Efficient dual treatment of the hormone-refractory prostate cancer cell line DU145 with cetuximab and 1,25-dihydroxyvitamin D3

BACKGROUND: Targeting of the epidermal growth factor receptor (EGFR) pathway is a promising treatment strategy for aggressive androgen-refractory prostate cancer (PCa). The effect of treating the androgen-resistant PCa cell line DU145 with a combination of the anti-EGFR drug cetuximab and 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) was evaluated. MATERIALS AND METHODS: DU145 cells were treated with 5 nM cetuximab, 100 nM 1,25(OH)2D3 or a combination of both. The effect of the treatments on cell growth, cell-cycle and apoptosis was evaluated. RESULTS: Single-drug treatments decreased DU145 cell growth by up to 25% and caused a 1.5-to 1.7-fold increase of apoptosis, but did not affect the cell-cycle distribution. However, dual treatment with a combination of cetuximab and 1,25(OH)2D3 inhibited DU145 cell proliferation by 40%, caused considerable cell-cycle arrest in the Go/Gl-phase, and enhanced apoptosis by 2.5-fold (compared to the control, p < 0. 0001, p <0. 006 and p <0. 0001, respectively). CONCLUSION: A combination of cetuximab and 1,25(OH)2D3 efficiently suppresses hormone-resistant PCa cell growth and could provide a basis for its clinical application.     

ZD1839: targeting the epidermal growth factor receptor in cancer therapy

The erbB family of receptors, which includes the epidermal growth factor receptor, has been widely implicated in promoting proliferation of malignant cells. The critical role played by epidermal growth factor receptor in cancer has resulted in extensive research for selective inhibitors of the epidermal growth factor receptor signalling pathway. Selective small molecule epidermal growth factor receptor-tyrosine kinase inhibitors, such as ZD1839 (Iressa^?), block signal transduction pathways implicated in proliferation and survival of cancer cells and other host-dependent processes promoting cancer cell growth. In preclinical studies, ZD1839, alone and in combination with other agents, has demonstrated antitumour activity in a range of tumour types. Results from Phase I trials, in healthy volunteers and in patients with advanced disease, have shown that ZD1839 has excellent bioavailability and an acceptable tolerability profile. In these studies, ZD1839 has also shown promising clinical activity in patients with a variety of tumour types. Furthermore, Phase II studies confirmed clinically meaningful antitumour activity and have demonstrated symptom relief in the second- and third-line treatment of non-small cell lung cancer. Phase III trials are currently evaluating ZD1839 in combination with gemcitabine/cisplatin or paclitaxel/carboplatin as first-line treatment of non-small cell lung cancer and an ongoing clinical trial programme is investigating other tumours (i.e., head and neck, prostate, colon and breast) and other combinations. This article provides an overview of the current profile of ZD1839.     

Epidermal growth factor receptor inhibitors: a new prospective in the treatment of lung cancer

Lung cancer is the leading cause of death worldwide. Current treatment modalities, including chemotherapy, radiotherapy and surgery, provide only limited improvement in the natural course of this disease. Therefore, the development of new therapeutic strategies is highly awaited. This review focuses on recent achievements on a novel class of anticancer drugs targeting the EGFR (Epidermal Growth Factor Receptor). The EGFR family is a group of four structurally similar growth factor receptors with tyrosine-kinase activity (EGFR, HER2/neu, ErbB-3, ErbB-4), which dimerize upon binding with a number of ligands, including EGF (Epidermal Growth Factor) and TGF (Transforming Growth Factor), allowing downstream transduction of mitogenic signals. Overexpression of EGFR and HER2 is frequently found in non-small-cell lung cancer (NSCLC), which accounts for over 80% of all malignant lung tumors, and has been associated with a worse clinical outcome. New agents developed to inhibit EGFR function include monoclonal antibodies and small-molecule receptor tyrosine-kinase inhibitors. In this review, results of most recent clinical with EGFR inhibitors including monoclonal antibodies, such as Trastuzumab (Herceptin), IMC-C225 (Cetuximab) and others (ABX-EGF, EMD 72000), and tyrosine-kinase inhibitors, such as ZD1839 (Gefitinib, Iressa), OSI-774 (Erlotinib, Tarceva) and others (CI-1033, GW2016), are summarized. In particular, final results of phase II (IDEAL 1 and 2) and III (INTACT 1 and 2) studies of ZD1839 are reported. In IDEAL trials (ZD1839 single agent in patients pre-treated with chemotherapy) there was clear evidence of tumor regression, symptoms improvement and overall clinical benefit, whereas in the two INTACT trials (ZD1839 in combination with standard platinum-based chemotherapy in chemo-naive patients) ZD1839 did not improve either survival or other clinical endpoints. Possible explanations for these contradictory results and future perspectives are discussed.     

