Grant: 5464-18 | Career Development Program (CDP):
Location:Sanford Burnham Prebys Medical Discovery Institute, La Jolla, California 92037
Project Title: The Impact Of Immunoglobulin Isotype (IgM Vs IgG) On B Cell Lymphomagenesis And Progression.Project Summary:
B-cell lymphomas continuously multiply because growth regulation signaling is altered to stay ‘on’ all the time. This signaling is normally triggered by activation of proteins called receptors on the cell surface. Most lymphomas require signaling from a group of receptors called the B-cell receptor (BCR). Normal B-cells produce a type of BCR, called immunoglobulin (Ig), in a process called the germinal center (GC) reaction. There are several types of Ig subtypes – IgM, IgG, and IgA isotypes—which are produced during the GC in response to stimuli generated by agents foreign to the body. B-cells with different Ig isotypes behave and function differently. Diffuse large B-cell Lymphoma (DLBCL) frequently arises during the GC reaction and express either IgM, IgG or IgA types of BCR. However, the signaling potential and role of Ig isotypes in B-cell lymphoma is poorly understood.
DLBCL can be further sub-categorized into two subgroups: the Germinal Center B-cell type (GCB) DLBCL and Activated B-cell type (ABC) DLBCL. While ABC-DLBCL is a more aggressive disease with a poorer clinical outcome, the GCB-DLBCL is a less aggressive disease with better patient outcomes. Importantly, both subgroups of DLBCL depend on B-cell receptor (BCR) signaling for their survival and proliferation. Interestingly, GCB-DLBCL cells primarily express the IgG isotype of BCR, whereas the majority of ABC-DLBCL cells express IgM. Consistent with these findings, the two subgroups of DLBCL respond differently to BCR signaling inhibition by ibrutinib. In a recent study, there was only a 5% response rate among GCB-DLBCL patients treated with ibrutinib, whereas there was a 38% response rate among ABC-DLBCL patients. These findings, along with the expression of different Ig isotypes, are suggestive of the involvement of differential BCR signaling cascades in the pathogenesis of GCB- versus ABC-DLBCL. To address this idea, we analyzed changes in protein activation between IgM and IgG positive cells in normal B-cells from mice. Remarkably, we observed differential activation of common signaling components among the different Ig isotypes. We are currently seeking to elucidate the molecular mechanisms by which different Ig isotypes impact the survival and proliferation of lymphoma cells. We will also study the effect that these differences may have on regulating the sensitivity of lymphoma cells to approved therapies. Lastly, during these studies we hope to uncover novel therapeutic targets, which may eventually lead to better treatments for DLBCL.
Grant: 5463-18 | Career Development Program (CDP):
Location:La Jolla Institute for Allergy and Immunology, La Jolla, California 92037
Project Title: TET Proteins In B Cell Function And Malignancy.Project Summary:
DNA methylation is a fundamental biological process that controls the activation state of genes, which represent basic modules of biological systems. TET proteins were recently identified as enzymes that mediate the removal of DNA methylation and have been shown to play vital roles in normal development of organisms. Moreover, the activity of TET enzymes is often deregulated during the pathogenesis of various hematological and non-hematological cancers. Consistent with these observations, recent studies have identified loss of TET protein functions in B-cell-derived malignancies, which are the most common hematological malignancies among adults in Western countries. However, the precise role of TET proteins in the regulation of normal B-cell functions and the mechanisms by which loss of TET activity contributes to development of B-cell-derived malignancies remains unclear. Interestingly, my preliminary studies suggest that TET protein shave important roles in the normal functions of B-cells and that loss of TET expression and activity in B-cells is associated with increased instability of the genetic material, which may cause a predisposition to cancer. I am currently investigating the mechanisms by which TET proteins control the physiological functions of B-cells as important components of the immune system. I will further extend these findings to understand mechanisms by which loss of TET proteins contributes to the development of B-cell lymphoma. Because TET protein functions are frequently deregulated in various cancers, the insights gained from these studies may be important in deciphering the origins of not just B-cell malignancies but also other kinds of cancers associated with deregulation of TET protein functions. This knowledge can be further applied in future designing of specific therapies that may potentially be efficacious across a broad spectrum of hematological and non-hematological malignancies.
