Tumor inhibitory effect of gefitinib (ZD1839, Iressa) and taxane combination therapy in EGFR-overexpressing breast cancer cell lines (MCF7/ADR, MDA-MB-231)

Some kinds of breast cancer cell lines, similar to several types of solid tumors, express epidermal growth factor receptor (EGFR). However, gefitinib, an EGFR tyrosine kinase inhibitor, is not effective for all these cell lines. Similarly, taxane is effective for many of the cell lines, although some, such as the multidrug-resistant MCF7/ADR cell line, show taxane-resistance. Here, we examined the growth inhibitory effect of combination treatment with gefitinib and taxane on the breast cancer cell lines MDA-MB-231 (EGFR-positive) and MCF7/ADR (EGFR- and HER2-positive). To estimate the combined effect, a Combination Index was calculated for each cell line. The combination of gefitinib and taxane showed a strong synergistic effect on MCF7/ADR cells, but an invitro additive-antagonistic effect on MDA-MB-231 cells. Similarly, the combination treatment showed a significantly increased tumor inhibitory effect on MCF7/ADR xenografts, but not on MDA-MB-231 xenografts. Regarding the mechanism of the synergistic effect, Western blotting analysis revealed that taxane activated the EGFR-Akt pathway in MCF7/ADR cells but not in MDA-MB-231. To determine the optimal sequential administration of gefitinib and taxane for MCF7/ADR cells, we used flow cytometry to analyze the cell cycle and apoptosis; finding that taxane treatment followed by gefitinib produced a higher rate of G2 arrest and apoptosis than gefitinib treatment followed by taxane. These results suggest gefitinib overcomes the drug-resistance of these cells, thereby increasing the effects of taxane on MCF7/ADR cells. Further, activation of the EGFR-Akt pathway by taxane is related to this synergistic effect.     

ZD1839 (Iressa), a novel epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor, potently inhibits the growth of EGFR-positive cancer cell lines with or without erbB2 overexpression.

Overexpression of the growth factor receptors EGFR and erbB2 occurs frequently in several human cancers and is associated with aggressive tumour behaviour and poor patient prognosis. We have investigated the effects of ZD1839 (Iressa), a novel EGFR tyrosine kinase inhibitor, on the growth, in vitro and in vivo, of human cancer cell lines expressing various levels of EGFR and erbB2. Proliferation of EGFR-overexpressing A431 and MDA-MB-231 cells in vitro was potently inhibited (50%-70%) by ZD1839 with half-maximally effective doses in the low nanomolar range. In parallel, ZD1839 blocked autophosphorylation of EGFR and prevented activation of PLC-gamma 1, ERK MAP kinases and PKB/Akt by EGF. It also inhibited proliferation in EGFR(+) cancer cell lines overexpressing erbB2 (SKBr3, SKOV3, BT474) by between 20% and 80%, effects which correlated with inhibition of EGF-dependent erbB2 phosphorylation and activation of ERK MAP kinase and PKB/Akt in SKOV3 cells. Oral administration of ZD1839 inhibited the growth of MDA-MB-231 and SKOV3 tumours, established as xenografts in athymic mice, by 71% and 32%, respectively. Growth inhibition coincided with reduced proliferation but no change in apoptotic index. Collectively, these results show that ZD1839, at the doses studied, is a potent inhibitor of proliferation not only in cells overexpressing EGFR but also in EGFR(+) cells that overexpress erbB2.     

表皮生长因子受体酪氨酸激酶抑制剂ZD1839对鼻咽癌细胞系的作用

ZDl839是一种喹唑啉类化合物,它是一种口服的选择性的表皮生长因子受体酪氨酸激酶抑制剂,已被美国食品药品管理局批准用于治疗晚期非小细胞肺癌。鼻咽癌是一种存在表皮生长因子受体表达的上皮源性的恶性肿瘤,ZDl839对其作用如何还未见有报道。本实验通过体外研究初步探讨ZDl839在鼻咽癌治疗中的可能价值。     

Enhancement of tumor radioresponse by combined treatment with gefitinib (Iressa, ZD1839), an epidermal growth factor receptor tyrosine kinase inhibitor, is accompanied by inhibition of DNA damage repair and cell growth in oral cancer

