Chapter 11: Research Projects: Working on an HIV Vaccine; Lipocalin 24p3; How CML Causes Uncontrolled Growth of Blood Cells

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Chapter 11: Research Projects: Working on an HIV Vaccine; Lipocalin 24p3; How CML Causes Uncontrolled Growth of Blood Cells

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In this chapter, Dr. Arlinghaus describes the research project he has conducted since returning to MD Anderson in 1986.He provides context by describing work on an HIV vaccine work he conducted at Johnson and Johnson then talks about his work on lipocalin 2.He explains how his research into the role of lipocalin 2 has evolved and how he is addressing the research hypothesis that when a tumor forms, an environment is created that is also invaded by normal cells.Some of these battle the tumor, and other come in and secrete lipocalin 24p3, killing other normal cells.Dr. Arlinghaus then turns to his work on how CML causes uncontrolled growth of blood cells and what this demonstrates about the host's involvement in the disease (a paper was published in 1995).He explains connections between this work and his other studies of Janus kinase 2, and the BCR-ABL.He goes on to describe unpublished work on how the BCR-ABL protein phosphorylates and thereby activates Janus kinase 2 in an uncontrolled mechanism.

Identifier

ArlinghausR_02_20140402_C11

Publication Date

4-2-2014

Publisher

The Making Cancer History® Voices Oral History Collection, The University of Texas MD Anderson Cancer Center

City

Houston, Texas

Topics Covered

The Interview Subject's Story - The Researcher; The Researcher; Evolution of Career; Character, Values, Beliefs, Talents; Discovery, Creativity and Innovation; Professional Practice; MD Anderson Impact; Definitions, Explanations, Translations; Understanding Cancer, the History of Science, Cancer Research; The Professional at Work; Discovery and Success

Disciplines

History of Science, Technology, and Medicine | Oncology | Oral History

Transcript

Tacey Ann Rosolowski, PhD:

Now last time you talked about going to Johnson & Johnson and then your return as Department Chair to MD Anderson in 19 --- in 1986 and you talked some about the work that you did but we --- we didn’t really get caught up with your --- your more current research. So maybe you’d like to tell me a bit about that.

Ralph B. Arlinghaus, PhD:

Yeah.

Tacey Ann Rosolowski, PhD:

We talked about the ABL kinases. We talked about your work on vaccines at the --- at Johnson & Johnson.

Ralph B. Arlinghaus, PhD:

See at Johnson & Johnson I was asked to work on HIV.

Tacey Ann Rosolowski, PhD:

Oh, okay. You hadn’t mentioned that part. Yeah.

Ralph B. Arlinghaus, PhD:

So I start working on vaccine strategies for HIV and it became clear at the time that antibody mediated vaccines for HIV were not protective. Still the case. There is no effective vaccine for HIV. I proposed that to make an effective vaccine that you had to make fragments of HIV proteins and induce killer T-cells. I became a self-made immunologist, if you will, while I was out there in California. I kep --- always kept learning and so my method, which I still believe in that method --and nobody’s taken me up on it, although I do have a patent application issued on that strategy. Basically the strategy is --- is induce killer T-cells to kill the infected cells, because the antibody mediated inactivation of HIV wasn’t working. So I wanted --- My patent talks about an antibody-negative approach to an HIV vaccine and that’s never really --- never been disproved but other people have taken up that mantra, but there isn’t an HIV vaccine that’s killer T-cell only. And what’s the reason for that? At the time --- this is going to get pretty deep for you. At the time, we were using short fragments of proteins to induce killer T-cells in mice and rabbits. That approach was faulty, because if you want to be --- have a protective approach, either antibody binding to the proteins or killer T-cells, lysine HIV infected cells or flu infected cells. You have to generate a response that sees many parts of the proteins in question and not just short peptides, which only activate the immune system with one or two sites on a peptide. So that was the faulty scenario. That was --- That was a false lead for me. So I --- I got a grant to study this from NIH and I didn’t get very far for reasons I just told you. So --- But remember when I came back to MD Anderson, I came back working on HIV and chronic myeloid leukemia and I decided I couldn’t be in both camps. So I turned over my HIV work to my postdoc. His name is Jacob Sastry [phonetic, ?Jagannadha K. Sastry, PhD,? ]. He’s a full professor in another department. He took and ran with that and is studying various aspects of HIV immunology and I got out of the HIV business to focus on chronic myeloid leukemia.

Tacey Ann Rosolowski, PhD:

I was just noticing that in 2005 you made a discovery about lipocalin 24P3 which …

Ralph B. Arlinghaus, PhD:

24P3, yeah.

