Chemoresistance induces enhanced adhesion and transendothelial penetration of neuroblastoma cells by down-regulating NCAM surface expression.
BACKGROUND
Drug resistance to chemotherapy is often associated with increased malignancy in neuroblastoma (NB). One explanation for the link between resistance and malignancy might be that resistance facilitates cancer progression and invasion. To investigate this hypothesis, adhesion, transendothelial penetration and NCAM (CD56) adhesion receptor expression of drug-resistant versus drug-sensitive NB tumor cells were evaluated.


METHODS
Acquired drug resistance was mimicked by exposing parental UKF-NB-2, UKF-NB-3 or IMR-32 tumor cells to increasing concentrations of vincristine- (VCR) or doxorubicin (DOX) to establish the resistant tumor cell sublines UKF-NB-2VCR, UKF-NB-2DOX, UKF-NB-3VCR, UKF-NB-3DOX, IMR-32VCR and IMR-32DOX. Additionally, the malignant behaviour of UKF-NB-4, which already possessed the intrinsic multidrug resistance (MDR) phenotype, was analyzed. UKF-NB-4 exposed to VCR or DOX were designated UKF-NB-4VCR or UKF-NB-4DOX. Combined phase contrast - reflection interference contrast microscopy was used to separately evaluate NB cell adhesion and penetration. NCAM was analyzed by flow cytometry, western blot and RT-PCR.


RESULTS
VCR and DOX resistant tumor sublines showed enhanced adhesion and penetration capacity, compared to their drug naïve controls. Strongest effects were seen with UKF-NB-2VCR, UKF-NB-3VCR and IMR-32DOX. DOX or VCR treatment also evoked increased invasive behaviour of UKF-NB-4. The process of accelerated tumor invasion was accompanied by decreased NCAM surface and protein expression, and down-regulation of NCAM coding mRNA. Transfection of UKF-NB-4VCR cells with NCAM cDNA led to a significant receptor up-regulation, paralleled by diminished adhesion to an endothelial cell monolayer.


CONCLUSIONS
It is concluded that NB cells resistant to anticancer drugs acquire increased invasive capacity relative to non-resistant parental cells, and that enhanced invasion is caused by strong down-regulation of NCAM adhesion receptors.
Blaheta
Roman A
RA
Zentrum der Chirurgie, Klinik für Urologie und Kinderurologie, Johann Wolfgang Goethe-Universität, Frankfurt am Main, Germany. blaheta@em.uni-frankfurt.de
Daher
Frederick H
FH
Michaelis
Martin
M
Hasenberg
Christoph
C
Weich
Eva M
EM
Jonas
Dietger
D
Kotchetkov
Rouslan
R
Doerr
Hans Willhelm
HW
Cinatl
Jindrich
J
D000970:Antineoplastic Agents / Q000494:pharmacology; D015153:Blotting, Western; D019002:CD56 Antigen / Q000235:genetics / Q000378:metabolism*; D002448:Cell Adhesion / Q000187:drug effects; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation; D015536:Down-Regulation; D004317:Doxorubicin / Q000494:pharmacology; D018432:Drug Resistance, Multiple; D019008:Drug Resistance, Neoplasm; D042783:Endothelial Cells / Q000378:metabolism*; D005434:Flow Cytometry; D006801:Humans; D008857:Microscopy, Interference; D009361:Neoplasm Invasiveness; D019006:Neural Cell Adhesion Molecules / Q000235:genetics / Q000378:metabolism*; D009447:Neuroblastoma / Q000276:immunology* / Q000473:pathology; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D014162:Transfection; D014750:Vincristine / Q000494:pharmacology
D000970:Antineoplastic Agents; D019002:CD56 Antigen; D019006:Neural Cell Adhesion Molecules; D012333:RNA, Messenger; D014750:Vincristine; D004317:Doxorubicin