Soft Tissue Sarcoma Cells Are Highly Sensitive to AKT Blockade: A Role for p53-Independent Up-regulation of GADD45α

Abstract

The AKT signaling pathway is activated in soft tissue sarcoma (STS). However, AKT blockade has not yet been studied as a potential targeted therapeutic approach. Here, we examined the in vitro and in vivo effects of AKT inhibition in STS cells. Western blot analysis was used to evaluate the expression of AKT pathway components and the effect of AKT stimulation and inhibition on their phosphorylation. Cell culture assays were used to assess the effect of AKT blockade (using a phosphatidylinositol 3-kinase inhibitor and a specific AKT inhibitor) on STS cell growth, cell cycle, and apoptosis. Oligoarrays were used to determine gene expression changes in response to AKT inhibition. Reverse transcription-PCR was used for array validation. Specific small inhibitory RNA was used to knockdown GADD45A. Human STS xenografts in nude mice were used for in vivo studies, and immunohistochemistry was used to assess the effect of treatment on GADD45A expression, proliferation, and apoptosis. Multiple STS cell lines expressed activated AKT. AKT inhibition decreased STS downstream target phosphorylation and growth in vitro; G 2 cell cycle arrest and apoptosis were also observed. AKT inhibition induced GADD45a mRNA and protein expression in all STS cells treated independent of p53 mutational status. GADD45A knockdown attenuated the G 2 arrest induced by AKT inhibition. In vivo, AKT inhibition led to decreased STS xenograft growth. AKT plays a critical role in survival and proliferation of STS cells. Modulation of AKT kinase activity may provide a novel molecularly based strategy for STStargeted therapies. [Cancer Res 2008;68(8):2895-903]

GADD45A expression is correlated with patient prognosis in esophageal cancer

“…Thus, the mechanism behind the regulation of GADD45A expression in ESCC remains unclear. Other factors, such as myc, phosphoinositide 3-kinase/AKT (23,24), activating transcription factor 2 (25) and Quercetin (26), which have been shown to regulate GADD45A expression, should be investigated in future studies.…”

Dual targeting of AKT and mammalian target of rapamycin: A potential therapeutic approach for malignant peripheral nerve sheath tumor

“…Immunohistochemistry was done using 5-μm-thick tissue microarray sections as described previously (24). Briefly, paraffin sections were dewaxed and rehydrated before antigen retrieval.…”

Anti-proliferative and pro-apoptotic actions of a novel human and mouse ovarian tumor-associated gene OTAG-12: downregulation, alternative splicing and drug sensitization

“…Addressing the molecular nature of cellular inhibition, we have found that hOTAG-12b stimulates expression of several pro-apoptotic genes such as BAD (Willis and Adams, 2005;Boisvert-Adamo and Aplin, 2008), CIDEB (Lugovskoy et al, 1999;Chen et al, 2000) and GADD45a (Thyss et al, 2005;Al-Romaih et al, 2008), and inhibits expression of anti-apoptotic genes such as NAIP (Liston et al, 2003;O'Riordan et al, 2008) and Akt1 (Zhu et al, 2008). hOTAG-12b, similar to p53, induces a set of overlapping genes such as GADD45a and distinct genes such as Akt1, BAD, CIDEB and NAIP.…”

“…Although the target gene(s) responsible for the primary effect of expression of hOTAG-12b remains unclear, stimulated expression of GADD45a as early as 12 h after DOX induction (Figure 8c) is noteworthy. GADD45a is also regulated by a p53-independent pathway (Chen et al, 2001;Zhu et al, 2008). In addition, GADD45a, a known transcriptional target of BRCA1 (Harkin et al, 1999), is responsible at least in part for the regulation of a G2 cell cycle checkpoint (Wagner et al, 2008).…”