Grant: 3376-18 | Career Development Program (CDP):
Location:The University of Utah, Salt Lake City, Utah 84112-9003
Project Title: Determining The Role Of SIRT5 In Acute Myeloid LeukemiaProject Summary:
Acute myeloid leukemia (AML) is the most deadly blood cancer, with more than 70% of patients dying from the disease within five years after diagnosis. The treatment option shave remained largely unchanged for the past 30 years. Chemotherapy and stem cell transplant are still the standard therapy for AML. The fact that most patients with AML will eventually relapse and succumb to their disease defines an urgent, unmet medical need for more effective drugs to treat this disease. To answer this call, we have taken a novel approach to identify new drug targets. We have methodically evaluated 1200 cancer-related genes one by one in patient samples to determine which genes are vital to the survival of AML cells. We have found that in approximately 70% of patient samples and cell lines, removal of SIRT5, a gene that regulates energy metabolism, kills AML cells but not healthy cells, suggesting that SIRT5 may be a new drug target in AML. The overall goal of my research is to better understand the role of SIRT5 in AML. Using a larger cohort of primary AML cells donated from patients, I will determine how common SIRT5 dependence is in this larger group of patients. My preliminary data also suggests that the dependence of AML cells on SIRT5 is not simply a matter of increased expression of the gene. Therefore, I will perform experiments to determine what other genes are involved in SIRT5-mediated dependence of AML cells. I will also determine how SIRT5 promotes a pro-tumor energy metabolism in AML cells and how inhibition of this abnormal metabolism kills AML cells. The aims of my proposal are to answer the following questions: 1) How common is SIRT5 dependence in AML, and what is the genetic basis of this dependence? 2) How does inhibition of SIRT5 kill AML cells? My proposal has the potential to establish SIRT5 as a novel drug target in AML and lead to the development of a new class of drugs that target SIRT5 to improve patient outcomes in AML.
Grant: 6529-18 | Translational Research Program (TRP):
Location:University of Minnesota, Twin Cities, Minneapolis, Minnesota 55414
Project Title: Innate Lymphoid Cell Type 2 Infusion For Graft-versus-host Disease (GVHD) Prevention And TreatmentProject Summary:
Hematopoietic stem cell transplantation (HSCT) using donor cells transplanted into patients can potentially cure patients with hematological malignancies. However, an immune response of the donor cells against the patient, termed graft-versus-host disease (GVHD) can lead to death in the patient limiting the use of stem cell transplantation. Here, we have discovered that a unique population of cells, known as innate lymphoid type 2 (ILC2s) have a potent capacity to prevent and treat murine acute GVHD. ILC2s are located at mucosal surface sites of exposure to foreign antigens and pathogens, especially the gut and the lung, which are key GVHD target organs. ILC2s control adverse immunological events by releasing anti-inflammatory cytokines and tissue reparative factors, as well as activate a network of immune regulatory cells in the recipient. We have found that conditioning regimens used to prepare patients for HSCT eliminates these cells before transplant and that such elimination predisposes the recipient to GVHD. Indeed, in HSCT patients, the slow tempo of recovery of ILC2s correlates with an increased likelihood of conditioning regimen toxicity and GVHD. In a mouse GVHD model, we have shown that interleukin-33, which increases ILC2s, when given prior to conditioning can markedly augment ILC2s and reduce GVHD. Further ILC2s given at the time of HSCT migrate to the gut, a primal GVHD target organ, and reduce GVHD. In mice with active acute GVHD, ILC2s given in 2 doses were remarkably effective in ameliorating lethality. Moreover, ILC2 infusion did not impair the graft-versus-tumor response . Thus the net result was control of GVHD without adverse effects on anti-leukemia responses. Using umbilical cord blood or cytokine mobilized peripheral blood, stem cell sources that have been used to replace the need for bone marrow harvests, we have developed a clinical relevant process to isolate and expand human ILC2s with validated ILC2 cell surface and functional characteristics. The present proposal will optimize the production of ILC2s from these 2 sources, ensure the process is FDA compatible, choose the preferred ILC2s source, and test the preferred product to suppress human peripheral blood cell mediated GVHD in vivo with mice that have been reconstituted with a human immune system. The project will then move to an FDA approved facility at one of the 2 sites (University of Minnesota for cord blood or University of North Carolina-Chapel Hill for peripheral blood) to perform the large scale validation runs required for FDA approval of a clinical trial. The competitive renewal will proceed with a combined two institution clinical trial of ILC2 infusion to treat patients with active acute GVHD that have failed to respond to standard of care as these patients have a very poor prognosis.
