Tag Archives: Auschwitz

The Life and Legacy of George Klein: Cancer Virus Pioneer and Witness to the Holocaust

George Klein, professor emeritus of tumor biology at the Karolinska Institute in Stockholm, where he worked with his wife Eva from the very beginning, passed away on December 10, 2016, at the age of 91. Klein was best known for discovering that Epstein-Barr virus (EBV)—the herpesvirus now known to cause infectious mononucleosis—causes two human cancers, Burkitt’s lymphoma and nasopharyngeal carcinoma. Moreover, Klein discovered that EBV triggers Burkitt’s lymphoma by facilitating a chromosomal translocation of the cellular c-myc oncogene, resulting in its constitutive expression. Klein also played pioneering roles in developing the concept of tumor-suppressor genes, and in opening the field of tumor immunology. Klein’s key discoveries are summarized below. But, first, Klein, like several other protagonists in these tales, was profoundly affected by events of the Second World War, and by the early days of the Cold War that followed.

From an announcement for a 2015 symposium at the Karolinska Institute, honoring George and Eva on the occasion of their 90th birthdays

George Klein’s Jewish family moved from Eastern Slovakia to Budapest in 1930. Nineteen-year-old George was working as an assistant secretary to the Jewish Council in Budapest when Nazi Germany began its occupation of Hungary in March 1944. Because George had been working for the Jewish Council, in April 1944 he chanced that to see the Vrba-Wetzler Report, known at the time as the “Auschwitz Report.” It was written by, and was secretly transmitted to the Jewish Council by Rudolf Vrba and Alfred Wetzler, two escapees from Auschwitz. It described firsthand the fate of Jews arriving at Auschwitz, and was meant to warn Hungary’s Jews, so that they might hide from, or rebel against their Nazi oppressors.

The Auschwitz report was not publicized in Hungary for reasons explained below. However, George’s supervisor at the Jewish Council gave him permission to tell his relatives and friends of what the report revealed. But they, like most Hungarian Jews, could not believe that such atrocities could actually be taking place. [During May, June, and July 1944, 437,000 Hungarian Jews were deported to Auschwitz; to be “resettled” according to the Nazis. But, in fact, most were murdered in the gas chambers.]

Klein was arrested and pressed into forced labor by the Nazis. Afterwards, since he knew the contents of the Auschwitz Report, he fled when he was about to be ordered to board one of the deportation trains to Auschwitz. Having escaped from almost certain death, he lived underground until January 1945, when the Russian Army liberated Budapest.

Forty-three years later, Klein was watching, Shoa, the monumental (nine-hour-long) French documentary film about the holocaust. Watching the movie, Klein chanced to see a man named Vrba (one of the six principal holocaust witnesses in the film) describe his experiences as a prisoner in Auschwitz. The events that Vrba recounted horrified Klein.

Later in the film, as Vrba described his escape from Auschwitz, Klein suddenly realized, “the report I had been given to read under a promise of secrecy in Budapest in May 1944—at the age of nineteen and at a time when deportations from the Hungarian countryside were at their peak—was identical to the Auschwitz Report of Vrba and Wetzler (1).”

Next in this remarkable tale, Klein decided to try to find Vrba, to “tell him of what enormous help his report had been to me. If I had not known what was awaiting me at the other end of the train trip, I would never have dared to risk an escape. It was not difficult to find Vrba, for it turned out that we were scientific colleagues. He is a professor of neuropharmacology in Vancouver, and I am now (in the Spring of 1987) sitting in a comfortable armchair in the faculty club at a Canadian university, talking with someone who, at first glance, seems quite ordinary. He impresses me as being relaxed and jovial. By now I have also read his book (Escape from Auschwitz, 1964), and I am aware that he has survived more death sentences than anyone else I have ever met (1).”

Vrba (1924–2006), was indeed a professor of pharmacology at the University of British Columbia; a position he held from 1976 until the early 1990s. Note that he and Wetzler were the first prisoners ever to escape from Auschwitz. Vrba’s real name was Walter Rosenberg. Rudolf Vrba was the nom de guerre he used after joining the resistance in his native Czechoslovakia. Afterwards, he made the change legal.

The horrors of the holocaust remained an obsession for Klein, although he was uncertain as to why that was so. “Was it to honor my murdered family, my murdered classmates? Or was it rather to steel myself against the darkest side of our human heritage?” In any case, Auschwitz and the holocaust were the main topics of conversation when Klein met with Vrba.

Vrba took Budapest’s Jewish Council to task for not widely broadcasting the warnings in the Auschwitz Report. He, and others, have alleged that Dr. Kastner, a well-known Zionist leader in Budapest, decided to keep the Report secret, in return for a promise from the Germans to allow sixteen-hundred people, as selected by Kastner, to safely emigrate from Hungary. Klein retorted that he knew Kastner from his work for the Jewish Council, and considered him to be a hero, because he had rescued many, while others tried to rescue only themselves or their own families. [In 1957, Kastner was murdered in Israel by a young man whose family was exterminated by the Nazis. Kastner remains a controversial figure to this day.]

Klein and Vrba next discussed whether dissemination of the Auschwitz Report might have caused Budapest’s Jews to revolt against the Nazi program of annihilation.  Klein argued that of the dozen or so people that he warned, no one believed him. Vrba countered, “You were a mere boy. Why would anyone believe what you were saying? The Jews would certainly have believed their responsible leaders (1).” Nonetheless, Vrba conceded that even the prisoners at Auschwitz were in denial of what they could see with their own eyes: “…prisoners, who knew full well that no one ever returned from the gas chambers, repressed such knowledge as they themselves lined up for execution in front of the chamber doors.”

