General MM

BOB1 intracellular transcription factor is a target for TCR-based therapy in MM

In January 2017 Jahn L. and colleagues from Leiden University Medical Center, The Netherlands published a paper in Blood, describing a pre-clinical study in which TCR gene transfer of BOB1 (intracellular B-cell specific transcription factor) conferred reactivity to recipient T cells and potent in-vivo anti-tumor activity in a mouse model of Multiple Myeloma (MM).

Key Findings

Identification of BOB1 epitopes and isolation of BOB1 reactive T cell clones

  • POU2AF1 (which encodes BOB1) was isolated as a B-cell specific gene
  • Four MHC-presented peptides of BOB1 were identified using the B-LCL ligandome, 3 of which bound to HLA-B*07:02
  • Stable pMHC tetramers for each BOB1-derived peptide were used to isolate/enrich T-cell clones from HLA-A2-/B7- individuals
  • Two T-cell clones: 3C10 recognizing BOB1245: A2 and 4G11 recognizing BOB144:B7, demonstrated specific recognition of peptide-pulsed target cells and reactivity against endogenously processed and presented peptide
  • T-cell clone 4G11 efficiently recognized all HLA-B7+ primary (BOB1-expressing) B-cell malignancies tested, including 5 CLLs, 4 ALLs, 3 MCLs, and 4 MMs, as well as K562 cells transduced with BOB1 (3C10 had insufficient avidity at this stage)
  • The 4G11 T-cell clone lacked reactivity towards healthy hematopoietic non-B cells and non-hematopoietic cells, indicating no off-target effects

TCR gene transfer confers reactivity to recipient T cells

  • A codon-optimized 4G11 TCR clone was developed along with NGF-R as a marker gene for selection/enrichment of transduced T cells
  • Expression of 4G11, on retrovirally transduced CD8+ T cells from an HLA-B7+ healthy individual, was confirmed by tetramer binding.
  • These clones:
    • readily recognized BOB1-expressing HLA-B7+ stimulator cells (MM cell lines – U9, U266, LCL-JY) as well as two ALL (acute lymphoblastic leukemia) cell lines
    • efficiently lysed several primary HLA-B7+ malignant cell samples
    • readily lysed HLA-B7+ primary MM samples at low effector-to-target ratios
    • did not lyse autologous activated T cells or CD14+ monocytes
    • proliferated upon stimulation with various BOB1-expressing HA-B7+ primary samples (including several MM cell lines)
  • No lysis was observed for mock transduced cells, indicating specificity of the TCR-4G11 clone

In-vivo mouse model for MM

  • U266 cells (HLA-B7+) were expanded in an NSG mouse for 3 weeks before introduction of TCR-4G11 T cells
  • Significant reduction of tumor cells in the bone marrow (BM) was observed with TCR-4G11 T cells but not with mock transduced cells

In conclusion, a chimeric BOB1-specific TCR clone conferred high anti-tumor reactivity to transduced T cells when tested in an in-vivo mouse xenograft model for established MM, and therefore paves the way for engineered T cells as a future treatment option for MM.

Abstract

Immunotherapy for hematological malignancies or solid tumors by administration of monoclonal antibodies or T cells engineered to express chimeric antigen receptors or T-cell receptors (TCRs) has demonstrated clinical efficacy. However, antigen-loss tumor escape variants and the absence of currently targeted antigens on several malignancies hamper the widespread application of immunotherapy. We have isolated a TCR targeting a peptide of the intracellular B cell-specific transcription factor BOB1 presented in the context of HLA-B*07:02. TCR gene transfer installed BOB1 specificity and reactivity onto recipient T cells. TCR-transduced T cells efficiently lysed primary B-cell leukemia, mantle cell lymphoma, and multiple myeloma in vitro. We also observed recognition and lysis of healthy BOB1-expressing B cells. In addition, strong BOB1-specific proliferation could be demonstrated for TCR-modified T cells upon antigen encounter. Furthermore, clear in vivo antitumor reactivity was observed of BOB1-specific TCR-engineered T cells in a xenograft mouse model of established multiple myeloma. Absence of reactivity toward a broad panel of BOB1- but HLA-B*07:02+ nonhematopoietic and hematopoietic cells indicated no off-target toxicity. Therefore, administration of BOB1-specific TCR-engineered T cells may provide novel cellular treatment options to patients with B-cell malignancies, including multiple myeloma.

References
  1. Jahn L.et al. TCR-based therapy for multiple myeloma and other B-cell malignancies targeting intracellular transcription factor BOB1. Blood. 2017 Mar 9;129(10):1284-1295. DOI: 10.1182/blood-2016-09-737536. Epub 2017 Jan 4.
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