532

Livin (ML-IAP, BIRC7) is a member of the IAP family of anti-apoptosis proteins, and itsexpression has been reported in melanoma and some carcinomas. We have evaluated livin expression in primary pediatric neuroblastoma (NB) tumors; fresh, cryopreserved NB tumors; and NB cell lines. In an immunohistochemical (IHC) study of archival, formalin-fixed biopsies from 68 NB patients, we detected elevated expression of livin in 27% of cases, low-intermediate expression in 53%, and no expression in 20% of cases. Livin mRNA expression in archival tissues (as measured by quantitative RT-PCR) was correlated with protein levels (IHC). In cryopreserved NB tumors, livin mRNA level was similarly correlated with protein level (immunoblot). Similar percentages of both fresh and archival tumors expressed high, intermediate-low and barely detectable or absent levels of livin. Furthermore, livin expression was confirmed in all eight NB lines tested, although expression varied markedly among lines. We further analyzed the relationship between livin expression, clinical/biological features with prognostic significance, and outcome. Livin expression alone did not appear to impact survival; however, patients with high livin and amplified MYCN had significantly decreased survival compared to patients lacking one or both markers. To evaluate the effect of elevated livin expression on drug-resistance, we stably transfected livin into NB line IMR32. Livin-transfected cells showed increased resistance to doxorubicin but not vincristine, relative to the neo-transfected control. Conversely, we used siRNA to knockdown endogenous livin in LA155N, a high-livin-expressing, doxorubicin-resistant NB line. Synthetic siRNA repressed livin by 90% (as measured by qRT-PCR and immunoblot) for at least 72 hours. Interestingly, livin knockdown induced apoptosis in the absence of other apoptotic stimuli and sensitized cells to doxorubicin. These results suggest that: a) livin is expressed in primary and cultured NB cells, b) high livin expression may identify a subset of NB patients with particularly poor-prognosis among those with MYCN-amplified tumors c) high livin expression may protect NB cells from some genotoxic agents, and resistance may be reversed by targeting this protein.

98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA