Local administration of dendritic cells inhibits established breast tumor growth: implications for apoptosis-inducing agents

Dendritic cells (DCs) can efficiently acquire foreign antigen(s) from apoptotic cells and induce MHC class I-restricted, antigen-specific CTLs. An accumulation of DCs within solid tumor masses in situ has been associated indirectly with a more favorable prognosis. Therefore, DCs may offer an efficient means for triggering immune responses within tumors, particularly in those masses containing significant apoptosis. We examined whether delivery of DCs could, alone, impact on the progressive growth of a tumor with a relatively high apoptotic index. We detected significant early apoptosis within the mass of a s.c. growing murine MT-901 breast carcinoma. DCs could efficiently engulf MT-901 tumor apoptotic cells in vitro. Intratumoral injections of syngeneic but not allogeneic DCs resulted in significant inhibition of MT-901 tumor growth. Histological examination of the tumor revealed intense mononuclear cell infiltration during and after DC injections. Tumor growth inhibition was relatively radiosensitive and dependent on host-derived CD8+ T cells. The baseline level of tumor apoptosis could be increased substantially by tumor necrosis factor alpha administration, leading to a greater DC-mediated antitumor effect. The antitumor effect could also be enhanced by first pulsing DCs with the foreign helper protein, keyhole limpet hemocyanin, prior to intratumoral delivery and combining it with the systemic administration of interleukin 2. Splenocytes from treated animals showed heightened levels of specific CTL activity and production of cytokines. The level of in situ tumor apoptosis appears to play a critical role in DC-mediated antitumor effects. The potential implication of these findings in DC-based tumor therapy strategies is discussed.     

Enhanced antiangiogenic therapy of squamous cell carcinoma by combined endostatin and epidermal growth factor receptor-antisense therapy.

PURPOSE: We tested the combined effects of antiangiogenic endostatin and epidermal growth factor receptor (EGFR) antisense gene therapy on squamous cell carcinoma (SCC). EXPERIMENTAL DESIGN and Results: The 1483 cell line of human head and neck SCC (HNSCC) and SCC-VII/SF murine SCC cells was used to establish tumors in nude mice and immunocompetent C3H mice, respectively. Tumor-bearing mice were treated with endostatin (20 mg/kg/day, s.c.), liposomal EGFR-antisense expression plasmid (25 microg/mouse, three times/week, intratumoral), a combination of both agents, or liposomal EGFR-sense plasmid as a control. Endostatin or EGFR-antisense alone significantly, yet partially, inhibited the growth of 1483 and SCC-VII/SF tumors, and a combination of both treatments completely blocked tumor growth. Immunohistochemistry analysis demonstrated that a complete suppression of tumor angiogenesis was achieved by the combination treatment. Down-regulation of vascular endothelial growth factor was shown in EGFR-antisense-treated tumors. These results suggest that the EGFR-antisense treatment, in addition to its inhibitory activity on tumor cell proliferation, might have a synergistic effect with endostatin on SCC-induced angiogenesis. In vitro studies demonstrated that EGFR inhibition by antisense oligonucleotides or EGFR-specific tyrosine kinase inhibitor down-regulated the production of VEGF in HNSCC cells. Additional experiments demonstrated that these EGFR inhibition approaches also directly suppressed the growth of endothelial cells. CONCLUSION: A combination of endostatin and EGFR targeting strategies profoundly inhibited the angiogenesis and growth of SCC in vivo. EGFR-antisense therapy might have multiple inhibitory effects against both tumor cells and endothelial cells, leading to enhanced antitumor efficacy. Such a combination strategy might represent a novel and promising approach for HNSCC therapy.     

Enhancement of antitumor effect by intratumoral administration of bax gene in combination with anticancer drugs in gastric cancer