Grant: 6549-18 | Translational Research Program (TRP):
Location:The University of Iowa, Iowa City, Iowa 52242
Project Title: Clinical Investigation Of CD38+/ Light Chain+/CD24+ As Putative Multiple Myeloma Stem Cell MarkersProject Summary:
Treatment failure in cancers, including, multiple myeloma (MM), is due to persistence of a minor population of cancer stem cells (CSCs), which are mainly non-cycling and very drug-resistant tumor cells. One major clinical observation supporting the existence of MMSCs is that we have shown that gene expression profiles (GEP) remain abnormal in many MM patients with long-lasting complete remission (CR) (> 10 years), suggesting the persistence of a cancer cell population with very low proliferative capacity and very limited sensitivity to our most intensive therapies. Different groups have reported on the presumed identity of MMSCs; however, a unique MMSC phenotype has not yet been established. To further identify phenotypic markers of MMSC, gene expression profiles were analyzed using primary MM cells of both MM stem cells and the bulk MM cells. The cell surface protein CD24 was found to be significantly upregulated in the MMSCs compared to the bulk MM cells. We have confirmed that CD24+ MM cells showed stem cell features, such as increased clonogenic potential, drug resistance and tumor formation capacity with as little as 10 cells. Importantly, we also confirmed that CD24+ MM cells exist in clinical MM samples, which are CD38+/CD24+/k+ or λ+ . Therefore, we have designed two Specific Aims to accomplish this project: (1) We will characterize the role of CD24 in maintaining stem cell features and investigate its clinical relevance in MM; and (2) We will determine whether CD38+/CD24+/k+ or λ+ primary MM cells have cancer stem cell features. Improved understanding of MMSCs biology will likely lead to the development of novel therapeutic targets that can be tested in the laboratory and in the clinic. Evidence from clinical trials and correlative studies should provide proof that that inhibition of MMSCs leads to improvement in long-term clinical outcomes. Such critical studies can only be conducted if we are able to determine and analyze if CD38+/CD24+/k+ or λ+ are indeed consistent MMSC markers. Eventually, we hope this work can define MMSC markers in clinical samples and to use this knowledge to develop a novel therapy to prevent MM relapse.
Grant: 6542-18 | Translational Research Program (TRP):
Location:Oregon Health & Science University, Portland, Oregon 97239
Project Title: Novel Approach To Thwart MYC In B-cell Neoplasia By Selective Targeting Cyclin-dependent Kinase 9Project Summary:
Diffuse large B-cell lymphoma (DLBCL), a disease of the lymph nodes, is the most common subtype of non-Hodgkin lymphoma accounting for >10,000 deaths in the United States annually. While “targeted therapy” has made significant progress in treatment of blood cancers, we continue to treat DLBCL with standard chemotherapy regimens, which are associated with significant side effects and high rate of failure. When DLBCL recurs after initial therapy, it often becomes incurable with chemotherapy. The novel class of agents called inhibitors of cyclin-dependent kinases (or “CDK inhibitors”) has shown promise in therapy of cancer. Cyclin-dependent kinases are proteins which exist in cells under normal conditions. A whole range of them exist and ensure that cells can make all the necessary components for their survival, as well as reproduce. Cancer cells co-opt CDKs in their unlimited growth, and thus CDKs are attractive targets in cancer therapy. However, drugs which inhibit multiple CDKs are toxic, probably because they inhibit he function of multiple CDKs in normal (non-cancerous) cells. By contrast, emergent selective CDKs which target only one-two specific proteins hold promise to be more efficacious and have fewer adverse events. Here we propose to study how selective CDK inhibitors work in cancer, specifically focusing on inhibition of CDK9. Our preliminary experiments suggest that selective CDK9 inhibition stems lymphoma growth by disrupting the function of another protein, called MYC. MYC regulates synthesis of many cellular constituents which ultimately ensure tumor survival and growth. MYC levels are high in DLBCL and it contributes to therapy resistance. Here we will study the effect of CDK inhibitors on MYC function. We will determine how MYC function is affected, and whether MYC disruption is the dominant mechanism of how cells die in response to CDK inhibitors. We will also search for drug partners which will make CDK inhibitors more toxic specifically to tumor cells. Together, this study will help develop novel targeted therapies with the goal of eradicating DLBCL.