Molecular blockade of EGFR with either an EGFR MAb or an EGFR TKI enhances the radiosensitivity of human SCCs. In the present study, we investigated whether treatment with the EGFR TKI gefitinib (Iressa, ZD1839) improves the response to radiotherapy in the OSCC cell lines HSC2 and HSC3. We examined potential mechanisms that may contribute to the enhanced radiation response induced by gefitinib. Growth inhibition was observed in vitro with radiation or gefitinib. A cooperative antiproliferative effect was obtained when cancer cells were treated with radiation followed by gefitinib. Cells treated with a combination of radiation and gefitinib arrested in G(1) and G(2)-M phases, with a decrease in the S-phase population. While radiation alone did not significantly affect MEK1/2 and p38 MAPK autophosphorylation, the combination of gefitinib and radiation completely inhibited the downstream signaling of EGFR. Results from DNA damage repair analysis in cultured OSCC cells demonstrated that gefitinib had a strong inhibitory effect on DNA-PKc pathways after radiation. Tumor xenograft studies demonstrated that the combination of gefitinib and radiation caused growth inhibition and tumor regression of well-established OSCC tumors in athymic mice; tumor volume was reduced from 1,008.2 to 231.4 mm(3) in HSC2 cells (p < 0.01) and from 284.2 to 12.4 mm(3) in HSC3 cells (p < 0.01). Immunohistochemical analysis of OSCC xenografts revealed that gefitinib caused a striking decrease in tumor cell proliferation when combined with radiotherapy. Overall, we conclude that gefitinib enhances tumor radioresponse by multiple mechanisms that may involve antiproliferative growth inhibition and effects on DNA repair after exposure to radiation.     

Analysis of epidermal growth factor receptor expression as a predictive factor for response to gefitinib ('Iressa', ZD1839) in non-small-cell lung cancer

Gefitinib ('Iressa', ZD1839) is an orally active epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor that has demonstrated antitumour activity and favourable tolerability in Phase II studies. We investigated whether EGFR expression levels could predict for response to gefitinib in patients with advanced non-small-cell lung cancer (NSCLC), who received gefitinib (250 mg day(-1)) as part of a worldwide compassionate-use programme. Tissue samples were analysed by immunohistochemistry to assess membrane EGFR immunoreactivity. Of 147 patients enrolled in our institution, 50 patients were evaluable for assessment of both clinical response and EGFR expression. The objective tumour response rate was 10% and disease control was achieved in 50% of patients. Although high EGFR expression was more common in squamous-cell carcinomas than adenocarcinomas, all objective responses were observed in patients with adenocarcinoma. Response and disease control with gefitinib were not associated with high EGFR expression. Overall, median survival was 4 months, and the 1-year survival rate was 18%. Strong EGFR staining correlated with shorter survival time for all patients. Gefitinib demonstrated promising clinical activity in this group of patients with NSCLC. These results have also shown that EGFR expression is not a significant predictive factor for response to gefitinib.     

Induction of cell cycle arrest and apoptosis in human nasopharyngeal carcinoma cells by ZD6474, an inhibitor of VEGFR tyrosine kinase with additional activity against EGFR tyrosine kinase

ZD6474 is a vascular endothelial growth factor receptor (VEGFR) and epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor. The present study was undertaken to investigate the direct antiproliferative effect of ZD6474 on human nasopharyngeal carcinoma (NPC) in vitro and the antitumor activity on NPC xenografts in vivo. Results indicated that ZD6474 treatment inhibited EGFR phosphorylation and led to a dose- and time-dependent decrease in NPC cell (CNE-1, CNE-2 and C666-1) proliferation. Further investigation demonstrated G0/G1 cell cycle arrest in all 3 cell lines, which was associated with an upregulation of p21 and/or p27, and downregulation of CDK4, CDK6 and CDK2. ZD6474 treatment also induced apoptosis in CNE-1 and CNE-2 cells. The apoptosis mechanisms involved reduction of Bcl-2 and/or Bcl-XL, induction of Bak and/or Bax, and activation of caspases-3, -9 and/or -8. The in vivo antitumor activity was evaluated in CNE-2 and C666-1 xenografted nude mice. Administration of ZD6474 (25-100 mg/kg/day, once-daily, p.o.) produced a dose-dependent inhibition of tumor growth and prolonged survival in both models. This study suggests that ZD6474 exerts direct antiproliferative effects on NPC cell lines in vitro by inducing G0/G1 arrest and apoptosis, and potent antitumor effects on NPC xenografts in vivo. It indicates that ZD6474 may offer a new and effective treatment for human NPC.     