Proapototic gene is an important target to enhance the chemotherapeutic effect in cancer cells. Based on in vitro study showing that the introduction of bax gene enhanced the sensitivity to anticancer drugs, we examined whether the intratumoral administration of bax gene could enhance the anti-tumor effect in combination with anticancer drugs in gastric cancer. The human gastric cancer cell line, MKN45 was transplanted into nude mice, and the intratumoral administration of bax gene was performed using the bax cDNA plasmid complexed with a cationic lipopolyamine. The enhancement of antitumor effect was examined in the combination with 5-fluorouracil (5-FU) and cisplatin (CDDP). The anticancer drugs were administered intraperitoneally with one third of LD50 four times at 4-day intervals, and the antitumor effect was assessed by the NCI protocol. The expression of bax gene was analyzed by RT-PCR and the apoptotic cell death was assessed by TUNEL method. The intratumoral administration of bax gene alone showed slight anti-tumor effect as compared to that of control tumor injected with vector alone. The antitumor effect of 5-FU and CDDP was significantly enhanced in the combination with intra-tumoral administration of bax gene as compared to that of CDDP and 5-FU alone (p<0.05, Student's t-test). The enhancement of antitumor effect was associated with the constitutive overexpression of bax gene and with the induction of apoptosis in the tumor treated with anticancer drug and bax gene. These results indicate that the combination therapy of intratumoral administration of bax gene complexed with a cationic lipopolyamine and anticancer drugs may provide us a new strategy for cancer chemotherapy to enhance its therapeutic efficacy in gastric cancer as termed with gene-chemotherapy.     

Histone deacetylase inhibitors enhance the apoptotic activity of insulin-like growth factor binding protein-3 by blocking PKC-induced IGFBP-3 degradation

Overexpression of insulin-like growth factor binding protein (IGFBP)-3 induces apoptosis of cancer cells. However, preexisting resistance to IGFBP-3 could limit its antitumor activities. This study characterizes the efficacy and mechanism of the combination of recombinant IGFBP-3 (rIGFBP-3) and HDAC inhibitors to overcome IGFBP-3 resistance in a subset of non-small cell lung cancer (NSCLC) and head and neck squamous cell carcinoma (HNSCC) cells. The effects of the combination of rIGFBP-3 and a number of HDAC inhibitors on cell proliferation and apoptosis were assessed in vitro and in vivo by using the MTT assay, a flow cytometry-based TUNEL assay, Western blot analyses and the NSCLC xenograft tumor model. Combined treatment with HDAC inhibitors and rIGFBP-3 had synergistic antiproliferative effects accompanied by increased apoptosis rates in a subset of NSCLC and HNSCC cell lines in vitro. Moreover, combined treatment with depsipeptide and rIGFBP-3 completely suppressed tumor growth and increased the apoptosis rate in vivo in H1299 NSCLC xenografts. Evidence suggests that HDAC inhibitors increased the half-life of rIGFBP-3 protein by blocking protein kinase C (PKC)-mediated phosphorylation and degradation of rIGFBP-3. In addition, combined treatment of IGFBP-3 with an HDAC inhibitor facilitates apoptosis through upregulation of rIGFBP-3 stability and Akt signaling inhibition. The ability of HDAC inhibitors to decrease PKC activation may enhance apoptotic activities of rIGFBP-3 in NSCLC cells in vitro and in vivo. These results indicated that combined treatment with HDAC inhibitor and rIGFBP-3 could be an effective treatment strategy for NSCLC and HNSCC with highly activated PKC.     

Small interfering RNA targeting epidermal growth factor receptor enhances chemosensitivity to cisplatin, 5-fluorouracil and docetaxel in head and neck squamous cell carcinoma

Overexpression of epidermal growth factor receptor (EGFR) has been found in various epithelial malignancies, including head and neck squamous cell carcinoma (HNSCC), and is associated with increased tumor growth, metastasis, resistance to chemotherapeutic agents and poor prognosis. As such, EGFR is a potential target for antitumor therapy and several EGFR inhibitors have been investigated in preclinical or clinical settings. In the present study, we used small interfering RNA (siRNA) to downregulate EGFR expression while evaluating the effect of EGFR siRNA on cell proliferation, and the combined effects with cisplatin, 5-fluorouracil (5-FU) and docetaxel in HNSCC. Furthermore, HNSCC xenografts were treated with EGFR siRNA alone or in combination with cisplatin, and tumor growth was examined. EGFR expression, proliferation, angiogenesis and apoptosis index were evaluated by immunohistochemistry. The results showed that EGFR siRNA efficiently downregulated EGFR expression and inhibited cell growth of HNSCC. Treatment with EGFR siRNA in combination with cisplatin, 5-FU and docetaxel enhanced chemosensitivity with a significant increase in apoptosis. EGFR siRNA delivered by atelocollagen enhanced the antitumor effect of cisplatin in the HNSCC xenograft model. These cumulative results suggest that EGFR siRNA combined with cisplatin, 5-FU and docetaxel may be a feasible strategy to enhance the effects of chemotherapy in patients with HNSCC.     

Insulin-like growth factor-I receptor signaling pathway induces resistance to the apoptotic activities of SCH66336 (lonafarnib) through Akt/mammalian target of rapamycin-mediated increases in survivin expression.

