The overall goal of the Phase III MT-COBRE is to ensure that the novel targets and compounds that have been identified by MT-COBRE faculty are translated to pivotal human clinical trials.  To that end, the Pilot Project Program has, as its major objective, the identification and initial funding of research projects to develop novel pre-clinical therapeutic agents identified by MT-COBRE Program and other cancer investigators.  Although the theme of this funding mechanism will be translational development of novel therapeutics and priority will be given to translational studies with a high potential for advancement to clinical studies, other meritorious grants may be funded as well.  It is the intent that these developmental projects will lead to R01 and other competitive grant funding through expansion or re-direction of their COBRE projects.  The Pilot Project Program has a total 5-year budget of $1,725,000; comprised of $575,000 from the MT-COBRE grant, $500,000 from the Executive Vice-President for Research at UofL, and $650,000 from discretionary funds at the James Graham Brown Cancer Center.  Applications are solicited by an RFP process and reviewed by the MT-COBRE Steering Committee.

Year 1:  July 1, 2013 – June 30, 2014

Haribabu Bodduluri, Ph.D. and John O. Trent, Ph.D.

Geoffrey J. Clark, Ph.D.

Brian F. Clem, Ph.D.

Uma Sankar, Ph.D.

Sucheta Telang, M.B.B.S.

Kavitha Yaddanapudi, Ph.D.

Year 2:  July 1, 2014 – June 30, 2015

Due to the degree of progress in the five grants listed here, and described above, they were renewed for a 2^nd year of funding.

See above:  Haribabu Bodduluri, Ph.D. and John O. Trent, Ph.D.; Geoffrey J. Clark, Ph.D.; Brian F. Clem, Ph.D.; Sucheta Telang, M.B.B.S.; and Kavitha Yaddanapudi, Ph.D.

Year 3:  July 1, 2015 – June 30, 2016

Paula J. Bates, Ph.D.

Chi Li, Ph.D.

Nobuyuki Matoba, Ph.D.

Robert A. Mitchell, Ph.D. and Kavitha Yaddanapudi, Ph.D. 

Abstracts

Year 1 (July 1, 2013 – June 30, 2014)

Leukotriene B₄ Receptor Agonists and Antagonists for Cancer Immunotherapy – Haribabu Bodduluri, Ph.D. and John O. Trent, Ph.D.

Chronic Inflammation is known to promote a wide range of diseases including cancers of lung, prostate, breast and colon.  Recent studies from our laboratory in a mouse model of spontaneous lung cancer (K-ras^LA1) showed that exposure to crystalline silica resulted in a significant increase in lung tumor burden.  Moreover, absence of the leukotriene B4 (LTB₄) receptor BLT1 attenuated this increase.  In previous studies, our laboratories developed molecular models that were 100% predictive in defining the binding pocket and in delineating an activation mechanism for BLT1.  Our modeling studies also validated the binding of inflammation resolving lipids derived from the omega-3 fatty acids (resolvinE1) to BLT1 and their role as weak agonists of BLT1.  Therefore, targeting the LTB₄/BLT1 pathway offers an excellent opportunity for developing novel immunotherapeutics.  Preliminary results on structure- and mechanism-based virtual screening identified an agonist and an antagonist for BLT1.  Activity of the agonistic compound was further enhanced by combinatorial chemical synthesis of related compounds.  One such compound was found to be a high affinity partial agonist for BLT1 that also acts as an antagonist leading to the hypothesis that “targeting BLT1 with weak partial agonists will inhibit chronic inflammation without creating immunodeficiency”.  The current proposal will test this hypothesis in two specific aims.  In aim 1, we will prepare additional compound related agonists (#14 and #63) as well as the antagonist (#9) and identify compounds with low toxicity and high efficacy in cellular models.  In aim 2, we will test the in vivo efficacy of selected compounds in an air pouch model of silica-induced inflammation.  Those compounds that display low toxicity and high efficacy in this model will be tested in the silica promoted lung cancer in K-ras^LA1 mice. These studies will identify novel compounds that block the BLT1-mediated chronic inflammation and will lead to the development of therapeutics for targeting inflammation associated with lung cancers.