Klein asked Vrba how he is able to live and function in Vancouver, a pleasant and friendly place, where no one has the slightest concept of what he endured: “…you must go back constantly to those days. You are called in as a witness at trials of old Nazis or their followers, people who claim that the holocaust never happened. You try to describe something that cannot be described in any human language, you try to explain the incomprehensible, you want people to listen to something they do not want to hear (1).” Vrba, in fact, never did reveal his Auschwitz experience to his colleagues. Vrba explained: “What would have been the use? No one who has not experienced it can understand.” Their conversation went on for almost ten hours. Afterwards, they parted like old friends, despite any differences in their views.

In the Fall of that year, Klein was reunited with Vrba in Paris, together with another newfound friend, German scientist Benno Muller-Hill. In 1966, Muller-Hill was a graduate student in Walter Gilbert’s Harvard laboratory, when he purified the lac repressor; the first genetic control protein to be isolated. Muller-Hill then began a second career lecturing and writing about the role of Nazi doctors and scientists in the holocaust. Klein met Müller-Hill for the first time at a meeting at the Institute for Genetics in Cologne, and the two immediately developed a close friendship.

Muller-Hill was in Paris to visit colleagues at the Pasteur Institute, as well as to meet Vrba. Klein was visiting Paris after attending a scientific meeting in Lyon. Vrba was in Paris at the invitation from the French radio service to refute claims of the ultra-right French leader, Jean Marie Le Pen, that the Nazi gas chambers never existed, and that if the Nazis indeed had any intent to annihilate the Jews, it was merely one of many episodes of the war. [Marine Le Pen, currently a leader of France’s ultra-right National Front, and a candidate for the presidency of France, is Jean Marie’s daughter. She was recently taken to task for denying that French officials and police were complicit in the Nazi roundup of more than 13,000 French Jews in July 1942 (they were later deported to Auschwitz). Le Pen also calls for the deportation of all immigrants from France; a stance that mainly targets Muslims.]

Klein and his two companions ambled about Paris on a beautiful Fall afternoon. They strolled around the Luxembourg Gardens, then continued along the banks of the Seine, turned toward the Latin Quarter, and then stood before the façade of Notre Dame. Yet their minds were elsewhere. “Vrba suggested that we visit the holocaust memorial behind Notre Dame…That walk of only a few minutes took us from the noisy tourist crowd to the silence of the museum’s rooms, where you feel alone and isolated among the symbolic chains and barbed wire. A faint glow of sunlight came in through the narrow openings in the wall. We were surrounded by the voices of the victims…We were all completely speechless. Even Vrba’s macabre sense of humor and his sharp sarcasm had fallen silent for the moment (1).”

After they exited from the memorial, they sat down in a small bistro, where Klein asked his two companions whether German scientists and doctors were actual architects of the holocaust or, instead, merely passive followers. “Benno had concluded from his exhaustive documentation that, contrary to what many wanted so desperately to believe, the ‘euthanasia programs’…and the horrible human experiments… could not be ascribed to a small minority of madmen, opportunists, or charlatans. On the contrary, they had been carried out by quite ordinary and in some instances, eminent physicians and scientists… He (Verba) thought … that would not explain why so many apparently ordinary people took part in the murders without showing any signs of remorse, or how the annihilation program could have been carried out with such efficiency… The discussions between Benno and Vrba continued for several hours (1).”

The day became even more notable later, since Klein had arranged for the threesome to have dinner that evening with Francois Jacob. After a glass of sherry in Jacob’s Latin Quarter apartment, the foursome went to a small restaurant around the corner.

Francois Jacob, and fellow Pasteur Institute scientist Jacques Monod, were awarded Nobel Prizes for their work together on the regulation of lactose metabolism in E. coli (2). More apropos the current episode, Jacob and Monod each received France’s highest military honors for his service during the Second World War—Jacob for his heroism serving with the Free French forces, and Monod for his heroism in the Resistance (2). Yet Jacob’s harrowing escape from Nazi-occupied France at 19-years in age, and his wartime exploits as one of Charles De Gaul’s most highly decorated volunteers, were barely known to his three dinner companions.

At first, Klein was somewhat worried that his friends might not like each other. Jacob often found conversation to be difficult; partly because the thousands of pieces of shrapnel that he carried in his body from the war, made it hard for him to sit comfortably. [Jacob’s wartime wounds prematurely ended his surgical career, and led him to turn to a career in science (2).] But, the get-together didn’t go badly at all.

Conversation eventually turned to the issue of holocaust deniers, as well as to those who would put the past completely behind them. As they talked, the incongruity of the scene suddenly struck Klein. They were sitting in a “first-class Parisian restaurant, surrounded by elegant people, having a very nice dinner in the best French tradition.” “…why did the three of us, with Jacob listening, choose to spend that beautiful Saturday in Paris compulsively focusing our attention on the black birds? We were all citizens of free countries, living well in peaceful times. Were we haunted by feelings of guilt toward the dead? Were we afraid that the whole experience would recur if we let go? We knew that the wide and relentless river of history is rarely influenced by knowledge of the past. In no more than one or two generations, archives of extreme horror turn into scraps of faded paper, with no more influence than dried leaves. I suddenly felt that we were like a traveler with a fear of flying, forcing himself to stay awake and keep his seatbelt buckled during the entire flight, obsessed with the idea that the plane would surely crash if he were to fall asleep. But perhaps we had other motives. Perhaps we wanted to feel a solidarity with each other by selecting a more or less taboo subject for our conversation, one avoided by most others. Or did we try to perform a kind of autopsy, using our brains to understand what human minds are capable of at their worst? Have we appointed our brains to serve as the pathologist and the cadaver at the same time?”

