Lymphoma SPORE Research Projects
The University of Iowa/Mayo Clinic Lymphoma SPORE Grant has four translational research projects designed to advance the science of lymphoid malignancies:
Project 1: The role of monocytes in non-Hodgkin lymphoma
- Co-leaders: Stephen M. Ansell, M.D., Ph.D., Clive S. Zent, M.D.
- Co-investigators: Allan B. Dietz, Ph.D.; Brian K. Link, M.D., University of Iowa
The tumor microenvironment plays an important role in non-Hodgkin lymphoma, and the role that intratumoral immune cells play in the pathology of lymphoma has been significantly understated. Intratumoral monocytes and macrophages are particularly important. Project data demonstrate that intratumoral monocytes in non-Hodgkin lymphoma are highly immunosuppressive and support malignant cell growth.
In preliminary work, the project team found that suppressive monocytic cells (SMCs) are abundant within the peripheral blood and tumor microenvironment in lymphoma patients and promote the survival of lymphoma cells. SMCs protect lymphoma cells from chemotherapy-induced cell death and promote lymphoma cell engraftment into severe combined immunodeficient (SCID) mice.
The team also found that SMCs within lymph nodes express immunosuppressive ligands, including B7-H1 (PD-L1, CD273); inhibit normal T-cell proliferation; and promote the induction of FoxP3+ regulatory T cells.
These preliminary studies suggest that suppressive monocytic cells have an effect on both malignant non-Hodgkin lymphoma cells and nonmalignant intratumoral T cells.
Based on these results, this lymphoma research project hypothesizes that suppressive monocytic cells are the intersection between the immune system and the malignant cell in non-Hodgkin lymphoma.
The project team plans to determine whether monocytes are specifically recruited to sites of lymphoma and which specific chemokines could be inhibited to prevent SMC migration; how lymphoma cells induce SMCs to support their malignant cell growth and to suppress the host's anti-tumor immunity; and whether promoting monocyte/macrophage maturation or inhibiting their interaction with other cells, particularly in the presence of monoclonal antibodies, improves their anti-tumor function.
This project has three aims:
- Aim 1. Define the mechanisms that result in the recruitment of monocytes to sites of lymphoma. The project is studying whether intratumoral monocytes/macrophages migrate in response to chemotactic ligands and establish a role for malignant cells in their recruitment. The project is also investigating whether monocytes undergo phenotypic and functional changes when recruited to lymphoma sites.
- Aim 2. Determine the mechanisms by which monocytes promote malignant B-cell survival and suppress intratumoral immune effector cells. The project is identifying the mechanisms responsible for the support of malignant cell growth and viability and the mechanisms responsible for suppression of effector T-cell function. The project is also investigating whether intratumoral monocytes remain immunosuppressive in the presence of monoclonal antibodies targeting lymphoma cells.
- Aim 3. Block suppressive monocyte formation to improve immunity and impact clinical outcome. The project is researching whether maturation of intratumoral monocytes suppresses their ability to support lymphoma cell growth and survival, reverses their immunosuppressive properties, and increases antibody-dependent cell-mediated cytotoxicity.
The research team expects this project to lead to a greater understanding of the role of monocytes and their progeny in non-Hodgkin lymphoma. This project's findings are likely to lead to an effective monocyte-directed therapeutic approach for patients with lymphoma.
Project 2: In situ immunization using nanoparticles
- Co-leaders: George J. Weiner, M.D., Brian K. Link, M.D., and Aliasger K. Salem, Ph.D., all with the University of Iowa
- Co-investigator: Alicia Olivier, D.V.M., Ph.D., University of Iowa
Immunotherapy of lymphoma through in situ manipulation of an involved lymph node can allow for the development of an active anti-lymphoma immune response without the need for ex vivo handling or immunization with a specific antigen.
A comprehensive and successful approach to in situ immunization in lymphoma would require the effective presentation of antigen by the lymphoma cells or professional antigen-presenting cells, the activation of lymphoma-specific T cells, and the suppression of the regulatory arm of the immune response to enhance the development of a sustained anti-lymphoma T-cell response.
This project's overall hypothesis is that in situ immunization with nanoparticles will allow for the induction and maintenance of a robust anti-lymphoma immune response with acceptable toxicity.