The Development of Novel Ras Antagonists to Inhibit Cancer – Geoffrey J. Clark, Ph.D.

The Ras oncoproteins are activated by mutation in almost one-third of human cancers.  Even in the absence of structural mutation, Ras proteins can be activated in tumors by defects in upstream Ras regulators.  An enormous body of work has been amassed over the last three decades supporting a key role for Ras activation in the development of metastatic cancer.  Yet Ras has defied efforts to develop effective targeted therapies directed against it.

Ras acts by binding and activating a panel of effector proteins, many of which are oncoproteins in their own right.  The best characterized of these are the Raf kinases, PI-3 kinases and the RalGDS family.  Considerable evidence recently has been presented showing that the RalGDS pathway may be the most important for the transforming effects of Ras in vivo.  There are currently no inhibitors of this pathway.  The crystal structure of Ras bound to RalGDS has been described.  Based on this structure, we have performed an in silico screen to identify compounds that are predicted to bind to the Ras effector domain when it is in the correct conformation to bind RalGDS.

In preliminary studies, several molecules have demonstrated inhibition of transformation and Ras signaling at 5 µM concentration in multiple cell lines.  This concentration was not growth inhibitory to the cells growing normally on plastic.  This action is exactly what we would predict if the molecules were specifically inhibiting the Ras/RalGDS pathway by binding to Ras and preventing a productive interaction with its effector RalGDS.  We propose that these molecules may serve as the basis for novel anti-Ras agents that could be highly effective against a range of cancer cells.  In our first two specific aims, we propose to further optimize the first generation molecules in silico, and confirm that they act to bind and block the interaction of Ras with its effectors.  We then seek to use xenograft assays to determine if the molecules can antagonize tumor formation and metastasis of two different cell lines driven by aberrant Ras activity.

We propose two further aims for the second year of the project.  The first seeks to further enhance the activity of the molecule in vivo by employing Medicinal Chemistry.  Finally, the fully optimized derivatives of our original molecules will be tested for the ability to inhibit the growth of mutant Ras containing human primary tumor grafts.

Aim 1: Optimize the inhibitors and define their mechanism of action.

Aim 2: Determine effects of the inhibitors against human tumor cell lines in vivo.

Aim 3:  Medicinal Chemistry Optimization of the lead compound.

Aim 4:  Determine if the best two optimized inhibitors are effective against Ras-driven human primary tumor xenografts (pdx).

Targeting PhosphoSerine Aonptransferase (PSAT1) in the Treatment of Lung Cancer – Brian F. Clem, Ph.D.

