High throughput screen for selective inhibitors of cyclophilin B

Summary

Principal Investigator: Richard J Bram
Abstract: DESCRIPTION (provided by applicant): Current treatments for cancer are manifestly unsatisfactory in the extreme. Development of improved therapies will be accelerated by the identification of compounds that specifically target novel cellular proteins that contribute to tumr cell survival. The endoplasmic reticulum (ER) resident prolyl isomerase Cyclophilin B (CypB) is highly up-regulated in multiple types of cancers. We recently discovered that it plays an essential role in supporting the proliferation and survival of glioblastoma multiforme (GBM), an important type of brain tumor, as well as several other types of malignancies. Other groups have independently confirmed our observations on the importance of CypB for cancer cell survival. Biochemical analysis revealed that knockdown of CypB enhanced ER stress, and led to elevated reactive oxygen species (ROS) and decreased levels of Chk1 and mutant p53, as well as decreased activation of Stat3, all of which have been implicated in regulation of cancer cell proliferation, suggesting a mechanism underlying the requirement for CypB in these tumor cells. Inhibiting CypB to kill cancer cells would not be useful if normal tissues were equally dependent upon it for viability. Importantly, while suppression of CypB is deleterious for tumor cell surviva, we found that mice completely lacking CypB have normal development and fertility with a minor phenotype involving collagen processing, demonstrating that loss of CypB is not required for viability of the organism. We propose, therefore, that CypB is a novel target in cancer that can be inhibited therapeutically. There exist some small molecule inhibitors of cyclophilins, and several have been tested in clinical trials (thus far for the treatment of hepatitis). Using these inhibitors in vitro, we find that several of these compounds efficiently killed GBM cell lines and tumor cells freshly isolated from mice. However, there are a multitude of highly related cyclophilins in cells, and the known inhibitors typically bind and block the action of most of them Global inhibition of cyclophilins likely engenders multiple off-target and potentially conflicting effects on tumor cells, particularly in the case of the D isoform (CypD), which is known to participate in necrotic cell death. In addition, the abundance of cellular cyclophilins requires tht very high amounts of inhibitor be used in order to interfere with the activity of the single target CypB. These finding lead to our central hypothesis that potent compounds with the ability to selectively inhibit CypB will have improved efficacy and reduced toxicity in the clinical treatment of GBMs and other cancers. We propose here to leverage our novel findings and expertise in the cyclophilin system, in synergy with the high-throughput screening and medicinal chemistry expertise at the Conrad Prebys Center for Chemical Genomics (CPCCG) at the Sanford-Burnham Medical Research Institute (SBMRI), to identify and refine compounds that specifically target CypB and that do not bind to CypD. The proposed critical path testing funnel is already in place, so we anticipate that we can rapidly obtain and evaluate selective in vitro hits for their activity against GBM cell lines and freshly isolated tumor cells, and ultimately their suitability s starting points for hit-to-lead studies and for future in vivo evaluation in animal models and eventually patients. The goals of this proposal are to perform a high throughput primary screen of a ~370,000-compound library based on binding to recombinant CypB, through the use of a protein thermal shift assay. Secondary screening will use GBM cell cytotoxicity and CypD binding assays. Hits will be validated and then tested for effects on intracellular signaling pathways. We anticipate that these novel chemical probes that selectively suppress CypB intracellular functions will lead to proof-of-concept anti-cancer compounds. These compounds will ultimately lead to establishing a novel approach for therapeutic treatment of GBM. In addition, they will have great utility as molecular probes to enhance studies defining the critical functions of CypB.
Funding Period: 2013-09-01 - 2016-06-30
more information: NIH RePORT

Top Publications

  1. pmc Cyclophilin B supports Myc and mutant p53-dependent survival of glioblastoma multiforme cells
    Jae Won Choi
    Authors Affiliations Departments of Immunology, Radiation Oncology, and Pediatric and Adolescent Medicine, Mayo Clinic College of Medicine, Rochester, Minnesota
    Cancer Res 74:484-96. 2014

Detail Information

Publications1

  1. pmc Cyclophilin B supports Myc and mutant p53-dependent survival of glioblastoma multiforme cells
    Jae Won Choi
    Authors Affiliations Departments of Immunology, Radiation Oncology, and Pediatric and Adolescent Medicine, Mayo Clinic College of Medicine, Rochester, Minnesota
    Cancer Res 74:484-96. 2014
    ..Our findings link chaperone-mediated protein folding in the ER to mechanisms underlying oncogenic transformation, and they make CypB an attractive and immediately targetable molecule for glioblastoma multiforme therapy...

Research Grants30

  1. M. D. Anderson Cancer Center SPORE in Multiple Myeloma
    ROBERT ZYGMUNT ORLOWSKI; Fiscal Year: 2013
    ..abstract_text> ..
  2. University of Maryland Greenebaum Cancer Center Support Grant
    Kevin J Cullen; Fiscal Year: 2013
    ..Reflecting our remarkable and continued growth, UMGCC seeks to renew its CCSG to enhance and expand its efforts in high-quality and clinically relevant cancer research. ..
  3. Optical Technologies and Molecular Imaging for Cervical Neoplasia
    Michele Follen; Fiscal Year: 2013
    ..abstract_text> ..
  4. Glioblastoma stem cells therapy
    Bakhos A Tannous; Fiscal Year: 2013
    ..We will validate the most promising drug hits in our experimental glioma stem cells-bioluminescent model in vivo. ..
  5. Signaling in Inflammation, Stress, and Tumorigenesis
    GEORGE ROBERT STARK; Fiscal Year: 2013
    ..abstract_text> ..
  6. Chicago Prevention and Intervention Epicenter (Chicago PIE)
    ROBERT ALAN WEINSTEIN; Fiscal Year: 2013
    ..The impact on ICU infection and prescribing characteristics of doctors will be assessed. To further assess the interventions, costs of averted outcomes and of the interventions will be compared. OPRIONAL OBEJCTIVE SCORE: 2 ..
  7. Nanoplatform Delivery of MDA-7 and Brachytherapy for Treatment of Brain Tumors
    Michael D Shultz; Fiscal Year: 2013
    ..The skills and knowledge obtained through the proposed mentoring will be invaluable to the success of the applicant and lead to unique and innovative projects in future endeavors. ..
  8. Mechanistic Pharmacology of Anti-Mitotics and Apoptosis Regulation
    Timothy J Mitchison; Fiscal Year: 2013
    ..In aim 4 we will pursue several approaches towards translating mechanistic understanding from aims 1-3 into improved patient care. ..
  9. Characterization of Pathways Controlling Cancer at the Level of Gene Regulation
    Phillip A Sharp; Fiscal Year: 2013
    ..The interactions and involvement of Rb and miRNAs in induction of cell death following DNA damage will also be studied. ..
  10. UNMC EPPLEY CANCER CENTER SUPPORT GRANT
    Kenneth H Cowan; Fiscal Year: 2013
    ....
  11. Cyclophilin B as a therapeutic target in glioblastoma
    Richard J Bram; Fiscal Year: 2013
    ..Together, these studies will test the feasibility of CypB as a novel target for therapy of GBM, and set the stage for transition to clinical studies in humans. ..
  12. Mayo Clinic Breast Cancer SPORE
    JAMES NEWELL INGLE; Fiscal Year: 2013
    ..abstract_text> ..
  13. Experimental Therapeutics of Leukemia
    John C Byrd; Fiscal Year: 2013
    ..We believe that this SPORE group, as a multidisciplinary, highly interactive and accomplished team, will have a substantial impact on improving the clinical outcome of leukemia patients. ..