The B-cell tumor–associated antigen ROR1 can be targeted with T cells modified to express a ROR1-specific chimeric antigen receptor

Abstract

Monoclonal antibodies and T cells modi- IntroductionB-cell chronic lymphocytic leukemia (B-CLL) and mantle cell lymphoma (MCL) are common B-cell malignancies that respond to chemotherapy but are rarely cured. Allogeneic hematopoietic stem cell transplantation (HCT) enables a T cell-mediated graft-versus-leukemia (GVL) effect and induces durable remissions in a subset of patients with chemotherapy-refractory B-CLL and MCL, demonstrating that these malignancies are susceptible to recognition and elimination by T cells. 1,2 In a previous study, we identified tumor-reactive CD8 ϩ T cells directed against minor histocompatibility (H) and tumor-associated antigens (TAA) expressed by B-CLL in patients with sustained tumor regression after allogeneic HCT. 3 These results have encouraged the development of T cell-adoptive immunotherapy to augment the GVL effect after HCT. However, major challenges for therapy with ␣␤ ⌻-cell receptor (TCR)-bearing T cells include the need to identify antigens with restricted expression on malignant cells to avoid graft-versus-host disease, and the population distribution and requirement for human leukocyte antigen (HLA)-restriction for both minor H antigens and TAA. 4 An approach that could overcome these challenges and also enable T-cell therapy for B-CLL and MCL in the nontransplant setting is to genetically modify T cells to express a chimeric antigen receptor (CAR) that is specific for a cell surface protein expressed by malignant cells. CARs consists of a single-chain antibody fragment (scFv) that is derived from the variable heavy (VH) and variable light (VL) chains of a monoclonal antibody (mAb) linked to the TCR CD3 chain that mediates T-cell activation and cytotoxicity. 5 Costimulatory signals can also be provided through the CAR by fusing the costimulatory domain of CD28 or 4-1BB to the CD3 chain. 5,6 CARs are specific for cell surface molecules independent from HLA, thus overcoming the limitations of TCR-recognition including HLA-restriction and low levels of HLA-expression on tumor cells. B-cell lineage differentiation molecules such as CD19 and CD20 are retained on most B-cell tumors, and T cells modified with CD19-and CD20-specific CARs are currently being evaluated in clinical trials. 7,8 However, targeting B-cell lineage-specific antigens with immunotherapy has the disadvantage of eliminating normal mature B cells, which can increase the risk of infection. 9,10 Here, we evaluate a strategy to selectively eliminate malignant B cells without damaging the mature normal B-cell compartment by targeting the receptor tyrosine kinase-like orphan receptor 1 (ROR1). ROR1 was identified as a highly expressed gene in B-CLL by expression profiling and it has been shown that ROR1-protein is uniformly expressed on the cell surface of B-CLL. 11-14 The ROR1-gene encodes a 105-kDa protein with a Submitted May 3, 2010; accepted August 4, 2010. Prepublished online as Blood First Edition paper, August 11, 2010; DOI 10.1182 DOI 10. /blood-2010 An Inside Blood analysis of this article appears at...

Mass spectrometry-based identification of a naturally presented receptor tyrosine kinase-like orphan receptor 1-derived epitope recognized by CD8 + cytotoxic T cells

“…3,4 In agreement with these studies, we identified the HLA-B*07:02-restricted ROR1 peptide NPRYPNYMF as a component of the HLA ligandome of CLL samples (Figure 1 A,B and Online Supplementary Table S1). In contrast, the ROR1 peptide could not be found in the HLA ligandome of normal cells (data not shown).…”

“…It has been demonstrated that ROR1 messenger Ribonucleic Acid (mRNA) is highly expressed in CLL cells (Online Supplementary Figure S1) and a uniform expression of ROR1 protein is detected on their cell surface. 3,4 In contrast, ROR1 expression was not found in other blood cells or on bone marrow-derived cells, except on a small subset of precursor B cells.…”

“…4 When investigating ROR1 protein expression of normal tissues, contradictory results were obtained: according to two studies and data obtained from a public proteomics repository (Human Integrated Protein Expression Database), ROR1 was not or only marginally expressed in normal tissues (Online Supplementary Figure S2B). 5,6 However, Balakrishnan et al reported that ROR1 is expressed in the parathyroid, pancreatic islets, and regions of the gastrointestinal tract.…”

Chimeric antigen receptor modified T cell therapy for B cell malignancies

“…Multiple cell surface antigens expressed by B cell malignancies, including CD19, CD20, CD22, CD23, CD38, ROR1, and kappa light chain, have been targeted with T cells engineered to express scFv-containing CAR constructs [13,15,[20][21][22][23][24][25], and some of these antigens are now being targeted in phase I clinical trials of CAR-modified T cell therapy (Table 1). Although targeting using a scFv incorporated into a CAR has remained the most prevalent strategy to target B cell malignancies, other approaches have been used to target CAR-modified T cells to solid tumors, for example by incorporation of IL-13 into a CAR to redirect engineered T cells to IL-13Ra2-expressing tumors [26].…”

Investigating the Molecular Specific Interactions on Cell Surface Using Atomic Force Microscopy

“…Then, a drop of cell suspension was placed on a poly-L-lysine-coated glass slide , these experiments indicate that Raji cells express ROR1. ROR1 is the receptor tyrosine kinase-like orphan receptor 1 and functional data suggest that ROR1 may act in Wnt-signaling and promote the survival of malignant cells [49,50]. Raji cell line is from a Burkitt's lymphoma patient, which is a subtype of B-cell NHL.…”