Differential radiosensitisation by ZD1839 (Iressa), a highly selective epidermal growth factor receptor tyrosine kinase inhibitor in two related bladder cancer cell lines

The epidermal growth factor receptor (EGFR) is expressed in a wide variety of epithelial tumours including carcinoma of the bladder. Stimulation of the EGFR pathway is blocked by ZD1839 (Iressa), a highly selective EGFR tyrosine kinase inhibitor. Radical radiotherapy is an established organ sparing treatment option for muscle invasive bladder cancer and this study has explored the possibility for the use of ZD1839 as a radiosensitiser in this scenario. The effect of combination treatment with ZD1839 (0.01 microM) and ionising radiation in the established bladder cancer cell lines MGH-U1 and its radiosensitive mutant clone S40b was measured by clonogenic assays. A highly significant radiosensitising effect was seen in both cell lines (P < 0.001 for MGH-U1 and S40b cell lines). This effect was independent of the concentration of the drug and the duration of exposure prior to treatment with ionising radiation. Cell cycle kinetics of both cell lines was not significantly altered with ZD1839 (0.01 microM) as a single agent. A modest induction of apoptosis was observed with ZD1839 (0.01 microM) as a single agent, but a marked induction was observed with the combination treatment of ZD1839 and ionising radiation. These results suggest a potentially important role for ZD1839 in combination with radiotherapy in the treatment of muscle invasive bladder cancer.     

ZD1839 ('Iressa'), a specific oral epidermal growth factor receptor-tyrosine kinase inhibitor, potentiates radiotherapy in a human colorectal cancer xenograft model

The effect of ZD1839 ('Iressa'), a specific inhibitor of the tyrosine kinase activity of the epidermal growth factor receptor, on the radiation response of human tumour cells (LoVo colorectal carcinoma) was evaluated in vitro and in vivo. ZD1839 (0.5 microM, incubated days 1-5) significantly increased the anti-proliferative effect of fractionated radiation treatment (2 Gy day(-1), days 1-3) on LoVo cells grown in vitro (P=0.002). ZD1839 combined with either single or fractionated radiotherapy in mice bearing LoVo tumour xenografts, also produced a highly significant increase in tumour growth inhibition (P< or = 0.001) when compared to treatment with either modality alone. The radio-potentiating effect of ZD1839 was more apparent when radiation was administered in a fractionated protocol. This phenomenon may be attributed to an anti proliferative effect of ZD1839 on tumour cell re-population between radiotherapy fractions. These data suggest radiotherapy with adjuvant ZD1839 could enhance treatment response. Clinical investigation of ZD1839 in combination with radiotherapy is therefore warranted.     

表皮生长因子受体酪氨酸激酶抑制剂ZD1839与奥沙利铂联合对人肺癌A549细胞的作用

背景与目的:ZD1839是一种表皮生长因子受体酪氨酸激酶的小分子抑制剂,是目前肺癌分子靶向治疗中较为成熟的药物,因其单药临床有效率低,如何与化疗联合提高抗癌效应受到关注.本实验通过研究ZD1839联合奥沙利铂的不同给药方案对人肺腺癌细胞A549的杀伤作用,探讨两药联合的最佳模式.方法:以药物联合效应测定方法,评价ZD1839和奥沙利铂不同给药顺序对A549细胞的抑制作用,以流式细胞仪测定不同联合方案对A549细胞周期分布及凋亡率的影响.观察ZD1839、奥沙利铂不同给药方案作用时,裸鼠A549细胞移植瘤生长情况,测定肿瘤生长抑制率.结果:先奥沙利铂后ZD1839的序贯给药方案表现出明显的协同作用,药物联合指数为0.51±0.01;而先ZD1839后给予奥沙利铂时,药物联合指数为1.56±0.03,两药间为拮抗作用.先奥沙利铂后ZD1839组G2/M期细胞比例与其他组相比明显增加,达到37.9%(P＜0.05),细胞凋亡率达到22.3%.体内实验表明,先奥沙利铂后ZD1839组抑瘤率最高,达到58.9%;而先奥沙利铂 ZD1839 24 h后ZD1839 48 h组的抑瘤率为52.4%,ZD1839后奥沙利铂组的抑瘤率为30.6%.结论:先奥沙利铂后ZD1839序贯给药对A549细胞增殖的抑制作用更强.     

