Grant: 5462-18 | Career Development Program (CDP):
Location:Yale University, New Haven, Connecticut 06520-8327
Project Title: A Protein Degradation Approach For The Treatment Of Acute Myeloid LeukemiaProject Summary:
Many cancers result from a genetic mutation causing an “always on” protein. Current treatments are based on the deactivation of the proteins by blocking that protein’s active site. Herein I propose an alternative approach in which proteins are permanently degraded rather than temporarily deactivated, which may prove to be a more favourable form of therapy. To do this, I will take advantage of the cell’s own natural ability to degrade its own proteins when they are in excess or no longer needed. I will design and prepare compounds which recruit the native protein degradation machinery to the target proteins by creating a bridge between protein degradation components and the target protein. This approaches uses a two-headed molecule called a Proteolysis Targeting Chimera (PROTAC).
The potential advantages of protein degradation over protein inhibition are three fold:
1. Constant and complete deactivation of proteins is necessary for a treatment to be successful. Protein inhibition, as happens with standard targeted drugs, is often a reversible process, allowing previously inhibited proteins to again be functional. Protein degradation, as happens with PROTACs, is irreversible, therefore resulting in complete deactivation.
2. PROTACs have shown the potential to be catalytic, meaning one PROTAC molecule could destroy more than one protein molecule, preventing cells from simply producing more protein to overcome the deactivation of the existing population.
3. An issue arising from current treatments is the development of resistance after treatment for a relatively short period of time. The proposed PROTAC compounds may be able to circumnavigate such resistance mechanisms.
I propose to prepare PROTACs containing recognition elements for target proteins involved in blood cancers. Specifically, I am focusing on FLT3, which is a protein important in about 1/3 of all AMLs. I will assess PROTACs for the ability to degrade target proteins in cell-based models. The resulting compounds will then be optimised before progression into animal models. It is conceivable that by employing protein degradation, it may be possible to completely remove all disease causing protein. The ultimate goal is to produce a drug that may be useful for the treatment of AML containing FLT3 mutations. Importantly, PROTAC technology has applicability in a number of different cancers.
Grant: 1344-18 | Career Development Program (CDP):
Location:Fred Hutchinson Cancer Research Center, Seattle, Washington 98109-1024
Project Title: The Biological And Therapeutic Consequences Of SF3B1 Mutations In Myelodysplastic SyndromesProject Summary:
Myelodysplastic syndromes (MDS) are a group of blood disorders characterized by impaired differentiation of hematopoietic stem cells into functional blood cells. MDS frequently has a poor prognosis and is associated with a high risk of transformation into acute myeloid leukemia. There are few treatment options for MDS, largely because the underlying molecular changes that drove MDS were not known until recently.
Recent genome sequencing studies revealed that MDS and related diseases are associated with specific mutations (genetic changes) in hematopoietic stem cells. These mutations most commonly affect genes that control a molecular process termed "RNA splicing." RNA splicing is critical to the process by which genetic information in DNA is "read" to make proteins. We now know that MDS-associated mutations that affect RNA splicing cause mistakes during the transfer of genetic information from DNA to protein. However, we do not yet know precisely which mistakes ultimately give rise to MDS.
We plan to use both experimental and computational methods to determine how mutations that affect RNA splicing give rise to MDS. Understanding the specific molecular changes that occur in MDS cells carrying these mutations will enable us to identify potential new therapeutic opportunities for treating MDS. Because the same mutations affecting RNA splicing are found in other blood diseases as well, such as chronic lymphocytic leukemia, we hope that our discoveries will improve the treatment of many different blood diseases.
Grant: 5465-18 | Career Development Program (CDP):
Location:The Regents of the University of California, San Francisco, San Francisco, California 94143
Project Title: Inhibiting The Palmitoylation/Depalmitoylation Cycle As A Selective Therapeutic Strategy In NRAS Mutant Leukemia.Project Summary:
Acute myeloid leukemia (AML) is an aggressive blood cancer that affects children and adults. Recent advances for sequencing the DNA of leukemia cells have greatly advanced our understanding of the genetic causes of AML; however, this new knowledge has not yet resulted in better treatments.