To test this hypothesis, this project is assessing the effect of intratumoral injection of nanoparticles (NPs) containing doxorubicin on lymphoma cells, the immune microenvironment, and the anti-lymphoma immune response in animal models and in a phase I clinical trial in patients with lymphoma.
The project is also assessing the effect of intratumoral injection of nanoparticles containing both doxorubicin and the toll-like receptor 9 (TLR9) agonist CpG ODN (CpG dox NPs) in mice and people.
Finally, the project is assessing how agents capable of maintaining the T-cell response impact the success of in situ immunization with nanoparticles.
This project has three aims:
- Aim 1. Assess the effect of in situ injection of nanoparticles containing doxorubicin on lymphoma cel
- Aim 1a. Refine the structure of doxorubicin nanoparticles to optimize the concentration and release of doxorubicin
- Aim 1b. Assess the toxicity, anti-tumor activity and immune effects of doxorubicin nanoparticles in mouse models of lymphoma
- Aim 1c. Conduct a phase I clinical trial of doxorubicin nanoparticles in patients with lymphoma and evaluate for toxicity the local effects of the injected doxorubicin nanoparticles on the treated node and the development of an anti-lymphoma immune response
- Aim 2. Assess the effect of in situ injection of nanoparticles containing both doxorubicin and the TLR9 agonist CpG ODN (CpG dox NPs) on lymphoma cells, the immune microenvironment and the anti-lymphoma immune response
- Aim 2a. Produce nanoparticles containing doxorubicin and various concentrations of CpG ODN
- Aim 2b. Assess the toxicity, anti-tumor activity and immune effects of CpG dox NPs in mouse models of lymphoma
- Aim 2c. Conduct phase I and II clinical trials of CpG dox NPs in patients with lymphoma based on results of studies with the doxorubicin nanoparticles
- Aim 3. Assess how agents capable of maintaining the T-cell response impact in situ immunization
- Aim 3a. Identify and incorporate the most effective agents that maintain T-cell activation, block T-cell checkpoints or block immunosuppressive cells into in situ immunization strategies
- Aim 3b. Assess whether a systemic or a local blockade of T-cell checkpoints or suppression of T-regulatory cells enhances the efficacy of in situ immunization
In situ immunization holds promise as a way to induce a long-term, anti-lymphoma immune response that results in clinical benefit for patients. This project is designed to explore a novel approach to in situ immunization that, if successful, could represent a novel approach to therapy for lymphoma and other cancers.
Project 3: Targeting JAK2 kinase in lymphoma
- Co-leaders: Mamta Gupta, Ph.D., Thomas E. Witzig, M.D.
This project's overall goal is to identify the molecular mechanisms underlying activation of the JAK/STAT pathway in lymphoma and to learn if inhibitors of this pathway can produce clinical benefit. The project team has identified several novel missense mutations in JAK2 and STAT3 genes.
The project's preliminary data demonstrate aberrantly activated JAK2 and STAT3 in more than 50 percent of diffuse large B-cell lymphoma patient samples. In vitro inhibition of JAK2 with the novel JAK2 inhibitor TG101348 (TG) inhibited JAK2 and STAT3 phosphorylation and induced apoptosis in a variety of lymphoma cell lines and patient samples.
This project has four aims:
- Aim 1. To characterize the biological significance of novel mutations in the JAK/STAT pathway in lymphoma. The project team is assessing the function of the novel L393V and M206K JAK2/STAT3 missense mutations. Suppressors of cytokine signaling (SOCS1) and protein tyrosine phosphatases (SHP1) are known key negative regulators of the JAK/STAT pathway. The project's preliminary data demonstrate silencing of SHP1 and SOCS1 genes in 33 percent and 86 percent of diffuse large B-cell lymphoma samples.
- Aim 2. To delineate the mechanisms of loss of negative regulation of the JAK/STAT pathway in lymphoma. The project team is delineating the mechanisms of silencing and how this regulates JAK/STAT pathway activation. The JAK/STAT signaling pathway is utilized by a number of growth factors and cytokines. The project has identified increases in several JAK/STAT pathway-specific cytokines (IL-2, IL-6, IL-10 and EGF) in serum samples from patients with diffuse large B-cell lymphoma compared to patients without lymphoma. In vitro, the team found that JAK2 and STAT3 are rapidly activated in response to IL-10 in lymphoma cells.
