The Expression of Human Leukocyte Formin Protein, a New Protein That Associates with AKT, Is Correlated with the Expression of ZAP-70 in CLL.

Recently, we have cloned a new human gene (GenBank Accession No. AY278319), belonging to the formin family. This new gene, called for us human leukocyte formin, presents the common domains found in formin-related proteins: FH1, FH2 and FH3 domains. Western Blotting analysis has demonstrated that the protein encoded by this gene is overexpressed in lymphoid malignancies and cancer cell lines. Based on this pattern of expression, our objective was to investigate the expression of human leukocyte formin protein, by Western blotting analysis, in mononuclear cells from chronic lymphocytic leukemia (CLL) patients, isolated on a Ficoll-Hypaque gradient. We studied 18 CLL patients with median age of 65 y.o. (range, 45 to 86) out of treatment for at least three months (Rai 0 n=8; Rai 1 n=6; Rai 2 n=1; Rai3 n=1; Rai 4 n=2). As normal control we used 6 blood donors. Our data showed an overexpression of the human leukocyte formin in the CLL group when compared with the control group (p= 0.0354), as well as a positive correlation of this protein and ZAP-70 in the CLL group (Spearman test, p= 0.0107). The expression of ZAP-70 has been associated with rapid progression and poor survival and can be used as a prognostic marker. Previously we had described that human leukocyte formin protein associates with Akt, a critical survival regulator in many different cell types. The association was observed in a protein extract of Jurkat cell line and in peripheral blood leukocytes from CLL patients. In an attempt to confirm the association between Akt and human leukocyte formin, we performed cotransfections in 293 cells using an expression vector pEGFP containing FH2 or FH3 domains, and an expression vector of pCMV-HA containing the full length of Akt. The results showed that both FH2 and FH3 domains are involved in the association with Akt. The correlation of human leukocyte formin and ZAP-70 expression, and the association of human leukocyte formin protein with Akt suggest that this new protein may be involved in the signaling pathway of leukemia cell survival and is possibly a new prognostic marker.