We isolated cDNA clones containing the entire coding region of the putative oncogene AKT2. Sequence analysis and in vitro translation demonstrated that AKT2 encodes a 56-kDa protein with homology to serine/threonine kinases; moreover, this protein contains a Src homology 2-like domain. AKT2 was shown to be amplified and overexpressed in 2 of 8 ovarian carcinoma cell lines and 2 of 15 primary ovarian tumors. AKT2 was mapped to chromosome region 19q13.1-q13.2 by fluorescence in situ hybridization. In the two ovarian carcinoma cell lines exhibiting amplification of AKT2, the amplified sequences were localized within homogeneously staining regions. We conclude that AKT2 belongs to a distinct subfamily of protein-serine/threonine kinases containing Src homology 2-like domains and that alterations of AKT2 may contribute to the pathogenesis of ovarian carcinomas. akt, the protooncogene transduced by the acute transforming retrovirus AKT8 (1, 2), encodes a protein-serine/threonine kinase containing a Src homology 2-like (SH2-like) domain (3). Two putative human cellular homologs, AKTI and AK12, were cloned by screening a human genomic DNA library with a v-akt probe under conditions of reduced stringency (4). Recently, we obtained AKTI clones from a normal human thymus cDNA library by using an AKTI genomic probe. Sequence analysis of portions of AKTI cDNA clones revealed that AKTI is the true human homolog of v-akt and is identical to the recently cloned RAC gene, which has been shown to encode a kinase related to members of the protein kinase C (PKC) family and the cyclic adenosine monophosphate-dependent protein kinase (cAMP-PK) (5). AKTI has been mapped to human chromosome band 14q32 (6), proximal to the immunoglobulin heavy-chain locus, and has been shown to be amplified in a gastric adenocarcinoma (4).In this communication, we report the cDNA cloning, sequence analysis,t and chromosomal mapping ofAKT2 and demonstrate that this putative oncogene encodes a protein belonging to a subfamily of serine/threonine kinases containing SH2-like domains. Moreover, we show that AKT2 is amplified and overexpressed in some human ovarian carcinoma cell lines and primary tumors, suggesting that it contributes to the development of common epithelial tumors of the ovary.

Imatinib mesylate targets mutated KIT oncoproteins in gastrointestinal stromal tumor (GIST) and achieves a clinical response in 80% of patients. The mechanism is believed to depend predominantly on the inhibition of KIT-driven signals for tumor cell survival and proliferation. Using a mouse model of spontaneous GIST, we found that the immune system contributes substantially to the anti-tumor effects of imatinib. Imatinib therapy activated CD8+ T cells and induced regulatory T cell (T reg) apoptosis within the tumor by reducing tumor cell expression of the immunosuppressive enzyme indoleamine 2,3-dioxygenase (Ido). Concurrent immunotherapy augmented the efficacy of imatinib in mouse GIST. In freshly obtained human GIST specimens, the T cell profile correlated with imatinib sensitivity and IDO expression. Thus, T cells are critical to the anti-tumor effects of imatinib in GIST and concomitant immunotherapy may further improve outcome in human cancers treated with targeted agents.

Large intergenic noncoding RNAs (lincRNA) have been less studied than miRNAs in cancer, although both offer considerable theranostic potential. In this study, we identified frequent upregulation of miR-196a and lincRNA HOTAIR in high-risk gastrointestinal stromal tumors (GIST). Overexpression of miR-196a was associated with high-risk grade, metastasis and poor survival among GIST specimens. miR-196a genes are located within the HOX gene clusters and microarray expression analysis revealed that the HOXC and HOTAIR gene were also coordinately upregulated in GISTs which overexpress miR-196a. In like manner, overexpression of HOTAIR was also strongly associated with high-risk grade and metastasis among GIST specimens. RNA interference-mediated knockdown of HOTAIR altered the expression of reported HOTAIR target genes and suppressed GIST cell invasiveness. These findings reveal concurrent overexpression of HOX genes with noncoding RNAs in human cancer in this setting, revealing miR-196a and HOTAIR as potentially useful biomarkers and therapeutic targets in malignant GISTs.