The above recounts only a small sampling of Klein’s conversations with Vrba, Muller-Hill, and Jacob, during their day together in Paris. For more, see reference 1.

In January 1945, 19-year-old George Klein emerged from the Budapest cellar where had been hiding during the last weeks of the German occupation. He gazed on the dead soldiers, civilians, and horses that were frozen in the snow, and was struck by the thought that he had survived, despite the likelihood that he would have ended his 19 years in a Nazi gas chamber or a slave labor camp. However, with the city now in Russian hands, George faced a new threat to his freedom; the Russian patrols that were exporting young Hungarians to labor camps in Russia.

Mindful of the danger on the streets, George was yet eager to begin his medical studies. So, he cautiously dodged the Russian patrols as he made his way to Budapest’s medical school, only to find war-torn deserted buildings and dead soldiers there.

Undeterred by the situation in Budapest, George and a friend set out to Szeged, with the hope of attending the medical school there. The journey of 160 miles took the pair five days, by way of a variety of vehicles, including a Russian military truck. In any case, they were admitted to the Szeged university on the same day that they arrived. And while the school was a shadow of its former self, with all the professors having fled to the West, to George, it was a “previously forbidden paradise (3).”

George spent two years in Szeged, and then returned to Budapest when the University reopened there. Back in Budapest, George fell “desperately” in love with Eva Fisher, a fellow medical student. [George describes their whirlwind romance in reference 3.] George now faced a dilemma. Before he met Eva, he finalized plans to visit Stockholm (under the sponsorship of the Jewish Student Club there). But going to Stockholm would mean leaving Eva behind, under conditions in which travel back into Hungary could be risky. Nonetheless, George went to Stockholm, with Eva believing she would never see him again. Yet the trip would be a defining experience for George and, eventually, would be important for Eva too. In Stockholm, George would learn of, and be riveted by the research of renowned cell biologist Torbjörn Caspersson, at Stockholm’s Karolinska Institute.

Caspersson’s research so enthralled George that he diligently pressed Caspersson for a junior research assistantship in his laboratory. But once George had been accepted by Caspersson, he viewed his situation with a “mixture of ecstatic happiness and enormous anxiety.” “I knew virtually nothing…I was halfway through my medical studies…I was desperately in love with a girl whom I had only known during a summer vacation of eight days and who was on the other side of an increasingly forbidding political barrier (3).”

Despite these misgivings, George knew that his future lay in Sweden, rather than Hungary. He had been accepted into Caspersson’s laboratory, and Hungary was falling increasingly under totalitarian Soviet domination. But Eva was still in communist Hungary. So, George risked returning there with one goal; to marry Eva, and then to leave Hungary for good. “The reunion with Eva confirmed what we both already knew: we wanted to live and work together (3).”

But George and Eva didn’t have the necessary documents to get married, nor did Eva have a passport to leave Hungry. Moreover, communist bureaucrats made it increasingly difficult to obtain these documents. In some instances, up to six weeks might be needed. However, George and Eva were daring and resourceful. When told by a police officer that it would take at least three weeks to obtain a marriage license, George suddenly acted on impulse: “I had always heard others tell of such things but I myself had neither seen nor done it. I pulled a fairly modest bill out of my pocket and put it in the policeman’s hand. ‘Pardon me, how much time was it you said?’ ‘I’ll go get it at once,’ he answered (3).”

With similar persistence and ingenuity, George and Eva obtained all their necessary documents, and they were married that very day! One document, a certificate asserting that neither George nor Eva had a venereal disease, would normally require a three-week lab test. But they beseeched an older colleague, now a doctor at a children’s hospital, to write the certificate for them. Their colleague did so, on his Children’s Hospital stationary. George and Eva then went to the prefecture to be married, only to find a disagreeable marriage official, who was determined to leave work for the day. But, when the official leafed through their papers, and saw the venereal disease certificate written on Children’s Hospital stationary: “He laughed until tears ran down his cheeks. This was the funniest thing he had seen during his whole time in service.” He then gladly married the couple.

As the Iron Curtain descended about Hungary, George and Eva left for Sweden, where they would now continue their medical studies. What’s more, Eva joined George in Caspersson’s laboratory at the Karolinska Institute. The couple would work together at the Karolinska until George’s death at the age of 91. [Eva was born to Jewish parents in Budapest in 1925. In 1944 and 1945, she and several members of her family hid from the Nazis at the Histology Institute of the University of Budapest. Encouraged by Caspersson, Eva had an independent research career, while also collaborating with George. She is best known for discovering natural killer cells, and for generating the Burkitt’s lymphoma cell lines, which she and George studied together (see below).]

George and Eva, at the Karolinska Institute, 1979

We conclude with a brief review of some of George Klein’s contributions to virology and to cancer research.