ZD1839: targeting the epidermal growth factor receptor in cancer therapy

The erbB family of receptors, which includes the epidermal growth factor receptor, has been widely implicated in promoting proliferation of malignant cells. The critical role played by epidermal growth factor receptor in cancer has resulted in extensive research for selective inhibitors of the epidermal growth factor receptor signalling pathway. Selective small molecule epidermal growth factor receptor-tyrosine kinase inhibitors, such as ZD1839 (Iressa), block signal transduction pathways implicated in proliferation and survival of cancer cells and other host-dependent processes promoting cancer cell growth. In preclinical studies, ZD1839, alone and in combination with other agents, has demonstrated antitumour activity in a range of tumour types. Results from Phase I trials, in healthy volunteers and in patients with advanced disease, have shown that ZD1839 has excellent bioavailability and an acceptable tolerability profile. In these studies, ZD1839 has also shown promising clinical activity in patients with a variety of tumour types. Furthermore, Phase II studies confirmed clinically meaningful antitumour activity and have demonstrated symptom relief in the second- and third-line treatment of non-small cell lung cancer. Phase III trials are currently evaluating ZD1839 in combination with gemcitabine/cisplatin or paclitaxel/carboplatin as first-line treatment of non-small cell lung cancer and an ongoing clinical trial programme is investigating other tumours (i.e., head and neck, prostate, colon and breast) and other combinations. This article provides an overview of the current profile of ZD1839.     

Characterization of sequence-dependent synergy between ZD1839 ("Iressa") and oxaliplatin

ZD1839 ("Iressa"), a selective epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI), is currently undergoing preclinical and clinical evaluation in several solid tumors. The present study aimed to assess the effect of ZD1839 in combination with oxaliplatin in the colon cancer cell lines HT-29 and LoVo. For in vitro chemosensitivity testing, cells were treated with serial dilutions of each drug sequentially at a fixed ratio of doses that corresponded to 1/20, 1/10, 1/5, 1/2, 1, 1.5 and 2 times the individual IC(50) values. Oxaliplatin followed by ZD1839 produced a synergistic effect. In contrast, oxaliplatin following ZD1839 exhibited an additive effect at best. Mass spectrometry examination revealed that ZD1839 modestly enhanced cellular oxaliplatin accumulation and platinum-DNA (Pt-DNA) adducts (P>0.05). In additional studies, high-performance liquid chromatography revealed that oxaliplatin had no effect on ZD1839 accumulation. In contrast, ZD1839 markedly inhibited removal of Pt-DNA adducts (P<0.05). With oxaliplatin treatment (1 day) followed by ZD1839 (1 day), then incubation with drug-free medium (1 day), 90% of Pt-DNA adducts remained. Apoptosis examination revealed that oxaliplatin-induced apoptosis was prolonged by sequential oxaliplatin followed by ZD1839 treatment compared with oxaliplatin alone. Further experiments revealed that ZD1839 decreased cellular gamma-glutamyltransferase activity.CONCLUSIONS: The above observations provide a mechanistic explanation for the synergy of oxaliplatin followed by ZD1839, and suggest that this treatment combination warrants further preclinical and clinical investigation.     

