Abstract
Activation of phosphatidylinositol 3'-kinase pathway is implicated in the pathogenesis of mantle cell lymphoma (MCL). The genetic change in phosphatidylinositol 3'-kinase catalytic subunit alpha (PIK3CA) in MCL has not been identified. Thirty-five primary MCL cases and 2 MCL cell lines (GRANTA-519 and Rec-1) were used to investigate somatic mutation and gene copy number of PIK3CA. Gene copy number was determined using quantitative real-time PCR and fluorescence in situ hybridization. We used quantitative real-time reverse transcription-PCR to measure PIK3CA transcription levels. Phosphatase and tensin homologue deleted on chromosome 10 (PTEN) and phoshorylated AKT protein levels were analyzed using Western blotting and immunohistochemistry. Flow cytometry was used to assess apoptosis after treatment of MCL cell lines and one control cell line with LY294002, a specific inhibitor of PI3KCA. Fifteen of 22 (68%) MCL cases and the MCL cell lines harbored a gain (> or =3) of PIK3CA gene copy number. In addition, cases with increased PIK3CA gene copy number had elevated PIK3CA mRNA levels. Furthermore, amplification of PIK3CA correlated with the status of AKT phosphorylation in 7 of 12 (58%) primary MCL cases. Inhibition of PIK3CA induced increased apoptosis in the MCL cell lines. PTEN protein expression was present in all 14 primary MCL cases and cell lines by Western blotting, whereas 5 of 33 (15%) cases tested by immunohistochemistry had loss of PTEN expression. We conclude that a gain of gene copy number of PIK3CA is frequent genetic alteration that contributes to MCL progression. PIK3CA is a promising therapeutic target in MCL.
Highlights
Activation of phosphatidylinositol 3′-kinase pathway is implicated in the pathogenesis of mantle cell lymphoma (MCL)
PIK3CA gene copy number was determined in two MCL cell lines, GRANTA-519 and Rec-1, and found to be amplified (3 and 4.7 copies, respectively; Fig. 2)
To investigate the functional implications of PIK3CA gain, we evaluated immunohistochemically the expression of active AKT and found that amplification of PIK3CA was associated with AKT phosphorylation in 7 of 12 (58%) primary MCL cases and the 2 MCL cell lines
Summary
Activation of phosphatidylinositol 3′-kinase pathway is implicated in the pathogenesis of mantle cell lymphoma (MCL). Results: Fifteen of 22 (68%) MCL cases and the MCL cell lines harbored a gain (≥3) of PIK3CA gene copy number. PTEN protein expression was present in all 14 primary MCL cases and cell lines by Western blotting, whereas 5 of 33 (15%) cases tested by immunohistochemistry had loss of PTEN expression. Dissecting the molecular events associated with activation of this pathway in mantle cell lymphoma presents an important challenge that has implications for the development and clinical testing of PI3K pathway inhibitors. Determination of PIK3CA gene and PTEN protein status may have value as biomarkers for patient selection in clinical trials testing PI3K or AKT inhibitors, but this remains a conjecture. Identification of molecular markers other than the underlying cyclin D1 overexpression might be useful for classifying patients in terms of prognosis, for better predicting response to treatment, and for assisting in the development of novel therapeutic approaches [8]
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