Tumor immunology: In 1960, George and Eva used methylcholanthrene to induce tumors in mice. Next, they surgically removed the tumors, killed them with irradiation, and inoculated them back into genetically compatible mice. Next, they challenged these mice with cells from a variety of different tumors, and showed that the immune systems of the inoculated mice rejected only those cancer cells that came from the original tumor. Thus, there are tumor-specific antigens that can be recognized by the immune system. See Aside 1.

[Aside 1: Importantly, the tumor resistance seen in these experiments did not arise spontaneously in the original tumor-bearing animals. Instead, it developed in the test mice, in response to sensitization with killed tumor cells. Thus, these experiments per se do not point towards an immune mechanism of tumor surveillance. Nonetheless, harnessing such a mechanism is currently a promising means of cancer therapy, and was a major theme in Klein’s thinking.]

The following year, Klein’s group showed that polyoma virus-induced tumors share a common antigen. Importantly, polyoma virus-induced tumors, and polyoma virus-transformed cells, were rejected irrespective of whether they released virus. Thus, antiviral immunity as such was neither necessary nor sufficient for tumor rejection. This was the first demonstration that tumors caused by a virus might share a common antigen. The Kleins, and others, later found similar “group-specific” transplantation antigens on other virus-induced tumors, including retrovirus-induced lymphomas.

Burkitt’s lymphoma: “Sometime in the mid-1960s, Eva suggested that we should use our experi­ence on virus-induced murine lymphomas to examine a human lymphoma with a presumptive viral etiology. Could we detect group specific antibody responses that might be helpful in tracing a virus? Burkitt’s lymphoma (BL) was the obvious choice (3).” [Burkitt’s lymphoma, originally described by Dennis Burkitt in 1958, is a malignant B-cell lymphoma that is most prevalent in tropical Africa and New Guinea. It is the most common childhood cancer in equatorial Africa. Burkitt first proposed that the lymphoma might have a viral etiology, since its geographic distribution is like that of yellow fever, which is caused by a flavivirus. In 1964, Tony Epstein and Yvonne Barr, by means of electron microscopy, discovered a virus in cells which they cultured from BL tissue, thereby giving credence to Burkitt’s premise.]

Klein’s group identified a membrane antigen (MA) that was expressed in some BL-derived cell cultures. Werner and Gertrude Henle had previously discovered that the MA antigen is a structural protein from a newly discovered herpesvirus—the virus that Epstein and Barr first saw in 1964. Klein decided to call that virus the Epstein Barr virus (EBV). The MA antigen is now known to be one of the EBV envelope glycoproteins. Klein and collaborators later identified complement receptor type 2 (CR2), also known as the complement C3d receptor, as the cell surface attachment protein for the viral MA glycoprotein. CR2 receptors on B cells play a role in enabling the complement system to activate B cells.

By 1970, Klein’s group, in collaboration with Harald zur Hausen, found that the subset of BL-derived cell lines that express MA are, in fact, those that produce EBV. However, more than 90% of the BL cell lines, and all nasopharyngeal carcinomas, were found to contain multiple EBV genomes per cell, irrespective of whether they produced virus. Thus, only a subset of BL and nasopharyngeal carcinoma cells that harbor EBV genomes, actually produce the virus. During this time, the Henles discovered that EBV is the cause of infectious mononucleosis, and that EBV could immortalize normal B cells in culture.

Oncogene activation by chromosomal translocation: A sero-epidemiological study, begun in Uganda in 1971 by Geser and de-The, showed that children with a high EBV load are more likely to develop BL than are children with a low EBV load. Thus, the presence of EBV genomes in a B cell increases the likelihood of it turning into a BL. “But this is still not a satisfactory explanation; some essential element is obviously missing (3).”

What then is the missing event that gives rise to BL? A 1972 study by Manolov and Manolova, Bulgarian scientists working with the Kleins, found that a particular chromosomal marker, 14q+, was present in about 80% of BL tumors. After the Manolovs returned to Bulgaria, the Kleins, in collaboration with Lore Zech, used the chromosomal banding technique recently developed by Caspersson and Zech to examine the BL-cell chromosomes more precisely. They showed that the 14q+ marker was derived from chromosome 8, which broke at the same site (8q24) and underwent a reciprocal translocation with the short arm of either chromosome 2 or chromosome 22. All BLs carried one of the translocations.

Meanwhile, another research group found that carcinogen-induced mouse plasmacytomas are associated with an almost homologous chromosomal translocation. Thus, a common mechanism seemed to underlie two distinct types of tumors, in two distinct species. In each instance, a putative oncogene was translocated to an immunoglobulin locus, which might then have caused the oncogene to be constitutive expressed. A somewhat similar mechanism was reported earlier for the induction of bursal lymphomas in chickens by the avian leukosis virus (ALV) . In that instance, the cellular c-myc gene came under the control of the ALV provirus promotor. What’s more, Michael Cole’s group identified the transposed gene in BL, and in the mouse plasmacytomas, as c-myc. It is not yet clear how EBV infection promotes the chromosomal translocation.

Tumor suppressor genes: In the early 1970s, Klein, and collaborator Henry Harris, played a pioneering role in developing the concept of tumor suppressor genes. They found that when highly malignant mouse cells are fused with normal mouse cells, the hybrid cells are non-malignant when inoculated into genetically compatible mice. That is, tumorgenicity is suppressed by fusion with normal cells. However, tumorgenicity reappears after some apparently important chromosomes, contributed by the normal cell, are lost from the hybrid cells.

