Chapter 06: Increasing Patients' Sensitivity to Taxol; Understanding the Function of ARHI (DISRAS3) and Autophagy

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Chapter 06: Increasing Patients' Sensitivity to Taxol; Understanding the Function of ARHI (DISRAS3) and Autophagy

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In this chapter, Dr. Bast discusses two other important areas of his research. He begins with a sketch of his work on reducing patients' resistance to Taxol by manipulating the overproduction of certain kinases. Next Dr. Bast talks about his work on the gene ARHI or DIRAS3. Reexpression of this gene inhibits cell proliferation and also induces autophagy of cancer cells as well as their dormancy. Dr. Bast explains autophagy, a process that provides energy for starving cells. He also explains that after surgery and chemotherapy, some ovarian cancer cells remain. His laboratory is currently looking at drugs that can promote autophagy to rid the body of these cells. Dr. Bast goes on to explain other animal studies in progress to reveal other mechanisms of DIRAS1, DIRAS2, and DIRAS3.

Identifier

BastRC_01_20140707_C06

Publication Date

7-7-2014

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; Discovery and Success; Understanding Cancer, the History of Science, Cancer Research; The History of Health Care, Patient Care; On Research and Researchers; Overview; Definitions, Explanations, Translations; Discovery and Success; MD Anderson Impact

Transcript

Robert Bast, MD:

In addition to early detection, for many years our laboratory pursued the idea of labeling or conjugating antibodies with the toxic A chain of ricin, a poison from castor beans, to try to deliver ricin A chain just to ovarian cancers and not to normal tissues. Radionuclide conjugates can also be delivered to ovarian cancer cells for very local radiotherapy. We were able to show that there was additive anti-tumor activity when ricin A chain conjugates and radionuclide conjugates were administered together.

Tacey A. Rosolowski, PhD:

Now, just so I’m understanding, this is the RAS? Am I hearing you properly? Because these are technical terms I may not have actually heard before. I mean, I may have read them. Is this what you’re—

Robert Bast, MD:

Not really Ras. Antibodies were directed against cell surface proteins on breast and ovarian cancer cells.

Tacey A. Rosolowski, PhD:

Oh, interesting.

Robert Bast, MD:

Over the last 10-15 years, we’ve focused our research in two additional areas: making paclitaxel (Taxol) chemotherapy more effective by modulating the sensitivity of ovarian cancer cells to the drug; and studies of ARHI (DIRAS3) an imprinted tumor suppressor that modulates cell growth, motility, autophagy and tumor dormancy. In the first area, we have studied more than a dozen kinases that regulate primary resistance to paclitaxel that is present before treatment with the drug. Knocking down each of these kinases has increased the drug’s effect on ovarian cancer cells. The most interesting has been SIK2. Ahmed Ahmed [MD, PhD], a postdoctoral fellow in our laboratory, had found that SIK2 was elevated in about thirty percent of ovarian cancers and that its overexpression was associated with a poor prognosis. SIK2 was required for the splitting of centrioles during cell division. When he knocked down SIK2, cancer cells became polyploid with more than two copies of each gene in each cancer cell. Knockdown of SIK2 enhanced sensitivity to paclitaxel. These observations formed the basis of a paper in Cancer Cell. Rationale for these studies goes back to a clinical trial that was done when paclitaxel was first being developed for ovarian cancer. The GOG [Gynecological Oncology Group] 132 clinical protocol tested cisplatin alone to paclitaxel alone, to a combination of paclitaxel and cisplatin in patients who had just had primary surgery for ovarian cancer, but had a very poor prognosis, because there was still a large amount of cancer that could not be resected. Seventy percent of patients responded either to platinum or to platinum plus paclitaxel, but only forty-two percent of patients responded to paclitaxel alone. In two of three large studies, patients lived longer with carboplatin and paclitaxel than with paclitaxel alone. Consequently we give all ovarian cancer patients carboplatin and paclitaxel, but no more than half of the patients benefit from the paclitaxel. There are two ways to improve care: one is to try to get a better predictive test to determine who does or does not respond to paclitaxel; the second is to make ovarian cancer cells more sensitive to the drug, so that we could raise that forty-two percent to a much higher number. And so we have identified kinases that regulate sensitivity to paclitaxel. Currently we are evaluating 14 kinase candidates funded by a grant from CPRIT [Cancer Prevention Research Institute of Texas]. One of the most promising targets is SIK2 and a small company named Arien has made an orally available inhibitor of the kinase. ARN3236 is a drug that inhibits growth of 80% of the ovarian cancer cell lines tested and that enhances sensitivity of ovarian cancer xenorafts to paclitaxel. So, this drug or one similar to it may actually be evaluated in clinical trials in the next year or two.

