Introduction: The homeobox transcription factor OTX2 plays a critical role in very early neurogenesis, but can become oncogenic when aberrantly expressed later in life. Previous work has identified OTX2 as an oncogene in medulloblastoma; however, little is known about the role of OTX2 in retinoblastoma, another childhood malignant tumor.
Methods: OTX2 amplification and expression levels were assessed in primary retinoblastoma tumors and retinoblastoma cell lines Y79 and WERI. To study the oncogenic role of OTX2 in vitro, we disrupted OTX2 expression in retinoblastoma cell lines via OTX2-specific siRNAs and all-trans retinoic acid (ATRA), and we assessed the amount cell apoptosis and proliferation after knockdown. To examine the oncogenic effects of OTX2 in vivo, we carried out RNA interference experiments in retinoblastoma mice xenografts and recorded tumor size and tumor growth after OTX2 knockdown. We also examined the expression levels of MYC, CRX, and phosphorylated RB after OTX2 knockdown.
Results: This study showed that OTX2 was frequently amplified and/or over-expressed in retinoblastoma primary tumors and cell lines. Knockdown of OTX2 expression by siRNA and pharmacologic inhibition by ATRA both repressed OTX2 expression. This significantly increased apoptosis and decreased cell proliferation in vitro. Knockdown of OTX2 also significantly decreased tumor size and growth in vivo. Loss of OTX2 resulted in decreased expression of MYC and CRX, but increased the phosphorylation of RB.
Conclusions: This study shows that the oncogene OTX2, previously demonstrated in medulloblastoma, may play an important oncogenic role in retinoblastoma via regulation of MYC and CRX and the phosphorylation of RB. These tumors may also be amenable to therapy targeting OTX2.
Patient Care: The opportunity to develop new targeted therapeutic agents against retinoblastoma is of paramount importance. Current treatments of enucleation followed by nonspecific adjuvant therapies may cure the tumor, but at grave costs. Loss of vision is universal and the long-term toxicities of these adjuvant therapies on this young patient population can be devastating. More specific agents may be less toxic. This study showed that ATRA repressed OTX2-expression and inhibited retinoblastoma cell growth. Although ATRA and other retinoids may affect multiple molecular pathways, the connection between OTX2 repression and growth inhibition effect of ATRA suggests that OTX2 expressing tumors may be amenable to therapy with retinoids. Our studies of OTX2, in conjunction with the studies of others, lay the conceptual framework for clinical trials of retinoids and OTX2-specific agents in the treatment of this challenging pediatric tumor.
Learning Objectives: By the conclusion of this session, participants should be able to:
1) Describe the importance of the oncogenic role of OTX2 in retinoblastoma, 2) Discuss, in small groups, the effects of OTX2 on other known cancer pathways and possible mechanisms of tumorigenesis, 3) Identify a potential new target for effective treatment of retinoblastoma.
References: Adamson DC, et al. 2010. OTX2 is critical for the maintenance and progression of shh-independent medulloblastomas. Cancer Res 70: 181-191.
Di C, et al. 2005. Identification of OTX2 as a medulloblastoma oncogene whose product can be targeted by all-trans retinoic acid. Cancer Res 65: 919-924.