly was a giant in the history of virology. But, if you have only a vague idea of who Hilleman was or of his achievements, you are not alone. Anthony Fauci, director of the U.S. National Institutes of Allergy and Infectious Diseases, who was present at the gathering, noted: “Very few people, even in the scientific community, are even remotely aware of the scope of what Maurice has contributed….I recently asked my post-docs whether they knew who had developed the measles, mumps, rubella, hepatitis B and chickenpox vaccines. They had no idea,” Fauci said. “When I told them that it was Maurice Hilleman, they said, ‘Oh, you mean that grumpy guy who comes to all of the AIDS meetings?’”

hillemanMaurice R. Hilleman: The greatest vaccinologist.

Consider this. Hilleman developed nine of the 14 vaccines routinely recommended in current vaccine schedules. These are the vaccines for the measles, mumps, rubella, hepatitis A, hepatitis B, and chickenpox viruses, and for meningococcal , pneumococcal, and Haemophilus influenzae bacteria. Moreover, he was the first to forecast the arrival of the 1957 Asian flu and, in response, led the development of a flu vaccine that may have saved hundreds of thousands or more lives worldwide (2). And, independently of Robert Huebner and Wallace Rowe, he discovered cold-producing adenoviruses, and developed an adenovirus vaccine. Overall, Hilleman invented nearly 40 vaccines. And, he was a discoverer of simian virus 40 (SV40). If the above accomplishments were not enough to ensure his fame, he also was the first researcher to purify interferon, and the first to demonstrate that its expression is induced by double-stranded RNA.

[Aside: I first became aware of Maurice Hilleman 44 years ago. It was in the context of his 1959 discovery of SV40, which I came across only because I was beginning my post-doctoral studies of the related murine polyomavirus. Bernice Eddy, at the U. S. National Institutes of Health (NIH), was probably the first to discover SV40, which she detected in early lots of the Salk polio vaccine (3). Hillman, then at Merck & Co, independently discovered the same virus in rhesus monkey kidney cell cultures, in which the polio vaccine was being produced. Hilleman gave SV40 its name. It was the 40th simian virus the Merck lab found in the monkey kidney cells. In 1961, both Eddy and Hilleman found that inoculating SV40 into hamsters causes tumors in the animals. Merck withdrew its polio vaccine from the market. But, by then, live SV40 had been unknowingly injected into hundreds of millions of people worldwide! More on this in a future posting.]

We begin our account of Hilleman’s achievements with his development of the mumps vaccine. In the days before the vaccine, mumps struck about 200,000 children in the United States, annually. Yet except in rare circumstances, the infection was mild, and was generally regarded as a childhood rite of passage. There is a sweetness to the story of the mumps vaccine that I hope you might enjoy.

The tale began at about 1:00 AM, on March 21, 1963, when 5-year-old Jeryl Lynn Hilleman ambled into her father’s bedroom complaining of a sore throat. Jeryl Lynn’s father felt his daughter’s swollen glands, and knew in a flash that it was mumps. And, while I suspect that many lay parents back in the day would also have recognized Jeryl Lynn’s symptoms, few would have done what her father did after first comforting his daughter. Although it was already past midnight, Maurice hopped into his car and drove the 20 minutes to his lab at Merck & Co. to pick up some cotton swabs and beef broth. Returning home, he then awakened Jeryl Lynn, gently swabbed her throat, and immersed the swabs in the nutrient broth. Next, he drove back to his lab and put the inoculated broth in a freezer.

Hilleman made the early A.M. dashes to his lab and back because he had to leave in the morning for a conference in South America, and his daughter’s infection might have cleared by the time he returned home from there. So, upon his return from South America, Hilleman, thawed the frozen sample from his daughter’s throat and inoculated it into chick embryos. Serial passage of the mumps virus in the chick embryos eventually generated attenuated mumps virus that in 1967 would serve as a live mumps vaccine.

The virus in the vaccine was dubbed the Jeryl Lynn strain, in honor of its source. Years later, an adult Jeryl Lynn Hilleman noted that her father had a need to be “of use to people, of use to humanity.” She added: “All I did was get sick at the right time, with the right virus, with the right father.”

We’ll have a bit more to say about the mumps vaccine shortly. But first, a few words about measles and rubella.