In the absence of glucose oxidation, a result of a metabolic shift towards aerobic glycolysis, cancer cells increase glutamine consumption primarily to fulfill the need for anaplerotic carbon metabolism within the TCA cycle.  This alteration ultimately drives these cells to glutamine dependency for cell survival as glutamine withdrawal is selectively cytotoxic to neoplastic cells harboring oncogenic or tumor suppressor mutations and suggests that the enzymes mediating these regulated processes may be cellular targets for anti-cancer strategies.  In particular, phosphoserine aminotransferase (PSAT1) converts glutamate to the TCA intermediate, a-ketoglutarate, while catalyzing the production of phosphoserine from phosphohydroxy pyruvate within the serine biosynthetic pathway.  Importantly, both elevated glutaminolysis and glucose-derived carbon flux through the serine pathway has been observed in lung cancer cells indicating that these neoplasms may increase glucose shunting into serine production to directly facilitate glutamine metabolism for anaplerosis.  However, the requirement for the enzymes within this pathway, specifically PSAT1, in support of glutamine metabolism and lung tumor growth has not yet been established.  Using Oncomine analyses and immunohistochemistry on primary human lung tissue, we found that PSAT1 is significantly elevated in lung tumors compared to matched normal lung.  We also observed increased PSAT1 transcripts and protein levels in two separate human lung cancer cell lines compared to normal bronchial epithelial cells and suppression of PSAT1 resulted in decreased proliferation of H460 lung carcinoma.  To determine the potential consequence of PSAT1 inhibition on glutamine metabolism, we examined glutamine uptake and observed a decrease after RNAi exposure.  In support of the specific targeting of PSAT1, administration of putative small molecule antagonists identified through in silico screening also decreased growth and glutamine uptake in the H460 cells.  From these preliminary studies, we hypothesize that PSAT1 activity is required for maintaining cell growth and survival of lung cancer cells, in part, through direct functional support for glutamine metabolism.  Using a combination of shRNA and small molecule inhibitor approaches, we will test this hypothesis by conducting the following Specific Aims:  1. determine the effect of PSAT1 inhibition on glutamine metabolism and mitochondrial function in lung cancer; and 2. examine the effect of PSAT1 inhibition on the growth and survival of lung cancer cells in vitro and in vivo.

CaMMK2 Inhibition s a Dual-Hit Strategy in the Prevention of Prostate Cancer Growth and ADT-induced Bone Loss – Uma Sankar, Ph.D.

Prostate cancer cells rely on sex steroids or androgens for cell proliferation and migration.  Consequently, androgen deprivation therapy (ADT) is the standard therapy against advanced-stage prostate cancer.  However, ADT results in a host of deleterious side-effects including rapid bone loss, putting patients at a significantly high risk of sustaining fragility fractures of the bone, including hip fractures.  A drug that can inhibit these two separate-but-related complications will alleviate a great deal of suffering and mortality associated with this malignancy.  Our previous studies indicate that the loss of Ca₂₊/calmodulin-dependent protein kinase kinase 2 (CaMKK2) or its pharmacological inhibition with a selective cell-permeable inhibitor STO-609 promotes robust bone growth while inhibiting bone resorption.  Moreover, administration of STO-609 confers protection from osteoporosis-induced by androgen deprivation in mice.  Further, several recent studies report CaMKK2 to be highly elevated in prostate cancer and its inhibition using STO-609 to restrict cancer cell growth in vitro and in vivo.  Thus, CaMKK2 represents a highly attractive drug target for the control of prostate cancer cell growth while preventing ADT-induced bone loss.  However, STO-609, which is the only pharmacological inhibitor of CaMKK2, is not specific to it.  Whereas it selectively inhibits CaMKK2 at low concentrations, it also inhibits CaMKK1 and other family members at higher concentrations.  Hence the development of a specific, cell permeable inhibitor of CaMKK2 is highly warranted and clinically urgent.  The objective of the proposed studies is to identify and develop a specific, cell-permeable pharmacological inhibitor of CaMKK2 for use as a “dual hit” therapeutic strategy to control cancer cell growth while protecting from ADT-mediated osteoporosis in patients with advanced-stage prostate cancer.  Specifically, the idea of getting a “double hit” on the cancer as well as on therapy-induced osteoporosis with the same drug is a highly innovative and appealing strategy in the treatment of patients with advanced forms of prostate cancer.

Targeting 6-Phosphofructo-2-Kinase/Fructose-2,6-Bisphosphatase-4 in Lung Cancer – Sucheta Telang, M.B.B.S.