Molecular mechanisms of ZD1839-induced G1-cell cycle arrest and apoptosis in human lung adenocarcinoma A549 cells

Epithelial growth factor receptor (EGFR) has been proposed as a target for anticancer therapy. ZD1839 (Iressa) is a quinazoline derivative that selectively inhibits the EGFR tyrosine kinase activity and is under clinical use in cancer patients. However, the molecular mechanisms involved in ZD1839-mediated anticancer effects remain largely uncharacterized. In this study, exposure of human lung adenocarcinoma A549 cells to ZD1839 caused G1 arrest, and subsequently induced apoptosis. Moreover, ZD1839 increased the protein levels of p27(KIP1) and retinoblastoma-related Rb2/p130 while decreased the expression of cyclin-dependent kinase-2 (CDK2), CDK4, CDK6 and cyclin-D1, cyclin-D3. In vitro kinase assay showed that ZD1839 decreased these CDKs expression in A549 cells, leading to significantly reduce their kinase activities. In addition, ZD1839-induced death of A549 cells with characteristics of apoptosis including apoptotic morphological changes, DNA fragmentation and enhancement of TUNEL-positive cell. These events were accompanied by a marked increase of Fas protein expression, and activation of caspase-2, -3, -8. Co-treatment of cells with Fas antagonist antibody significantly blocked ZD1839-induced apoptosis. Caspase-8 and caspase-3 inhibitors, but not a caspase-9 inhibitor, were also capable of restoring cell viability. Our results indicate that downregulation of the expression and function of CDK2, CDK4, CDK6, cyclin-D1 and cyclin-D3, as well as upregulation of p27(KIP1) and pRb2/p130, are strong candidates for the cell cycle regulator that arrests ZD1839-treated A549 cells at G1 phase. Furthermore, upregulation of Fas appears to play a major role in the initiation of ZD1839-induced apoptosis, activation of caspase-8/caspase-3 cascade is involved in the execution phase of this death program.     

Gallic Acid,an Active Constituent of Grape Seed Extract,Exhibits Anti-proliferative,Pro-apoptotic and Anti-tumorigenic Effects Against Prostate Carcinoma Xenograft Growth in Nude Mice

Purpose  Gallic acid, a natural agent present in a wide-range of fruits and vegetables, has been of potential interest as an anti-cancer agent; herein, we evaluated its efficacy in androgen-independent DU145 and androgen-dependent-22Rv1 human prostate cancer (PCa) cells. Materials and Methods  Cell viability was determined by MTT and apoptosis by Annexin V-PI assays. In vivo anti-cancer efficacy was assessed by DU145 and 22Rv1 xenograft growth in nude mice given normal drinking water or one supplemented with 0.3% or 1% (w/v) gallic acid. PCNA, TUNEL and CD31 immunostaining was performed in tumor tissues for in vivo anti-proliferative, apoptotic and anti-angiogenic effects of gallic acid. Results  Gallic acid decreased cell viability in a dose-dependent manner in both DU145 and 22Rv1 cells largely via apoptosis induction. In tumor studies, gallic acid feeding inhibited the growth of DU145 and 22Rv1 PCa xenografts in nude mice. Immunohistochemical analysis revealed significant inhibition of tumor cell proliferation, induction of apoptosis, and reduction of microvessel density in tumor xenografts from gallic acid-fed mice as compared to controls in both DU145 and 22Rv1 models. Conclusion  Taken together, our findings show the anti-PCa efficacy of gallic acid and provide a rationale for additional studies with this naturally-occurring agent for its efficacy against PCa.     

In vitro assessment of cytotoxic agent combinations for hormone-refractory prostate cancer treatment