Grant: 6550-18 | Translational Research Program (TRP):
Location:Dana-Farber Cancer Institute, Boston, Massachusetts 02215
Project Title: Prioritizing The In Vivo Therapeutic Relevance Of "myeloma-selective" Essential GenesProject Summary:
Multiple myeloma (MM) remains incurable because its malignant cells manage to eventually escape from even the most potent combinations of new therapies for this disease. Despite years of extensive research, the MM field has not yet comprehensively characterized which genes are essential for MM cells to survive and grow within the supportive microenvironmental niche of the bone marrow (BM). We applied, in our lab, the exciting recent technology of CRISPR/Cas9 genome editing to simultaneously investigate the function of thousands of genes. We identified in our studies a select set of genes that are considerably more important for survival and growth of MM cells in the laboratory, compared to tumor cells from other blood cancers or from solid tumors. Some of these "MM-selective" essential genes have known roles in the biology of MM pathophysiology, but several others have not been previously considered major therapeutic targets. This project will validate the therapeutic relevance of these "MM-selective" essential genes using a model in which biocompatible scaffolds are implanted into mice and are engineered, with the use of human stromal cells from the BM, to resemble the properties and local conditions of the BM in patients with MM. Building on our experience with the CRISPR/Cas9 system both in the lab and in mouse studies, we will examine in this "humanized" BM-like scaffold model which of our candidate genes remain essential for MM cells in this context and therefore represent promising therapeutic targets. Investigational drug-like small molecules are available for only a few of these "MM-selective" essential genes, while most of the latter are deemed currently "undruggable". We observed though that specific small molecule inhibitors against other targets involved in distinct components of the regulation of gene expression can significantly down-regulate the transcript levels of several of the most prominent "currently undruggable" MM-selective essential genes in MM cells, and indirectly can disrupt the molecular network of these genes. We will therefore examine if small-molecule inhibitors targeting MM-essential genes, directly or indirectly, exhibit activity against MM cells in the "humanized" BM-like scaffold-based model. By combining the powerful CRISPR technology with in vivo models simulating the human BM, and pharmacological agents with direct or indirect activity against individual or multiple candidate MM-selective essential genes, we hope to determine the therapeutic relevance of these gene and provide a framework to guide the translation of inhibitors for these targets towards the clinic for MM patients.
Grant: 1349-18 | Career Development Program (CDP):
Location:Washington University School of Medicine in St. Louis, St. Louis, Missouri 63112-1408
Project Title: Protection Of Proliferating B Lymphocytes From Transformation By A C-MYC-induced Tumor Suppressive ProgramProject Summary:
Lymphomas and leukemias are caused by uncontrolled proliferation of lymphocytes due to accumulating errors in the genome. However, cell proliferation is also an important biological activity across many different tissues and cell types. Specifically, proliferation of lymphocytes is essential for the immune responses that protect individuals from invading pathogens. Normal lymphocytes are able to proliferate even quicker than cancer cells in response to infection for extended periods of time. In the course of normal immune system development, lymphocytes also mutate their own genome to establish better immunity to infection. However, it is unknown how lymphocytes undergo normal proliferation and mutation events without increasing the risk of cancers. We hypothesize that lymphocytes engage an unidentified, unique mechanism to minimize the risk of cancers while facing the demand to proliferate and mutate their genome during their normal functional activities. To explore this hypothesis, we have studied sets of genes that are activated by a protein named c-Myc, which has been causally linked to many cancers in humans and other organisms. Since c-Myc is also essential for lymphocyte proliferation in response to infection, c-Myc may activate an unidentified pathway that is required to protect normally proliferating lymphocytes from becoming cancerous. Accordingly, mutations of genes in the pathway may increase the risk of lymphomas or leukemias. Indeed, we have identified a gene that is activated by c-Myc in proliferating lymphocytes and is necessary to suppress blood cancer development in an animal model.Moreover, a few mutations of this gene have been found in leukemic cells from human patients.Starting with these discoveries, we will define the molecular mechanisms by which this c-Myc initiated regulatory pathway suppresses leukemia and lymphoma. The knowledge from this study will contribute to the development of improved strategies for the risk assessment and diagnosis of leukemias and lymphomas and also to inventing new therapeutic approaches.
