Glaucocalyxin A (GLA), a major component isolated from Rabdosia japonica, has been proven to show anti-bacterial and anti-tumor biological characteristics according to previous studies. However, its potential effect on bladder cancer remains unknown. The present research aims to investigate the underlying mechanism in treating bladder cancer in vivo and in vitro. Cell proliferation was analyzed by CCK-8 assay and colony formation. Flow cytometry was used to measure the cell cycle distribution and apoptosis. The expressions of the cell cycle and apoptosis-related proteins were detected by western blotting and immunofluorescence staining. Meanwhile, the in vivo study was performed to evaluate the anti-tumor effect on a UMUC3 subcutaneous tumor of NOD/SCID mice model. GLA suppressed colony-formation ability, triggered G2/M arrest and promoted apoptosis of UMUC3 cells in a dose-dependent manner. Furthermore, western blotting showed that GLA downregulated the expressions of PI3K p85, p-Akt, Bcl-2, CDK1, Cyclin B1 whereas upregulated the levels of PTEN, Bax, Cleaved Caspase-3. In vivo, GLA at a dosage of 20 mg/kg significantly inhibited tumor growth compared with the control group by intraperitoneal injection. These results suggested that GLA-related G2/M arrest and apoptosis in UMUC3 cells were mediated by a suppressed PI3K/Akt signaling pathway, which regulated p21Waf1/Cip1 as well as intrinsic caspase cascade. Collectively, our observations could help to develop new drugs targeting the PI3K/Akt pathway for the treatment of bladder cancer.

Bufalin, one of the active ingredients of the Chinese drug Chan su, exhibits significant antitumor activity against various cancer types. However, the role of bufalin in renal cell carcinoma (RCC) remains unclear. In the present study, it was demonstrated that bufalin inhibited cell proliferation, blocked the cell cycle in the G2/M phase, and reduced the metastasis of human RCC ACHN cells via the upregulation of p21waf/cip1 and E-cadherin and the downregulation of cyclin dependent kinase 1, cyclin B1, N-cadherin, and hypoxia-inducible factor-1α (HIF-1α). Further mechanistic study revealed that bufalin reduced the expression of phosphorylated (phospho)-Akt and phospho-mammalian target of rapamycin (mTOR). Moreover, HIF-1α expression may be regulated through the inhibition of the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt)/mTOR signaling pathway. Thus, the present results suggest that bufalin induces cell cycle arrest and suppresses metastasis; this process may be associated with the PI3K/Akt/mTOR signaling pathway. Accordingly, it is suggested that bufalin is a therapeutic agent for RCC.

The management of castration-resistant prostate cancer (CRPC) is challenging, attributable to a lack of efficacious therapies. Chemotherapy is one of the most important treatments for CRPC. Doxorubicin has been extensively used in many different tumors and is often combined with other drugs to enhance effects and reduce toxicity. Costunolide is a natural sesquiterpene lactone with anti-cancer properties. In this study, we first demonstrated that the combination of costunolide and doxorubicin induced apoptosis significantly more than either drug alone in prostate cancer cell lines. Costunolide combined with doxorubicin induced mitochondria-mediated apoptosis through a loss of mitochondrial membrane potential and modulation of Bcl-2 family proteins. We found that this drug combination significantly increased the production of reactive oxygen species (ROS), as well as phosphorylation of c-jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinases, which play upstream roles in mitochondria-mediated apoptosis. Further studies showed that N-acetyl cysteine blocked JNK and p38 phosphorylation, suggesting that ROS were upstream activators of JNK and p38. However, a JNK inhibitor, but not a p38 inhibitor, blocked the increase in ROS observed in cells treated with a combination of costunolide and doxorubicin, suggesting that ROS and JNK could activate each other. In vivo, inhibition of tumor growth and induction of apoptosis were greater in mice treated with the costunolide and doxorubicin combination than in mice treated with either drug alone, without an increase in toxicity. Therefore, we suggested that costunolide in combination with doxorubicin was a new potential chemotherapeutic strategy for treating prostate cancer.