References:

  1. George Klein. Pieta, MIT Press, 1992.
  1. The Converging Lives of Jacques Monod, Francois Jacob, Andre Lwoff, and Albert Camus in Wartime France, Posted on the blog March 27, 2017.
  1. George Klein and Eva Klein. 1989. How One Thing Led to Another, Annual Review of Immunology, 7:1-33.

Vaccine Research using Children

Children have been used in vaccine research since its very beginning, usually said to have been in 1796, when Edward Jenner inoculated 8-year-old James Phipps with cowpox, and then challenged young James with actual smallpox (1). However, earlier, in 1789, Jenner inoculated his own 10-month-old son, Edward Jr., with swinepox. Edward Jr. then came down with a pox disease, which he fortunately recovered from. His father then challenged him with smallpox.

Edward Jr. survived his exposure to smallpox. But, since Edward Sr. wanted to determine the duration of young Edward’s protection, he again challenged his son with smallpox in 1791, when the boy was two.  Edward Sr. inoculated his son yet again with smallpox when the boy was three. Fortunately, young Edward was resistant to each of the smallpox challenges his father subjected him to.

Jenner used several other young children in his experiments, including his second son, Robert, who was 11-months-old at the time. One of the children in Jenner’s experiments died from a fever; possibly caused by a microbial contaminant in an inoculum. [Microbes were not known in the late 18th century.]

We have no record of how Jenner (or his wife) felt about his use of his own children. However, there is reason to believe that Jenner felt some remorse over his use of James Phipps, who he referred to as “poor James.” Jenner looked after Phipps in later years, eventually building a cottage for him; even planting flowers in front of it himself.

By the 20th century, some of the most esteemed medical researchers were using children—in institutions for the mentally deficient—to test new drugs, vaccines, and even surgical procedures. These institutions were typically underfunded and understaffed. Several of them were cited for neglecting and abusing their residents. Moreover, their young patients were usually from poor families, or were orphans, or were abandoned. Thus, many of the children had no one to look out for their interests. In addition, research at these institutions was hidden from the public. [The goings-on at these institutions were, in general, hidden from the public, and most of the public likely preferred it that way.] Federal regulations that might have protected the children were not yet in existence, and federal approval was not even required to test vaccines and drugs.

In the early 1940s, Werner Henle, of the University of Pennsylvania, used children at Pennhurst—a Pennsylvania facility for the mentally deficient—in his research to develop an influenza vaccine. [Pennhurst was eventually  infamous for its inadequate staffing, and for neglecting and abusing its patients (2). It was closed in 1987, after two decades of federal legal actions.] Henle would inoculate his subjects with the vaccine, and then expose them to influenza, using an oxygen mask fitted to their faces.

Pennhurst, a state-funded Pennsylvania facility for the mentally deficient, was one of the most shameful examples of the neglect and mistreatment that was common at these institutions. It was the site of Werner Henle’s research in the 1940s to develop an influenza vaccine.
Pennhurst, a state-funded Pennsylvania facility for the mentally deficient, was one of the most shameful examples of the neglect and mistreatment that was common at these institutions. It was the site of Werner Henle’s research in the 1940s to develop an influenza vaccine.

Henle’s vaccine did not protect all of his subjects. Moreover, it frequently caused side effects. Additionally, Henle maintained (correctly?) that a proper test of a vaccine must include a control group (i.e., a group exposed to the virus, but not to the vaccine). Thus, he deliberately exposed unvaccinated children to influenza. Children who contracted influenza had fevers as high as 104o F, as well as typical flu-like aches and pains.

Despite Henle’s investigations at Pennhuerst, he was a highly renowned virologist, best known for his later research on Epstein Barr virus. See Aside 1.

      [Aside 1: While Henle was researching his influenza vaccine at Pennhurst, Jonas Salk concurrently worked on an influenza vaccine, using adult residents (ranging in age from 20 to 70 years) at the Ypsilanti State School in Michigan.]

Next, consider Hilary Koprowski, an early competitor of Jonas Salk and Albert Sabin in the race to develop a polio vaccine (3). By 1950, Koprowski was ready to test his live polio vaccine in people. [That was four years before Sabin would be ready to do the same with his live polio vaccine.] Koprowski had already found that his vaccine protected chimpanzees against polio virus. And, he also tested his vaccine on himself. Since neither he nor the chimpanzees suffered any ill effects, Koprowski proceeded to test his vaccine on 20 children at Letchworth Village for mentally disabled children, in Rockland County, NY.  [Like Pennhurst, Letchworth Village too was cited for inadequately caring for its residents.]  Seventeen of Koprowski’s inoculated children developed antibodies to the virus, and none developed complications.

Koprowski did not initiate his association with Letchworth. Actually, Letchworth administrators, fearing an outbreak of polio at the facility, approached Koprowski, requesting that he vaccinate the children. Koprowski gave each child “a tablespoon of infectious material” in half a glass of chocolate milk (4). Koprowski never deliberately infected the Letchworth children with virulent virus.

Koprowski reported the results of his Letchworth studies at a 1951 conference of major polio researchers, attended by both Salk and Sabin. When Koprowski announced that he actually had tested a live vaccine in children, many conferees were stunned, even horrified. Sabin shouted out: “Why did you do it? Why? Why (4)?” See Aside 2.