Activity of a specific inhibitor, gefitinib (Iressa, ZD1839), of epidermal growth factor receptor in refractory non-small-cell lung cancer.

BACKGROUND: Gefitinib (Iressa(TM), ZD1839) is an orally active, selective epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor. Phase I studies showed that it is well tolerated, with evidence of tumor regression in patients with advanced non-small-cell lung cancer (NSCLC). Therefore, we aimed to assess the antitumor activity and tolerability of gefitinib in a series of patients with previously treated, advanced NSCLC, as a part of a compassionate use program. PATIENTS AND METHODS: To be eligible, all patients were required to have histologically or cytologically proven advanced or metastatic NSCLC, prior chemotherapy with at least one cisplatin-containing chemotherapy regimen or contraindication to cytotoxic drugs, Eastern Cooperative Oncology Group performance status < or =2, and adequate hematological, renal and hepatic parameters. All patients provided signed informed consent. Patient re-evaluation was performed every 4-6 weeks. RESULTS: Seventy-three consecutive patients were enrolled. Response rate, including complete and partial response, was 9.6%; an additional 43.8% of patients achieved stable disease, for an overall disease control of 53.4%. EGFR1 status was evaluated by immunocytochemistry in 25 patients. According to EGFR1 immunoreactivity all responses were observed with medium/strong imunoreactivity while three out of four responses were observed in high expressive patients. Median survival for all patients was 4 months while it reached 6 months for patients with disease control. The 1-year survival rate was 13.1% for the entire series and 23.2% for patients with disease control. Non-hematological toxicity was generally mild. CONCLUSION: Gefitinib has promising activity with a good toxicity profile in patients with progressive NSCLC who have received one or two prior chemotherapy regimens. A possible relationship within response and EGFR1 expression is suggested.     

Antitumor activity of the selective epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) Iressa (ZD1839) in an EGFR-expressing multidrug-resistant cell line in vitro and in vivo

Selective tyrosine kinase inhibitors are regarded as promising antitumor agents for cancer treatment. Iressa (ZD1839) is an orally active, selective EGFR-TKI (epidermal growth factor receptor-tyrosine kinase inhibitor) that blocks signal transduction pathways implicated in cancer cell proliferation, survival and other host-dependent processes promoting cancer growth. The cellular mechanisms of ZD1839 action against human malignant cells and drug-resistant cells were evaluated in vitro. Among the cell lines tested, ZD1839 showed a strong growth-inhibitory effect in vitro on human leukemic cells resistant to phorbol ester. This cell line, K562/TPA, shows a non-P-glycoprotein-mediated multidrug-resistant phenotype. The IC50 value of ZD1839 on K562/TPA was approximately 400-fold lower than that on the parental K562 cell (K562 = 12 +/- 2 microM; K562/TPA = 0.025 +/- 0.002 microM) in vitro as determined by a dye formation assay. The expression of EGFR and EGFR mRNA was clearly present in K562/TPA but not in parental K562 cells as determined by Western blotting and RT-PCR. EGFR was autophosphorylated in K562/TPA detected by the antiphosphotyrosine antibody. The in vivo antitumor effects of ZD1839 on K562 and K562/TPA cells were also investigated in BALB/c nude mice. K562/TPA cells transplanted subcutaneously into mice disappeared completely with ZD1839 treatment (20 mg/kg/day, days 3-9). This was not the case in K562 cells. These results suggest that ZD1839 is highly active against tumor cells with non-P-glycoprotein-mediated multidrug resistance that express EGFR. Iressa is a trademark of AstraZeneca (Cheshire, UK).     

