Ovarian Cancer SPORE Grant Research Projects

The Mayo Clinic Ovarian Cancer SPORE's major research themes focus on pathogenesis (immunosuppression) of ovarian cancer and translational approaches to improve clinical outcomes. Each project in the Ovarian Cancer SPORE is codirected by interdisciplinary investigative teams, including basic scientists, population scientists and clinician scientists with expertise in conducting translational research.

The four research projects in the Ovarian Cancer SPORE at Mayo Clinic are:

Project 1: Poisoning of PARP and topoisomerase I (topo I) to treat ovarian cancer

Principal investigators: Scott H. Kaufmann, M.D., Ph.D., and Harry J. Long III, M.D.

Although most women initially respond to platin-based chemotherapy, the high relapse rate and poor response to subsequent therapy underscore the need for more effective therapy for platin-resistant ovarian cancer.

Our previous studies examining the action of topotecan (TPT), a topoisomerase I-directed agent that is approved for the second line treatment of ovarian cancer, have demonstrated that TPT induces replication fork stalling followed by activation of a kinase cascade involving ATR and checkpoint kinase 1. Building on reports that poly (ADP-ribose) polymerase-1 (PARP1) is involved in restarting stalled replication forks and in reversing trapped topo I-DNA complexes, we have more recently demonstrated that the PARP inhibitor veliparib enhances TPT cytotoxicity in ovarian cancer cell lines. Even though this sensitization is more prominent in BRCA1/2-deficient cells, it also occurs in ovarian cancer cells with wild-type BRCA1 and BRCA2. Moreover, this sensitization occurs at veliparib concentrations that are 20-fold lower than those required to kill BRCA1/2-deficient cells directly. Additional results indicate that veliparib is sensitizing cells through a base excision repair pathway that involves XRCC1 and suggest that PARP1 must be present for this sensitization.

To further study this interaction between TPT and veliparib, and to extend the potential benefits of PARP inhibitor therapy to as large a subset of ovarian cancer patients as possible, the Mayo Clinic Ovarian Cancer SPORE investigators have several aims:

• Aim 1. Identify the mechanism by which PARP1 inhibition enhances the antiproliferative effects of TPT in ovarian cancer cells by further elucidating the DNA repair pathway that is critical for veliparib mediated sensitization and determining how inhibited PARP1 actively sensitizes cells to TPT.
• Aim 2. Determine the mechanism of veliparib-induced anti-proliferative effects in BRCA1/2-deficient cells by identifying the endogenous DNA lesions that contribute to the cytotoxicity of veliparib and examining how PARP inhibition contributes to this demise.
• Aim 3. Evaluate the ability of a series of markers to predict response to the TPT/veliparib combination in a CTEP-sponsored phase II trial in patients with relapsed ovarian cancer. Samples from patients enrolled in this trial provide a unique opportunity to search for potential predictive markers of response to this regimen.

These studies, which make extensive use of the Biospecimens and Patient Registry Core and the Biostatistics Core of the Mayo Clinic Ovarian Cancer SPORE, are designed to increase understanding of the action of the PARP inhibitor veliparib, both alone and in combination with TPT, in ovarian cancer, thereby advancing clinical development of the PARP inhibitor as a potentially promising ovarian cancer therapy.

Project 2: Mechanisms of immunosuppression in ovarian cancer

Principal investigators: Keith L. Knutson, Ph.D., and Ellen L. Goode, Ph.D.

Ovarian cancer has a high mortality rate, and despite some improvements in survival with new chemotherapies, the cure rate has not improved in decades. While clinical features, such as cancer stage, are excellent indicators of outcome, significant variability in outcomes remains. Although it is recognized as an immune reactive malignancy, ovarian cancer eludes immunity because of a tumor-induced, complex immune suppressive network. In this project within the Ovarian Cancer SPORE, we are examining determinants of immune suppression at an inherited level and at the level of the tumor microenvironment. We are integrating inherited variation and novel tumor phenotypes with rich clinical annotation in biological models of these microenvironmental relationships and their impact on survival. We are studying three unique elements of this network — CD4+ T-regulatory cells (Tregs), CD8+ Tregs and PD-1+ expression on intratumoral dendritic cells (DCs).

Hypothesis
Our general hypothesis is that immune suppression, which reflects a sophisticated interaction of a network of genes, molecules and cells, contributes to ovarian cancer pathogenesis. This hypothesis is examined in the following aims:

• Aim 1. To evaluate the association between inherited variation in 32 immune regulatory genes and overall survival among invasive ovarian cancer cases
• Aim 2. To determine the role of CD8+ Tregs and PD-1+ DC in the ovarian cancer immune microenvironment
• Aim 3. To assess the role of inherited variation in immune regulatory genes and intermediate tumor phenotypes in a multivariate model of survival in ovarian cancer

Our proposed transdisciplinary project integrates population and basic research to increase our understanding of the immunological mechanisms guiding relationships between host factors, immunological tumor characteristics and ovarian cancer outcome. Results of this work will direct subsequent immune therapies in our already-existing immunotherapy program. The clinically useful information being developed in this Ovarian Cancer SPORE project includes:

• Identification of targets to block the suppressive mechanisms that ovarian cancer employs to evade immune surveillance and eradication
• Identification of ways to better personalize use of novel immune-based therapies for the prevention of ovarian cancer recurrence
• Identification of novel immune targets not currently addressed with immune-based therapies

Project 3: Optimizing measles virotherapy in the treatment of recurrent ovarian cancer

Principal investigators: Evanthia Galanis, M.D., and Kah Whye Peng, Ph.D.