      [Aside 2: In the 1930s, Canadian scientist Maurice Brodie tested a killed polio vaccine in twelve children, who supposedly had been “volunteered by their parents (4).” For a short time Brodie was hailed as a hero. However, too little was known at the time for Brodie to ensure that his formaldehyde treatment had sufficiently inactivated the live polio virus. Consequently, Brodie’s vaccine actually caused polio in several of the children. After this incident, most polio researchers could not conceive of ever again testing a polio vaccine, much less a live one, in children.]

Neither Koprowski nor Letchworth Village administrators notified New York State officials about the tests. Approval from the state would seem to have been required, since Koprowski later admitted that he was certain he would have been turned down. And, it is not clear whether Koprowski or the school ever got consent from the parents to use their children. However, recall there were not yet any federal regulations that required them to do so.

Koprowski was untroubled by the uproar over his use of the Letchworth children, arguing that his experiments were necessary. Yet he later acknowledged: “if we did such a thing now we’d be put on jail…” But, he added, “If Jenner or Pasteur or Theiler (see Aside 2) or myself had to repeat and test our discoveries [today], there would be no smallpox vaccine, no rabies vaccine, no yellow fever vaccine, and no live oral polio vaccine.”  Moreover, he maintained that, secret or not, his use of the Letchworth children fit well within the boundaries of accepted scientific practice.

   [Aside 2: Nobel laureate Max Theiler developed a vaccine against yellow fever in 1937; the first successful live vaccine of any kind (5). Theiler formulated a test for the efficacy of his vaccine, which did not involve exposing humans to virulent virus. Sera from vaccinated human subjects were injected into mice, which were then challenged with the Yellow Fever virus.]

Koprowski referred to the Letchworth children as “volunteers (6).” This prompted the British journal The Lancet to write: “One of the reasons for the richness of the English language is that the meaning of some words is continually changing. Such a word is “volunteer.” We may yet read in a scientific journal that an experiment was carried out with twenty volunteer mice, and that twenty other mice volunteered as controls.” See Aside 3.

     [Aside 3: Koprowski was a relatively unknown scientist when he carried out his polio research at Letchworth. He later became a renowned virologist, having overseen the development of a rabies vaccine that is still used today, and having pioneered the use of therapeutic monoclonal antibodies. Yet, he is best remembered for developing the world’s first effective polio vaccine; several years before Salk and Sabin brought out their vaccines.

   Most readers of the blog are aware that the Salk and Sabin vaccines are credited with having made the world virtually polio-free. What then became of Koprowski’s vaccine? Although it was used on four continents, it was never licensed in the United States. A small field trial of Koprowski’s vaccine in 1956, in Belfast, showed that its attenuated virus could revert to a virulent form after inoculation into humans. Yet a 1958 test, in nearly a quarter million people in the Belgian Congo, showed that the vaccine was safe and effective. Regardless, the vaccine’s fate was sealed in 1960, when the U.S. Surgeon General rejected it on safety grounds, while approving the safer Sabin vaccine. Personalities and politics may well have played a role in that decision (3, 4).

  Interestingly, Sabin developed his vaccine from a partially attenuated polio virus stock that he received from Koprowski. It happened as follows. In the early 1950s, when Koprowski’s polio research was further along than Sabin’s, Sabin approached Koprowski with the suggestion that they might exchange virus samples. Koprowski generously sent Sabin his samples, but Sabin never reciprocated.

   Koprowski liked to say: “I introduce myself as the developer of the Sabin poliomyelitis vaccine (7).” He and Sabin had a sometimes heated adversarial relationship during the time when their vaccines were in competition. But they later became friends.]

Sabin was at last ready to test his polio vaccine in people during the winter of 1954-1955. Thirty adult prisoners, at a federal prison in Chillicothe, Ohio, were the subjects for that first test in humans. [The use of prisoners also raises ethical concerns.]

Recall Sabin’s public outcry in 1951 when Koprowski announced that he used institutionalized children to test his polio vaccine. In 1954, Sabin sought permission to do the very same himself; asserting to New York state officials: “Mentally defective children, who are under constant observation in an institution over long periods of time, offer the best opportunity for the careful and prolonged follow-up studies…”

Although Sabin had already tested his attenuated viruses in adult humans (prisoners), as well as in monkeys and chimpanzees, the National Foundation for Infantile Paralysis, which funded polio research in the pre-NIH days of the 1950s, blocked his proposal to use institutionalized children. Thus, Sabin again used adult prisoners at the federal prison in Ohio. With the concurrence of prison officials, virtually every inmate over 21 years-old “volunteered,” in exchange for $25 each, and a possible reduction in sentence. None of the prisoners in the study became ill, while all developed antibodies against polio virus.

Testing in children was still a necessary step before a polio vaccine could be administered to children on a widespread basis. But, Sabin’s vaccine could not be tested in children in the United States. Millions of American children had already received the killed Salk vaccine, and the National Foundation for Infantile Paralysis was not about to support another massive field trial of a vaccine, in children, in the United States (3).

Then, in 1959, after a succession of improbable events, 10 million children in the Soviet Union were vaccinated with Sabin’s vaccine (3). The Soviets were so pleased with the results of that massive trial that they next vaccinated all seventy-seven million Soviet citizens under 20 years-of-age with the Sabin vaccine. That figure vastly exceeded the number of individuals in the United States, who were vaccinated with the rival Salk vaccine during its field trials.

Next up, we have Nobel laureate John Enders who, in the 1950’s, oversaw the development of the first measles vaccine. Enders and co-workers carried out several trials of their attenuated measles vaccine; first in monkeys and then in themselves. Since the vaccine induced an increase in measles antibody titers, while causing no ill effects, they next tested it in severely handicapped children at the Walter E. Fernald State School near Waltham, Massachusetts.

