Current projects extend prior studies to answer several questions.
1. Cancer Personalized Profiling by Deep Sequencing (CAPP-Seq):
As part of a close collaboration with Max Diehn's lab at Stanford, we have developed a general method for ultrasensitive detection of circulating tumor DNA that is broadly applicable to human cancers. Projects involve specific application of CAPP-Seq to individual tumor types and for specific clinical questions.
2. A Genome-wide Landscape for Molecular Profiles and Clinical Outcomes Across Human Cancer:
Molecular and cellular profiles of individual tumors have identified several determinants of therapeutic responses and outcomes that distinguish tumor types, and also some that are shared between cancers. While several such features are known and have informed therapeutic targeting, there are likely many other as yet uncharacterized novel biological programs associated with the clinical behavior of diverse tumors. To address this need, we constructed a publicly available resource for Prediction of Cancer Outcomes from Gene expression profiles (PRECOG). This resource comprises a compendium of genome-wide expression profiles and clinical outcomes for 18k patients diagnosed with 39 cancer subtypes. These tools and resources will allow us to (1) characterize the genome-wide landscape of prognostic genes across cancers, including (2) shared and distinguishing features of individual tumor types, and define (3) specific sets of genes, (4) processes, and (5) cell types with influences on cancer-related survival.
In a related project, to resolve tumor-associated leukocyte subsets within heterogeneous specimens, we developed a novel method (CIBERSORT) to distinguish immune populations in corresponding bulk tumor transcriptomes. Unlike methods that require living cells as input (e.g., flow cytometry), CIBERSORT allows us to identify and quantify distinct types of immune cells from their RNA signatures in bulk tumor gene expression profiles, allowing both fresh and archival tumor specimens to be analyzed with relative ease and high-throughput.
3. Genetic Determinants of Immunological & Clinical Response in Follicular NHL:
Several ongoing projects involve work on indolent follicular lymphomas, which remain incurable. Here, immunological maneuvers such as vaccination of patients with unique patient-specific proteins derived from the immunoglobulin gene of each tumor (idiotypic vaccines) seems to only benefit a subset of patients who are able to mount such responses. We are working in trying to identify this subset of patients, using molecular predictors for vaccine outcomes that consider the immunoglobulin sequences used for the construction of the vaccine, gene expression profiles of the tumors, and genotypes of the host. Here, mechanistic studies are focused on a novel molecular feature we discovered within tumor immunoglobulin sequences as predicting humoral immune responses to idiotypic vaccines. These studies aim to glean an understanding of the biochemical and immunological basis for our finding in the laboratory using mice, and to validate it in other patients. Ultimately, the goal will be to use this knowledge to resume human trials examining the role of this immunological vaccination approach for an increasingly pervasive human disease.
The analytical framework we have developed consists of tools and resources for the discovery of molecular and cellular biomarkers of clinical outcomes, whether in specific cancers or across diverse tumors. Our approach allows molecular and cellular signatures to be compared across distinct malignancies, assessed for the predictive power they add to known risk factors, and used for construction of prognostic models integrating diverse genes or cell types. Since expression signatures of small numbers of genes and cells can have utility for predicting response to therapy, including anti-tumor agents, we are actively applying these methods toward the goal of discovering novel predictive and prognostic biomarkers, including immunotherapies.
4. Genetic basis for molecular variation in DLBCL:
We are interested in discovering the molecular underpinnings for the diversity among aggressive lymphomas, including expression profiles characterizing unique subsets. Specifically, we are examining lymphomas to understand the genetic basis for variation in expression profiles specifying cell-of-origin, and affecting the expression of genes such as LMO2 as a marker of germinal center B-cell derivation, as well as CD47 and CD137 as therapeutic targets for monoclonal antibodies that elicit novel immune effector mechanisms. Here, we are assaying whether these phenotypes reflect heritable or acquired genetic events. To answer these questions, we are applying next-generation sequencing technologies to examine the sequence of the exonic regions of the genomes of these tumors, and the corresponding transcriptome.
5. Modeling the Molecular Determinants of Induced Anti-Tumor Immune Responses in Mantle Cell Lymphoma:
In collaboration with the Davis, Levy and Elias Laboratories, we are currently conducting a clinical trial of therapeutic vaccination for patients with Mantle Cell Lymphoma (NCT00490529), a heretofore incurable hematologic malignancy. We are using a combination of novel technologies and bioinformatics platforms to discover the genetic and immunological determinants of the immune responses that we have induced. Our central hypothesis is that clinical and immunological responses in patients with this disease after their therapeutic vaccination are determined by the somatic mutations encoded in their tumor genomes. Alterations in the tumor proteome, such as novel (neo-) antigenic peptides generated in the process of somatic mutation, can serve as potent substrates for specific anti-tumor immune responses when appropriately presented in the context of major histocompatibility complex (MHC) to effector T-cells, and in turn recognized by their antigen receptors. We are testing these hypotheses in close collaboration with ICBP@Stanford, first taking advantage of systematic methodologies for interrogation of the tumor coding genome, transcriptome, and MHC-peptidome to discover somatic mutations that are predicted and/or observed to bind cognate MHC. We are synthesizing and assembling synthetic versions of these candidate peptide neoantigens with corresponding MHC molecules, and using them in large peptide-MHC 'tetramer' panels to interrogate the T lymphocyte responses induced by tumor vaccination in the patients. In molecularly profiling, and functionally tracking these dynamic responses in serial blood specimens from patients before and after immunization, we are aiming to differentiate patients with or without clinical responses to this therapy, and to better predict their distinct outcomes. We anticipate that this integrated approach will reveal the interplay between nascent and induced immune responses and genetic factors in control of disease progression in MCL and inform new ways of battling this deadly disease. This approach should also have relevance to other cancers.
6. Characterization of a Cancer Stem Cell Hierarchy in NHL:
Studies in our laboratory also focus on further elucidating whether a CSC hierarchy exists in lymphomas, in order to be able to develop novel therapies targeting these cells. We now have mouse several models of indolent and aggressive primary human lymphomas as transplanted tumors in immune deficient mice, and this represents a major biological advance. This should allow our group to functionally and genetically define whether lymphoma initiating cells are present in these tumors using methods of flow cytometry and selective xenotransplantation.
7. Validation of an Acute Myeloid Leukemia Stem Cell Signature for Risk Evaluation and Assignment of Therapy:
We are interested in translating discoveries regarding tumor stem cells into the clinic through novel assays that measure CSC frequency and response to therapy. Here, we plan to validate a signature of leukemic stem cells we recently defined in AML in a prospective manner in the clinical setting, as a predictor of patient response to therapy, minimal residual disease, relapse, and death. Ultimately, as with the planned trial to develop anti-CD47 monoclonal antibody therapies for AML and NHL, our goal is to develop additional clinical trials at Stanford which will test CSC-based therapies in patients.
8. Prospective validation of a novel molecular risk predictor for diffuse large B-cell lymphoma:
We have developed a novel and robust method for predicting risk of outcomes for patients with the commonest lymphoma [diffuse large B-cell lymphoma (DLBCL)]. Given its simplicity, reproducibility, and practicality, this novel method could be a cost-effective approach to identify novel drugs that improve outcomes of patients with DLBCL within risk-adapted trials. However, a key requirement preceding such trials is its validation in a prospective setting, and this multi-center trial proposes to address this aim.