Chapter 02: A New World of Research at University of Pennsylvania: Focusing a Research Career

Chapter 02: A New World of Research at University of Pennsylvania: Focusing a Research Career

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Description

Dr. Fidler details how he began to look more deeply into the question of cancer cell differentiation, a groundbreaking discovery that was ultimately published in Nature. (Dr. Fidler shows the interviewer a Plexiglas containing a unit he invented to facilitate injection of cells into the tail veins of up to 100 mice per hour, underscoring the resourcefulness that a researcher had to have to move ahead quickly with a study.) Through this success, he was recruited to join the National Cancer Institute at Frederick, Maryland. His wife-to-be, Margaret Kripke, known for her pioneering work in photoimmunology, was also hired in a concurrent recruitment, and he describes how Dr. Kripke challenged him with the question that inspired a new line of research: “How do you know whether the cells you are culturing from a line are a selection or an adaptation?” His discovery that the differentiation of metastasis cells is a priori “revolutionized the world,” he states, noting that “you cannot treat a heterogeneous disease with homogeneous therapy” –the origin of individualized therapy. Dr. Fidler then explains connections between Paget’s “seed and soil” theory and his next experiments with transplanting metastatic cells between the organs of mice.

Identifier

FidlerIJ_01_20110926_C02

Publication Date

9-26-2011

City

Houston, Texas

Topics Covered

The Interview Subject's Story - The ResearcherThe Researcher Overview Understanding Cancer, the History of Science, Cancer Research Definitions, Explanations, Translations Career and Accomplishments Discovery, Creativity and Innovation Professional Practice Evolution of Career

Transcript

Tacey Ann Rosolowski, PhD:

What was the world that opened up?

Isaiah J Fidler, DVM, PhD:

World that opened up was that there was virtually no research at the veterinary school. No basic research. I did clinical research. I wrote several papers about the incidence of breast cancer in dogs and how old are the dogs, what strain they are, reviewed records and wrote papers. OK? But if I wanted to do tissue culture, whatever, there wasn’t such a thing. And it was recommended to me to interview specifically in the department of pathology, which in fact had individual who were very interested in metastases. And my mentor was the late Irving Zeidman. All of these individuals are no longer with us unfortunately. Z-E-I-D-M-A-N. Irving Zeidman and Charles Breedis and others have written many many papers on cancer metastasis. They were pathologists. The other area of heavy heavy study in metastasis was in Pittsburgh by the surgeon Fisher, Bernard Fisher, who is still with us, thank God. He’s in his mid 90s. We still correspond. And there was also Leonard Weiss, W-E-I-S-S, Leonard, at Buffalo, New York, oh, God, the cancer center in Buffalo.

Tacey Ann Rosolowski, PhD:

Roswell.

Isaiah J Fidler, DVM, PhD:

Roswell Park. So these were the three areas of cancer metastasis research. Irving Zeidman took me, and it was amazing. I used to come at -- I’m used to 7:00, 7:30 to work. And these prima donnas, the pathology, show to work at 9:00. And they sat around drinking coffee. Arguing with each other till 10:00, etc. And I had my lunch with them when they had breakfast. But in any event, the question that came up. I wanted to know. We knew that tumor cells to get say from the breast cancer to the lung or to the brain, there were many publication on that, have to enter the bloodstream. Otherwise how do they get there? Or the lymphatics. So either the lymph vessels or the blood vessels. They had to circulate through the body. And what I wanted to know is how many cells can enter the circulation and how many cells survive in the circulation to give rise to the distant metastasis. At the time what I decided to do, and after consultation with many, was to take a mouse tumor, a mouse melanoma, which has never been in culture before by the way, I was the first one to put it in culture. Egotistical, I labeled it B16F for Fidler. Nobody knows. I keep telling people who lecture me about B16. I laugh. I say, “Do you know what the F stand for?” And they don’t but that’s OK.

Tacey Ann Rosolowski, PhD:

I’m curious why no one had cultured it before.