If mumps was not a major killer, measles certainly was. Before Hilleman and his colleagues introduced their measles vaccine (Rubeovax) in 1962, there were 7 to 8 million measles fatalities worldwide each year, and virtually all of the victims were children. Hilleman developed his attenuated measles vaccine from a measles strain isolated earlier by John Enders. Hilleman attenuated the Enders isolate by putting it through 80 serial passages in different cell types.

[Aside: In a previous posting, we noted that Enders, together with colleagues Thomas Weller and Frederick Robbins, shared a Nobel Prize in Physiology or Medicine for growing poliovirus in non-nervous tissue (3). Apropos the current story, bear in mind that Salk and Sabin developed polio vaccines that have nearly rid the world of this once dread virus. Nevertheless, the Nobel award to Enders, Weller, and Robbins was the only Nobel award ever given in recognition of polio research!]

Rubeovax was somewhat tainted by its side effects; mainly fever and rash. While these reactions were successfully dealt with by combining Rubeovax with a dose of gamma globulin, in 1968 Hilleman’s group developed a new, more attenuated measles strain by passage of the Rubeovax virus 40 more times through animal tissues. Hilleman dubbed the new measles strain “Moraten,” for “More Attenuated Enders.” The new measles vaccine, Attenuvax, was administered without any need for gamma globulin.

Our chronicle continues with the rubella vaccine. Rubella poses its greatest danger to fetuses of non-immune pregnant woman, particularly during the first trimester of pregnancy. In up to 85% of these women, infection will result in a miscarriage or a baby born with severe congenital abnormalities. An outbreak of rubella began in Europe in the spring of 1963, and quickly spread worldwide. In the United States, the 1963 rubella outbreak resulted in the deaths of 11,000 fetuses, and an additional 20,000 others born with birth defects (e.g., deafness, heart disease, cataracts).

Hilleman had been working on a rubella vaccine at the time of the 1963 outbreak. But, he was persuaded to drop his own vaccine and, instead, refine a vaccine (based on a Division of Biologics Standards’ rubella strain) that was at the time too toxic to inoculate into people. By 1969 Hilleman was able to attenuate the DBS strain sufficiently for the vaccine to be approved by the FDA.

Next, and importantly, Hilleman combined the mumps, measles, and rubella vaccines into the single trivalent MMR vaccine, making vaccination and, hence, compliance vastly easier. Thus, MMR was a development that should have been well received by many small children and their mothers, as well as by public health officials.

In 1978 Hilleman found that another rubella vaccine was better than the one in the trivalent vaccine. Its designer, Stanley Plotkin (then at the Wistar Institute), was said to be speechless when asked by Hilleman if his (Plotkin’s) vaccine could be used in the MMR. Merck officials may also have been speechless, considering their loss in revenues. But for Hilleman, it was simply the correct thing to do.

Like Jonas Salk and Albert Sabin before him (3), Maurice Hilleman was never awarded a Nobel Prize. There is no obvious reason for the slight in any of these three instances. In Salk’s case, it may have been because Alfred Nobel, in his will, specified that the award for Physiology or Medicine shall be for a discovery per se; not for applied research, irrespective of its benefits to humanity. But, Max Theiler received the Nobel Prize for producing a yellow fever vaccine. What’s more, the Nobel committee seemed to equivocate regarding the discovery that might have been involved in that instance. Regardless, the Nobel award to Theiler was the only Nobel Prize ever awarded for a vaccine! [A more complete accounting of the development of Theiler’s yellow fever vaccine can be found in The Struggle Against Yellow Fever: Featuring Walter Reed and Max Theiler, now on the blog.]

Sabin had done basic research that perhaps merited a Nobel Prize (3). But, the Nobel committee may have felt uneasy about giving the award to Sabin, without also recognizing Salk. Or, perhaps the continual back-and-forth carping between supporters of Salk and Sabin may have reduced enthusiasm in Stockholm for both of them.