We have investigated new drug combinations of potential clinical value for treatment of hormone-refractory prostate cancer. Combinations of paclitaxel, carboplatin and mitoxantrone, and combinations of these three drugs with compounds targeting important pathways for cancer progression, 13-cis-retinoic acid and chelerythrine, were assessed. The drugs combinations were incubated for 72 h in steroid-free conditions with two androgen-independent cell lines, DU145 and PC3. Cytotoxicity assay was performed using resazurin and Hoescht 33342. Synergism and antagonism were measured by the combination index, and calculated for each combination by the median-effect method. All six compounds exhibited cytotoxic effects when tested alone. Paclitaxel exhibited the highest and 13-cis-retinoic acid the lowest effect on both cell lines. Paclitaxel demonstrated synergism or additivity with 13-cis-retinoic acid in both cell lines, whereas antagonistic effects were observed when it was tested in combination with carboplatin. Chelerythrine showed additive effects with mitoxantrone in both cell lines and with paclitaxel in PC3 cells. Our results suggest that combination of paclitaxel and 13-cis-retinoic acid, and of chelerythrine with mitoxantrone and paclitaxel, may have clinical value for the treatment of hormone-refractory prostate cancer.     

Silibinin Impairs Constitutively Active TGFα-EGFR Autocrine Loop in Advanced Human Prostate Carcinoma Cells

PURPOSE: Epidermal growth factor (EGF) and transformation growth factor-alpha (TGFalpha) are potent mitogens that regulate proliferation of prostate cancer cells via autocrine and paracrine loops, and promote tumor metastasis. They exert their action through binding to the cell surface receptor, epidermal growth factor receptor (EGFR), and cause activation of Erk1/2 mediated mitogenic signaling in human prostate cancer (PCA) at both advanced and androgen-independent stages. Thus, we rationalized that inhibiting this mitogenic pathway could be useful in controlling advanced PCA growth. METHODS: LNCaP and DU145 human PCA cells were treated with silibinin (100-200 muM) for different time points, and the levels of TGFalpha, activated signaling molecules (EGFR, Erk1/2 and Jnk1/2) and Erk1/2 kinase activity were analyzed employing ELISA, immunoprecipitation and/or immunoblotting techniques. The mRNA levels of TGFalpha were analyzed by RT-PCR. RESULTS: Treatment of cells (LNCaP and DU145) with silibinin resulted in a decrease in TGFalpha protein at both secreted and cellular levels together with a decrease in its mRNA level. Silibinin also caused an inhibition of EGFR activation followed by that of Erk1/2 without any change in their protein levels. The kinase activity of Erk1/2 to Elk1 was also inhibited by silibinin at least in DU145 cells. In other study, silibinin caused strong inhibition of Jnk1/2 activation in LNCaP cells while in DU145 cells, a strong induction in Jnk1/2 activation was observed. These results suggest that silibinin impairs TGFalpha-EGFR-Erk1/2 signaling in both androgen-dependent (LNCaP) and -independent (DU145) advanced human prostate carcinoma cells. CONCLUSIONS: This study, for the first time, identifies the inhibitory effect of silibinin on constitutively active TGFalpha-EGFR autocrine loop in advanced human PCA cells, which plausible contributes to the strong efficacy of silibinin in PCA prevention and intervention, as reported in recent studies.     

Epidermal growth factor receptor tyrosine kinase inhibitors as anticancer agents

The epidermal growth factor receptor (EGFR)-driven autocrine growth pathway has been implicated in the development and progression of the majority of the most common human epithelial cancers, making the blockade of this growth pathway a promising anticancer therapeutic strategy. Different approaches have been developed to block EGFR activation and/or function in cancer cells. In the past 15 years, various anti-EGFR blocking monoclonal antibodies (MAb), recombinant proteins containing transforming growth factor-alpha (TGFalpha) or EGF fused to toxins, and tyrosine kinase inhibitors (TKIs) have been generated and their biological and potentially therapeutic properties characterised. One of these agents, MAb IMC-C225, a chimeric human-mouse IgG1 MAb, is the first anti-EGFR agent to enter phase II to III clinical trials in patients with cancer. Several small compounds that block the ligand-induced activation of the EGFR tyrosine kinase have been developed. Among these EGFR-TKIs, various quinazoline-derived agents have been synthesised and have shown promising activity as anticancer agents in preclinical models. ZD1839 ('Iressa'), an anilinoquinazoline, is an orally active, selective EGFR-TKI which is currently under clinical evaluation in phase II to III clinical trials in patients with cancer. Preclinical data for ZD1839 strongly support the possibility of potentiating the antitumour activity of conventional chemotherapy with agents that selectively block the EGFR.     

