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Expression of Pro-apoptotic Caspase-3 Correlates With Lung Cancer Cell Growth Behavior Among Phenotypes With Different Intrinsic Doubling Times FREE TO VIEW

King F. Kwong, MD; Lindsay B. Cooper, B.S.; Mark J. Krasna, MD
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Greenebaum Cancer Center, University of Maryland, Baltimore, MD


Chest. 2003;124(4_MeetingAbstracts):197S. doi:10.1378/chest.124.4_MeetingAbstracts.197S-a
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PURPOSE:  Different lung cancer sub-types exhibit highly variable growth rate phenotypes. Apoptosis pathways are highly conserved, even among cancer cells. Caspase-3 is a critical down-stream effector that promotes programmed cell-death when activated. The purpose of this study is to identify in lung cancer cells of varying growth phenotypes whether Caspase-3 expression correlates with cancer cell growth.

METHODS:  Non-small cell lung cancer cell lines (A549, Calu, and NCI-H596) were selected because of their divergent genetic mutational characteristics and cultured in appropriate media and incubation conditions. Intrinsic cancer cell proliferation rates were measured using a modified MTT growth assay. Cytosolic fractions were isolated from cell extracts and were assayed for total protein concentration as well as Capase-3 protease enzymatic activity. Equivalent protein amounts from each lung cancer cell line were separated by 1-D gel electrophoresis and subjected to Western immunoblotting using an antibody monospecific for Caspase-3 protein.

RESULTS:  Growth rates varied between the three lung cancer cell lines as represented by their estimated cell doubling times (A549- 8 hr ; Calu- 30 hr ; H596- 27 hr). Intrinsic Caspase-3 activities for A549, Calu, and H596 were 2.17+/−0.85 x 10−3 OD/ug, 3.42 +/− 1.38 x 10−3 OD/ug, and 4.5 +/− 1.1 x 10−3 OD/ug, respectively. Differences in intrinsic Caspase-3 activity (slow- vs. fast-growing) were significant between H596 and A549 (p=0.04) but not between Calu and A549 (p=0.26). Western immunoblotting confirmed the presence of greater Caspase-3 protein quantity in H596 than in A549.CONCLUSIONS: Intrinsic Caspase-3 protein quantity and activity appear to be greater in the slower growing H596 lung cancer cell line as compared to the much faster, and possibly less cell-cycle regulated, A549. This same relationship was also seen morphologically in Calu versus A549, although their data did not reach statistical significance.

CLINICAL IMPLICATIONS:  Down regulation of Caspase-3 may represent a critical defect in the mechanistic control pathway of lung cancer cells. Greater expression of Caspase-3 activity may possibly inhibit cellular proliferation in lung cancer.

DISCLOSURE:  K.F. Kwong, None.

Wednesday, October 29, 2003

12:30 PM - 2:00 PM




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