Tacey Ann Rosolowski, PhD:

Yeah which is using killer cells.

Ralph B. Arlinghaus, PhD:

We’re still working on that. Still working on that. So we published an important paper on that so we could show in mouse models that if you could --- What we found is leukemia cells, CML cells induced lipocalin 2 formation and caused its secretion from the cell into the environment. We showed in this 2008 Oncogene paper that if you knocked out the lipocalin 2 gene and induced leukemia in mice, that didn’t make lipocalin 2. The disease was --- was disabled. So what the steps of CML leukemia are is --- see what happens in people that get leu --- CML or most other leukemias, the bone marrow and spleen get replaced by leukemia cells. Our data, which we published using this knockout model, that if you removed, not inhibited or reduced, but removed lipocalin 2 from the environment that _______ ) can no longer induce in its cells lipocalin 2 and could no longer secrete it. Then those leukemia cells did not cause leukemia in mice over many, many weeks. So wh --- why is that lipocalin 2 so important? Lipocalin 2 was first published by one of the people who advanced the field of lipocalin 2. His name is Michael Green. He sent me the CDNA to start working on lipocalin 2. He found out that lipocalin 2 kills normal T-cells and B-cells in mice and humans. So our data argued that chronic myeloid leukemia cells induce secretion of lipocalin 2 into the environment to kill normal spleen cells to make room in the spleen to allow the leukemia cells to overgrow the spleen, to kill bone marrow cells to allow the leukemia cells to overgrow. Because we showed the leukemia cells could not compete with normal cells to outgrow and overgrow cells in the bone marrow and spleen.

Tacey Ann Rosolowski, PhD:

Yeah, that is a really different theory then. It pushes ce --- normal cells out of the way.

Ralph B. Arlinghaus, PhD:

In fact it’s killing them with lipocalin 2.

Tacey Ann Rosolowski, PhD:

Really interesting. Wow.

Ralph B. Arlinghaus, PhD:

I’m very proud of that work.

Tacey Ann Rosolowski, PhD:

) Yeah. Fascinating.

Ralph B. Arlinghaus, PhD:

And I’m still working on it.

Tacey Ann Rosolowski, PhD:

So what are the --- what are the directions you’re working on now with it?

Ralph B. Arlinghaus, PhD:

With lipocalin 2? Well, it took a while to come to the next part but it --- it’s not published so it’s --- this is hypothesis now. It appears that the host supplies lipocalin 2 just as the leukemia cells produce lipocalin 2, that both steps are necessary for stable leukemia. So it’s not just the leukemia cells that are producing lipocalin 2, but it looks like its these normal bone marrow cells that go to the tumor sites --not published-- secrete lipocalin 2. So, for example, when leukemia has to spread to the spleen, something has to go into the spleen to prepare the way for the leukemia cells to overgrow it. And that turns out that there are normal cells that sort of partner with the leukemia cells to --- in this case, kill normal spleen cells to make space for the leukemia cells to grow. Leukemia cells are doing its part, but these other cells are also participating. So it’s much more complicated than just the leukemia cells producing this killer molecule, because it looks like normal immune cells, which go to the site and also assist in that killing normal cells.

Tacey Ann Rosolowski, PhD:

So the question of --- is how the leukemia cells are harnessing or co-opting the activity of those normal cells to partner --- make them come into partnership.

Ralph B. Arlinghaus, PhD:

Well I don’t know the answer to that. I don’t know the answer.

Tacey Ann Rosolowski, PhD:

Yeah. How interesting. How scary.

Ralph B. Arlinghaus, PhD:

It is. So you think of the normal host as being able to repress.

Tacey Ann Rosolowski, PhD:

Absolutely.

Ralph B. Arlinghaus, PhD:

But in this case our data says at some point for either breast ca --- we’ve done it in breast cancer models mouse and CML mouse models --that the host also produces lipocalin 2, which plays a role in the progression of the disease. I don’t like that result, but that’s the way it is. So how they partner, how they communicate, the breast cancer cell or the leukemia cell, I don’t know the answer to that. But you know when a tumor forms there’s a --- generally speaking there’s a tumor, what could we call it, a tumor environment and that environment is invaded by normal bone marrow cells some of which are meant to kill the tumor. Others like these cells I was talking about are supplying lipocalin 2 to help out in the breast cancer formation or the leukemia formation. That’s the way it looks right now.

Tacey Ann Rosolowski, PhD:

So very, very complicated process.

