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Hairy cell leukemia (HCL) is a chronic lymphoproliferative disease, the cause of which is unknown. Diagnostic of HCL is abnormal expression of the gene that encodes the β2 integrin CD11c. In order to determine the cause of CD11cgene expression in HCL theCD11cgene promoter was characterized. Transfection of theCD11cpromoter linked to a luciferasereporter gene indicated that it is sufficient to direct expression in hairy cells. Mutation analysis demonstrated that of predominant importance to the activity of the CD11cpromoter is its interaction with the activator protein-1 (AP-1) family of transcription factors. Comparison of nuclear extracts prepared from hairy cells with those prepared from other cell types indicated that hairy cells exhibit abnormal constitutive expression of an AP-1 complex containing JunD. Functional inhibition of AP-1 expressed by hairy cells reducedCD11cpromoter activity by 80%. Inhibition of Ras, which represents an upstream activator of AP-1, also significantly inhibited the CD11cpromoter. Furthermore, in the hairy cell line EH, inhibition of Ras signaling through mitogen-activated protein kinase/extracellular signal–regulated kinase kinases 1 and 2 (MEK1/2) reduced not only CD11cpromoter activity but also reduced both CD11c surface expression and proliferation. Expression in nonhairy cells of a dominant-positive Ras mutant activated the CD11cpromoter to levels equivalent to those in hairy cells. Together, these data indicate that the abnormal expression of the CD11cgene characteristic of HCL is dependent upon activation of the proto-oncogenes rasand junD.

Hairy cell leukemia (HCL) is a chronic lymphoproliferative disease, the cause of which is unknown. Diagnostic of HCL is abnormal expression of the gene that encodes the β2 integrin CD11c. In order to determine the cause of CD11c gene expression in HCL theCD11c gene promoter was characterized. Transfection of theCD11c promoter linked to a luciferase reporter gene indicated that it is sufficient to direct expression in hairy cells. Mutation analysis demonstrated that of predominant importance to the activity of the CD11c promoter is its interaction with the activator protein-1 (AP-1) family of transcription factors. Comparison of nuclear extracts prepared from hairy cells with those prepared from other cell types indicated that hairy cells exhibit abnormal constitutive expression of an AP-1 complex containing JunD. Functional inhibition of AP-1 expressed by hairy cells reducedCD11c promoter activity by 80%. Inhibition of Ras, which represents an upstream activator of AP-1, also significantly inhibited the CD11c promoter. Furthermore, in the hairy cell line EH, inhibition of Ras signaling through mitogen-activated protein kinase/extracellular signal–regulated kinase kinases 1 and 2 (MEK1/2) reduced not only CD11c promoter activity but also reduced both CD11c surface expression and proliferation. Expression in nonhairy cells of a dominant-positive Ras mutant activated the CD11cpromoter to levels equivalent to those in hairy cells. Together, these data indicate that the abnormal expression of the CD11cgene characteristic of HCL is dependent upon activation of the proto-oncogenes ras and junD.

We hypothesized that P2Y12 antagonism would lead to a decrease in serum thromboxane generation. We tested our hypothesis by employing both pharmacological and molecular genetic approaches.Serum thromboxane levels are a cogent marker for platelet hyperreactivity. It is produced when thrombin, which is generated during coagulation cascades, acts on PAR receptors on platelets. Serum was collected from non-anticoagulated blood after clot formation, following which serum thromboxane levels were measured with ELISA. The serum thromboxane levels in mice dosed with clopidogrel (n=4) (30 mg/kg body weight) were inhibited 88.04% compared to untreated mice. In human blood treated with the active metabolites of clopidogrel (R138727) or prasugrel (R361015) (n=3) were inhibited by 83.7% and 95.1%, respectively, compared to untreated human serum (100%). We also evaluated serum thromboxane levels in P2Y receptor null mice (n=4). Where as serum thromboxane levels in P2Y1 null mice were similar to those in wild type littermates, those in the P2Y12 null mice were inhibited by 78.9%. Furthermore serum thromboxane levels in P2Y12 deficient patients previously described in France and Japan were reduced to 25.3% and 11.7%, respectively, compared to normal volunteers (100%). We had one patient who was only on clopidogrel whose serum thromboxane levels were inhibited by 92.1%, compared to normal volunteers. In a pilot study, we recruited healthy volunteers (n=6), who were given clopidogrel for a week. Serum thromboxane levels were measured before and after the administration of clopidogrel. On an average, the serum thromboxane levels in these volunteers were inhibited by 72.3% after receiving clopidogrel.In conclusion, P2Y12 receptor antagonism alone could lead to decrease in serum thromboxane levels. Hence in future novel P2Y12 antagonists alone might offer better protection without an additional need for aspirin.No relevant conflicts of interest to declare.

Abstract 4013