Although preclinical studies have suggested that farnesyltransferase inhibitors (FTI) have promising antitumor activity, clinical trials have shown that FTI activity in patients is actually limited. The mechanism that induces resistance to FTI treatment is still not fully understood. The FTI SCH66336 has been shown to induce apoptotic and antiangiogenic activities in a subset of head and neck squamous cell carcinoma (HNSCC) and non-small cell lung cancer (NSCLC) cell lines. We therefore investigated the mechanisms mediating resistance to the therapeutic activities of SCH66336 in HNSCC and NSCLC. Our various analyses showed that insulin-like growth factor-I receptor (IGF-IR) activation interferes with the antitumor activity of SCH66336 in HNSCC and NSCLC cells. Treatment with SCH66336 activated the IGF-IR/phosphatidylinositol 3-kinase/Akt pathway, leading to increased mammalian target of rapamycin (mTOR)-mediated protein synthesis of survivin in a subset of HNSCC and NSCLC cell lines that were insensitive to the apoptotic activities of the drug. Inhibition of IGF-IR, Akt, or mTOR or the knockdown of survivin expression abolished resistance to SCH66336 and induced apoptosis in the cells. Overexpression of survivin by the use of adenoviral vector protected SCH66336-sensitive HNSCC cells from the apoptotic activities of the drug. Our results suggest that expression of phosphorylated IGF-IR, phosphorylated Akt, phosphorylated mTOR, and survivin serves as biological markers of SCH66336 responsiveness in HNSCC and NSCLC cells and that SCH66336 induces survivin expression through an IGF-IR/Akt/mTOR-dependent pathway. Thus, combining inhibitors of IGF-IR, phosphatidylinositol 3-kinase/Akt, mTOR, or survivin with SCH66336 may be an effective anticancer therapeutic strategy for patients with HNSCC or NSCLC.     

Hepatocyte growth factor sensitizes human ovarian carcinoma cell lines to paclitaxel and cisplatin

The hepatocyte growth factor (HGF) receptor, encoded by the MET oncogene, is expressed in approximately 70% of human ovarian carcinomas and overexpressed in 30% of cases. Because HGF is known to protect cells from apoptosis, we investigated whether receptor expression modifies ovarian cancer cell response to chemotherapy. The apoptotic effect of the front-line chemotherapeutic drugs paclitaxel and cisplatin on cells treated with HGF was studied. In ovarian cancer cell lines, pretreatment with HGF surprisingly enhances the apoptotic response to low doses of paclitaxel and cisplatin. HGF empowers specifically the intrinsic apoptotic pathway, whereas it protects cells from extrinsic Fas-induced apoptosis. Chemotherapy sensitization is specific for HGF because another growth factor (e.g., epidermal growth factor) increases ovarian cancer cell survival. In nonovarian cancer cell models, as expected, HGF provides protection from drug-induced apoptosis. These data show that HGF sensitizes ovarian carcinoma cells to low-dose chemotherapeutic agents. This suggests that HGF may be used to improve response to chemotherapy in a set of human ovarian carcinomas molecularly classified based on the MET oncogene expression.     

Enhancing antimelanoma immune responses through apoptosis

We examined the feasibility of using tumor apoptosis at accessible sites to enhance antimelanoma immune responses in a model of spontaneous canine melanoma. We show that priming peripheral blood mononuclear cells with apoptotic melanoma cells significantly enhanced autologous and allogeneic lymphokine-activated killing of tumor cells. Since various pathways required for intrinsic apoptosis are often inactivated in melanoma, we used Fas ligand (FasL) overexpression to promote extrinsic apoptosis. FasL induced apoptosis in five of six cell lines. Each of the susceptible lines, but not the resistant one, expressed Fas mRNA. In addition, direct intratumoral administration of FasL DNA to tumor-bearing dogs was safe, with no adverse events reported over 7 days of observation. A reduction of tumor burden was seen in three of five dogs treated. The reduction of tumor volume was correlated with Fas expression by the tumors, although one dog with a Fas-negative tumor survived for 82 weeks after treatment. Our data show that overexpression of FasL is suitable to promote apoptosis of Fas(+) melanomas, and support the notion that priming immune responder cells with apoptotic tumor cells may enhance antitumor responses. The results also suggest that intratumoral administration of FasL offers a safe route for therapeutic gene delivery.     