Although gastrointestinal stromal tumors (GISTs) harboring activating KIT or platelet-derived growth factor receptor A (PDGFRA) mutations respond to treatment with targeted KIT/PDGFRA inhibitors such as imatinib mesylate, these treatments are rarely curative. Most often, a sizeable tumor cell subpopulation survives and remains quiescent for years, eventually resulting in acquired resistance and treatment failure. Here, we report that imatinib induces autophagy as a survival pathway in quiescent GIST cells. Inhibiting autophagy, using RNAi-mediated silencing of autophagy regulators (ATGs) or antimalarial lysosomotrophic agents, promotes the death of GIST cells both in vitro and in vivo. Thus, combining imatinib with autophagy inhibition represents a potentially valuable strategy to promote GIST cytotoxicity and to diminish both cellular quiescence and acquired resistance in GIST patients.imatinib | targeted therapy | quiescence G astrointestinal stromal tumor (GIST) is the most common mesenchymal neoplasm of the gastrointestinal tract (1). Importantly, GISTs commonly harbor receptor tyrosine kinase mutations resulting in ligand-independent, constitutive activation that drives tumor cell proliferation; 85% have activating KIT mutations and an additional 7% have mutually exclusive platelet-derived growth factor receptor A (PDGFRA) mutations (1). (1). As a result, imatinib mesylate, a small molecule tyrosine kinase inhibitor that abrogates KIT and PDGFR activity, is highly effective as a treatment for metastatic GIST (1). Before imatinib, recurrent or metastatic GIST was uniformly fatal (2).Although imatinib is quite effective in stabilizing disease progression in GIST, it is generally not curative. Less than 2% of patients experience complete radiographic regression (3). Even upon prolonged imatinib treatment, most are left with a substantial and stable tumor burden consisting of viable nonproliferating tumor cells, indicating that significant numbers of GIST cells can survive imatinib and remain quiescent. Moreover, imatinib withdrawal in such individuals results in rapid disease progression, necessitating lifelong imatinib therapy (4). The inability of imatinib to fully eradicate GIST cells also contributes to acquired imatinib resistance, mostly due to intraallelic second-site KIT mutations that interfere with imatinib binding (5). Hence, from a therapeutic standpoint, it is critical to identify new agents or strategies to kill GIST cells either as single agents or in combination with imatinib.As a result, we sought to dissect the contributions of macroautophagy (hereafter called autophagy), an evolutionarily conserved lysosomal self-digestion process, to the survival of GIST cells during imanitib-induced quiescence (6). Autophagy is a key mechanism to recycle energy and nutrients during starvation or stress (7). Although the precise role of autophagy in cell survival versus death is highly context dependent (8, 9), growing evidence indicates that autophagy can promote tumor cell survival in response to both cyt...

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Gastrointestinal stromal tumour (GIST), originating from the interstitial cells of Cajal (ICCs), is characterized by frequent activating mutations of the KIT receptor tyrosine kinase. Despite the clinical success of imatinib that targets KIT, most advanced GIST patients develop resistance and eventually die of the disease. The ETS family transcription factor, ETV1, is a master regulator of the ICC lineage. Using mouse models of Kit activation and Etv1 ablation, we demonstrate that Etv1 is required for GIST initiation and proliferation in vivo, validating it as a therapeutic target. We further uncover a positive feedback circuit where MAP kinase activation downstream of KIT stabilizes the ETV1 protein and ETV1 positively regulates KIT expression. Combined targeting of ETV1 stability by imatinib and MEK162 resulted in increased growth suppression in vitro and complete tumour regression in vivo. The combination strategy to target ETV1 may provide an effective therapeutic strategy in GIST clinical management.

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Human herpesvirus 6 (HHV-6) genome has been detected in several human lymphoproliferative disorders with no signs of active viral infection, and found to be integrated into chromosomes in some cases. We previously reported a woman with HHV-6–infected Burkitt’s lymphoma. Fluorescence in situ hybridization showed that the viral genome was integrated into the long arm of chromosome 22 (22q13). The patient’s asymptomatic husband also carried HHV-6 DNA integrated at chromosome locus 1q44. To assess the possibility of chromosomal transmission of HHV-6 DNA, we looked for HHV-6 DNA in the peripheral blood of their daughter. She had HHV-6 DNA on both chromosomes 22q13 and 1q44, identical to the site of viral integration of her mother and father, respectively. The findings suggested that her viral genomes were inherited chromosomally from both parents. The 3 family members were all seropositive for HHV-6, but showed no serological signs of active infection. To confirm the presence of HHV-6 DNA sequences, we performed polymerase chain reaction (PCR) with 7 distinct primer pairs that target different regions of HHV-6. The viral sequences were consistently detected by single-step PCR in all 3 family members. We propose a novel latent form for HHV-6, in which integrated viral genome can be chromosomally transmitted. The possible role of the chromosomally integrated HHV-6 in the pathogenesis of lymphoproliferative diseases remains to be explained.