Bryostatin Enhances the Cytotoxic Effects of Anti-CD22 Immunotoxins in CLL by Two Distinct Mechanisms: Implications for a Sequential Therapy.

Abstract 3429

Poster Board III-317

Recently, treatment of CLL has been improved by the addition of Rituximab, a monoclonal antibody directed against CD20. Although Rituximab displays modest activity when used as monotherapy, the combination of Rituximab and chemotherapy has demonstrated impressive efficacy in CLL. However, even though remissions can be achieved in the majority of patients, CLL still remains an incurable disease for most patients. Therefore, alternative treatment options are needed. BL22 is a recombinant protein composed of the variable region of a monoclonal antibody, which binds to CD22 on the surface of normal and malignant B-cells and of PE38, a truncated Pseudomonas exotoxin. Previously, we demonstrated that BL22 induced cell death in CLL, involving the intrinsic apoptotic pathway. However, apoptosis induction correlates with expression of CD22 on the surface of CLL cells and was only moderate in CD22low expressing cells. The aim of this study was to increase BL22 cytotoxicity by modulating the surface expression of CD22 on CLL cells. Bryostatin is a macrocyclic lactone which structurally mimics the PKC-activating second messenger diacylglycerol. This PKC modulator has demonstrated anti-leukemic effects in CLL by itself in vivo. Here we show that Bryostatin up-regulates the expression of CD22 on CLL cells in a PKC dependent pathway, thereby increasing the cytotoxic effects of BL22. However, PKC-mediated up-regulation of Mcl-1 attenuated the anti-leukemic effects of BL22. We unravel that CD22 and Mcl-1 up-regulation by Bryostatin occurs in a dose dependent manner. Importantly, higher doses of Bryostatin prevented the undesirable up-regulation of Mcl-1 by inhibition of PKC in spite of CD22 up-regulation. Therefore, combined treatment of BL22 and Bryostatin seems to be a promising approach to enhance the cytotoxic effects of the immunotoxin by modulating PKC activity and CD22 up-regulation. In addition, our data provide evidence for a sequential therapy with both drugs, since CLL cells can be primed with Bryostatin. After a single dose of Bryostatin, PKC-modulation lasts for several days, allowing to administer BL22 at later time points, thereby decreasing the risk of potential drug interactions. Finally we demonstrate that stromal cell mediated drug resistance in CLL can be overcome by Bryostatin/ BL22 therapies. Conclusively, Bryostatin enhances the anti-leukemic effects of BL22 in CLL. Clinical trails are needed to prove if such an approach has clinical impact on the treatment and prognosis of B-CLL.