Ralph B. Arlinghaus, PhD:

Yes ma’am.

Tacey Ann Rosolowski, PhD:

Very complicated process.

Ralph B. Arlinghaus, PhD:

I haven’t published this so you --- this is hypothesis yet. The data that says the normal cells that move to the tumor mark environment and produce lipocalin 2 is true in my lab.

Tacey Ann Rosolowski, PhD:

Very, very interesting. So what other projects are you working on?

Ralph B. Arlinghaus, PhD:

Well I’m still working on chronic myeloid leukemia. In the mid-1990s I published a paper. I’m trying to --- Where was it published? I have to look it up. I can look it up for you now, but it was in 1995 and I was interested --- Remember chronic myeloid leukemia takes over the blood cell and causes the blood cell to grow uncontrol --- out of control? I wondered what that must mean, since again, I would call it unique thinking. That must mean BCR-ABL leukemia cells need help from the host and it turns out that leukemia cells have a protein. When you --- when you put the oncoprotein into leukemia cells they produce this protein called Janus kinase 2.

Tacey Ann Rosolowski, PhD:

You talked about that last time.

Ralph B. Arlinghaus, PhD:

And Janus kinase 2 is a major factor in how the cells make more copies of themselves if they are blood cells. So Janus kinase 2 is a major contributor to making blood cells. So I started thinking in 1995 that maybe BCR-ABL cooperates with jak2 to cause the disease, and that guess turns out to be right, because I published several papers along the way. The latest one in 2013 --- 2011 says that Janus kinase 2 and BCR-ABL function together to [?dry?] the leukemic phenotype in CML cells. So if you will remember the Philadelphia chromosome? I told you about that? Philadelphia chromosome forms just by mistake in people who have CML. That means the end of chromosome 9 and the end of chromosome 22 --- I think I said this last time. I don’t know if I did or not.

Tacey Ann Rosolowski, PhD:

I don’t know if you did.

Ralph B. Arlinghaus, PhD:

The end of 9 and 22 exchange. So now the end of 9 is on 22 and the end of 22 is on 9. That hybrid chromosome produces this hybrid protein called BCR-ABL. BCR is from chromosome 22 and ABL is from chromosome 9. Make a hybrid BCR-ABL protein. So that event is --- is critical for the deed but --- but then we’ve just found, again not published, that the BCR-ABL oncoprotein activates Janus kinase 2 to become --- to do what it does. Now remember Janus kinase 2, like all cellular enzymes, are usually silent, and they become activated from signals generally from outside the cell. But in the case of CML, BCR-ABL is activating Janus kinase 2 inside the leukemia cell constantly. Constantly driving jak2 to do things, to make more leukemia cells and have leukemia cells --- leukemia cells spread to other parts of the body. So some of this is published. The latest was 2013 in a leukemia journal. So …

Tacey Ann Rosolowski, PhD:

Quite the puzzle to unravel.

Ralph B. Arlinghaus, PhD:

So what I’ve found now, and haven’t published, is that BCR-ABL phosphorylates --- Does that term mean anything to you? --- phosphorylates jak2.

Tacey Ann Rosolowski, PhD:

I recognize the term.

Ralph B. Arlinghaus, PhD:

It’s a --- It’s --- It modifies jak2. It’s puts a phosphate on one of the tyrosines in jak2 that activates jak2. So jak2 is silent. Then BCR-ABL activates it, but it turns out that a normal ABL protein, again not published, --- the normal ABL protein activates jak2 in a normal situation. So jak2 in normal blood cells uses normal ABL to activate it. Again, under very highly controlled system, but when BCR-ABL becomes eternally activated in that cell, it’s always phosphorylating and activating jak2. So jak2 is always on, always doing things. Whereas in you and I our jak2 is active, silent, active, silent. You need more blood cells? Jak2 is active. You don’t need any more? Jak2 is shut off. So that control mechanism in CML doesn’t exist. It’s out of control. So we have preliminary data. What residue in jak2 is phosphorylated by either normal ABL kinase or the hybrid BCR-ABL kinase? So that’s the next paper I want to publish --- that --- that site, that BCR-ABL or ABL phosphorylates. Then I want to work on how does that phosphorylation event activate jak2? How does it make jak2 from an inactive enzyme to an active enzyme? That will be a next step. So if I can get grant support, I know what I’ll be working on two years from now. I just need to train people to do my experiments.

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Chapter 11: Research Projects: Working on an HIV Vaccine; Lipocalin 24p3; How CML Causes Uncontrolled Growth of Blood Cells

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