- Aim 3. Identify the role of JAK2 specific cytokine-mediated signaling in lymphoma. The project is investigating the role of signaling mediated through their receptors for these interleukins with a focus on IL-10.
- Aim 4. Determine the clinical activity of JAK inhibitors in a phase II trial. These agents are in clinical trials at Mayo Clinic for myeloproliferative neoplasms and have been found to be safe with clinical activity. The project team is using a unique trial design to test the hypothesis that preselection will improve the overall response rate. Specifically, the team is stratifying patients based on tumor cell STAT3 activation status: Group A (tumors with activation of JAK/STAT) and Group B (those without activation). Translational studies are using patient serum and tumor samples to investigate the relationship of tumor STAT3 activation, serum cytokines, and other biomarkers with tumor response and clinical outcome.
These studies are designed to understand the mechanisms of JAK/STAT pathway activation and to study a new JAK2 kinase inhibitor in a clinical trial for relapsed lymphoma.
Project 4: Genetic epidemiology and function of germline and somatic variants in diffuse large B-cell lymphoma
Diffuse large B-cell lymphoma (DLBCL) is the most common non-Hodgkin lymphoma subtype. It is well-established that there are driver somatic mutations in DLBCL etiology and prognosis, as well as a role for germline genetic susceptibility.
Early results from candidate gene association studies have shown a promising role for common genetic variants in immune and apoptotic genes in non-Hodgkin lymphoma. Re-sequencing of these genes in the tumors of 40 patients with diffuse large B-cell lymphoma identified novel genomic alterations. This project is characterizing the etiologic and therapeutic significance of these novel mutations and identifying additional risk variants.
The project team is leveraging the Lymphoma SPORE's involvement in the large International Lymphoma Epidemiology (InterLymph) Consortium genome-wide association study (GWAS) of DLBCL and the SPORE's whole-exome next-generation sequencing study of paired tumor and germline DLBCL cases.
This project has three aims:
- Aim 1. Characterize the therapeutic significance of novel mutations in genes that regulate the intrinsic apoptosis pathway. This project hypothesizes that mutations in genes identified through re-sequencing work (CASP9, BCL2L11 and APAF1) will have functional impacts on the apoptosis pathway. The project team is assessing the functional significance of four novel mutations in CASP9, BCL2L11 and APAF1 by determining if there is an inherent defect in the intrinsic apoptosis pathway and apoptosome formation in cells that express mutant forms of these proteins.
- Aim 2. Identify novel germline low-frequency variants associated with DLBCL risk. The project hypothesizes that germline variants of low frequency will be associated with a risk of developing diffuse large B-cell lymphoma. In stage 1, the project team is using whole-exome sequencing data from the 1000 Genomes Project to impute genotypes and then test for association. The team is conducting a technical validation on the top 1,536 significant imputed genotypes using the samples from stage 1. In stage 2, the team is conducting an in silico validation of all technically validated SNPs from stage 1 using InterLymph GWAS data. In stage 3, the team is re-sequencing the top 20 gene regions from stage 2 and testing for association. In secondary analyses, the team is testing whether these validated variants impact prognosis (event-free and overall survival).
- Aim 3. Identify somatically acquired driver mutations in genes critical to diffuse large B-cell lymphoma pathogenesis. The project hypothesizes that somatically acquired mutations will be associated with a risk of developing DLBCL. Using exome data from 77 paired tumor-normal DLBCL samples, the project team is identifying novel and likely functional driver mutations by filtering against the paired germline DNA and bioinformatics analysis. The top 20 genes are being sequenced to validate regions of interest in 500 DLBCL patients with paraffin tumor tissue in the SPORE. In secondary analyses, the project team is testing whether these mutations impact prognosis.
As the first comprehensive study of both germline and somatic genetic variants in diffuse large B-cell lymphoma, this project is likely to provide new insights into lymphomagenesis. These findings may help with risk assessment, prognostic stratification and identification of new treatment targets.