Growth inhibitory activity of a novel lectin from Cliona varians against K562 human erythroleukemia cells.
OBJECTIVE
In this study, the antitumoral potential of a novel lectin (CvL) purified from the marine sponge Cliona varians was studied in different cancer cell lines.


METHODS
CvL cytotoxicity was evaluated in mammalian tumor cells and in normal human peripheral blood lymphocytes by the MTT assay using the same range of concentrations (1-150 microg ml(-1)). The mechanisms involved in K562 cell death were investigated by confocal fluorescence microscopy, flow cytometry and immunoblot.


RESULTS
CvL inhibited the growth of human leukemia cells, with IC(50) values of 70 and 100 microg ml(-1) for K562 and JURKAT cells, respectively, but it was ineffective on blood lymphocytes and solid tumor cell lines. K562 cell death occurred 72 h after exposure to the lectin and with signs of apoptosis, as analyzed by DAPI and annexin V/PI staining. Investigation of the possible mediators of this process showed that cell death occurred via a caspase-independent pathway. Confocal fluorescence microscopy indicated a pivotal role for the lysosomal protease cathepsin B in mediating cell death. Accordingly, pre-incubation of K562 cells with the cathepsin inhibitor L-trans-epoxysuccinyl-L-leucylamido-(4-guanidino)butane (E-64) abolished CvL cytotoxic effect. Furthermore, we found upregulation of tumor necrosis factor receptor 1 (TNFR1) and down-modulation of p65 subunit of nuclear factor kappa B (NFkappaB) expression in CvL-treated cells. These effects were accompanied by increased levels of p21 and reduced expression of pRb, suggesting that CvL can induce cell cycle arrest.


CONCLUSIONS
Collectively, these findings indicate an antileukemic effect for CvL and suggest that cathepsin B acts as a death mediator in CvL-induced cytotoxicity possibly in an uncharacterized connection with the membrane death receptor pathway.
Queiroz
Alexandre F S
AF
Departamento de Biofísica e Farmacologia, Centro de Biociências, Universidade Federal do Rio Grande do Norte, CEP 59072-970, Natal, RN, Brazil.
Silva
Rodrigo A
RA
Moura
Raniere M
RM
Dreyfuss
Juliana L
JL
Paredes-Gamero
Edgar J
EJ
Souza
Ana C S
AC
Tersariol
Ivarne L S
IL
Santos
Elizeu A
EA
Nader
Helena B
HB
Justo
Giselle Z
GZ
de Sales
Maurício P
MP
D000818:Animals; D000970:Antineoplastic Agents / Q000302:isolation & purification / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D020169:Caspases / Q000378:metabolism; D018929:Cell Culture Techniques; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D049852:Clione / Q000737:chemistry*; D005434:Flow Cytometry; D006801:Humans; D015151:Immunoblotting; D020128:Inhibitory Concentration 50; D019169:Jurkat Cells; D020014:K562 Cells; D037102:Lectins / Q000302:isolation & purification / Q000494:pharmacology*; D008214:Lymphocytes / Q000166:cytology / Q000187:drug effects
D000970:Antineoplastic Agents; D037102:Lectins; D020169:Caspases