Tacey A. Rosolowski, PhD:

Mm-hmm.

Robert Bast, MD:

The third area that we’re involved in is in understanding the function of a gene that was originally called NOEY2 (Normal ovarian epithelium Yinhua-Yu 2), then ARHI (Aplesia Ras Homology I) and most recently DIRAS3. (laughter) So, maybe for today’s discussion, we’ll call it DIRAS3.

Tacey A. Rosolowski, PhD:

DIRAS3, yeah.

Robert Bast, MD:

But ARHI is more in the literature.

Tacey A. Rosolowski, PhD:

Okay.

Robert Bast, MD:

This gene is expressed by normal ovarian epithelium, but down-regulated in about sixty percent of ovarian cancers. DIRAS3 is also down-regulated in breast cancer, lung cancer, prostate cancer, pancreatic cancer, liver cancer and thyroid cancer. So, what we’re discovering in ovarian cancer might have much broader applicability. When you re-express DIRAS3 at physiologic levels, you inhibit proliferation. You inhibit motility. We’ve gone in the mechanisms of both of those. But most interesting, we’ve found that DIRAS3 induces a process called autophagy, and establishes tumor dormancy. We have genetically modified ovarian cancer cells, so that we can increase the levels of DIRAS3 with doxycycline. In culture, re-expression of DIRAS3 kills ovarian cancer cells within 3 days. But in xenografts in immunosuppressed nude mice, re-expression of DIRAS3 doesn’t kill cancer cells. They just sit there. And when you take the mice off of the doxycycline, the DIRAS3 goes back down in the cancer, the tumor xenografts grow out like nothing has happened. So, we’ve got a way to control dormancy. Dormant cells also undergo a process of autophagy or self-eating. If we inhibit autophagy ovarian cancer cells grow out more slowly and if we increase autophagy to a point where cancer cells die, the mice are cured.

Tacey A. Rosolowski, PhD:

Please.

Robert Bast, MD:

Straight technical material or some explanations.

Tacey A. Rosolowski, PhD:

I mean, a mixture helps, actually.

Robert Bast, MD:

Okay.

Tacey A. Rosolowski, PhD:

Because it is a varied audience for the material.

Robert Bast, MD:

Autophagy is a mechanism that’s used by normal cells as well as cancer cells to survive when they are starving without adequate amino acids or adequate glucose. During autophagy, vesicles formed within the cells that surround mitochondria, bits of endoplasmic reticulum, and high molecular weight proteins. Vesicles carrying this intracellular cargo then fuse with lysosomes. They autophagolysosomes then become acidified, activating proteases and lipases that break down proteins and lipids to amino acids and fatty acids which provide energy for starving cells. In the short run, this is provides a protective mechanism that saves cancer cells, but prolonged autophagy will kill cancer cells.

Tacey A. Rosolowski, PhD:

Mm-hmm.

Robert Bast, MD:

Chloroquine can block autophagy functionally by neutralizing the contents of autophagolysosomes so that the proteases and lipases are no longer active. You still have the autophagic vesicles, but they’re just not producing fatty acids and amino acids that are need for energy to maintain the viability of starving cancer cells. And so, but if you feed chloroquine to mice with xenografts that are dormant, you have increased DIRAS3 chloroquine. You markedly delay the outgrowth of tumors. Similarly, we found in cell culture that autophagic cells died, but if you restored some of the growth factors in the xenograft environment, you can partially rescue the autophagic cancer cells in culture. If you add antibodies against these growth factors to the culture, you don’t rescue them anymore. If you treat mice with antibodies against VEGF [Vascular Endothelial Growth Factor], against IL8 [Interleukin-8], and also against the receptor for IGF [Insulin-like growth factor receptor], you can cure a fraction of mice, completely inhibiting the outgrowth of the dormant cells.

Tacey A. Rosolowski, PhD:

Wow.