One of the most common mutations found in AML alters a type of RAS gene called NRAS. The protein made by NRAS works like an “on” and “off” switch that instructs cells to grow in response to growth factors. RAS gene mutations found in AML and other cancers lock these switches in the “on” position, which drives abnormal growth. Recent studies of AML cells have shown that NRAS gene mutations are absent when patients are in remission and frequently reappear when the leukemia relapses. Therefore, NRAS mutations are likely very important for the growth of AML cells, and inhibiting abnormally active N-Ras proteins (proteins created by the NRAS gene) may be of great benefit for patients. Unfortunately, developing drugs that can directly turn abnormal N-Ras proteins “off” is extremely difficult.
We are testing a new approach for inhibiting mutant N-Ras by exploiting a potential “Achilles heel” in the protein. It is likely that N-Ras must be located at the cell surface to stimulate growth. This localization depends on two chemical modifications that are regulated by different enzymes: the addition of a lipid group (palmitoylation) and its subsequent removal (depalmitoylation). We think that inhibiting this cycle will kill AML cells with NRAS mutations but will not affect normal cells. We will test this using a mouse model in which we engineered a mutation of the NRAS gene so it cannot be palmitoylated. Next, we will investigate chemical inhibitors of the enzyme that depalmitoylate the N-Ras protein as a possible treatment for AMLs with NRAS mutations. Finally, we will try to define the enzymes responsible for N-Ras palmitoylation, with the long-term goal of blocking this reaction as an alternative to inhibiting depalmitoylation. Altogether, I anticipate that my project will advance our understanding of NRAS mutant AML and will identify novel strategies to treat this aggressive blood cancer.
Grant: 3380-18 | Career Development Program (CDP):
Location:Dana-Farber Cancer Institute, Boston, Massachusetts 02215
Project Title: Interrogating The Sf3b1 Mutated/Atm Deleted Mouse As A Novel Faithful Model Of Chronic Lymphocytic LeukemiaProject Summary:
Human genomic analyses have defined the complex genetic heterogeneity of chronic lymphocytic leukemia (CLL) as the most common indolent B-cell malignancy. These studies have revealed that selection of certain genetic alterations occurs throughout disease progression and correlates with therapy failure. Despite the remarkable efficacy of a number of recently introduced therapies, CLL remains incurable, and resistance to these novel drugs is challenging the clinical management of CLL patients.
Genetically engineered mouse models represent a promising approach to studying the functional impact of novel cancer-associated gene alterations and are useful to developing preclinical platforms for testing the efficacy of novel drug combinations. The main challenge with CLL modeling is the lack of animal models that faithfully recapitulate the genetic changes discovered in patients. Through novel genetic engineering strategies, we therefore seek to introduce mutations typical of human CLL in mice and to characterize disease features in these novel models, including, but not limited to, aberrancies in B cells (the cell of origin of this leukemia), and T lymphocytes (the cells which generally control immune responses but are notably dysfunctional in CLL patients, thus favoring disease progression).
We recently observed CLL development in animals bearing two of the most common gene alterations found in patients, that is mutations in the genes Sf3b1 and Atm, whose functionality is critical for CLL survival and responsiveness to therapy. We took advantage of this model to create a transplantable platform, whereby leukemias harvested from a donor animal can be expanded into recipients, which are then treated with different drugs (and/or their combinations). The first class of compounds that we will test is splicing modulators, which are drugs capable of interfering with alternative splicing – the main process regulated by Sf3b1 Alternative splicing is a core cellular process involved in the regulation of gene function. Preliminary studies have already shown efficacy of splicing modulators when tested alone or combined with FDA-approved agents for the treatment of CLL.
The overall goal of my studies is to establish robust preclinical platforms to test new therapies and to facilitate the optimization of treatment strategies tailored to the genetic makeup of individual CLL patients, with the aim of obtaining deeper clinical remissions and potentially allowing treatment discontinuation in these patients.