Yet by virtually any measure, Hilleman’s achievements vastly exceeded those of Salk, Sabin, Theiler, and just about everyone else. His basic interferon work alone should have earned him the Prize. Hilleman’s group demonstrated that certain nucleic acids stimulate interferon production in many types of cells, and detailed interferon’s ability to impede or kill many viruses, and correctly predicted its efficacy in the treatment of viral infections (e.g., hepatitis B and C), cancers (e.g., certain leukemias and lymphomas), and chronic diseases (e.g., multiple sclerosis). What’s more, Hilleman developed procedures to mass-produce and purify interferon. And, regarding his unmatched achievements as a vaccinologist, he did more than merely emulate Pasteur’s procedures for developing attenuated viral vaccines. His hepatitis B vaccine was the first subunit vaccine produced in the United States. It was comprised of the hepatitis B surface antigen (HBsAg), which Hilleman purified from the blood of individuals who tended to be infected with hepatitis B virus (e.g., IV drug abusers). Subsequently, to avoid the potential danger of using human blood products in the vaccine, Hilleman developed recombinant yeast cells that produced the HBsAg. And, Hilleman’s meningococcal vaccine was the first vaccine to be based on polysaccharides, rather than on a whole pathogen or its protein subunits.

So, why then was Hilleman bypassed by the Nobel committee? John E. Calfree, in The American, wrote: “As the 80-plus-year-old Hilleman approached death, Offit and other academic scientists lobbied the Nobel committee to award Hilleman the Nobel Prize for Medicine, based partly on his vaccine work and partly on his contributions to the basic science of interferons. The committee made clear that it was not going to award the prize to an industry scientist.” (4) [Paul Offit, referred to here, is the co-developer of the rotavirus vaccine, Rotateq, and a biographer of Hilleman.]

Calfree also notes that Hilleman’s tendency towards self effacement, and his absence from the academic and public spotlight, may also have worked against him. And, unlike Salk, whose name was closely linked to his polio vaccine (3), Hilleman’s name was never associated with any of his nearly forty vaccines. [Yet in the case of Jonas Salk, his public acclaim is generally believed to have hurt him in the eyes of his colleagues and of the Nobel committee.]

Considering the enormity of Hilleman’s contributions, his anonymity was really quite remarkable. As Calfree relates: “In one of the most striking of the dozens of anecdotes told by Offit, Hilleman’s death was announced to a meeting of prominent public health officials, epidemiologists, and clinicians gathered to celebrate the 50th anniversary of the Salk polio vaccine. Not one of them recognized Hilleman’s name!”

With Hilleman’s public anonymity in mind, we conclude our account with the following anecdote. In 1998, a Dr. Andrew Wakefield became a celebrity and hero in the eyes of the public. How this happened, and its consequences are troubling for several reasons, one of which is that it brought undeserved suffering to the self-effacing and benevolent Maurice Hilleman. The Wakefield incident merits, and will have a full-length blog posting of its own. But for now, in 1998 Wakefield authored a report in the prestigious British journal The Lancet, in which he claimed that the MMR vaccine might cause autism in children. The story had a bizarre series of twists and turns, with Wakefield and co-authors eventually issuing a retraction. The immediate cause of the retraction was the disclosure that Wakefield, on behalf of parents of autistic children, had accepted funding to investigate a link between the MMR vaccine and autism. The purpose of the investigation was to determine whether a legal case against the vaccine manufacturer might have merit. In addition to the obvious conflict of interest, Wakefield’s paper had serious technical flaws as well. At any rate, a number of independent studies subsequently demonstrated that there is no causal link between the MMR vaccine and autism. And, in 2010 Wakefield was barred by the British Medical Society from the practice of medicine. But the harm had been done. Hilleman had become the recipient of hate mail and death threats. And, more important to Hilleman I expect, many worried parents, even today, prevent their children from receiving the MMR vaccine (5). Ironically, the very success of the MMR vaccine enabled people to forget just how devastating measles and rubella could be.  Maurice Hilleman succumbed to cancer on April 11, 2005.

1. Nature Medicine 11, S2 (2005)
2. Opening Pandora’s Box: Resurrecting the 1918 Influenza Pandemic Virus and Transmissible H5N1 Bird Flu  On the blog.
3. Jonas Salk and Albert Sabin: One of the Great Rivalries of Medical Science  On the blog
4. Calfree, J.E., Medicine’s Miracle Man , The American, January 23, 2009
5. Reference 4 contains a somewhat similar tale, in which a 1992 article in Rolling Stone attributed the emergence of HIV to Hillary Koprowski’s polio vaccine. It created a sensation but, as might be expected, there was no evidence to support its premise.