Robert Bast, MD:

So, we’ve discovered a couple of different ways to eliminate dormant cells; one would be chloroquine, although chloroquine, clinically, is, there are a number of toxic side effects. Hydroxychloroquine is another possibility.

Robert Bast, MD:

You could imagine, again, translating this work to try to eliminate dormant cells in ovarian cancer patients, because at the present time, about half to two thirds of patients with ovarian cancer after chemotherapy will have a normal CA-125 and PET CT. But if you perform “second look” surgery after chemotherapy, about half of those patients will have small nodules of cancer that are destined to grow back. Over the last 20 years, few second look operations have been performed as there was no curative therapy for persistent disease, but we are considering reinstituting “second looks” as part of the Moon Shot project at MD Anderson. Years ago, many second look procedures were performed at [Memorial] Sloan Kettering [Cancer Center]. From their pathology archives, we’ve obtained samples of tumor from primary cancers and from the second looks after platinum based therapy. Only 20% of the primary cancers had ARHI-positive autophagic cells, whereas 80% of the second looks contained numerous ARHI-positive autophagic cells consistent with our model for tumor dormancy.

Tacey A. Rosolowski, PhD:

Mm-hmm. Mm-hmm.

Robert Bast, MD:

It looks like our model with our inducible cell lines and xenografts actually mimics what’s actually happening in ovarian cancer patients. So, we have a really exciting opportunity to functionally inhibit autophagy and eliminate dormant cancer cells, with drugs like chloroquine and hydroxychloroquine, or new agents that are being developed or to eliminate growth/survival factors that are required to keep the autophagic cells from self-destructing after consuming their own body parts.

Tacey A. Rosolowski, PhD:

Right, right. Because how do you, I mean, how does—how do you force the autophagic cells to target the cancer cells?

Robert Bast, MD:

Well, it turns out that the cancer cells are autophagic.

Tacey A. Rosolowski, PhD:

Okay.

Robert Bast, MD:

Themselves.

Tacey A. Rosolowski, PhD:

Right, okay.

Robert Bast, MD:

And it looks like at least our experiments so far point to the fact that you’ve got to have autophagy to maintain dormancy.

Tacey A. Rosolowski, PhD:

So, so the ARHI is actually specifically linked to the autophagy of the cancer cell?

Robert Bast, MD:

Yes.

Tacey A. Rosolowski, PhD:

And no other kinds of cells?

Robert Bast, MD:

Well, no, to normal cells, as well.

Tacey A. Rosolowski, PhD:

Oh, okay.

Robert Bast, MD:

But presumably, the cancer cells depend more on autophagy than most normal cells.

Tacey A. Rosolowski, PhD:

Interesting. Oh, okay.

Robert Bast, MD:

And so, for—

Tacey A. Rosolowski, PhD:

So for especially—so that’s the sort of shift between cancers providing their own energy sources, and then going to needing angiogenesis to be fed by the blood supply?

Robert Bast, MD:

Yes, because the other thing about the small nodules on the peritoneal cavity that you can find at second look surgery is that they’re usually very poor in blood vessels. So, these cells are almost certainly nutrient deprived and likely to be autophagic.

Tacey A. Rosolowski, PhD:

Okay. Oh, interesting. Wow!

Robert Bast, MD:

We are also doing some more fundamental research. Zhen Lu [MD], an assistant professor, and Margie Sutton, a graduate student, in our lab are looking at the importance of more fundamental mechanisms related to DIRAS3 (ARHI). Mouse cells can undergo autophagy, but they don’t have DIRAS3.

Tacey A. Rosolowski, PhD:

Hmm.

Robert Bast, MD:

Mice split from the human lineage about sixty million years ago. Pigs and cows are on the human side; they have DIRAS3, but mice don’t. Mice and humans do have DIRAS1 and 2, and Margie is looking at the possibility that DIRAS1 and 2 may be doing for mice what DIRAS3 is doing for humans in terms of mediating autophagy. Another discovery that she has made is that DIRAS3 binds to Ras, and so we’re looking at the possibility that we might have a Ras-specific inhibitor. So far there’s no drug that actually specifically targets Ras.

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Chapter 06: Increasing Patients' Sensitivity to Taxol; Understanding the Function of ARHI (DISRAS3) and Autophagy

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