ZD1839 ('Iressa') as an anticancer agent

ZD1839 ('Iressa') is an orally active, selective epidermal growth factor receptor-tyrosine kinase inhibitor which blocks signal transduction pathways implicated in the proliferation and survival of cancer cells and other host-dependent processes promoting cancer growth. In preclinical studies, ZD1839 produced reversible growth inhibition and growth delay in a wide range of tumour cell lines and human tumour xenografts. Moreover, this activity was enhanced when ZD1839 was coadministered with cytotoxic agents. Preliminary results from phase I trials in patients with advanced disease and a wide variety of tumour types suggest that ZD1839 has an acceptable tolerability profile and promising clinical efficacy, particularly in non-small cell lung cancer (NSCLC). ZD1839 is currently in phase III clinical development for the treatment of advanced NSCLC. In addition, further trials are ongoing or planned in a number of other tumour types.     

Induction of remission in a patient with metastatic breast cancer refractory to trastuzumab and chemotherapy following treatment with gefitinib ('Iressa', ZD1839)

Despite new therapies and several treatment options, metastatic breast cancer (MBC) remains incurable. One reason for the low median survival rate may be intense cross-talk between growth factor receptors such as the epidermal growth factor receptor (EGFR/HER1) and the HER2 growth factor receptor. This report describes the case history of a patient with MBC whose disease had progressed despite surgery, radiotherapy and four different chemotherapy regimens, including trastuzumab (a monoclonal antibody that specifically blocks HER2) combined with docetaxel. However, treatment with 500 mg/day gefitinib ('Iressa', ZD1839), an EGFR tyrosine kinase inhibitor, and trastuzumab (2 mg/kg/week) caused a rapid and sustained regression of breast cancer metastases in skin and lymph nodes. Thus, for patients with MBC whose tumors co-express EGFR and HER2, gefitinib in combination with trastuzumab may prevent receptor cross-talk, improving the outcome of MBC.     

Enhanced suppression of prostate tumor growth by combining C-CAM1 gene therapy and angiogenesis inhibitor TNP-470

We have previously shown that C-CAM1-based gene therapy effectively suppressed prostate tumor growth in nude mice xenograft models. In this study, we examined the effects of combining C-CAM1-based therapy and TNP-470, a potent angiogenesis inhibitor, on prostate cancer in a xenografted tumor model. The direct cytotoxic effects of Ad-C-CAM1 (recombinant adenovirus containing C-CAM1 cDNA) and TNP-470 on DU145 cells in vitro were determined by microculture tetrazolium assay. The in vivo antitumor effects of either agent alone were studied in a DU145 xenografted tumor model. Cells were infected with Ad-C-CAM1 or the control virus at multiplicities of infection (m.o.i.) of 5 or 10 and then inoculated onto nude mice 48 h later. TNP-470 (0, 17 or 35 mg/kg) was given 15, 17 and 19 days after inoculation. Combined treatments in vivo were carried out to determine whether there were synergistic antitumor effects. Both Ad-C-CAM1 and the control virus were minimally toxic to DU145 in vitro. There was evident dose-dependent suppression of xenografted tumor growth by either Ad-C-CAM1 or TNP-470. By the median-effect analysis, combination of the two agents generated strong synergistic antitumor effects as shown by marked tumor suppression as compared to either treatment alone. The novel strategy may have clinical implications for the treatment of prostate cancer.     