A role for p38 MAPK in head and neck cancer cell growth and tumor-induced angiogenesis and lymphangiogenesis

We have recently gained a remarkable understanding of the mutational landscape of head and neck squamous cell carcinoma (HNSCC). However, the nature of the dysregulated signaling networks contributing to HNSCC progression is still poorly defined. Here, we have focused on the role of the family of mitogen activated kinases (MAPKs), extracellular regulated kinase (ERK), c-Jun terminal kinase (JNK) and p38 MAPK in HNSCC. Immunohistochemical analysis of a large collection of human HNSCC tissues revealed that the levels of the phosphorylated active form of ERK1/2 and JNK were elevated in less than 33% and 16% of the cases, respectively. Strikingly, however, high levels of active phospho-p38 were observed in most (79%) of hundreds of tissues analyzed. We explored the biological role of p38 in HNSCC cell lines using three independent approaches: treatment with a specific p38 inhibitor, SB203580; a retro-inhibition strategy consisting in the use of SB203580 combined with the expression of an inhibitor-insensitive mutant form of p38α; and short-hairpin RNAs (shRNAs) targeting p38α. We found that specific blockade of p38 signaling significantly inhibited the proliferation of HNSCC cells both in vitro and in vivo. Indeed, we observed that p38 inhibition in HNSCC cancer cells reduces cancer growth in tumor xenografts and a remarkable decrease in intratumoral blood and lymphatic vessels. We conclude that p38α functions as a positive regulator of HNSCC in the context of the tumor microenvironment, controlling cancer cell growth as well as tumor-induced angiogenesis and lymphangiogenesis.     

Honokiol inhibits the growth of head and neck squamous cell carcinoma by targeting epidermal growth factor receptor

Here, we report the chemotherapeutic effect of honokiol, a phytochemical from Magnolia plant, on human head and neck squamous cell carcinoma (HNSCC). Treatment of HNSCC cell lines from different sub-sites, SCC-1 (oral cavity), SCC-5 (larynx), OSC-19 (tongue) and FaDu (pharynx) with honokiol inhibited their cell viability, which was associated with the: (i) induction of apoptosis, (ii) correction of dysregulatory cell cycle proteins of G0/G1 phase. Honokiol decreased the expression levels of epidermal growth factor receptor (EGFR), mTOR and their downstream signaling molecules. Treatment of FaDu and SCC-1 cell lines with rapamycin, an inhibitor of mTOR pathway, also reduced cell viability of HNSCC cells. Administration of honokiol by oral gavage (100 mg/kg body weight) significantly (P < 0.01-0.001) inhibited the growth of SCC-1 and FaDu xenografts in athymic nude mice, which was associated with: (i) inhibition of tumor cell proliferation, (ii) induction of apoptosis, (iii) reduced expressions of cyclins and Cdks, and (iv) inhibition of EGFR signaling pathway. Molecular docking analysis of honokiol in EGFR binding site indicated that the chemotherapeutic effect of honokiol against HNSCC is mediated through its firm binding with EGFR, which is better than that of gefitinib, a commonly used drug for HNSCC treatment.     

Antitumor effects of the tumor necrosis factor (PAC-4D) against human gynecological carcinoma transplanted into nude mice

The antitumor effects of human recombinant TNF (PAC-4D) were examined on three human gynecological carcinomas transplanted into CD-1 nude mice (uterine cervical carcinoma: UZ-1-N; ovarian carcinoma: OCl-1-N and OS-4-N). PAC-4D was administered intratumorally at a dose of 1,000 U, 3,000 U or 10,000 U/head from day 0 to day 4. Tumor size, body weight and peripheral WBC were measured on days 0, 4, 8, 12 and 16 and histological studies were made on day 6. The results were as follows: With UZ-1-N, intratumoral administration of PAC-4D at doses of 3,000 U and 10,000 U/head caused a marked inhibition of the tumor growth. Similarly, antitumor activity of PAC-4D against OCl-1-N was remarkable at a dose of 10,000 U/head. The administration of PAC-4D at doses of 3,000 U and 10,000 U/head was not effective against OS-4-N. The histological changes of grade II A-B by Ohboshi-Shimosato criteria of response were observed in the tumor of the effective groups against PAC-4D. There were no influences on the body weight and WBC after administration of PAC-4D. Although tumor cells possessed different sensitivities to PAC-4D, PAC-4D is strongly recommended for the treatment of gynecological malignancies.     