Lung cancer accounts for the majority of cancer-related mortality in the world – in 2013, over 160,000 deaths are predicted in the United States alone.  Non-small cell lung cancers (NSCLC), the most frequent (85%) of lung tumors, exhibit markedly increased glucose uptake and glycolysis over normal lung and this metabolic phenotype serves to satisfy their increased requirement for energy and biosynthetic intermediates.  Oncogenic alterations, overexpression of HIF-1α and the loss of p53 function each stimulate glycolysis by activating a family of bifunctional 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatases (PFKFB1-4).  The PFKFBs produce fructose-2,6-bisphosphate (F2,6BP) which is an allosteric activator of 6-phosphofructo-1-kinase, a rate limiting step in the glycolytic pathway.  Until recently, the PFKFB3 isoform has been considered the dominant source of F2,6BP in tumor tissue due to a high kinase activity coupled with expression in multiple tumors.  However, roles of the other isoforms have not been fully explored and recent studies indicate that PFKFB4 expression is increased in several solid tumors.  In examination of the expression of the PFKFB isoforms in lung cancer and normal lung, we found that PFKFB4 expression was significantly increased relative to the other PFKFB isoforms in lung tumors.  We also determined that PFKFB4 mRNA and protein expression were strongly induced by hypoxia, through upregulation of HIF-1α, and that decreasing PFKFB4 expression, particularly in hypoxia, markedly reduced F2,6BP production, glucose uptake, glycolytic flux to lactate and the TCA cycle and increased apoptosis.  We now have identified a novel small molecule inhibitor of PFKFB4 (termed 5MPN) that inhibits recombinant PFKFB4 activity (without affecting PFKFB3 activity).  5MPN decreased F2,6BP and suppressed glucose uptake in vitro, was cytostatic to several NSCLC cell lines, did not affect the growth or viability of normal bronchial epithelial cells and, importantly, caused a marked reduction in growth of established Lewis lung carcinoma tumors in mice in vivo.  Based on our preliminary results, we hypothesize that PFKFB4-catalyzed F2,6BP may be critical for survival and glycolytic flux of tumors in hypoxia and that inhibition of PFKFB4 with a small molecule inhibitor may prove to be an effective anti-neoplastic strategy.  The objectives of our application are to confirm the mechanism of action of 5MPN and to examine the ability of 5MPN to attenuate tumor growth in a physiologically relevant mouse model of lung cancer. We plan to accomplish these objectives by pursuing the following aims:  1. determine the effects of 5MPN on metabolism, growth, invasiveness, and resistance to apoptosis of normal, immortalized and ras-transformed human bronchial epithelial cells in vitro and in vivo; and 2. examine the effect of 5MPN on the growth of ras-dependent lung adenocarcinomas in vivo.

A Broad Spectrum Lung Cancer Stem Cell Vaccine – Kavitha Yaddanapudi, Ph.D.

Lung cancer is a prevalent disease that consumes many lives every year.  Disease relapse, invasion and metastases are the main causes of death.  Recent discoveries provide compelling evidence that at least some types of cancer are initiated and maintained by a small population of malignant cells called cancer-initiating stem cells (CICs).  Relapse, invasion, and metastases are explained by the fact that CICs have a different biology than all other tumor cells and, importantly, are resistant to chemotherapies and radiation.  Lung CICs have been shown to represent about 1-15% of all tumor cells and can form tumors with injections as low as 100 cells.  Evidence from published studies have demonstrated that human, as well as rodent, cancers contain populations of cells that express embryonic stem (ES) cell antigens.  Cells containing these proteins also express markers used to identify lung CICs; therefore, we hypothesized that ES cells and CICs share several common molecular traits.  To test this hypothesis, we vaccinated mice with irradiated, allogeneic murine ES cells in combination with a source of granulocyte-macrophage colony stimulating factor (GM-CSF) as an immunostimulatory adjuvant (ES cell vaccine) and investigated whether an anti-tumor immune response was elicited.  We discovered that ES cell vaccination is very effective in preventing both implantable and carcinogen-induced lung adenocarcinoma development without any detectable toxicity or signs of autoimmunity.  Preliminary studies from our laboratory reveal that splenocytes from ES cell-immunized mice are preferentially cytotoxic to lung CICs.  Experiments proposed in this application seek to expand these novel findings to convincingly demonstrate that ES cells immunize against lung cancer associated CICs and that anti-CIC immunity is responsible for preventing lung adenocarcinoma development.  A major goal of this study is to assess the potential of this novel, CIC-targeting ES cell vaccine as a treatment option for lung cancer patients.  We will test the translational potential of our approach using a modified and novel ES cell vaccination strategy consisting of irradiated ES cells chemically fused with normal dendritic cells (DC).  We will test the ES/DC fusion vaccine in conjunction with GM-CSF (ES/DC-GM).  Experiments proposed in this study will address the in vivo efficacy and safety of ES/DC-GM vaccine in prophylactic as well as therapeutic settings against both primary and metastatic lung cancer.  To fulfill the stated objectives, the following aims are proposed: 1) Investigate whether lung cancer-initiating cells are targets of ES cell vaccination-induced anti-tumor immunity; and 2) Evaluate the translational potential of a novel ES cell/Dendritic cell fusion vaccine in a spontaneous pulmonary metastasis mouse model of lung cancer.  Our proposed study will provide important insights towards developing a safe and effective immunotherapeutic vaccine for lung cancer onset and/or recurrence.

