Faculty Bibliography

BACKGROUND:Adaptive CD19-targeted chimeric antigen receptor (CAR) T-cell transfer has become a promising treatment for leukemia. Although patient responses vary across different clinical trials, reliable methods to dissect and predict patient responses to novel therapies are currently lacking. Recently, the depiction of patient responses has been achieved using in silico computational models, with prediction application being limited. METHODS:) relapse. Real-time CAR T-cell and tumor burden data of 209 patients were collected from clinical studies and standardized with unified units in bone marrow. Parameter estimation was conducted using the stochastic approximation expectation maximization algorithm for nonlinear mixed-effect modeling. RESULTS:relapse. Furthermore, we predicted patient responses by combining the peak and accumulated values of CAR T-cells or by inputting early-stage CAR T-cell dynamics. A clinical trial simulation using virtual patient cohorts generated based on real clinical patient datasets was conducted to further validate the prediction. CONCLUSIONS:Our model dissected the mechanism behind distinct responses of leukemia to CAR T-cell therapy. This patient-based computational immuno-oncology model can predict late responses and may be informative in clinical treatment and management.

Clonal hematopoiesis (CH) is an aging-associated condition characterized by the clonal outgrowth of mutated pre-leukemic cells. Individuals with CH are at an increased risk of developing hematopoietic malignancies. Here, we describe a novel animal model carrying a recurrent TET2 missense mutation, frequently found in CH and leukemic patients. In a fashion similar to CH, animals show signs of disease late in life when they develop a wide range of myeloid neoplasms, including acute myeloid leukemia (AML). Using single cell transcriptomic profiling of the bone marrow, we show that disease progression in aged animals correlates with an enhanced inflammatory response and the emergence of an aberrant inflammatory monocytic cell population. The gene signature characteristic of this inflammatory population is associated to poor prognosis in AML patients. Our study illustrates an example of collaboration between a genetic lesion found in CH and inflammation, leading to transformation and the establishment of blood neoplasms.

Radium 223 (Ra223) is a bone-seeking, alpha-particle-emitting radionuclide approved for the treatment of patients with metastatic prostate cancer and is currently being tested in a variety of clinical trials for primary and metastatic cancers to bone. Clinical evaluation of Ra223 hematologic safety showed a significantly increased rate of neutropenia and thrombocytopenia in patients, hinting at myelosuppression as a side effect. In this study we investigate the consequences of Ra223 treatment on bone marrow biology. Ra223 accumulated in bones and induced zonal radiation damage confined at the bone interface, followed by replacement of the impaired areas with adipocyte infiltration, as monitored by three-dimensional multiphoton microscopy, ex vivo. Flow cytometry and single cell transcriptomic analyses on bone marrow hematopoietic populations revealed transient, non-specific Ra223-mediated cytotoxicity on resident populations, including stem, progenitor and mature leukocytes. This was paralleled by a significant decrease of white blood cells and platelets in peripheral blood, which was overcome within 40 days post-treatment. Ra223 exposure did not impair full hematopoietic reconstitution, suggesting that the bone marrow function is not permanently hampered. Our results provide a comprehensive explanation of Ra223 reversible effects on bone marrow cells and exclude long-term myelotoxicity, supporting its safety for patients.

The mutualistic relationship of gut-resident microbiota and the host immune system promotes homeostasis that ensures maintenance of the microbial community and of a largely non-aggressive immune cell compartment^1,2. The consequences of disturbing this balance include proximal inflammatory conditions, such as Crohn's disease, and systemic illnesses. This equilibrium is achieved in part through the induction of both effector and suppressor arms of the adaptive immune system. Helicobacter species induce T regulatory (T_reg) and T follicular helper (T_FH) cells under homeostatic conditions, but induce inflammatory T helper 17 (T_H17) cells when induced T_reg (iT_reg) cells are compromised^3,4. How Helicobacter and other gut bacteria direct T cells to adopt distinct functions remains poorly understood. Here we investigated the cells and molecular components required for iT_reg cell differentiation. We found that antigen presentation by cells expressing RORγt, rather than by classical dendritic cells, was required and sufficient for induction of T_reg cells. These RORγt⁺ cells-probably type 3 innate lymphoid cells and/or Janus cells⁵-require the antigen-presentation machinery, the chemokine receptor CCR7 and the TGFβ activator α_v integrin. In the absence of any of these factors, there was expansion of pathogenic T_H17 cells instead of iT_reg cells, induced by CCR7-independent antigen-presenting cells. Thus, intestinal commensal microbes and their products target multiple antigen-presenting cells with pre-determined features suited to directing appropriate T cell differentiation programmes, rather than a common antigen-presenting cell that they endow with appropriate functions.

