Abstract
Abstract 375
B cell receptor (BCR) signaling promotes survival of the malignant clone in chronic lymphocytic leukaemia (CLL) through its ability to stimulate NFkB pathway signaling. In lymphoid cells, antigen receptor stimulation of this pathway is achieved by engaging the Carma-1 – Bcl10 – MALT1 (CBM) complex for eventual activation of I-kB kinases (IKKs). In B cells, protein kinase C beta (PKCbeta) is an important mediator of CBM complex activation. However, in CLL cells we found that PKCs do not appear to have a role in BCR-mediated NFkB pathway signaling, despite high expression levels of PKCbeta, because the presence of specific inhibitors of this kinase (LY379196 and bisindolylmaleimide-I) has no effect on the induction of IKK phosphorylation during BCR crosslinking. Examination of CBM complex expression suggests an explanation for this phenomenon; the expression levels of Carma-1 and MALT-1 are largely similar in CLL and normal B cells, but the expression of Bcl10 is much reduced in CLL cells. These findings, taken together with the established role of Bcl10 in the pathway of BCR-induced NFkB activation, suggest that CLL cells may employ a different mechanism to activate this pathway during BCR stimulation. Tyrosine kinases are known to play a role in BCR-induced IKK activation in CLL cells because compounds like dasatinib and PP2 inhibit NFkB pathway activation by BCR. One possible tyrosine kinase is c-Abl because we have shown this protein to be overexpressed in CLL cells, where it plays a role in activation of the NFkB pathway. To investigate the role of c-Abl in BCR-induced IKK activation, we used the inhibitor imatinib and found that the presence of this compound partially inhibited IKK phosphorylation in BCR-stimulated CLL cells. However, imatinib can also inhibit Lck, a T cell-specific src-family tyrosine kinase that is expressed by CLL cells. To differentiate between Lck- and c-Abl-mediated BCR signals we used the specific inhibitor 4-amino-5-(4-phenoxyphenyl)-7H-pyrrolo[3,2d] pyrimidin-7-yl-cyclopentane (Lck-i). We found that the presence of this compound in CLL cell cultures undergoing BCR stimulation almost completely inhibited the induction of IKK activation. Investigation of Lck-i specificity revealed this compound did not inhibit either c-Abl or Lyn at the concentration used to inhibit Lck in CLL cell cultures. Further investigation of the effects of Lck-i showed that this compound was also effective in inhibiting BCR-induced activation of the Akt and ERK signaling pathways. Taken together, these data suggest a major role for Lck in BCR-mediated signaling in CLL cells, and question the existing paradigm on the importance of Lyn.
Disclosures:
No relevant conflicts of interest to declare.
Molecular cloning of human Syk. A B cell protein-tyrosine kinase associated with the surface immunoglobulin M-B cell receptor complex.