Year 2 (July 1, 2014 – June 30, 2015)

Due to the degree of progress in the five grants listed here, and described above, they were renewed for a 2^nd year of funding.

See above:  Haribabu Bodduluri, Ph.D. and John O. Trent, Ph.D.; Geoffrey J. Clark, Ph.D.; Brian F. Clem, Ph.D.; Sucheta Telang, M.B.B.S.; and Kavitha Yaddanapudi, Ph.D.

Year 3 (July 1, 2015 – June 30, 2016)

Targeting SOX9 as a Novel Approach to Treating Intractable Cancers – Paula J. Bates, Ph.D.

It has become clear lately that many solid tumors contain a heterogeneous population of cancer cells, often including a subpopulation of tumor initiating cells (TICs) that drive cancer progression and metastasis.  The failure of current treatments to eradicate TICs (also referred to as cancer stem cells) appears to be a major contributor to clinical relapse and recurrence; thus, combining therapies that destroy TICs with conventional regimens could lead to greatly improved clinical outcomes for patients with intractable cancers.  SOX9 is a developmental transcription factor that is frequently reactivated in many aggressive cancer types.  There is compelling evidence that SOX9 is a master regulator of TICs and that inhibiting SOX9 activity would be an excellent strategy for treating cancer.  However, SOX9 appears to have been overlooked as a molecular target, perhaps because, as a transcription factor, it is (incorrectly) assumed to be “undruggable”.  Our group has now developed two new approaches to selectively target and destroy SOX9-high cancer cells.  The first involves XB05, a synthetic small molecule that we recently discovered to have a strong preference for killing SOX9-high cancer cells.  SOX9 is probably not a direct molecular target of XB05, but we hypothesize that SOX9-high TICs are exquisitely sensitive to the disruption of redox homeostasis caused by this molecule.  The second approach involves small molecules discovered by a virtual screen for compounds that target the DNA-binding domain of SOX9.  These molecules have been tested in a variety of cell lines and exhibit the expected selectivity (i.e., highly cytotoxic to SOX9-high cancer cells with much less activity against normal cells and SOX9-low cancer cells), but have not yet been assessed in functional assays.  The objective for this pilot project is to obtain definitive data regarding whether these small molecules can preferentially inhibit SOX9 activity and target SOX9-high cancer cells without toxicity to non-malignant cells.  Our specific aims are: (1) determine the effect of candidate SOX9 inhibitors on SOX9 activity in vitro, (2) evaluate the ability of candidate SOX9 inhibitors to target SOX9-high cells in an ex vivo system, and (3) synthesize and characterize analogs of SOX9-inhibitors to generate structure-activity relationships.  This is a highly collaborative project involving a PI who co-discovered the candidate SOX9 inhibitors, a team of co-investigators with expertise in cancer drug development and medicinal chemistry, and a collaborator who has elucidated the important role of SOX9 in cancer stem cell biology.  There is a high level of innovation due to the novelty of the target, the entirely unexplored class of compounds represented, and the clinical relevance of the ex vivo cancer model.  Moreover, the translational potential of this research is high; we envision that the ultimate product will consist of a therapeutic agent (an optimized analog of our best SOX9 inhibitor) plus a companion biomarker assay to identify patients with SOX9-driven cancers who are most likely to benefit from therapy. By the end of this pilot project, we expect to have compelling data that will allow us to secure substantial additional funding and move forward with these promising compounds.

