Involvement of tyrosine kinase p56/Lck in apoptosis induction by anticancer drugs.
Induction of apoptosis is a hallmark of the cellular response of human lymphocytes and lymphoma cells to treatment with anticancer drugs and irradiation. Both treatment modalities trigger apoptosis through intrinsic, mitochondrial apoptosis pathways resulting in the activation of caspases. We and others have shown that the tyrosine kinase p56/Lck is involved in the regulation of apoptosis induced by irradiation or treatment with ceramide but dispensable for death receptor triggered cell death. However, the role of p56/Lck for apoptosis induction in response to anticancer drugs is unclear. To elucidate the putative requirement of p56/Lck for apoptosis signaling of cytotoxic drugs, activation of caspases and alteration of mitochondrial functions were determined in Jurkat T cells, the p56/Lck deficient JCaM1.6 cells and the p56/Lck retransfected JCaM1.6/Lck cells in response to chemotherapeutic drugs with different targets of their primary action. Treatment with Doxorubicin, Paclitaxel or 5-Fluorouracil induced a breakdown of the mitochondrial membrane potential and apoptotic cell death in p56/Lck expressing Jurkat and the retransfected JCaM1.6/Lck cells within 48h of treatment. However, almost no mitochondrial alterations and no induction of apoptosis could be detected in the p56/Lck deficient JCaM1.6 cells. Correspondingly, activation of caspases-9, -8, and -3 and cleavage of the caspase-3 substrate PARP (poly-(ADP-ribose)-polymerase) were almost completely absent in JCaM1.6 cells while present in p56/Lck positive Jurkat and JCaM1.6/Lck cells. In contrast, retransfection of the cells with the p56/Lck-related tyrosine kinase Src could not restore sensitivity to the treatment with cytotoxic drugs indicating a specific role of the tyrosine kinase p56/Lck in apoptosis signaling. Importantly, kinase-activity of p56/Lck may be dispensable for its pro-apoptoptic action since preincubation with the Src-kinase inhibitor PP2 did not reduce apoptosis induced by cytotoxic drugs. In conclusion, the tyrosine kinase p56/Lck is essential for apoptosis induction by Doxorubicin, Paclitaxel and 5-Fluorouracil regulating early steps of the mitochondrial apoptosis signaling cascade, including alteration of mitochondrial functions and caspase-activation.
Gruber
Charlotte
C
Department of Radiooncology, University of Tübingen, Hoppe-Seyler Strasse 3, 72076 Tübingen, Germany.
Henkel
Marco
M
Budach
Wilfried
W
Belka
Claus
C
Jendrossek
Verena
V
D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D004317:Doxorubicin / Q000494:pharmacology; D005472:Fluorouracil / Q000494:pharmacology; D006801:Humans; D019169:Jurkat Cells; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck) / Q000172:deficiency / Q000235:genetics / Q000502:physiology*; D008928:Mitochondria / Q000187:drug effects / Q000201:enzymology; D017239:Paclitaxel / Q000494:pharmacology; D011505:Protein-Tyrosine Kinases / Q000502:physiology; D015398:Signal Transduction / Q000187:drug effects
D000970:Antineoplastic Agents; D004317:Doxorubicin; D011505:Protein-Tyrosine Kinases; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck); C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D017239:Paclitaxel; D005472:Fluorouracil