The effect of gefitinib (Iressa,ZD1839) in combination with oxaliplatin is schedule-dependent in colon cancer cell lines

Background Clinical trials of gefitinib (Iressa, ZD1839) in combination with cytotoxic agents have been carried out or are ongoing in several varieties of tumor. To provide a rationale for future clinical trials, the effects of combining gefitinib with oxaliplatin in different sequences of administration and different dose ratios in two colon cancer cell lines were evaluated.Materials and methods The colon cancer cell lines HT-29 and LoVo were used. The methods consisted of median effect and combination index analysis, Western blot, mass spectrometry, and a cell death ELISA.Results In vitro analysis demonstrated that the combination effects of the two agents were sequence-dependent. Changing the sequence of administration from gefitinib first to gefitinib last changed the combination effect from antagonism to synergy. The dose ratio between the two agents affected the combination effects. When equiactive doses of the two agents were used with the sequence gefitinib following oxaliplatin, the greatest level of synergism was obtained (CI=0.6±0.2, P=0.032). Further evaluation revealed that gefitinib significantly inhibited removal of Pt-DNA adducts (P<0.05), providing a potential explanation for the sequence-dependent synergy observed with gefitinib following oxaliplatin. However, this effect was not dose-dependent. Additional studies demonstrated that gefitinib enhanced the effects of oxaliplatin by maintaining oxaliplatin-induced apoptosis, and equiactive dose of gefitinib following oxaliplatin induced prominent enhancement of apoptosis.Conclusions Oxaliplatin followed by an equiactive relative dose of gefitinib is an appropriate combination for evaluation in colon cancer.     

表皮生长因子受体酪氨酸激酶抑制剂ZD1839与伊利替康联合效应的实验研究

目的 探讨ZD1839与伊利替康的活性代谢产物7-乙基-10羟基-喜树碱（SN38）联合的最佳方案及其机制。方法 以药物联合效应测定方法,评价ZD1839和SN38不同给药顺序对人结肠癌细胞HT-29和LoVo的抑制作用;以Western blot和免疫共沉淀方法,分析ZD1839与化疗不同联合方案对各自靶蛋白及其下游分子表达的影响;以流式细胞仪测定不同联合方案对细胞周期的影响;以组蛋白相关的DNA碎片分析,比较不同方案对细胞凋亡指数的影响。结果 先SN38后ZD1839的序贯给药方案表现出明显的协同作用;反之,则表现为拈抗作用。SN38明显抑制细胞拓扑异构酶-1（Topo-1）活性;ZDl839不影响表皮生长因子受体（EGFR）的表达,但能抑制EGFR的磷酸化。与SN38单药相比,SN38联合ZDl839对Topo-1的抑制无增强;与单药ZD1839相比,联合方案对EGFR、MAPK磷酸化的抑制作用无增强,但ZD1839、SN38同时给药,和先SN38后ZD1839序贯给药对AKT的抑制有所增强。同时,联合方案对细胞周期分布的改变影响明显。ZD1839可明显维持化疗诱导的DNA损伤和细胞凋亡。结论 先SN38后ZD1839序贯给药可能是治疗结肠癌的最佳联合模式。     

ZD1839, a novel,oral epidermal growth factor receptor-tyrosine kinase inhibitor,as salvage treatment in patients with advanced non-small cell lung cancer. Experience from a single center participating in a compassionate use program

Objective: We evaluated the efficacy and tolerability of the orally active, selective epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) ZD1839 in patients with pretreated advanced non-small cell lung cancer (NSCLC) participating in a compassionate use program. Patients and methods: Thirty-one patients with advanced, unresectable and progressive NSCLC, previously treated with one or two chemotherapy regimens, received ZD1839 250 mg orally once daily. Patients who had received only one prior chemotherapy regimen had to be considered unsuitable for second-line chemotherapy. Results: The disease control rate was 32% (95% CI: 15.8–48.7) (1/31 patients had a partial response and 9/31 patients had stable disease) and the median overall survival 23 weeks (range 4–40). Symptom improvement was reported by 39% of patients overall and by 83% of patients who achieved disease control. The median time to symptom improvement was 3 weeks (range 2–4). Adverse events were generally mild (grade I or II) and reversible and consisted mostly of skin rash, diarrhea and fatigue. Conclusions: ZD1839 demonstrated clinically meaningful antitumor activity with significant improvement in symptoms in this heavily pretreated group of patients with advanced NSCLC. Furthermore, ZD1839 showed a favorable toxicity profile, with the majority of adverse events being mild and reversible.     