Novel Antitumor Activity of a Bacterial Homoserine Lactone – Chi Li, Ph.D.

An important task in cancer research is to develop new treatments that overcome the resistance to cell death displayed by tumor cells due to impaired apoptotic pathways. The opportunistic bacterium, Pseudomonas aeruginosa, produces the quorum sensing molecule N-(3-oxododecanoyl)-homoserine lactone (C12).  We recently have found that C12 preferentially induces tumor cell apoptosis in vitro and inhibits transplanted tumor growth in vivo independent of Bcl-2 proteins, probably through direct damage to the mitochondrial outer membrane.  Importantly, C12 cytotoxicity is mediated through the lactonase activity of paraoxonase 2 (PON2).  PON2 is upregulated in many types of cancer, including lung, enabling cancer cells to resist conventional therapeutic drugs.  Decreasing PON2 expression in human lung tumor cells reduces cell proliferation, lowering the likelihood of drug-induced resistance.  We propose that PON2 cleaves C12 into a secondary, cytotoxic metabolite(s) that preferentially induces lung tumor cell apoptosis without the involvement of Bcl-2 proteins.  The experiments in this grant are designed to explore the potential of C12 or C12 metabolite(s) as an anti-lung cancer therapeutic treatment.  Two specific aims are envisioned:  1) Investigate the mechanism of apoptosis mediated by PON2/C12 interaction; and 2) Investigate the role of PON2 in inhibitory effects of C12 on lung tumors.  We believe that PON2/C12 interaction represents a novel therapeutic target for discovering drugs that induce apoptosis in PON2-overexpressing lung tumors in a fashion independent of the Bcl-2 protein profile.  Triggering apoptosis in tumor cells by C12 or C12 metabolite(s) through directly permeabilizing mitochondria in tumor cells holds promise as a novel therapy for lung cancer.

Plant-made Lectibody Targeting Tumor-associated High-Mannose-Glycan Antigens as a Novel Cancer Immunotherapeputic/Diagnostic Agent – Nobuyuki Matoba, Ph.D.

Recent advances in glycobiology have revealed that high-mannose-type glycans (HMG), which are immaturely processed, mannose-rich oligosaccharides attached to asparagine residues of glycoproteins, are elevated on the cell surface glycome of various tumors.  This finding indicates that tumor-associated HMG constitute a new therapeutic target.  However, currently there is no approved anti-cancer agent specifically targeting these glycans.  Our long-term goal is to develop novel therapeutic strategies targeting tumor-associated HMG.  A clinically proven therapeutic approach targeting tumor surface biomarkers is the development of monoclonal antibodies that are capable of inducing immune-mediated anti-tumor activities, such as antibody-dependent cellular cytotoxicity (ADCC).  However, development of HMG-specific monoclonal antibodies has been challenging.  To overcome this issue, we have developed a novel lectibody, Avaren-Fc (AvFc), which consists of the HMG-binding lectin Avaren fused to the fragment crystallizable (Fc) region of human immunoglobulin (Ig)G1.  Our preliminary data showed that AvFc binds with high affinities to a broad spectrum of cancer cell lines, but not to normal human cells.  Additionally, AvFc induced potent ADCC against many of the cancer cells tested.  Moreover, AvFc exhibited outstanding safety profiles in rats upon intravenous administration.  AvFc is efficiently bioproduced in Nicotiana plants, enabling us to obtain large quantities of AvFc quickly and more efficiently than any other conventional recombinant production systems.  Based on these results, we hypothesize that the lectibody AvFc could be developed as a first-in-class anti-cancer immunotherapeutic agent targeting tumor-associated HMG.  To prove this hypothesis we propose two Specific Aims in this application.  In Aim 1, we will investigate AvFc’s anti-tumor activity in murine tumor challenge models.  We will use syngeneic and human tumor xenograft models to determine the effectiveness of AvFc.  In Aim 2, we will evaluate AvFc’s tumor specificity based on immunohistochemistry of human tumor tissue sections and in vivo biodistribution analysis using microPET/CT imaging in mouse tumor models.  In summary, the proposed research should reveal the cancer immunotherapeutic potential of our novel tumor-associated HMG-targeting lectibody AvFc.  The data generated in this project will strongly justify a federal grant proposal for additional preclinical studies and support the design of pre-IND and IND-enabling studies towards a Phase I clinical trial.