Synergistic antitumor activity of epidermal growth factor receptor tyrosine kinase inhibitor gefitinib and IFN-alpha in head and neck cancer cells in vitro and in vivo.

PURPOSE: Epidermal growth factor receptor (EGFR) overexpression has been implicated in the development of head and neck squamous cell carcinomas (HNSCC) and represents a potential therapeutic target for this disease. We have reported previously that growth inhibitory concentrations of IFN-alpha enhance the expression and activity of EGFR and that this effect could represent an escape mechanism to the growth inhibition and apoptotic cell death induced by IFN-alpha. In this study, we investigate whether the combination of IFN-alpha and gefitinib (Iressa, AstraZeneca Pharmaceuticals, Macclesfield, United Kingdom), a selective EGFR tyrosine kinase inhibitor, might have a cooperative antitumor effect on HNSCC-derived cell lines. EXPERIMENTAL DESIGN: The interaction of IFN-alpha and gefitinib was evaluated in vitro on HNSCC-derived cell lines by median drug effect analysis calculating a combination index with CalcuSyn software and in vivo by using HNSCC xenografts in nude mice. The mechanism of gefitinib and IFN-alpha interactions was also studied by analysis of cell cycle kinetics, apoptosis assays, and Western blotting of EGFR signal transduction components. RESULTS: Simultaneous exposure to gefitinib and IFN-alpha produced synergistic antiproliferative and proapoptotic effects compared with single drug treatment. Furthermore, daily treatment of gefitinib (50 mg/kg p.o.) in combination with an IFN-alpha regimen (50,000 units s.c. three times weekly) induced tumor growth delay and increased survival rate on established HNSCC xenografts in nude mice. Moreover, the concomitant treatment with gefitinib suppressed the stimulation of extracellular signal-regulated kinase phosphorylation/activity induced by IFN-alpha both in vitro and in vivo. CONCLUSION: The observed cooperative antitumor effects could be, at least in part, explained by the inhibition exerted by gefitinib of an IFN-alpha-induced EGF-dependent survival pathway, which involves extracellular signal-regulated kinase activation. These results provide a rationale for the clinical evaluation of gefitinib in combination with IFN-alpha in HNSCC.     

Antitumor effect of intratumoral administration of bone marrow-derived dendritic cells transduced with wild-type p53 gene.

PURPOSE: Dendritic cells (DCs) are attractive effectors for cancer immunotherapy because of their potential to function as professional antigen-presenting cells for initiating cellular immune responses. The tumor suppressor gene p53 is pivotal in the regulation of apoptosis, and approximately 50% of human malignancies exhibit mutation and aberrant expression of p53. We investigated the antitumor effect of intratumoral administration of bone marrow-derived dendritic cells transduced with wild-type p53 gene. EXPERIMENTAL DESIGN: We examined whether intratumoral administration of DCs infected with recombinant adenovirus expressing murine wild-type p53 (Ad-mp53) could induce systemic antitumor responses against mutant p53-expressing tumors, highly immunogenic MethA, or weakly immunogenic MCA-207 implanted in syngeneic mice. RESULTS: Accumulation of wild-type p53 protein in bone marrow-derived murine DCs could be successfully achieved by Ad-mp53 infection. Treatment with intratumoral injection of Ad-mp53-transduced DCs caused a marked reduction in the in vivo growth of established MethA and MCA-207 tumors with massive cellular infiltrates. Administration of p53-expressing DCs suppressed the growth of both injected MCA-207 tumors and untreated distant MCA-207 tumors, but not unrelated Lewis lung carcinoma tumors, suggesting the augmentation of systemic immunogenicity against MCA-207 tumor cells. Moreover, intratumoral injection of p53-expressing DCs had a greater antitumor effect than did s.c. immunization. CONCLUSIONS: Our results indicate that intratumoral administration of DCs expressing murine wild-type p53 leads to significant systemic immune responses and potent antitumor effects in mutant p53-expressing murine cancer models. These findings raise the possibility of using this strategy of intratumoral injection of p53-expressing DCs for human cancer treatment.     

