Institution Short Name: Johns Hopkins University

Effectiveness of cancer vaccine to elicit immunity against tumor cells has been fundamentally proved in experimental animals or in some clinical conditions. Currently, one of the most important subjects of cancer vaccine is how to maximize its potency. In this regard, recent studies have suggested that LIGHT, a molecule belonging to tumor necrosis factor superfamily, induces potent anti-tumor immunity by a unique mechanism that facilitates both migration and activation of lymphocytes at the site of tumor.  

In this study, we will delineate the molecular-based mechanism of this phenomenon and to develop further efficient cancer vaccine using LIGHT. To this end, we will first take advantage of mutant protein of LIGHT or neutralizing monoclonal antibodies against HVEM or LTbetaR, two functional receptors of LIGHT. These experiments will elucidate the relative importance of HVEM versus LTbetaR or membrane versus soluble LIGHT in association with T cell trafficking and activation as well as anti-tumor efficacy.  

In the following experiments, we will attempt to develop innovative and more potent strategies of LIGHT cancer vaccine. By molecular engineering techniques, we will express stimulatory anti-HVEM antibody on the surface of tumor cells as cancer vaccine or generate pentameric constructs of this antibody to facilitate the signal delivery. In addition, attenuated measles virus vector that is specifically targeted to tumor cells will be utilized for tumor-selective LIGHT expression in vivo.  

Successful completion of this project will strongly support the potency of LIGHT as a target for advanced cancer immunotherapy, and thus lay the foundation of translational studies on LIGHT-based cancer gene therapy aiming at clinical applications.  

The myelodysplastic syndromes (MDS) are a group of clonal neoplastic hematologic disorders characterized by varying degrees of bone marrow failure, abnormal hematopoiesis, and proliferation of myeloid blast cells. Impaired maturation of hematopoietic progenitors is manifest clinically by peripheral cytopenias and morphologic abnormalities in the marrow (“dysplasia”). Thought to be disorders of hematopoietic stem cells, clonal cytogenetic abnormalities are frequently identified. Although the disease can evolve toward acute leukemia, morbidity and mortality most frequently result from a marrow failure syndrome.  

Evidence exists that immune activation against hematopoietic elements frequently occurs in MDS patients, based on the identification of lymphocytic infiltrates in the marrow, oligoclonal expansion of T cells, and excessive production of tumor necrosis factor alpha. Whether this represents a secondary event in response to cell injury and the generation of neo-antigens, or an initiating event inducing immunopathology, remains controversial. Nevertheless, MDS are thought to be immunologically responsive diseases, as immunomodulatory drugs can induce remissions, and allogeneic bone marrow transplantation can be curative in the small fraction of patients for whom this is an option.  

Recently, three classes of therapeutic agents have been shown to have activity in MDS; a) DNA methyltransferase inhibitors, b) histone deacetylase inhibitors (HDACi), and c) immunomodulatory derivatives (IMiDs) of thalidomide. Central to this proposal is the observation that all three classes of drugs augment discrete elements of host immunity, making their integration with therapeutic cancer vaccines ripe for exploration.  

 
We have developed a genetically modified tumor cell vaccine for the treatment of myeloid malignancies. The human erythroleukemia cell line K562 has been stably transfected to secrete GM-CSF. K562 cells express many of the antigens shown to be overexpressed in myeloid leukemias and MDS. In early phase clinical trials for both acute and chronic myeloid leukemias, we have observed the induction of anti-tumor immunity and associated clinical responses following K562/GM-CSF vaccination (see preliminary data). In this proposal, we seek to evaluate the integration of K562/GM-CSF vaccination with systemic therapies for MDS that alter host immunity and/or hematopoietic cellular differentiation. Specifically, we will:  

 
1. Examine the in vivo effect of: a) DNA methyltransferase inhibitors, b) HDACi, and c) IMiDs on the response to GM-CSF tumor vaccines in a mouse model (year 1).  


2. Conduct a clinical trial in MDS testing K562/GM-CSF vaccination integrated with the systemic agent(s) identified in aim 1 as being most active in combination with GM-CSF tumor vaccines (years 2 and 3). 

 
3. Evaluate immune responses specific for autologous MDS cells as well for as candidate antigens overexpressed in MDS using pre and post vaccination blood and marrow samples (years  

Immunotherapy using gene-transduced tumor cells has emerged as a potentially plausible approach for the control of advanced stage ovarian cancer. We have recently demonstrated that vaccination with irradiated murine allogeneic ovarian cancer cells secreting heat shock protein 70 (Hsp70) were capable of generating potent tumor antigen-specific immune responses and strong anti-tumor effects against ovarian cancers (Chang et al. Cancer Research 2007; 67, 10047-10057).  

Hsp70 has been shown to bind antigenic peptides from tumors. The secretion of Hsp70 from the ovarian cancer cells will allow the Hsp70-associated antigenic peptide to be concentrated and targeted to dendritic cells (DCs) with subsequent DC activation. Based on the encouraging preclinical data, we propose to generate a clinical grade tumor cell-based vaccine engineered to secrete high levels of Hsp70. We have access to several high grade serous carcinoma cell lines, that express several known tumor antigens, such as CA-125 and mesothelin.  

The generation of the cGMP clinical lots of a tumor cell-based vaccine using high grade serous carcinoma cell lines that secrete high levels of Hsp70 represents an innovative approach that may create an opportunity for the therapy of these devastating ovarian cancers. This product will form the basis for the development of a pipeline of immunotherapeutic approaches at Johns Hopkins University. Thus, in the current proposal, we propose the following. 

  •  Specific Aim 1: To generate and characterize a high grade serous carcinoma cell line that stably secretes high levels of Hsp70. 
  •  Specific Aim 2: To cGMP manufacture and release Master Cell Banks and a clinical lot, per FDA CBER guidelines, of the Hsp70-secreting ovarian cancer cell-based vaccines.  
  • Specific Aim 3: To perform phase I clinical studies using clinical grade Hsp70-secreting ovarian cancer cell-based vaccines in patients with high grade serous carcinoma.  
  • Specific Aim 4: To characterize tumor antigen-specific CD8+ T cell immune responses in vaccinated individuals.  

Successful implementation of the current proposal may lead to the development of an innovative therapeutic vaccine against high grade ovarian serous carcinoma. We are currently testing two distinct types of cancer vaccines developed at Johns Hopkins, including a mesothelin-expressing Listeria-based vaccine, which is being developed by Anza Therapeutics and the GM-CSF secreting K562 cell line, which is being developed by Cell Genesys.  

The mesothelin-expressing Listeria-based vaccine is currently being tested in patients with pancreatic cancer and mesothelioma, while the GM-CSF secreting K562 cell line is being tested in chronic myeloid leukemia patients. Given the suitable safety of the proposed Hsp70-secreting ovarian cancer cell-based vaccine, we will also examine its application in combination with mesothelin- expressing Listeria and/or GM-CSF secreting K562 cell lines to further enhance the tumor specific immune responses.