ZD6474, an inhibitor of VEGFR and EGFR tyrosine kinase activity in combination with radiotherapy

Radiation enhances both epithelial growth factor receptor (EGFR) and vascular endothelial growth factor (VEGF) expression, which are a part of key pathways for tumor progression. Some tumors may not respond well to EGFR inhibitors alone or may develop resistance to EGFR inhibitors. Therefore, drug therapy targeted to VEGF receptors and EGFRs, when combined with radiotherapy (RT), may improve tumor control and provide wider applicability. This article focuses on ZD6474, an inhibitor of EGFR and VEGF receptor signaling in combination with RT. We discuss preclinical and clinical studies with RT and inhibitors of VEGF or EGFR signaling first. We then address issues associated with ZD6474 pharmacokinetic dosing, and scheduling when combined with RT. We also discuss ZD6474 in the context of anti-EGFR therapy resistance. Dual inhibition of EGFR and VEGF receptor signaling pathways shows promise in enhancing RT efficacy.     

Radiosensitization of human breast cancer cells by a novel ErbB family receptor tyrosine kinase inhibitor

Purpose: Overexpression of the ErbB family of growth factor receptors is present in a wide variety of human tumors and is correlated with poor prognosis. The purpose of this study was to determine the effects of a novel small molecule ErbB tyrosine kinase inhibitor, CI-1033, in combination with ionizing radiation on breast cancer cell growth and survival.

Materials & Methods: Growth assays were performed on ErbB-overexpressing human breast cancer cells developed in our laboratory in the presence of 0.1–1.0 μM CI-1033 (Parke Davis). Clonogenic survival assays were performed in the presence of ionizing radiation with or without CI-1033. For some experiments, clonogen numbers, defined as the product of surviving fraction and total number of cells, were calculated at each time point during a course of multifraction radiation.

Results: CI-1033 potently inhibited the growth of ErbB-overexpressing breast cancer cells. A single 48-h exposure of 1 μM CI-1033 resulted in growth inhibition for 7 days, whereas three times weekly administration resulted in sustained growth inhibition. Clonogenic survival was modestly decreased after a 7-day exposure to CI-1033. Exposure to both CI-1033 and radiation (6 Gy) yielded a 23-fold decrease in clonogenic survival compared to radiation alone. In a multifraction experiment, exposure to CI-1033 and three 5-Gy fractions of gamma radiation decreased the total number of clonogens in the population by 65-fold compared to radiation alone.

Conclusion: CI-1033 results in potent growth inhibition and modest cytotoxicity of ErbB-overexpressing breast cancer cells, and has synergistic effects when combined with ionizing radiation. These data suggest that CI-1033 may have excellent clinical potential both alone and in combination with radiation therapy.     

Sequence-dependent effects of ZD1839 ('Iressa') in combination with cytotoxic treatment in human head and neck cancer

Elevated levels of epidermal growth factor receptor in head and neck cancer have been extensively reported, and are correlated with poor prognosis. The combination of cisplatin and 5-fluorouracil is a standard treatment regimen for head and neck cancer, with radiation representing another therapeutic option. Six head and neck cancer cell lines were used to study the cytotoxic effects of combining ZD1839 ('Iressa'), a new selective epidermal growth factor receptor tyrosine kinase inhibitor, and radiation. Two of the cell lines were also used to study the combination of ZD1839 and cisplatin/5-fluorouracil. Cytotoxic effects were assessed by the MTT test. The results indicated that ZD1839 applied before radiation gave the best effects (P=0.002); an effect that was strongest in those p53-mutated cell lines that express the highest epidermal growth factor receptor levels. The effects of ZD1839 with cisplatin and/or 5-fluorouracil were sequence dependent (P<0.003), with the best results achieved when ZD1839 was applied first. For the triple combinations, ZD1839 applied before cisplatin and 5-fluorouracil resulted in a slight synergistic effect (P=0.03), although the effect was greater when ZD1839 was applied both before and during cytotoxic drug exposure. In conclusion, ZD1839 applied before radiation and before and/or during cisplatin/5-fluorouracil may improve the efficacy of treatment for head and neck cancer.     