Grant: 6530-18 | Translational Research Program (TRP):
Location:The Children's Hospital of Philadelphia, Philadelphia, Pennsylvania 19178-1457
Project Title: CBL Regulation Of Ubiquitination And Cytokine Signaling In Myeloid MalignanciesProject Summary:
CBL mutations are found in diverse myeloid malignancies, MDS (myelodysplastic syndrome), myeloproliferative neoplasm (MPN), acute myeloid leukemia (AML), and particularly MDS/MPN overlap syndrome, a disease with both MDS and MPN features. MDS/MPN overlaps are often diagnosed as juvenile myelomonocytic leukemia (JMML) or chronic myelomonocytic leukemia (CMML) in which CBL mutations have the most occurrence, ~20%. There remains a critical need for the development of effective therapies in CMML as it is associated with a median survival of 34 months and has no approved disease modifying therapies.
CBL family proteins are E3 ubiquitin ligases that are important in modifying proteins in a way that target them for degradation. CBL keeps its substrates at a proper level so that cell proliferation is appropriately controlled. Almost all mutations in CMMLs disrupt CBL function, thus alleviating the break and balance provided by normal CBL proteins. Therefore, it is imperative to understand what the relevant CBL targets are to effectively treat myeloid leukemias with CBL mutations.
This application is based on our recent discovery that identified a new mechanism that mediates CBL function. We found that CBL family proteins determine JAK2 kinase levels in the cell with the help of the adaptor protein LNK (also called SH2B3). Both JAK2 and LNK play critical roles in controlling blood stem cell expansion and blood development. JAK2 mutations are found in a large population of MPN patients. Recently, LNK is also found mutated in JMML and CMML, attesting the importance and relevance of our work. CMMLs often progress into AML with short latency, and currently there is no curative treatment other than bone marrow transplantation. This underscores the importance of CBL proteins in regulating blood stem cells, and supports the idea that they work in concert with JAK2. If we could better understand how these new molecules function in normal, pre-cancerous and leukemic blood cells, we might learn about ways to control them to intervene in these diseases. This is the goal of the present application.
For our studies we propose to use animal models that afford genetic perturbations of the CBL genes in order to study their role in blood stem and progenitor cells. We plan to carry out comprehensive screens with cutting-edge technologies to identify new CBL substrate proteins that may be important for better targeting CMML and genes that potentially confer therapy resistance. We will also carry out experiments in primary human CMML cells as well as patient-derived xenografts (PDXs) to elucidate the mechanisms by which CBL molecules regulate JAK2 and new substrates to explore them as targets for pharmacologic intervention.
Upon completion of the proposed experiments we hope to have gained valuable new insights into the CBL pathway and provide a blueprint for the design of new treatment modalities for CMMLs and myeloid malignancies in general.
Grant: 6557-18 | Translational Research Program (TRP):
Location:University of Perugia. Division of Hematology and Clinical Immunology, Department of Clinical and Experimental Medicine, Perugia, 06132
Project Title: HAIRY CELL LEUKEMIA (HCL): A CHEMOTHERAPY-FREE TREATMENT STRATEGY CENTERED AROUND BRAF INHIBITIONProject Summary:
We discovered a mutation of the BRAF gene as the genetic cause of Hairy Cell Leukemia (ref. 1). We also found in the laboratory that drugs designed to block mutated BRAF kill patients' hairy cells, while sparing cells laking this mutation (ref. 2), including normal cells. Thus, inhibiting mutated BRAF could represent a new approach to the treatment of HCL, which is currently based on myelotoxic chemotherapeutics (purine analogs) killing not only leukemic cells but also normal bone marrow (i.e., myeloid) cells.