Enders seemed somewhat more sensitive than either Henle or Koprowski to the ethics of using institutionalized children. Samuel L. Katz, the physician on Enders’ team, personally explained the trial to every Fernald parent, and no child was given the vaccine without written parental consent. [Federal guidelines requiring that step still did not exist.] Also, no child was deliberately infected with virulent measles virus.

Katz personally examined each of the inoculated Fernald children every day. None of these children produced measles virus, while all of them developed elevated levels of anti-measles antibodies. Also, the Fernald School had been experiencing severe measles outbreaks before the Enders team vaccinated any of its children. But, when the next measles outbreak struck the school, all of the vaccinated children were totally protected.

In 1963, the Enders vaccine became the first measles vaccine to be licensed in the United States. Several years later it was further attenuated by Maurice Hilleman (8) and colleagues at Merck. In 1971, it was incorporated into the Merck MMR (measles, mumps, and rubella) vaccine. See Aside 4.

    [Aside 4: Before Enders carried out his measles investigations he pioneered the growth of viruses in tissue culture. In 1949, Enders, and collaborators Thomas Weller and Frederick Robbins, showed that poliovirus could be cultivated in the laboratory. This development was crucial, allowing Salk and Sabin to grow a virtually unlimited amount of polio virus and, consequently, to develop their polio vaccines. In 1954, Enders, Weller, and Robbins were awarded the Nobel Prize for Physiology or Medicine for their polio virus work.]

It may surprise some readers that before the mid 1960s the so-called Nuremburg Code of 1947 comprised the only internationally recognized ethical guidelines for experimentation on human subjects. The Nuremburg Code was drawn up by an American military tribunal during the trial of 23 Nazi physicians and scientists for atrocities they committed while carrying out so-called “medical” experiments during World War II. [Sixteen of the 23 Nazis on trial at Nuremburg were convicted, and 7 of these were executed (see Note 1)].

The Nuremberg Code’s Directives for Human Experimentation contained strongly stated guidelines. Its tenets included the need to obtain informed consent (interpreted by some to prohibit research using children), the need to minimize the risks to human subjects, and the need to insure that any risks are offset by potential benefits to society.

But, despite the well-articulated principles of the Nuremberg Code, it had little effect on research conduct in the United States. Federal rules, with the authority to regulate research conduct, would be needed for that. So, how did our current federal oversight of research come to be?

A 1996 paper in the The New England Journal of Medicine, “Ethics and Clinical Research,” by physician Henry Beecher, brought to the fore the need for rules to protect human subjects in biomedical research (9). Beecher was roused to write the paper in part by the early 1960s experiments of Saul Krugman, an infectious disease expert at NYU. Krugman used mentally deficient children at the Willowbrook State School in Staten Island, New York, to show that hepatitis A and hepatitis B are distinct diseases (9). Also, before a hepatitis vaccine was available, Krugman inoculated the children with serum from convalescing individuals, to ask whether that serum might protect the children against hepatitis. Krugman exposed the children to live virus either by injection, or via milkshakes seeded with feces from children with hepatitis.

Krugman found that convalescent sera indeed conferred passive immunity to hepatitis. Next, he discovered that by infecting passively protected patients with live hepatitis virus he could produce active immunity. Krugman had, in fact, developed the world’s first vaccine against hepatitis B virus (HBV) (see Aside 4). [Although Krugman used mentally deficient institutionalized children in his experiments, his investigations were nonetheless funded in part by a federal agency; the Armed Forces Epidemiology Section of the U.S. Surgeon General’s Office.]

         [Aside 4: The first hepatitis B vaccine licensed for widespread use was developed at Merck, based on principles put forward by Nobel Laureate Baruch Blumberg, (10).]

Beecher was particularly troubled by two aspects of Krugman’s experiments. First, Krugman infected healthy children with live virulent virus. Beecher maintained that it is morally unacceptable to deliberately infect any individual with an infectious agent, irrespective of the potential benefits to society. [See reference 11 for an alternative view. “The ethical issue is the harm done by the infection, not the mere fact of infection itself.”]

Second, Beecher charged that the Willowbrook School’s administrators coerced parents into allowing their children to be used in Krugman’s research. The circumstances were as follows. Because of overcrowding at the school, Willowbrook administrators closed admission via the usual route. However, space was still available in a separate hepatitis research building, thereby enabling admission of additional children who might be used in the research.

Were the Willowbrook parents coerced into allowing their children to be used in the research there? Consider that the parents were poor and in desperate need of a means of providing care for their mentally impaired children. Making admission of the children contingent on allowing them to be used in the research might well be viewed as coercion. Yet even today, with federal guidelines now in place to protect human subjects, institutions such as the NIH Clinical Center admit patients who agree to participate in research programs. Is that coercion?

Beecher’s 1966 paper cited a total of 22 instances of medical research that Beecher claimed were unethical (9). Four examples involved research using children. Krugman’s work at Willowbrook was the only one of these four examples that involved vaccine research. Beecher’s other examples involved research using pregnant women, fetuses, and prisoners. But it was Beecher’s condemnation of Krugman’s hepatitis research at Willowbrook that is mainly credited with stirring debate over the ethics of using children in research.

