Cyproheptadine reduces expression of cyclin D2 in multiple myeloma lines. (A) KMS12, LP-1, OCI-MY5, MM1.R, OPM1, and U266 multiple myeloma cells and (B) OCI-AML2, OCI-AML3, HL60, OCI-M2, and NB4 leukemia cells (10⁶) were treated with 20 μM of cyproheptadine (CYP). Twenty-four hours after incubation, cell lysates were prepared, normalized for total protein, and analyzed by SDS-PAGE/immunoblotting using antibodies specific for cyclin D1, D2, D3, and anti–β-actin. CCND ratio represents (density of the D-cyclin)/(density of β-actin) relative to control cells. (C) OCI-MY5 and KMS11 myeloma cells (10⁶) were treated with increasing concentrations of cyproheptadine. Twenty-four hours after incubation, cell lysates were prepared, normalized for total protein, and analyzed by SDS-PAGE/immunoblotting using antibodies specific for cyclin D2 and anti–β-actin. (D) LP1 myeloma and OCI-AML2 leukemia cells (10⁶) were treated with 15 μM of cyproheptadine. At increasing times after incubation, cell lysates were prepared, normalized for total protein, and analyzed by SDS-PAGE/immunoblotting using antibodies specific for cyclin D2 and cyclin D3. CCND ratio represents (density of the D-cyclin)/(density of β-actin) relative to control cells. (E) LP1 myeloma and OCI-AML2 leukemia cells (10⁶) were treated with 10 and 20 μM of cyproheptadine. Twenty-four hours after incubation, total cellular RNA was isolated. Cyclin D2 (CCND2) and Cyclin D3 (CCND3) mRNA expression were measured relative to 18S RNA by real-time RT-PCR. Data represent the mean plus or minus SEM fold change of CCND2/18S or CCND3/18S expression relative to untreated controls (ΔΔCT normalization) from 4 independent experiments. *P = .017, **P = .004, #P = .015, ##P = .006, all by Student t test.