Indeed, we showed that a brief treatment by mouth with the BRAF inhibitor vemurafenib proved highly active in HCL patients relapsed after, or refractory to, chemotherapy, without the toxic effects of the latter (ref. 3). However, the disease tends to come back some time after stopping vemurafenib intake due to the persistence, at the end of treatment, of some residual hairy cells in the bone marrow. These residual hairy cells represent the reservoir for subsequent leukemia regrowth and, in about half of patients, they manage in some way to bypass BRAF and reactivate MEK, which is the target of BRAF and which should be deactivated when BRAF is inhibited by vemurafenib (ref. 3).
This project aims at improving on these results by rationally adding to vemurafenib other "intelligent", non-toxic drugs, in order to eradicate all leukemic cells and therefore achieve lont-term complete remssions. In particular, in a set of patients we will combine vemurafenib with cobimetinib, an inhibitor of MEK that is also taken by mouth, in order to impede MEK reactivation during therapy with vemurafenib. In another set of patients, we will combine vemurafenib with obinutuzumab, a monoclonal antibody given intravenously, that recognizes a molecular marker (CD20) on the surface of all hairy cells (i.e., irrespective of whether or not they managed to reactivate MEK); in so doing, obinutuzumab is expected to stimulate the immune system to attack leukemic cells from the outside, while vemurafenib is concurrently blocking mutated BRAF inside them (two-pronged strategy). Finally, if patients still relapse after either of these two drug combinations (vemurafenib plus cobimetinib; vemurafenib plus obinutuzumab), we will use all these three drugs together to try to get these patients in remission again.
We will accomplish this goal through a clinical trial that will be conducted in multiple centers throughout Italy, and that will be open to patients that have relapsed/refractory HCL or that are unfit for chemotherapy (e.g., due to old age, frail medical conditions, or ongoing infections which could be worsened by chemotherapy).
This project will help to clarify the most attractive combination of "intelligent" drugs to be used in relapsed/refractory HCL patients, and to be selected for potentially challenging purine analogs in newly diagnosed patient with this leukemia, in the prospect of a chemotherapy-free therapeutic strategy for all HCL patients.
Grant: 6536-18 | Translational Research Program (TRP):
Location:The University of North Carolina at Chapel Hill, Atlanta, Georgia 30384-2420
Project Title: Exploiting The Inducible Caspase9 To Pharmacologically Modulate CD19.CAR-T Cell Function In VivoProject Summary:
The administration of a subset of human immune cells cultured in the laboratory and known as T lymphocytes that have been engineered to express a chimeric molecule that recognizes leukemic cells (CD19.CAR-Ts) has shown remarkable antitumor effects in patients with a blood tumor known as acute lymphoblastic leukemia (ALL). We propose here to improve this therapy by reducing its potentially lethal side effects that are caused by the extreme expansion of these cells once they encounter tumor cells in patients. These side effects are known as 1) cytokine release syndrome, with symptoms varying from flu-like symptoms to more severe side effects such as cardiac arrest, renal failure and ultimately death; 2) seizure and coma that are often life threatening; 3) life-long inability to recover the normal cells of the blood that protect from infections. We have now developed and validated in the laboratory a strategy that allows a precise regulation of the expansion of the T cells in patients, by using an absolutely safe drug that only works on the T cells infused into the patients, thus significantly controlling the cytokine release syndrome. In addition, a slightly higher dose of the same drug can be used to completely eliminate the T cells infused when the patient has achieved a durable elimination of the tumor cells. We now propose to use this strategy to in a phase I clinical trial (LCCC1541-ATL) to determine whether receiving iC9-CAR-Ts cells is safe and tolerable in adult and pediatric subjects with relapsed/refractory ALL and to determine our capacity to control the expansion of the T cells once infused into the patients. We have written the clinical trial, which has received approvals from the various regulatory agencies and validated the production of cells in our specialized manufacturing facility.