Limited sampling models for CPT-11, SN-38, and SN-38 glucuronide

PURPOSE: To determine the ability of WMC26, a prototypic bisimidazoacridone (BIA), to induce apoptosis in sensitive colon adenocarcinoma cells and to advance the hypothesis that cancer cells that are growth-arrested by WMC26 are predisposed to undergo apoptotic death by abrogators of cell cycle checkpoints. METHODS: The antiproliferative activity of WMC26 was examined in detail by a 4-day MTT assay, cell counting, BrdU incorporation and a two-color LIVE/DEAD assay. To detect apoptosis a number of established techniques were used, including gel electrophoresis, flow cytometry, and confocal laser microscopy of treated cells. The activity of senescence-associated beta-galactosidase in treated cells was also analyzed. RESULTS: WMC26, at physiological concentrations, induced complete and longlasting growth arrest of HCT116 cells in culture but did not trigger cell death. The growth-arrested cells (blocked at G1 and G2/M cell cycle checkpoints) did not synthesize DNA but were metabolically active and had intact plasma membranes. Although they resembled the senescence-like phenotype reported to be induced by treatment with some antitumor agents, the cells did not express senescence-associated beta-galactosidase, an indicator of the senescence-like state. Treatment of WMC26 growth-arrested cells with 1 microM UCN-01, an abrogator of the G2/M checkpoint, caused a very rapid (1 h) change in morphology and cell death within 72 h. CONCLUSIONS: BIAs do not induce apoptosis in sensitive colon tumor cells. They are highly cytostatic but only marginally toxic to the cells even at concentrations 100-fold higher than those sufficient for complete growth arrest. In this respect WMC26 differs from some other DNA-interacting antitumor agents that produce cell growth arrest at low concentrations but are toxic at higher doses. The complete growth arrest induced by WMC26 in colon cancer cells sensitized them to apoptotic death induced by UCN-01. This finding suggests that a combination of WMC26 and cyclin-dependent kinase inhibitors may be an attractive treatment method for colon cancer that utilizes the highly tumor-selective activity of WMC26.     

Roles of JNK, p38 and ERK mitogen-activated protein kinases in the growth inhibition and apoptosis induced by cadmium

Cadmium (Cd), a human carcinogen, can induce apoptosis in various cell types. Three major mitogen-activated protein kinases (MAPKs), c-JUN N-terminal kinase (JNK), p38 and extracellular signal-regulated kinase (ERK), have been shown to regulate apoptosis. In this study we explore the ability of Cd to activate JNK, p38 and ERK, including their effects on Cd-mediated growth inhibition and apoptosis in a human non-small cell lung carcinoma cell line, CL3. The kinase activity of JNK was induced dose-dependently by 30-160 microM CdCl(2). High cytotoxic doses of Cd (130-160 microM) markedly activated p38, but low Cd doses did not. Conversely, the activities of ERK1 and ERK2 were decreased by low cytotoxic doses of Cd (相似文献   

Honokiol, a constituent of oriental medicinal herb magnolia officinalis, inhibits growth of PC-3 xenografts in vivo in association with apoptosis induction.

PURPOSE: This study was undertaken to determine the efficacy of honokiol, a constituent of oriental medicinal herb Magnolia officinalis, against human prostate cancer cells in culture and in vivo. EXPERIMENTAL DESIGN: Honokiol-mediated apoptosis was assessed by analysis of cytoplasmic histone-associated DNA fragmentation. Knockdown of Bax and Bak proteins was achieved by transient transfection using siRNA. Honokiol was administered by oral gavage to male nude mice s.c. implanted with PC-3 cells. Tumor sections from control and honokiol-treated mice were examined for apoptotic bodies (terminal deoxyribonucleotidyl transferase-mediated dUTP nick end labeling assay), proliferation index (proliferating cell nuclear antigen staining), and neovascularization (CD31 staining). Levels of Bcl-2 family proteins in cell lysates and tumor supernatants were determined by immunoblotting. RESULTS: Exposure of human prostate cancer cells (PC-3, LNCaP, and C4-2) to honokiol resulted in apoptotic DNA fragmentation in a concentration- and time-dependent manner irrespective of their androgen responsiveness or p53 status. Honokiol-induced apoptosis correlated with induction of Bax, Bak, and Bad and a decrease in Bcl-xL and Mcl-1 protein levels. Transient transfection of PC-3 cells with Bak- and Bax-targeted siRNAs and Bcl-xL plasmid conferred partial yet significant protection against honokiol-induced apoptosis. Oral gavage of 2 mg honokiol/mouse (thrice a week) significantly retarded growth of PC-3 xenografts without causing weight loss. Tumors from honokiol-treated mice exhibited markedly higher count of apoptotic bodies and reduced proliferation index and neovascularization compared with control tumors. CONCLUSION: Our data suggest that honokiol, which is used in traditional oriental medicine for the treatment of various ailments, may be an attractive agent for treatment and/or prevention of human prostate cancers.