Induction of apoptosis in renal cell carcinoma by combination of maitake mushroom extract (PDF) and vitamin C.

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Introduction and Objective: A bioactive extract (PDF) obtained from maitake mushroom (Grifola frondosa) has been demonstrated to have anticancer and immunostimulatory activity. Such bioactivity has also been shown to be synergistically activated by vitamin C (V.C). Although several conventional therapeutic options for renal cell carcinoma (RCC) are currently available, the unsatisfactory outcomes compellingly require for establishing a more effective treatment modality. Accordingly, we examined the combined effects of PDF and V.C on the growth of RCC in vitro as a possible complementary and alternative approach.
 Methods: The standardized form of mushroom extract, PDF, was obtained commercially. Human RCC, ACHN cell line, was employed and exposed to various concentrations of PDF or in combination with V.C, and cell viability/growth was assessed at specified times. To explore the growth inhibitory mechanism, cell cycle analysis and Western blot analysis on several apoptotic parameters were performed.
 Results: A dose-dependent study showed that PDF was capable of inducing a ~45% and ~70% growth reduction at 700 and 1000 μg/ml, respectively. When the lower, ineffective concentrations of PDF were combined with V.C (200 μM), ~90% cell death was attained with the combination of 300 µg/ml PDF and V.C. Yet, a given concentration (200 µM) of V.C itself had no effects. Cell cycle analysis revealed that compared to controls, there was an 88% decrease in the S phase cell population with a concomitant 80% increase in the G₁ cells following PDF/V.C treatment. Western blots further revealed that PDF/V.C-induced cell death was associated with a down-regulation of anti-apoptotic bcl-2, an up-regulation of pro-apoptotic Bax, and degradation of poly-(ADP-ribose)-polymerase (PARP).
 Conclusions: The present study demonstrates that PDF has antiproliferative and cytotoxic effects on renal carcinoma ACHN cells in vitro. Although the higher concentrations (>1000 µg/ml) of PDF alone are required to be significantly effective, the lower concentration (300 μg/ml) of PDF can be synergistically potentiated with V.C (200 μM), becoming extremely cytotoxic to these cancer cells (~90% cell death). Such a cytotoxic mechanism is associated with a G₁ cell cycle arrest (blocking the G₁-S phase transition) and the modulation of critical apoptotic regulators (bcl-2, Bax, and PARP), leading ultimately to apoptosis. Thus, PDF is a potential apoptosis-inducing agent that may have implications in a safer, more effective alternative therapeutic modality for RCC.