Emerging drugs for non-small cell lung cancer

Lung cancer is the leading cause of cancer death in the world. Therapeutic improvements caused by recent cytotoxic agents seem to have reached a plateau. New therapeutic strategies are, therefore, necessary to improve the cure rate. These include receptor-targeted therapy, signal transduction or cell-cycle inhibition, angiogenesis inhibitors, cyclooxygenase-2 (COX-2) inhibitors, gene therapy and vaccines. The antiepidermal growth factor receptor (EGFR) group includes compounds acting on the extracellular domain of EGFR, such as IMC-C225 and trastuzumab; small molecules inhibiting EGFR phosphorylation, such as ZD 1839 and OSI-774; or compounds that interfere with one of the downstream steps, such as mitogen-activated protein kinase kinase (MEK) inhibitors. Farnesyl transferase inhibitors, such as SCH66336, and protein kinase C inhibitors, such as ISIS 3521, have also shown antitumour activity. Antiangiogenesis inhibitors include matrix metalloprotease inhibitors (MMPIs), such as marimastat, AG3340, BAY 12-9566, BMS-275291 and Col-3. Antiangiogenic agents offer great potential for the treatment of lung cancer, as shown in preclinical models, whereas emerging data suggest that there are limits to their use as monotherapy in advanced disease. Molecules targeting vascular endothelial growth factor (VEGF) or its receptor (VEGFR) also seem to control tumour progression and may prolong survival. COX-2 inhibitors are another class of agents currently under evaluation in clinical trials for their chemoprevention role in subjects at high lung cancer risk, and also in patients with non-small cell lung cancer (NSCLC) in combination with standard chemotherapeutics. Genetic and immunologic therapies represent two additional promising modalities. All of these therapies are in different phases of clinical testing and have shown encouraging activity alone or in combination with chemotherapy drugs.     

Pharmacokinetics and tolerability of the orally active selective epidermal growth factor receptor tyrosine kinase inhibitor ZD1839 in healthy volunteers

OBJECTIVE: To investigate the pharmacokinetics and tolerability of ZD1839 (Iressa), an orally active selective epidermal growth factor receptor-tyrosine kinase inhibitor, in healthy volunteers. DESIGN: Two randomised, double-blind, placebo-controlled, parallel-group studies of pharmacokinetics and tolerability, followed by a nonblind, randomised, 2-period crossover study to assess the effect of food on bioavailability. SETTING: Two centres in the UK. STUDY PARTICIPANTS: Healthy male volunteers aged between 18 and 62 years. INTERVENTIONS: The first study investigated the pharmacokinetics and tolerability of ascending single oral doses of ZD1839 (1 to 75mg). The second study investigated the pharmacokinetics and tolerability of multiple doses of ZD1839 (100mg once daily for 3 days). The third study investigated the effect of food on the bioavailability of a single 50mg dose of ZD1839. OUTCOME MEASURES AND RESULTS: Peak plasma drug concentrations (Cmax) of ZD 1839 occurred between 3 and 7 hours after administration. Cmax and area under the concentration-time curve (AUC) were dose-proportional from 10 to 100mg. The terminal elimination half-life (t1/2beta) was 28 hours (range 12 to 51 hours). Cmax was reduced by 34% and AUC by 14% by ingestion of food; t1/2beta was not affected. Urinary recovery of ZD1839 was <0.5%, indicating that this was not a major route of elimination. The pharmacokinetics of ZD1839 during administration of multiple doses could be predicted from day 1 values. There were no serious adverse events or withdrawals, and the frequency of adverse events was similar that with placebo. CONCLUSIONS: These data support the further clinical investigation of ZD 1839. The elimination half-life suggests that once daily oral administration is appropriate.     

Emerging drugs for non-small cell lung cancer

Lung cancer is the leading cause of cancer death in the world. Therapeutic improvements caused by recent cytotoxic agents seem to have reached a plateau. New therapeutic strategies are, therefore, necessary to improve the cure rate. These include receptor-targeted therapy, signal transduction or cell-cycle inhibition, angiogenesis inhibitors, cyclooxygenase-2 (COX-2) inhibitors, gene therapy and vaccines. The antiepidermal growth factor receptor (EGFR) group includes compounds acting on the extracellular domain of EGFR, such as IMC-C225 and trastuzumab; small molecules inhibiting EGFR phosphorylation, such as ZD 1839 and OSI-774; or compounds that interfere with one of the downstream steps, such as mitogen-activated protein kinase kinase (MEK) inhibitors. Farnesyl transferase inhibitors, such as SCH66336, and protein kinase C inhibitors, such as ISIS 3521, have also shown antitumour activity. Antiangiogenesis inhibitors include matrix metalloprotease inhibitors (MMPIs), suchs as marimastat, AG3340, BAY 12-9566, BMS-275291 and Col-3. Antiangiogenic agents offer great potential for the treatment of lung cancer, as shown in preclinical models, whereas emerging data suggest that there are limits to their use as monotherapy in advanced disease. Molecules targeting vascular endothelial growth factor (VEGF) or its receptor (VEGFR) also seem to control tumour progression and may prolong survival. COX-2 inhibitors are another class of agents currently under evaluation in clinical trials for their chemoprevention role in subjects at high lung cancer risk, and also in patients with non-small cell lung cancer (NSCLC) in combination with standard chemotherapeutics. Genetic and immunologic therapies represent two additional promising modalities. All of these therapies are in different phases of clinical testing and have shown encouraging activity alone or in combination with chemotherapy drugs.     