Isaiah J Fidler, DVM, PhD:

Because nobody worked with the cell. The way you transferred tumor was from animal to animal to animal to animal. There was no -- at the time there were no laminar flow cabinets where everything is sterile. We had to work in a cabinet with UV light to kill the bugs at night. And in the summer you couldn’t culture anything because you’ll have bacteria and mold contamination. And we had to wash our hands with iodine. I mean for months my hands were brown all the time. And then another fellow in pathology who is no longer with us, John Kreider, a human pathologist who then went to do research, OK, so he had his clinical background, and he was researching alongside me. Was hired by NASA to help them with some mice that were -- to analyze tissue from mice that they were going to send to space. So he goes to NASA around Philadelphia, and he comes back and he says, “I can’t believe what I just saw. I saw this cabinet where there is airflow that keeps all the bugs out. And they did it so they can assemble parts for the space that will be dust-free, bug-free, mold-free.” So he contacted the company and said, “Will you give me a unit? I want to see if I can do tissue culture in it.” That was the first tissue culture cabinet. Now there must be hundreds of millions of those. And John never got a patent on his idea, which is a mistake. But in those days we didn’t think patent. Today everything is patent, patent. “Do you have a patent?” “No.” “Then you are a loser.” But at the time we only thought about knowledge. So tissue culture became, at least at the University of Pennsylvania in our department, we had two units that were given to us. Gift. They were $50,000 then. Today they’re a few thousand, nothing. They build so much. And started culturing cells. And talking, talking, talking to other people. And there was another fellow who was a radiotherapist who also had a Luther Terry. So we like had a Luther Terry brunch club if you know what I mean, talking to each other. And he said, “If you can label the DNA of the cells, label it with an isotope called -- that has iodine-125 in it. And as long as the cells are alive, they emit a signal for the I-125, which is a gamma emitter. And that gamma emitter is very strong. So if you take a lung or liver and you put it in a tube the emission will go through and you can measure it. If it was an alpha emitter or beta emitter, you have to make jelly out of it, which is impossible. And as long as the cell is alive they’ll emit that from the DNA. Once the cell is dead, the DNA will break down. The I-125 will disappear in the urine, and that’s it. So if you have a high count you have live cells, and a low count low cells. And you can run alongside, and you know exactly how many cells are in the animal.” I said, “Oh, I love you for that.” And I did the first experiment with -- iododeoxyuridine I-125. That’s too complicated. I-125-emitting DNA. And showed that after one month, OK, of all the cells that are introduced to the animal, maybe 0.01% survive. Now I will tell you it’s even less than that because we can do more sensitive things. But at the time I couldn’t. And shown also another interesting thing. That initially we injected the cells into the tail vein of the mouse. Initially there were counts everywhere. But ultimately when the animal died the melanoma was only in the lung. Not in the liver, not in the kidney, not in the bone. We couldn’t find tumor residues. But initially they were everywhere.

Tacey Ann Rosolowski, PhD:

Did you have any suspicion of such results when you --

Isaiah J Fidler, DVM, PhD:

No.

Tacey Ann Rosolowski, PhD:

-- began these tests? No.

Isaiah J Fidler, DVM, PhD:

No, of course not. I had no -- I just -- I couldn’t believe that every cell will survive, because then patient would not survive five or six years sometimes. Today in 2011, there’s a big deal about circulating tumor cells, etc. And I say, “Why don’t you read the literature? You will see that just because they circulate doesn’t mean that they’re going to survive and kill the patient.” But in any event the finding was so startling that I was told, “Why don’t you write your thesis based on one experiment?” So I wrote my thesis. It’s still there. And it’s the record. It’s the shortest thesis ever written at the University of Pennsylvania. Let me see if I can find it. Well, it’s one of those. By the way we didn’t have computer. Everything had to be typed. No, that’s not it. That’s my graduates list. That’s old, that’s another one. It looks just like that. Mine is thin. All of these are my graduate student who wrote theses. I’m sorry.

Tacey Ann Rosolowski, PhD:

No. That’s fine.

Isaiah J Fidler, DVM, PhD:

Here it is.

Tacey Ann Rosolowski, PhD:

That’s great. With photos and everything.

Isaiah J Fidler, DVM, PhD:

Yeah. Anyhow. And I say in 1970 I’m deeply in debt to the member of the department, Dr. Irving Zeidman, John Kreider for his initial suggestion which have encouraged me to undertake the work, that’s laminar airflow, financial support to the predoctoral fellowship from USPHS and my patient people, my wife and two children. Because to do that in a year and a half, I worked a long time. But anyhow, I was told, “Present it.” And I finished my PhD in a very short time, like in two years or two and a half years. And I insisted that my degree would be in human pathology, not veterinary but human pathology, since I was in the human, in Penn Medical School. I wanted to stay in the department of pathology at the medical school. But the young chairman of the department, Peter Nowell, who should have received the Nobel Prize for his work --

Tacey Ann Rosolowski, PhD:

What did he work on?