Overcoming Melanoma Immunotherapeutic Resistance by MIF Targeting – Robert A. Mitchell, Ph.D. and Kavitha Yaddanapudi, Ph.D.

Myeloid-derived suppressor cells (MDSCs) are potently immunosuppressive innate immune cells that accumulate in advanced cancer patients and actively inhibit anti-tumor T lymphocyte responses.  Increased numbers of circulating MDSCs directly correlate with melanoma patient mortality and reduced anti-tumor immune responses.  Our preliminary results demonstrate that circulating monocytic MDSCs are found in significantly higher numbers in chemotherapy naïve late stage melanoma patients and the immunosuppressive activity of these cells is highly reliant upon the innate immunomodulatory cytokine, macrophage migration inhibitory factor (MIF).  Importantly, inhibition of MIF in melanoma patient MDSCs results in a reversion of immunosuppressive MDSC phenotypes towards an immunostimulatory, antigen-presenting, dendritic cell (DC)-like phenotype.  We hypothesize that the aberrant accumulation of MDSCs dictates resistance to the adaptive immune checkpoint inhibitors, Ipilimumab (IPI) and/or Nivolumab (NIVO) and that therapeutic targeting of MIF represents a highly novel and clinically viable approach to targeting MDSC-associated immunotherapeutic resistance in melanoma patients.  To test these hypotheses, we will determine if baseline MDSC percentages (intrinsic resistance) and/or change in MDSC percentages as a function of time (acquired resistance) correlate with patient responses to IPI and/or NIVO immunotherapies.  We will also stringently evaluate whether a highly active, orally bioavailable MIF small molecule inhibitor (4-iodo-6-phenylpyrimidine – 4-IPP) attenuates melanoma patient MDSC-dependent immunosuppressive activities and/or induces MDSC → DC-like functional differentiation.  Finally, we will test proof-of-concept by combining orally delivered 4-IPP with IPI and/or NIVO immunotherapies in syngeneic mouse models of BRAFwt and BRAFmt melanoma.  Results from these studies will provide evidence for the inclusion of peripheral blood MDSC frequencies as a clinically relevant prognostic biomarker for melanoma patients undergoing immunotherapy.  Perhaps more importantly, the combined human and mouse data will provide key pre-clinical rationale and justification for developing combination strategies using current immune checkpoint inhibitors and MIF targeting therapies in late stage melanoma patients.

Year 4 (July 1, 2016 – June 30, 2017)

Due to the degree of progress on the four grants listed and described under Year 3, they were renewed for a 2^nd year of funding.

See above:  Paula J. Bates, Ph.D.; Chi Li, Ph.D.; Nobuyuki Matoba, Ph.D.; and Robert A. Mitchell, Ph.D. and Kavitha Yaddanapudi, Ph.D..

Year 5 (July 1, 2017 – June 30, 2018)

To be determined