Berubicin Hydrochloride

BACKGROUND: ;Neuroblastoma (NB) is one of the most common extracranial tumors in children. The chemotherapeutic doxorubicin (Dox) remains a mainstay for treatment. However, the emergence of drug resistance seriously limits treatment efficacy. Numerous Dox analogues have been designed to more potently kill drug naive as well as drug-resistant NB. We have shown that the Dox analogue WP744 has enhanced the killing activity of NB compared to Dox, but the mechanism(s) of action is unclear.;MATERIALS AND METHODS: ;MTT assays were used to characterize the relative potencies of Dox and WP744 against SH-SY5Y NB cells. Western blotting was used to assess activation of caspases-3, -9, anti-poly (ADP-ribose) polymerase, p53, p21, AIF, IkappaBalpha, Bcl-2, Bcl-X(L), and cyclin D1. Nuclear factor (NF-kappaB) activation was assessed by electrophoretic mobility shift assay.;RESULTS: ;After WP744 treatment, enhanced apoptosis and cell death were seen, associated with cleavage of caspases-3, -9, and anti-poly (ADP-ribose) polymerase, an increase in p53 protein levels, and the induction of p21. WP744 also induced translocation of apoptosis-inducing factor from mitochondria to nuclei. Most remarkably, WP744 was 50-fold more potent than Dox in activating NF-kappaB.

Several reports within the last 5 years have suggested that nuclear factor (NF)-kappaB activation suppresses apoptosis through expression of anti-apoptotic genes. In the present report, we provide evidence from four independent lines that NF-kappaB activation is required for the cytotoxic effects of doxorubicin. We used doxorubicin and its structural analogues WP631 and WP744, to demonstrate that anthracyclines activate NF-kappaB, and this activation is essential for apoptosis in myeloid (KBM-5) and lymphoid (Jurkat) cells. All three anthracyclines had cytotoxic effects against KBM-5 cells; analogue WP744, was most potent, with an IC(50) of 0.5 microM, and doxorubicin was least active, with an IC(50) of 2 microM. We observed maximum NF-kappaB activation at 1 microM with WP744 and at 50 microM with doxorubicin and WP631, and this activation correlated with the IkappaBalpha degradation. Because the anthracycline analogue (WP744), most active as a cytotoxic agent, was also most active in inducing NF-kappaB activation and the latter preceded the cytotoxic effects, suggests that NF-kappaB activation may mediate cytotoxicity. Second, receptor-interacting protein-deficient cells, which did not respond to doxorubicin-induced NF-kappaB activation, were also protected from the cytotoxic effects of all the three anthracyclines.

BACKGROUND: ;The synthetic 4'-O-benzylated doxorubicin analog WP744 was designed to abrogate transport by the multidrug resistance (MDR)-associated ATP-binding cassette (ABC) proteins P-glycoprotein (Pgp) and multidrug resistance protein (MRP-1). We compared its uptake and cytotoxicity with those of doxorubicin and daunorubicin in cell lines overexpressing Pgp, MRP-1 or breast cancer resistance protein (BCRP) and in acute myeloid leukemia (AML) cells.;METHODS: ;Cellular uptake was studied by flow cytometry and cytotoxicity in 96-h 96-well cultures in cell lines overexpressing Pgp, MRP-1 or wild type (BCRP(R482)) or mutant (BCRP(R482T), BCRP(R482G)) BCRP and in pre-treatment AML marrow cells.;RESULTS: ;Uptake and cytotoxicity of WP744 were consistently greater than those of doxorubicin and daunorubicin at equimolar concentrations in all cell lines studied and in AML cells.;CONCLUSION: ;WP744 overcomes transport by Pgp, MRP-1 and BCRP in cell lines and AML cells and is a promising agent for clinical development in AML and other malignancies with broad-spectrum multidrug resistance.