title: Schedule-Dependent Antiangiogenic and Cytotoxic Effects of Chemotherapy on Vascular Endothelial and Retinoblastoma Cells.
abstract: Current treatment of retinoblastoma involves using the maximum dose of chemotherapy that induces tumor control and is tolerated by patients. The impact of dose and schedule on the cytotoxicity of chemotherapy has not been studied. Our aim was to gain insight into the cytotoxic and antiangiogenic effect of the treatment scheme of chemotherapy used in retinoblastoma by means of different in vitro models and to evaluate potential effects on multi-drug resistance proteins. Two commercial and two patient-derived retinoblastoma cell types and two human vascular endothelial cell types were exposed to increasing concentrations of melphalan or topotecan in a conventional (single exposure) or metronomic (7-day continuous exposure) treatment scheme. The concentration of chemotherapy causing a 50% decrease in cell proliferation (IC50) was determined by MTT and induction of apoptosis was evaluated by flow cytometry. Expression of ABCB1, ABCG2 and ABCC1 after conventional or metronomic treatments was assessed by RT-qPCR. We also evaluated the in vivo response to conventional (0.6 mg/kg once a week for 2 weeks) and metronomic (5 days a week for 2 weeks) topotecan in a retinoblastoma xenograft model. Melphalan and topotecan were cytotoxic to both retinoblastoma and endothelial cells after conventional and metronomic treatments. A significant decrease in the IC50 (median, 13-fold; range: 3-23) was observed following metronomic chemotherapy treatment in retinoblastoma and endothelial cell types compared to conventional treatment (p<0.05). Metronomic topotecan or melphalan significantly inhibited in vitro tube formation in HUVEC and EPC compared to vehicle-treated cells (p<0.05). Both treatment schemes induced apoptosis and/or necrosis in all cell models. No significant difference was observed in the expression of ABCB1, ABCC1 or ABCG2 when comparing cells treated with melphalan or topotecan between treatment schedules at the IC50 or with control cells (p>0.05). In mice, continuous topotecan lead to significantly lower tumor volumes compared to conventional treatment after 14 days of treatment (p<0.05). Continuous exposure to melphalan or topotecan increased the chemosensitivity of retinoblastoma and endothelial cells to both chemotherapy agents with lower IC50 values compared to short-term treatment. These findings were validated in an in vivo model. None of the dosing modalities induced multidrug resistance mechanisms while apoptosis was the mechanism of cell death after both treatment schedules. Metronomic chemotherapy may be a valid option for retinoblastoma treatment allowing reductions of the daily dose.
authors.affiliation: Clinical Pharmacokinetics Unit, Hospital de Pediatría JP Garrahan, Buenos Aires, Argentina.|National Scientific and Technical Research Council, CONICET, Buenos Aires, Argentina.
authors.forename: Ursula
authors.initials: U
authors.lastname: Winter
authors.affiliation: Experimental Thrombosis Laboratory, IMEX, National Academy of Medicine, Buenos Aires, Argentina.
authors.forename: Hebe A
authors.initials: HA
authors.lastname: Mena
authors.affiliation: National Scientific and Technical Research Council, CONICET, Buenos Aires, Argentina.|Experimental Thrombosis Laboratory, IMEX, National Academy of Medicine, Buenos Aires, Argentina.
authors.forename: Soledad
authors.initials: S
authors.lastname: Negrotto
authors.affiliation: National Scientific and Technical Research Council, CONICET, Buenos Aires, Argentina.|Inmunogenetics Laboratory, INIGEM, University of Buenos Aires, Buenos Aires, Argentina.
authors.forename: Eloisa
authors.initials: E
authors.lastname: Arana
authors.affiliation: Developmental tumor biology Laboratory and Department of Pediatric Hematology and Oncology, Hospital Sant Joan de Deu, Barcelona, Spain.
authors.forename: Guillem
authors.initials: G
authors.lastname: Pascual-Pasto
authors.affiliation: Clinical Pharmacokinetics Unit, Hospital de Pediatría JP Garrahan, Buenos Aires, Argentina.
authors.forename: Viviana
authors.initials: V
authors.lastname: Laurent
authors.affiliation: Pathology, Hospital Sant Joan de Deu, Barcelona, Spain.
authors.forename: Mariona
authors.initials: M
authors.lastname: Suñol
authors.affiliation: National Scientific and Technical Research Council, CONICET, Buenos Aires, Argentina.|Hospital de Pediatría JP Garrahan, Buenos Aires, Argentina.
authors.forename: Guillermo L
authors.initials: GL
authors.lastname: Chantada
authors.affiliation: Developmental tumor biology Laboratory and Department of Pediatric Hematology and Oncology, Hospital Sant Joan de Deu, Barcelona, Spain.
authors.forename: Angel M
authors.initials: AM
authors.lastname: Carcaboso
authors.affiliation: Clinical Pharmacokinetics Unit, Hospital de Pediatría JP Garrahan, Buenos Aires, Argentina.|National Scientific and Technical Research Council, CONICET, Buenos Aires, Argentina.
authors.forename: Paula
authors.initials: P
authors.lastname: Schaiquevich
mesh_terms: D020533:Angiogenesis Inhibitors / Q000494:pharmacology / Q000627:therapeutic use*; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology / Q000627:therapeutic use*; D017209:Apoptosis / Q000187:drug effects; D045744:Cell Line, Tumor; D005260:Female; D064593:Heterografts; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D051379:Mice; D008819:Mice, Nude; D012175:Retinoblastoma / Q000188:drug therapy* / Q000473:pathology
chemical_list: D020533:Angiogenesis Inhibitors; D000970:Antineoplastic Agents