Absence of oncogenic AKT1 E17K mutation in prostate, esophageal, laryngeal and urothelial carcinomas, hepatoblastomas, gastrointestinal stromal tumors and malignant meningiomas

To the Editor

AKT1, which contains a pleckstrin homology (PH) domain, is a serine/threonine protein kinase. Mounting evidence indicates that activation of AKT1 is important in cancer development. An activating AKT1 mutation was recently identified in human breast (8.2%), colorectal (5.9%) and ovarian cancers (2.0%) [1]. The AKT1 mutation was a missense mutation that substituted an amino acid (E17K) in the PH domain. The AKT1 E17K mutation induces cellular transformation in vitro and produces leukemias in mice [1].

Because of the importance of AKT signaling activation in cancer development, following studies analyzed the AKT1 E17 mutation in a wide range of human cancers [2–5]. They found the AKT1 E17 mutation in breast, lung, and colon cancers, and malignant melanoma, but not in other cancers, including pancreas, liver, stomach cancers, and leukemias [2–5]. To see whether the AKT1 E17 mutation occurs in other cancers, we analyzed the AKT1 E17 mutation in cancers where the mutation has not been studied.

For this, we analyzed the AKT1 E17 mutation in methacarn-fixed tissues of 134 prostate adenocarcinomas, 61 esophageal squamous cell carcinomas, 31 laryngeal squamous cell carcinomas, 26 urothelial carcinomas, 38 hepatoblastomas, 22 gastrointestinal stromal tumors (GIST) and 10 malignant meningiomas by a polymerase chain reaction (PCR)-single strand conformation polymorphism (SSCP) assay. All of the patients of the cancers were Koreans. In the tumors, malignant cells and normal cells were selectively procured from hematoxylin and eosin-stained slides using a 30G1/2 hypodermic needle, as described previously [2], [6]. DNA extraction from the microdissected tissues was performed by a modified single-step DNA extraction method by proteinase K treatment [2], [6]. Because the AKT1 E17K mutation was detected in the exon 3, genomic DNA each from tumor cells and corresponding normal cells were amplified with one primer pair covering the exon 3. Radioisotope ([³²P]dCTP) was incorporated into the PCR products for detection by SSCP autoradiogram. As a positive control, we included a breast cancer in the SSCP that had been shown to harbor the AKT1 E17K mutation [2].

On the SSCP autoradiograms, all of the PCR products from the cancers were clearly seen. However, the SSCP from them did not reveal any aberrantly migrating band compared to wild-type bands from the normal tissues, indicating there was no evidence of the AKT1 E17K mutation in the cancers. The positive control showed aberrant bands in the SSCP. To confirm the SSCP results, we repeated the experiments twice, including tissue microdissection, PCR and SSCP to ensure the specificity of the results, and found that the data were consistent.

Many research groups have analyzed the AKT1 E17K mutation in many types of human cancers [1–5]. However, only breast cancers have been reported to harbor the mutation at moderate frequency (8.2%, 5.8% and 4.3%) by different researchers [1–3]. By contrast, other cancers showed very low or no incidences of the mutation [2–5]. Our study presented here did not reveal the AKT1 E17K mutation in the cancers analyzed, either. Our data indicate that the AKT1 E17K mutation is rare in prostate adenocarcinomas, esophageal squamous cell carcinomas, laryngeal squamous cell carcinomas, urothelial carcinomas, hepatoblastomas, GIST and malignant meningiomas, and suggest that the AKT1 E17K mutation may not contribute to AKT signaling activation in these cancers.

Acknowledgements

This study was supported by a grant from the Innovative Research Institute for Cell Therapy, Republic of Korea (A062260). The prostate cancer tissues were supplied from the Prostate Bank in Korea supported by Korea Science and Engineering Foundation.