Isaiah J Fidler, DVM, PhD:

He worked on leukemia. He showed the leukemia starts from a single cell. In any event told me, “No, you cannot stay here, because there’ll always be a suspicion that you’re not independent. You must leave.” But I didn’t want to leave Philadelphia. I already had a little home, etc. So I heard that the dental school is interested in recruiting me. And I went, had lunch with the dean, and over lunch I told him what it would take to recruit me. And when he said yes to everything I thought I should have asked for more. But anyhow I went to the department of pathology at the dental school, University of Pennsylvania. So here I am, a veterinarian who graduated from veterinary school, medical school, and now I’m in the dental school.

Tacey Ann Rosolowski, PhD:

So that was shortly after 1970 or that was 1970?

Isaiah J Fidler, DVM, PhD:

That was the end of ’70, beginning of ’71. And then I decided to look deeper into the finding. Just a few cells survive to give rise to metastasis. And the serious question of the day was are they different from all the cells that didn’t survive. That’s a selection process? Or is it adaptation? So I did -- now it looks like a simple experiment. At the time it was very unusual. I injected -- don’t remember, 10,000, 20,000 cells into a mouse. And the mouse developed let’s say ten isolated colonies in the lung. Since they were black, it was very easy to find them. Very carefully dissected those metastases. And put them in tissue culture and grew a culture out of them. And injected that culture into the tail vein of another syngeneic, in other words genetically identical, mouse. So there would not be immune rejection.

Tacey Ann Rosolowski, PhD:

Can I ask what the significance of the tail vein is?

Isaiah J Fidler, DVM, PhD:

It’s the easiest vein to reach in a mouse. They have a huge vein. For me. Now we can inject carotid artery and things, but you need a microscope for that. The tail vein I could, when I was good I could do probably 100 an hour.

Tacey Ann Rosolowski, PhD:

So it took a lot of hand skills too.

Isaiah J Fidler, DVM, PhD:

Yeah but I invented this thing. I didn’t take a patent on it unfortunately but this was the original. You put a mouse here. And here’s the tail, OK?

Tacey Ann Rosolowski, PhD:

For the recorder, we’re looking at a Plexiglas container for a mouse.

Isaiah J Fidler, DVM, PhD:

Yeah but this was even easier. You just put the mouse, hold the tail, and you inject.

Tacey Ann Rosolowski, PhD:

Oh I see. So and then it just stabilizes the mouse, keeps it from squirming.

Isaiah J Fidler, DVM, PhD:

When you collect. This is a sieve like tea. You collect your lesions from the lung. You take a syringe and you grind them. They come out as groups of cells. And you put that in culture. So at the time you had to be very resourceful to do your own things. Otherwise waiting for others to invent things for you would take too long. In any event I’ve done it several times. And every passage it was clearly more and more and more metastases in the lung. So after a few passages the whole lung was black. And it was very clear that I’m selecting for more and more and more malignant cells. This was published in Nature, made a big noise. And then there were many other. Obviously I’m giving you one major experiment. But at the time I probably published 20 papers. Not just on that but many other things.

Tacey Ann Rosolowski, PhD:

What were some of the related things you’re doing? Because I was really interested in the array of studies that you did.

Isaiah J Fidler, DVM, PhD:

In the array for example it became very clear that cells that circulate and survive do so because they can clump together. You have a clump, the internal cell will survive the bumps of the circulation. But the clump didn’t have to come just from tumor, tumor, tumor. We have shown that the clump could be lymphocytes that bind to the tumor cell and don’t kill it or platelets. A platelet is a cell that’s supposed to block hemorrhage when it happens. Unfortunately clumps of platelets can lead to a lot of problems in the human. Including -- trying to think how to say it in simple language. When platelet clump in the brain, they lead to total disaster. In the heart they can lead to cardiac failure. So on and so forth. We found that tumor cell clump with platelets. A colleague at the medical school asked me to help him to inject mice because he said that the best way to treat to prevent platelets from clotting is a simple aspirin. And Gabriel Gasic in fact published that paper, thanked me for injecting mice for him. But in any event showing that aspirin can reduce metastasis. You know that today aspirin is being recommended to individuals who have had heart failure, brain problem, etc. Simple aspirin.