Grant: 6531-18 | Translational Research Program (TRP):
Location:The Trustees of Columbia University in the City of New York, Columbia University Medical Center, New York, New York 10027
Project Title: New Therapies In Relapsed ALLProject Summary:
Acute lymphoblastic leukemia (ALL) is an aggressive hematological tumor resulting from the malignant transformation of early lymphoid progenitor cells. Despite intensive chemotherapy, 20% of pediatric and over 50% of adult ALL patients fail to achieve a complete remission or relapse after intensified chemotherapy. Importantly, relapsed ALL is frequently associated with chemotherapy resistance and, despite salvage therapy with intensified treatment, cure rates are still unsatisfactory low. Here we propose to explore the therapeutic role of inosine monophosphate dehydrogenase (IMPDH) inhibitors in the treatment of relapsed and chemotherapy resistant ALL. Specifically we will ask: what is the specific role and mechanisms that mediate the antileukemic effects of IMPDH inhibitors in relapsed leukemia?; What patients would benefit best from treatment with these drugs?; and finally, what drug combinations may enhance the antitumor activity of these therapeutics? These studies will be instrumental for the clinical development of IMPDH inhibitors in the treatment of high risk leukemia.
Grant: R0858-18 | Quest for CURES (QFC):
Location:University of Miami, Atlanta, Georgia 30384-5803
Project Title: The Aging Epigenome: Clues To The Pathogenesis Of MDSProject Summary:
Myelodysplastic syndromes (MDS) are diseases of the blood-producing cells in the bone marrow (BM) with a high risk for progression to an aggressive acute leukemia. While rare before the age of 50, its incidence increases significantly with every decade of age and thus it is likely that age-acquired changes in the BM may predispose to the development of MDS. However, the mechanism behind this increased incidence is not fully understood. We propose that as we age, cells in the bone marrow accumulate changes in the nuclear instructions that govern their behavior. These instructions are encoded not only on their genetic material (known as DNA), but also on a series of chemical modifications of the cell’s genetic material known as epigenetic modifications. These epigenetic modifications are what give cells the ability to “interpret” the information on the genetic code. Therefore, any abnormalities acquired at the epigenetic level can have serious consequences on a cell’s behavior. We hypothesize that cumulative changes in the epigenetic information of BM cells acquired during aging change the cells' behavior and susceptibility to other lesions, laying the foundation for the increased incidence of MDS. We will study the normal changes acquired during aging at both the genetic and epigenetic levels and compare them to the disease-associated patterns seen in MDS in order to identify those epigenetic changes that may predispose for the development of this disorder.
Grant: 3372-18 | Career Development Program (CDP):
Location:The Trustees of Columbia University in the City of New York, Columbia University Medical Center, New York, New York 10027
Project Title: The Role Of Diverse Cytokines Secreted By Myeloid-biased Multipotent Progenitors In Driving LeukemiaProject Summary:
Myelogenous leukemia is a type of blood cancer characterized by the abnormal production of white blood cells in the bone marrow. Abnormally produced white blood cells prevent the proper production of healthy blood cells and eventually lead to failure of the healthy blood system. There are several well-known disease-causing mutations, and many researchers are studying them to find out how the mutations cause disease and to develop treatments based on the targeting of those mutations. However, many cancers are characterized by the accumulation of several mutations, and targeting only one specific mutation is not the most efficient way to treat the disease. Therefore, my study aims to find a treatment that is applicable to a broad range of myelogenous leukemias and is not associated with an individual mutation. In a previous study, we identified a specific immature bone marrow cell population whose expansion is common throughout various myelogenous leukemia mouse models with variant disease-causing mutations. This indicates that expansion of this cell population may reflect the commonalities of the various myelogenous leukemia subgroups and suggests that this cell population is also a critical driver of disease progression in various subgroups. We also discovered ways to experimentally regulate the production of that specific population, which may provide potential therapeutic opportunities. Currently, I study the cellular characteristics of that cell population, with the long-term goal of understanding how expansion of these cells contributes to disease development. More specifically, I will focus on a protein secreted by that population, and I will investigate the function of the secreted protein in driving overproduction of abnormal white blood cells. To achieve my goals, I will use diverse experimental methods using cells isolated from mice as well as several mouse models. My study will provide insight into a mechanism common to the development of various forms of myelogenous leukemia and may contribute to the development broadly applicable therapeutic treatments.