We have demonstrated that engineered measles virus strains have significant anti-tumor activity against ovarian cancer lines and xenografts. Their tumor specificity is due to abundant expression of the measles virus receptor CD46 in ovarian cancer cells. The virus, upon entry into tumor cells, causes membrane fusion with neighboring cells, syncytia formation and death. Our group in the Mayo Clinic Ovarian Cancer SPORE was the first to translate this approach into a phase I clinical trial of a measles virus derivative producing human carcinoembryonic antigen, MV-CEA (CEA added to facilitate viral monitoring) in recurrent ovarian cancer patients. Despite low levels of viral replication, as evidenced by modest CEA elevation in a minority of patients, there was promising early evidence of anti-tumor activity, including CA-125 decreases and prolonged disease stabilization in heavily pre-treated patients.

Hypothesis
We hypothesize that by increasing the efficiency and extent of tumor cell infection we can further augment the anti-tumor activity of measles virotherapy in ovarian cancer. We propose to accomplish this by testing the translational potential of three novel approaches:

• A different measles virus strain, MV-NIS, which encodes the sodium iodide symporter (NIS) therapeutic transgene, thus allowing imaging of viral distribution in vivo and use of iodine-131
• Use of infected cell carriers for viral delivery
• Combining the measles virus with cyclophosphamide, an immunosuppressive medication with anti-tumor properties

This project has four aims:

• Aim 1. To perform a limited phase I trial of intraperitoneal (IP) administration of MV-NIS in patients with recurrent ovarian cancer
• Aim 2. To optimize the efficacy of IP measles virotherapy for ovarian cancer in measles-immune mice by employing virus-infected cell carriers and testing the added benefit of cyclophosphamide
• Aim 3. To test the efficacy of intravenous (IV) measles virotherapy for ovarian cancer and optimize it in measles-immune mice by using virus-infected cell carriers, with and without addition of cyclophosphamide following optimization of IP or IV delivery. The added value of Iodine-131 will also be tested
• Aim 4. To translate the most promising approach to a phase I trial in patients with recurrent ovarian cancer

Project 4: Mechanism-guided development of an innovative therapy for ovarian cancer

Principal investigators: Larry M. Karnitz, Ph.D., and Paul Haluska Jr., M.D., Ph.D.

Here, Mayo Clinic Ovarian Cancer SPORE investigators propose a novel drug combination based on our mechanistic studies analyzing responses activated by floxuridine (FUdR), an FDA-approved drug with activity in ovarian cancer. In cells, FUdR is converted to FdUMP, which inhibits thymidylate synthase (TS), causing accumulation of dUTP. FdUMP is also converted to FdUPT. Both dUTP and FdUTP are then incorporated as U and 5-fluorouracil (5-FU) lesions in the genome.

The genomically incorporated U and 5-FU are repaired by the base excision repair (BER) pathway, in which lesions are removed by uracil glycosylases (SMUG1, UNG, MBD4, TDG) generating abasic sites that are processed to single-stranded (ss) DNA breaks. The ssDNA breaks recruit poly (ADP-ribose) polymerase (PARP)-1 and PARP-2 (referred to as PARP), leading to PARP activation and the accumulation poly (ADP-ribose) polymers on PARP and other proteins that complete repair.

Ovarian Cancer SPORE investigators have discovered that PARP inhibitors (PARPi), including veliparib, remarkably sensitized ovarian cancer cells to FUdR (PARPi did not sensitize to 5-FU).

We have also discovered that defects in several DNA repair pathways, including some that are often altered in ovarian cancers — positively and negatively — affect the cytotoxicity of FUdR alone and FUdR+veliparib (Fd+A).

Hypothesis
We hypothesize that:

• A series of DNA repair enzymes converts the DNA structures induced by Fd+A into highly toxic lesions that kill cells.
• The levels of these repair proteins may correlate with patient responses to Fd+A, a combination that will undergo clinical testing.

Accordingly, we will test these hypotheses with these aims:

• Aim 1. Delineate the roles and regulation of uracil glycosylases in FUdR-treated ovarian cancer cells
• Aim 2. Mechanistically evaluate the role of ERCC1 in the toxicity of FD+A
• Aim 3. Evaluate the ability of potential biomarkers to predict responses of ovarian cancer to Fd+A