Did Krugman deserve Beecher’s condemnation? Before Krugman began his investigations at Willowbrook, he plainly laid out his intentions in a 1958 paper in the New England Journal of Medicine (12). Importantly, Krugman listed a number of ethical considerations, which show that he did not undertake his Willowbrook investigations lightly. In fact, Krugman’s ethical considerations, together with his plans to minimize risks to the children, were not unlike the assurances one might now submit to an institutional review board (11).

Many (but not all) knowledgeable biomedical researchers claimed that Beecher misunderstood Krugman’s research and, thus, unjustly vilified him. Krugman was never officially censored for his Willowbrook investigations. Moreover, condemnation of Krugman did not prevent his election in 1972 to the presidency of the American Pediatric Society, or to his 1983 Lasker Public Service Award.

To Beecher’s credit, his 1966 paper was instrumental in raising awareness of the need to regulate research using human subjects. Beecher was especially concerned with the protection of children and, apropos that, the nature of informed consent.

In 1974, the National Research Act was signed into law, creating the National Commission for the Protection of Human Subjects of Biomedical and Behavioral Research. The basic ethical principles identified by the Commission are summarized in its so-called Belmont Report, issued in 1978. Its tenets include minimizing harm to all patients, and the need to especially protect those with “diminished autonomy” or who are incapable of “self-determination.”  In addition, federal guidelines now require universities and other research institutions to have Institutional Review Boards to protect human subjects of biomedical research. [Reference 13 (available on line) contains a detailed history of the establishment of these policies.]  See Aside 6.

      [Aside 6: The infamous U.S. Public Health Service Tuskegee syphilis research program, conducted between 1932 and 1972, in which several hundred impoverished black men were improperly advised and never given appropriate treatment for their syphilis, also raised public awareness of the need to protect human subjects. More recently, research involving embryonic stem cells and fetuses has stoked an ongoing and heated public debate. Policies regarding this research are still not settled, with stem-cell research being legal in some states, and a crime in others. Other recent technological advances, such as DNA identification and shared databases, have been raising new concerns, such as the need to protect patient privacy. In response to these new developments, in June 2016, the US National Academies of Sciences, Engineering and Medicine released a report proposing new rules (indeed a complete overhaul of the 1978 Belmont Report) to deal with these circumstances. The Academy’s report has stirred debate in the biomedical community]

Note 1: The use of children in medical research makes many of us profoundly uneasy. We may be particularly troubled by accounts of the exploitation of institutionalized children, who comprised a uniquely defenseless part of society. Indeed, it was the very vulnerability of those children that made it possible for them to be exploited by researchers. Consequently, some readers may well be asking whether the activities of vaccine researchers Krugman, Koprowski, Sabin, Henle and others might have been comparable to that of the Nazis on trial at Nuremberg. So, I offer this cautionary interjection. While in no way condoning the vaccine researchers using institutionalized children, their work was carried out for the sole purpose of saving human lives. As Koprowski suggested above, if not for that work, we might not have vaccines against smallpox, rabies, yellow fever, and polio. Now, consider Josef Mengele, a Nazi medical officer at Auschwitz, and the most infamous of the Nazi physicians. [Mengele was discussed several times at Nuremberg, but was never actually tried. Allied forces were convinced at the time that he was dead, but he had escaped to South America.] At Auschwitz, Mengele conducted germ warfare “research” in which he would infect one twin with a disease such as typhus, and then transfuse that twin’s blood into the other twin. The first twin would be allowed to die, while the second twin would be killed so that the organs of the two children might then be compared. Mengele reputedly killed fourteen twin children in a single night via a chloroform injection to the heart. Moreover, he unnecessarily amputated limbs and he experimented on pregnant women before sending them to the Auschwitz gas chambers.

References:

  1. Edward Jenner and the Smallpox Vaccine, Posted on the blog September 16, 2014.
  2.  Pennhurst Asylum: The Shame of Pennsylvania, weirnj.com/stories/pennhurst-asylum/
  3.  Jonas Salk and Albert Sabin: One of the Great Rivalries of Medical Science, Posed on the blog March 27, 2014.
  4.  Oshinsky D, Polio: An American Story, Oxford University Press, 2005.
  5. The Struggle Against Yellow Fever: Featuring Walter Reed and Max Theiler, Posted on the blog May 13, 2014.
  6.  Koprowski H, Jervis GA, and Norton TW. Immune response in human volunteers upon oral administration of a rodent-adapted strain of poliomyelitis virus. American Journal of Hygiene, 1952, 55:108-126.
  7.  Fox M, Hilary Koprowski, Who Developed First Live-Virus Polio Vaccine Dies at 96, N.Y. Times, April 20, 2013.
  8. Maurice Hilleman: Unsung Giant of Vaccinology, Posted on the blog April 14, 2014.
  9. Beecher HK. Ethics and clinical research. The New England Journal of Medicine, 1966, 274:1354–1360.
  10.  Baruch Blumberg: The Hepatitis B Virus and Vaccine, Posted on the blog June 2, 2016.
  11.  Robinson WM, The Hepatitis Experiments at the Willowbrook State School. science.jburrougs.org/mbahe/BioEthics/Articles/WillowbrookRobinson2008.pdf
  12. Ward R, Krugman S, Giles JP, Jacobs AM, Bodansky O. Infectious hepatitis: Studies of its natural history and prevention. The New England Journal of Medicine, 1958, 258:407-416.
  13.  Ethical Conduct of Clinical Research Involving Children. http://www.ncbi.nlm.nih.gov/books/NBK25549/