The small GTPase ADP-Ribosylation Factor 1 mediates the sensitivity of triple negative breast cancer cells to EGFR tyrosine kinase inhibitors

The clinical use of EGFR-targeted therapy, in triple negative breast cancer patients, has been limited by the development of resistance to these drugs. Although activated signaling molecules contribute to this process, the molecular mechanisms remain relatively unknown. We have previously reported that the small GTPase ADP-Ribosylation Factor 1 (ARF1) is highly expressed in invasive breast cancer cells and acts as a molecular switch to activate EGF-mediated responses. In this study, we aimed at defining whether the high expression of ARF1 limits sensitivity of these tumor cells to EGFR inhibitors, such as gefitinib. Here, we show that the knock down of ARF1 expression or activity decreased the dose and latency time required by tyrosine kinase inhibitors to induce cell death. This may be explained by the observation that the depletion of ARF1 suppressed gefitinib-mediated activation of key mediators of survival such as ERK1/2, AKT and Src, while enhancing cascades leading to apoptosis such as the p38MAPK and JNK pathways, modifying the Bax/Bcl2 ratio and cytochrome c release. In addition, inhibiting ARF1 expression and activation also results in an increase in gefitinib-mediated EGFR internalization and degradation further limiting the ability of this receptor to promote its effects. Interestingly, we observed that gefitinib treatment resulted in the enhanced activation of ARF1 by promoting its recruitment to the receptor AXL, an important mediator of EGFR inhibition suggesting that ARF1 may promote its pro-survival effects by coupling to alternative mitogenic receptors in conditions where the EGFR is inhibited. Together our results uncover a new role for ARF1 in mediating the sensitivity to EGFR inhibition and thus suggest that limiting the activation of this GTPase could improve the therapeutic efficacy of EGFR inhibitors.     

Preclinical antitumor activity of S‐222611, an oral reversible tyrosine kinase inhibitor of epidermal growth factor receptor and human epidermal growth factor receptor 2

Epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER2) are validated molecular targets in cancer therapy. Dual blockade has been explored and one such agent, lapatinib, is in clinical practice but with modest activity. Through chemical screening, we discovered a novel EGFR and HER2 inhibitor, S‐222611, that selectively inhibited both kinases with IC₅₀s below 10 nmol/L. S‐222611 also inhibited intracellular kinase activity and the growth of EGFR‐expressing and HER2‐expressing cancer cells. In addition, S‐222611 showed potent antitumor activity over lapatinib in a variety of xenograft models. In evaluations with two patient‐oriented models, the intrafemoral implantation model and the intracranial implantation model, S‐222611 exhibited excellent activity and could be effective against bone and brain metastasis. Compared to neratinib and afatinib, irreversible EGFR/HER2 inhibitors, S‐222611 showed equivalent or slightly weaker antitumor activity but a safer profile. These results indicated that S‐222611 is a potent EGFR and HER2 inhibitor with substantially better antitumor activity than lapatinib at clinically relevant doses. Considering the safer profile than for irreversible inhibitors, S‐222611 could be an important option in future cancer therapy.     

Src激酶抑制剂ZD6474在乳腺癌MCF-7细胞转移中的作用和机制

目的:探讨Src激酶抑制剂ZD6474在乳腺癌MCF-7细胞转移中的作用和机制。方法:乳腺癌MCF-7细胞经ZD6474处理后,检测肿瘤细胞体外黏附、侵袭能力的改变,应用Western blot及Real-time PCR观察细胞Src激酶磷酸化水平及E-钙黏素和β-连环蛋白表达的变化,报告基因技术检测Snail在转录水平表达的变化。结果:ZD6474可抑制MCF-7细胞中Src激酶活性,在ZD6474浓度为1×10-5mol/L和1×10-4mol/L时,细胞的抑制率分别为12.2%和27.5%。Src酪氨酸激酶抑制剂可上调E-cadherin在mRNA和蛋白表达水平,下调β-catenin在mRNA和蛋白表达水平及Snail启动子活性。结论:Src激酶与乳腺癌MCF-7细胞肿瘤转移能力密切相关,阻断Src激酶活性可抑制肿瘤细胞转移能力。