Grant: 3378-18 | Career Development Program (CDP):
Location:Sloan Kettering Institute for Cancer Research, New York, New York 10087
Project Title: Optimizing CAR T Cell Therapy For Multiple MyelomaProject Summary:
While advances have recently been made in the management of multiple myeloma (MM), MM is still considered incurable, with most patients dying from their disease. Thus, there is a critical need to identify and evaluate new therapeutic modalities that may induce durable remissions or cures.
T-cells keep the body healthy by killing cells infected with bacteria or viruses. Chimeric antigen receptor (CAR) T-cell therapy involves collecting a patient’s T-cells and genetically reprogramming them in the lab to recognize and kill cancer cells instead of infected cells. The cells are then infused back to the same patient where they serve as a living drug. In relapsed or refractory acute lymphoblastic leukemia (ALL) this strategy led to dramatic complete responses in approximately 80% of patients treated.
To extend the effectiveness of this therapy to MM, we and others developed MM-specific CARs. Specifically, we screened tens of billions of human antibody fragments and identified a handful that bound to a specific protein target unique to MM cells. We then used the DNA encoding these ‘hits’ to make CARs. Subsequently, we compared how T-cells modified to contain these different CARs preformed in a variety of tests in cell cultures and in mice. We found one CAR, which conveyed very favorable properties on T-cells, to test in patients. We recently began treating MM patients on the clinical trial to test this novel CAR T-cell therapy.
Based on our experience with CAR T-cell therapy for ALL, we hypothesize that patients may develop resistance to our therapeutic approach. Potential sources of resistance may include loss of our target protein on the MM cells and/or suppression by other cells in the environment in which the MM cells reside. We will evaluate the effectiveness, including resistance mechanisms, of our CAR T-cell therapy by analyzing samples obtained through clinical trial. We are also developing “armored CARs,” which express a second protein that may alter the environment in a way that enhances anti-tumor immunity. Additionally, we will study patient samples in a unique mouse model that can support both MM tumor cells and cells found in the environment in which MM cells normally reside in patients. This model will be used to investigate whether our next generation CAR vectors, including CAR T-cells targeting MM-specific proteins (our armored CAR) may enhance long term anti-MM immunity.
Grant: 5469-18 | Career Development Program (CDP):
Location:Joan & Sanford I. Weill Medical College of Cornell University, New York, New York 10022
Project Title: Mutations Disrupting Gene Enhancer Epigenetic Complexes As Drivers Of Lymphomagenesis.Project Summary:
Lymphoma is a form of cancer that affects immune cells called lymphocytes, a type of white blood cell. There are many subtypes and maturation stages of lymphocytes and, therefore, many kinds of lymphomas. Follicular lymphomas (FL) develop from B lymphocytes (B-cells) and are the second most common subtype of non-Hodgkin lymphoma. Clinically, FL is an indolent lymphoma, characterized by slow progression and relatively high overall survival rate. However, FL is generally considered incurable and roughly half of all FL patients eventually progress into a more aggressive lymphoma with very poor prognoses.
During the normal immune response, B-cells undergo a stage where they grow much faster than usual and tolerate changes in their genetic material, allowing them to develop into fine-tuned functional immune effectors. Three proteins, called BCL6, SMRT and NCoR, work together to drive this process of terminal differentiation in a tightly controlled manner. Any alterations, such as those occurring in FL, can cause cells to become unresponsive to signals cueing exit from this program. B-cells thus become “frozen” at this stage of development; they are unable to complete their normal program and fail to die when instructed to, collecting in the lymph nodes and forming tumors.
Recently, through analyzing the genomes of FL patients, our laboratory has identified several mutations in a gene called TBL1XR1. Our preliminary data suggest that mutations in TBL1XR1 may result in increased stability of SMRT and NCoR. In this study, we plan to investigate whether the increased stability of SMRT and NCoR proteins by TBL1XR1 mutants can cause enhanced BCL6 activity and in turn lead to the development of FL. Unveiling the mechanisms by which B-cells cease to respond to their normal regulatory signals will profoundly contribute to our understanding of FL. In addition, our study will explore whether TBL1XR1 mutations can predict which FL patients have a BCL6-dependent disease and thereby help identify patients who could benefit from existing drugs targeting this protein.