Epidermal growth factor receptor tyrosine kinase inhibitors in late stage clinical trials

The epidermal growth factor receptor (EGFR) is a cell membrane receptor that plays a key role in cancer development and progression. Ligand-activated EGFR-dependent signalling is involved in cell proliferation, apoptosis, angiogenesis and metastatic spread. Targeting the EGFR, therefore, represents a promising molecular approach in cancer treatment. Several anti-EGFR agents are in clinical development. Three drugs are currently in Phase II and III development as single agents, or in combination with other anticancer modalities: IMC-225 (cetuximab/Erbitux; ImClone), a chimaeric human-mouse monoclonal IgG(1) antibody, which blocks ligand binding and functional activation of the EGFR; OSI-774 (erlotinib/Tarceva; Genentech/OSI/Roch) and ZD1839 (gefitinib/Iressa; AstraZeneca), two small molecule EGFR-selective inhibitors of tyrosine kinase enzymatic activity, which prevent EGFR autophosphorylation and activation. Iressa is the first EGFR-targeting agent to be registered as an anticancer drug in Japan, in Australia and in the US for the third-line treatment of chemoresistant non-small cell lung cancer (NSCLC) patients. This review will focus on the preclinical background and on the results from the first series of clinical trials with these drugs. Furthermore, continuing clinical trials and a series of open clinical issues for the development of optimal strategies of using EGFR-targeting agents will be discussed.     

Molecular mechanisms of ZD1839 （Iressa）-induced apoptosis in human leukemic U937 cells

Aim： To investigate the molecular mechanisms of ZD 1839-induced apoptosis in human leukemic U937 cells. Methods： The inhibition of human leukemic U937 cell growth was assessed by 3-（4,5-dimethyl-2-thiazolyl）-2,5-diphnyl-2H-tetrazolim bromide （MTT） assays, lactate dehydrogenase （LDH） release, and cell cycle distribution. The expression of anti- and pro-apoptotic proteins was detected by Western blot analysis. Results： This study demonstrated that ZD1839 induced apoptosis in leukemic U937 cells by the downregulation of Bcl-2, caspase activation and subsequent apoptotic features. Cotreatment with ZD 1839 and the caspase- 3 inhibitor z-DEVD-fmk blocked apoptosis, indicating that caspase-3 activation is at least partially responsible for ZD 1839-induced apoptosis. The ectopic expression of Bcl-2 attenuated caspase-3 activation, PARP cleavage, and subsequent indicators of apoptosis, including sub-G1 DNA content and LDH release. These results indicate that the downregulation of Bcl-2 plays a major role in the initiation of ZD1839-induced apoptosis, and that the activation of a caspase cascade is involved in the execution of apoptosis. Furthermore, ZD1839 treatment triggered the activation of p38 mitogen-activated protein kinase （MAPK） and the downregulation of c-Jun-N-terminal kinase （JNK）, extracellular signal-regulated kinase （ERK） and phosphatidyl inositol 3-kinase （PI3K）/Akt. The inhibition of the ERK and PI3K/Akt pathways also significantly increased cellular death. Conclusion： ZD 1839 activated caspase-3 and the inhibited Bcl-2 in human leukemic U937 cells through the downregulation of the ERK and PI3K/Akt pathways.