B cell progenitor acute lymphoblastic leukemia (B-ALL) treatment has been revolutionized by T cell-based immunotherapies-including chimeric antigen receptor T cell therapy (CAR-T) and the bispecific T cell engager therapeutic, blinatumomab-targeting surface glycoprotein CD19. Unfortunately, many patients with B-ALL will fail immunotherapy due to 'antigen escape'-the loss or absence of leukemic CD19 targeted by anti-leukemic T cells. In the present study, we utilized a genome-wide CRISPR-Cas9 screening approach to identify modulators of CD19 abundance on human B-ALL blasts. These studies identified a critical role for the transcriptional activator ZNF143 in CD19 promoter activation. Conversely, the RNA-binding protein, NUDT21, limited expression of CD19 by regulating CD19 messenger RNA polyadenylation and stability. NUDT21 deletion in B-ALL cells increased the expression of CD19 and the sensitivity to CD19-specific CAR-T and blinatumomab. In human B-ALL patients treated with CAR-T and blinatumomab, upregulation of NUDT21 mRNA coincided with CD19 loss at disease relapse. Together, these studies identify new CD19 modulators in human B-ALL.

Despite improvements in outcomes for children with B and T-cell acute lymphoblastic leukemia (B-ALL and T-ALL), patients with resistant or relapsed disease fare poorly. Previous studies have demonstrated the essential role of cyclin D3 in T-ALL disease initiation and progression and that targeting of the CDK4/6-cyclin D complex can suppress T-ALL proliferation, leading to efficient cell death in animal models. Studies in leukemia and other malignancies, suggest that schedule is important when combining CDK4/6 inhibitors (CDKis) with cytotoxic agents. Based on these observations, we broadened evaluation of two CDKis, palbociclib (PD-0332991, Pfizer) and ribociclib (LEE011, Novartis) in B and T-ALL as single agent and in combination with conventional cytotoxic chemotherapy, using different schedules in preclinical models. As monotherapy, CDKis caused cell cycle arrest with a significant decrease in S phase entry and were active in vivo across a broad number of patient-derived xenograft samples. Prolonged monotherapy induces resistance, for which we identified a potential novel mechanism using transcriptome profiling. Importantly, simultaneous but not sequential treatment of CDKis with conventional chemotherapy (dexamethasone, L-asparaginase and vincristine) led to improved efficacy compared to monotherapy in vivo. We provide novel evidence that combining CDKis and conventional chemotherapy can be safe and effective. These results led to the rational design of a clinical trial.

Image-based analysis as a method for mutation detection can be advantageous in settings when tumor tissue is limited or unavailable for direct testing. Here, we utilize two distinct and complementary machine learning methods of analyzing whole slide images (WSI) for predicting mutated BRAF. In the first method, WSI of melanomas from 256 patients were used to train a deep convolutional neural network (CNN) in order to develop a fully automated model that first selects for tumor-rich areas (Area Under the Curve AUC=0.96) then predicts for mutated BRAF (AUC=0.71). Saliency mapping was performed and revealed that pixels corresponding to nuclei were the most relevant to network learning. In the second method, WSI were analyzed using a pathomics pipeline that first annotates nuclei and then quantifies nuclear features, demonstrating that mutated BRAF nuclei were significantly larger and rounder nuclei compared to BRAF WT nuclei. Lastly, we developed a model that combines clinical information, deep learning, and pathomics that improves the predictive performance for mutated BRAF to AUC=0.89. Not only does this provide additional insights on how BRAF mutations affect tumor structural characteristics, machine learning-based analysis of WSI has the potential to be integrated into higher order models for understanding tumor biology.

tRNAs are central players in decoding the genetic code linking codons in mRNAs with cognate amino acids during protein synthesis. Recent discoveries have placed tRNAs as key regulators of gene expression during hematopoiesis, especially in hematopoietic stem cell (HSC) maintenance and immune development. These functions have been shown to be influenced by dynamic changes in tRNA expression, post-transcriptional base modifications, tRNA-interacting proteins, and tRNA fragmentation; these events underlie the complexity of tRNA-mediated regulatory events in hematopoiesis. In this review, we discuss these recent findings and highlight how deregulation of tRNA biogenesis can contribute to hematological malignancies.