Abstract

Chronic myelogenous leukemia (CML) is a myeloproliferative disorder, which originates from pluripotent hematopoietic bone marrow progenitor cells. It is characterized by excessive proliferation of the granulopoiesis in the bone marrow. In approximately 90% of the patients with CML the Philadelphia chromosome (Ph) is the characteristic cytogenetic hallmark. The Ph is a shortened chromosome 22, which arises from a acquired reciprocal translocation between chromosome 9 and 22 (t(9;22) (q34;q11)). This translocation fuses the bcr gene with part of the c-abl gene, which encodes a protein-tyrosine kinase. The chimeric bcr-abl oncogene is translated into a 210-kDa chimeric protein (p210) which exhibits constitutive ABL kinase activity. In the present study, we analysed the involvement of the BCR-ABL protein in the induction of antigen-specific cytotoxic T lymphocytes (CTLs) in order to develop an immunotherapeutic approach targeting this neo-antigen in patients with CML. To accomplish this, we generated dendritic cells (DCs) in vitro and electroporated them with various sources of RNA harbouring the chimeric bcr-abl transcript. These genetically engineered DCs where used as antigen presenting cells (APCs) for the in vitro induction of CTLs.

By applying this approach we found that the CTLs induced by DCs transfected with RNA extracted from bcr-abl-positive K-562 cells or CML blasts do not recognize epitopes derived from the chimeric BCR-ABL fusion protein. In contrast, they were able to lyse autologous DCs electroporated with RNA isolated from patients with acute myeloid leukemia (AML), indicating that antigens shared among these malignant cells are involved and recognized by these CTLs. However, we successfully generated BCR-ABL-specific CTLs by DCs electroporated with in vitro transcribed bcr-abl-RNA. In patients with CML in complete cytogenetic remission during IFN-α treatment there was some reactivity against BCR-ABL in IFN-γ ELISPOT assays, which was weaker as compared to proteinase 3 (PR3)- or Prame-directed responses supporting the observations obtained in healthy donors.

In summary, BCR-ABL is processed and presented by Ph+ cells but is not the immunodominat antigen that induces CD8+ T lymphocytes in competition with other tumor-associated antigens. Nevertheless, APCs transfected with pure bcr-abl-IVT in vitro are capable of inducing BCR-ABL specific CTLs that efficiently lyse Ph+ target cells.

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