Tacey Ann Rosolowski, PhD:

The idea that it’s a blood thinner.

Isaiah J Fidler, DVM, PhD:

Well, it prevent platelets from clumping. So that was the lymphocyte story. I worked on that. I have a very small group at the time. I wasn’t that advanced. But we were recruited. Margaret Kripke, who was -- and I were married, and we were recruited -- well, we were not married yet. But we were looking at a place to go together. She was at University of Utah. And I was at University of Pennsylvania. And we had two independent recruitments simultaneously to a new program that opened at Frederick, Maryland. By the National Cancer Institute. And the director of the program, who’s still one of our dearest friends, Michael Hanna H-A-N-N-A, from Oak Ridge, Tennessee became the director of the program. He recruited Margaret because he knew her work. And she’s the one that showed that UV light is immunosuppressive. Margaret was the chairman of immunology here. She became the executive vice president and the provost of the institute. When she retired DuBois took over. It was her. She did it all. But in any case we were recruited to go to Maryland. And about that time she challenged me and she said, “OK, so how do you know whether the cells that you are culturing from the lung, how do you know whether it’s selection or adaptation?” And I said, “Not only don’t I know what you mean, I don’t even know what’s the difference between the two. So give me -- I’m just a dumb veterinarian. I’ll go read about it.” So I read, read, read. And I told her, “I still don’t know how to do it.” And she said, “You have to do the Luria-Delbruck fluctuation analysis.” OK, Luria-Delbruck, which nobody knows who they are. Just shows you. They received the Nobel for medicine. In 1938, ’39 they were interested in resistance of bacteria to antibacterial agent. By now you know don’t use penicillin, it’s a waste of time, but at the time it was the cure. But there were always -- at the beginning it looked great. And all of a sudden you have a resistant strain in tissue culture. Was that strain resistant a priori? You just killed all the sensitive? And that one bacteria that was resistant now grows up? Or was the drug mutagenic? The drug created DNA infidelity or DNA failure or DNA genetic alteration and the drug created the resistance? What Luria-Delbruck did then. They took bacteria, microorganisms. And cloned them. They took a single bacteria per dish. And they grew cultures. Each one originating from a single bacteria. They had thousands of these dishes. And they flooded them with antibacterial agent. And they showed conclusively that in some dishes every bacteria was killed, every mold was killed. And in few, none were killed. And they were not exposed to the same drug, different application, but only once. In other words, the resistance preexisted. That’s called a fluctuation analysis. Since in my selection experiment that I published in Nature I did it six times, maybe it just was that the procedure that I’m using, the tissue culture, the sieve, somehow I’m mutagenizing the cells. So what Margaret and I did, for which we received every conceivable prize you can think of. We did the first to my knowledge fluctuation analysis with mammalian cells. We took this melanoma. We isolated single cell from the culture. And we grew cultures from single cells and injected ten mice from each culture. And each one came from a single cell. And with other cells, with others, we injected a mix, the parental. Well, the parental injected to 80 mice gave average number of metastases. Eight, nine, ten, between eight and 12, eight and 12, some eight, some 12. But the median was not significantly different. But when we injected cells that all were derived from this single cell or that single cell or that, some gave no metastases whatsoever. Some gave 200. Some gave 50. The variation was enormous. In other words, this proved that some cells in the tumor are metastatic and the majority are not. And we never played the game of again, again. They all were given the same chance. Once. It was published in Science and was immediately accepted to my great surprise and really in my opinion revolutionized the world. Because for the first time it said the tumors are heterogeneous. In other words they don’t consist of cells that are all alike. Tacey, you know what it means in 2011 that when you talk about individualized therapy, genetic therapy, bullshit therapy, some cells will respond and some cells will not. You cannot treat a heterogeneous disease with homogeneous therapy. The two words are not parallel. It has to be combination against this, against that. By now there are thousands and thousands of papers. I’m sorry. My eye is killing. Thousands of papers about heterogeneity for cell size, cell shape. Making enzyme this and enzyme that. And this protein and that protein. When you look carefully, you see that the world is very heterogeneous. So that really made a significant significant sort of revolution in the field.

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Chapter 02: A New World of Research at University of Pennsylvania: Focusing a Research Career

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