Intracellular trafficking as a determinant of AS-DACA cytotoxicity in rhabdomyosarcoma cells
<p>Abstract</p> <p>Background</p> <p>Rhabdomyosarcoma (RMS) is a malignant soft tissue sarcoma derived from skeletal muscle precursor cells, which accounts for 5-8% of all childhood malignancies. Disseminated RMS represents a major clinical obstacle, and the need for be...
| Main Authors: | , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
BMC
2011-08-01
|
| Series: | BMC Cell Biology |
| Online Access: | http://www.biomedcentral.com/1471-2121/12/36 |
| _version_ | 1831538104647811072 |
|---|---|
| author | Stewart Bernard W Wakelin Laurence PG Hambley Trevor W Bryce Nicole S Huynh Tony Wolf Steven J Catchpoole Daniel R |
| author_facet | Stewart Bernard W Wakelin Laurence PG Hambley Trevor W Bryce Nicole S Huynh Tony Wolf Steven J Catchpoole Daniel R |
| author_sort | Stewart Bernard W |
| collection | DOAJ |
| description | <p>Abstract</p> <p>Background</p> <p>Rhabdomyosarcoma (RMS) is a malignant soft tissue sarcoma derived from skeletal muscle precursor cells, which accounts for 5-8% of all childhood malignancies. Disseminated RMS represents a major clinical obstacle, and the need for better treatment strategies for the clinically aggressive alveolar RMS subtype is particularly apparent. Previously, we have shown that the acridine-4-carboxamide derivative AS-DACA, a known topoisomerase II poison, is potently cytotoxic in the alveolar RMS cell line RH30, but is 190-fold less active in the embryonal RMS cell line RD. Here, we investigate the basis for this selectivity, and demonstrate in these RMS lines, and in an AS-DACA- resistant subclone of RH30, that AS-DACA-induced cytotoxicity correlates with the induction of DNA double strand breaks.</p> <p>Results</p> <p>We show that inhibition of the multidrug-resistance associated protein (MRP1) has no effect on AS-DACA sensitivity. By exploiting the pH-dependent fluorescence properties of AS-DACA, we have characterized its intracellular distribution, and show that it concentrates in the cell nucleus, as well as in acidic vesicles of the membrane trafficking system. We show that fluorescence microscopy can be used to determine the localization of AS-DACA to the nuclear and cytoplasmic compartments of RMS cells grown as spheroids, penetrance being much greater in RH30 than RD spheroids, and that the vesicular signal leads the way into the spheroid mass. EEA1 and Rab5 proteins, molecular markers expressed on early-endosomal vesicles, are reduced by > 50% in the sensitive cell lines.</p> <p>Conclusion</p> <p>Taking the evidence as a whole, suggests that endosomal vesicle trafficking influences the toxicity of AS-DACA in RMS cells.</p> |
| first_indexed | 2024-12-16T23:19:02Z |
| format | Article |
| id | doaj.art-95a37adc2da04c5bbb14306d9e8d62c5 |
| institution | Directory Open Access Journal |
| issn | 1471-2121 |
| language | English |
| last_indexed | 2024-12-16T23:19:02Z |
| publishDate | 2011-08-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Cell Biology |
| spelling | doaj.art-95a37adc2da04c5bbb14306d9e8d62c52022-12-21T22:12:13ZengBMCBMC Cell Biology1471-21212011-08-011213610.1186/1471-2121-12-36Intracellular trafficking as a determinant of AS-DACA cytotoxicity in rhabdomyosarcoma cellsStewart Bernard WWakelin Laurence PGHambley Trevor WBryce Nicole SHuynh TonyWolf Steven JCatchpoole Daniel R<p>Abstract</p> <p>Background</p> <p>Rhabdomyosarcoma (RMS) is a malignant soft tissue sarcoma derived from skeletal muscle precursor cells, which accounts for 5-8% of all childhood malignancies. Disseminated RMS represents a major clinical obstacle, and the need for better treatment strategies for the clinically aggressive alveolar RMS subtype is particularly apparent. Previously, we have shown that the acridine-4-carboxamide derivative AS-DACA, a known topoisomerase II poison, is potently cytotoxic in the alveolar RMS cell line RH30, but is 190-fold less active in the embryonal RMS cell line RD. Here, we investigate the basis for this selectivity, and demonstrate in these RMS lines, and in an AS-DACA- resistant subclone of RH30, that AS-DACA-induced cytotoxicity correlates with the induction of DNA double strand breaks.</p> <p>Results</p> <p>We show that inhibition of the multidrug-resistance associated protein (MRP1) has no effect on AS-DACA sensitivity. By exploiting the pH-dependent fluorescence properties of AS-DACA, we have characterized its intracellular distribution, and show that it concentrates in the cell nucleus, as well as in acidic vesicles of the membrane trafficking system. We show that fluorescence microscopy can be used to determine the localization of AS-DACA to the nuclear and cytoplasmic compartments of RMS cells grown as spheroids, penetrance being much greater in RH30 than RD spheroids, and that the vesicular signal leads the way into the spheroid mass. EEA1 and Rab5 proteins, molecular markers expressed on early-endosomal vesicles, are reduced by > 50% in the sensitive cell lines.</p> <p>Conclusion</p> <p>Taking the evidence as a whole, suggests that endosomal vesicle trafficking influences the toxicity of AS-DACA in RMS cells.</p>http://www.biomedcentral.com/1471-2121/12/36 |
| spellingShingle | Stewart Bernard W Wakelin Laurence PG Hambley Trevor W Bryce Nicole S Huynh Tony Wolf Steven J Catchpoole Daniel R Intracellular trafficking as a determinant of AS-DACA cytotoxicity in rhabdomyosarcoma cells BMC Cell Biology |
| title | Intracellular trafficking as a determinant of AS-DACA cytotoxicity in rhabdomyosarcoma cells |
| title_full | Intracellular trafficking as a determinant of AS-DACA cytotoxicity in rhabdomyosarcoma cells |
| title_fullStr | Intracellular trafficking as a determinant of AS-DACA cytotoxicity in rhabdomyosarcoma cells |
| title_full_unstemmed | Intracellular trafficking as a determinant of AS-DACA cytotoxicity in rhabdomyosarcoma cells |
| title_short | Intracellular trafficking as a determinant of AS-DACA cytotoxicity in rhabdomyosarcoma cells |
| title_sort | intracellular trafficking as a determinant of as daca cytotoxicity in rhabdomyosarcoma cells |
| url | http://www.biomedcentral.com/1471-2121/12/36 |
| work_keys_str_mv | AT stewartbernardw intracellulartraffickingasadeterminantofasdacacytotoxicityinrhabdomyosarcomacells AT wakelinlaurencepg intracellulartraffickingasadeterminantofasdacacytotoxicityinrhabdomyosarcomacells AT hambleytrevorw intracellulartraffickingasadeterminantofasdacacytotoxicityinrhabdomyosarcomacells AT brycenicoles intracellulartraffickingasadeterminantofasdacacytotoxicityinrhabdomyosarcomacells AT huynhtony intracellulartraffickingasadeterminantofasdacacytotoxicityinrhabdomyosarcomacells AT wolfstevenj intracellulartraffickingasadeterminantofasdacacytotoxicityinrhabdomyosarcomacells AT catchpooledanielr intracellulartraffickingasadeterminantofasdacacytotoxicityinrhabdomyosarcomacells |