Sensitization of cervix cancer cells to Adriamycin by Pentoxifylline induces an increase in apoptosis and decrease senescence.
BACKGROUND
Chemotherapeutic drugs like Adriamycin (ADR) induces apoptosis or senescence in cancer cells but these cells often develop resistance and generate responses of short duration or complete failure. The methylxantine drug Pentoxifylline (PTX) used routinely in the clinics setting for circulatory diseases has been recently described to have antitumor properties. We evaluated whether pretreatment with PTX modifies apoptosis and senescence induced by ADR in cervix cancer cells.


METHODS
HeLa (HPV 18+), SiHa (HPV 16+) cervix cancer cells and non-tumorigenic immortalized HaCaT cells (control) were treated with PTX, ADR or PTX + ADR. The cellular toxicity of PTX and survival fraction were determinated by WST-1 and clonogenic assay respectively. Apoptosis, caspase activation and ADR efflux rate were measured by flow cytometry, senescence by microscopy. IkappaBalpha and DNA fragmentation were determinated by ELISA. Proapoptotic, antiapoptotic and senescence genes, as well as HPV-E6/E7 mRNA expression, were detected by time real RT-PCR. p53 protein levels were assayed by Western blot.


RESULTS
PTX is toxic (WST-1), affects survival (clonogenic assay) and induces apoptosis in cervix cancer cells. Additionally, the combination of this drug with ADR diminished the survival fraction and significantly increased apoptosis of HeLa and SiHa cervix cancer cells. Treatments were less effective in HaCaT cells. We found caspase participation in the induction of apoptosis by PTX, ADR or its combination. Surprisingly, in spite of the antitumor activity displayed by PTX, our results indicate that methylxantine, per se does not induce senescence; however it inhibits senescence induced by ADR and at the same time increases apoptosis. PTX elevates IkappaBalpha levels. Such sensitization is achieved through the up-regulation of proapoptotic factors such as caspase and bcl family gene expression. PTX and PTX + ADR also decrease E6 and E7 expression in SiHa cells, but not in HeLa cells. p53 was detected only in SiHa cells treated with ADR.


CONCLUSIONS
PTX is a good inducer of apoptosis but does not induce senescence. Furthermore, PTX reduced the ADR-induced senescence and increased apoptosis in cervix cancer cells.
Bravo-Cuellar
Alejandro
A
División de Inmunología, Centro de Investigación Biomédica de Occidente, Instituto Mexicano del Seguro Social, Sierra Mojada 800, Colonia Independencia, Guadalajara, Jalisco, CP 44340, México.
Ortiz-Lazareno
Pablo C
PC
Lerma-Diaz
Jose M
JM
Dominguez-Rodriguez
Jorge R
JR
Jave-Suarez
Luis F
LF
Aguilar-Lemarroy
Adriana
A
del Toro-Arreola
Susana
S
de Celis-Carrillo
Ruth
R
Sahagun-Flores
Jose E
JE
de Alba-Garcia
Javier E Garcia
JE
Hernandez-Flores
Georgina
G
D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D020169:Caspases / Q000187:drug effects / Q000378:metabolism; D045744:Cell Line, Tumor; D002469:Cell Separation; D002470:Cell Survival / Q000187:drug effects; D016922:Cellular Senescence / Q000187:drug effects*; D053938:DNA Fragmentation; D004317:Doxorubicin / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005434:Flow Cytometry; D015870:Gene Expression / Q000187:drug effects; D006367:HeLa Cells; D006801:Humans; D010431:Pentoxifylline / Q000494:pharmacology*; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016159:Tumor Suppressor Protein p53 / Q000096:biosynthesis / Q000187:drug effects; D002583:Uterine Cervical Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology
D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53; D004317:Doxorubicin; D020169:Caspases; D010431:Pentoxifylline