Acridine orange/ethidium bromide (AO/EB) staining showing apoptosis induction by TW-37. A, Apoptosis induction of TW-37 on 4 WSU-cell lines was assayed after 72-h exposure. WSU cell lines were seeded and treated with 500 nM of TW-37 and with inactive congener TW-37a [designated as "(-)"] in triplicate and apoptosis was determined by AO/EB staining after 72 h. B, Apoptosis induction of TW-37 on patient-derived NHL cells was determined on 3 selected samples. Apoptotic cells were assayed by AO/EB staining after exposure of TW-37 with concentrations ranging from 0 to 750 nM (0 is the same as TW-37a). C, Bax-to-Mcl-1 ratio positively correlates with induction of apoptosis by TW-37. The Bax/Mcl-1 ratio was plotted on the abscissa against this AO/EB metric on the ordinate for four cell lines (the filled diamonds represent 48 h and the empty squares represent 72 h treatments). Each line is calculated by linear regression using equal weighting of the four points; the lines described closely emanate from the origin (x-intercept = 0.046 to 0.084). Patient data (Patients-1 and 3, empty triangles) lie close to the lines fitted to the data for the four established NHL cell lines.