Activation of EGFR,HER2 and HER3 by neurotensin/neurotensin receptor 1 renders breast tumors aggressive yet highly responsive to lapatinib and metformin in mice

A present challenge in breast oncology research is to identify therapeutical targets which could impact tumor progression. Neurotensin (NTS) and its high affinity receptor (NTSR1) are up regulated in 20% of breast cancers, and NTSR1 overexpression was shown to predict a poor prognosis for 5 year overall survival in invasive breast carcinomas. Interactions between NTS and NTSR1 induce pro-oncogenic biological effects associated with neoplastic processes and tumor progression. Here, we depict the cellular mechanisms activated by NTS, and contributing to breast cancer cell aggressiveness.We show that neurotensin (NTS) and its high affinity receptor (NTSR1) contribute to the enhancement of experimental tumor growth and metastasis emergence in an experimental mice model. This effect ensued following EGFR, HER2, and HER3 over-expression and autocrine activation and was associated with an increase of metalloproteinase MMP9, HB-EGF and Neuregulin 2 in the culture media. EGFR over expression ensued in a more intense response to EGF on cellular migration and invasion. Accordingly, lapatinib, an EGFR/HER2 tyrosine kinase inhibitor, as well as metformin, reduced the tumor growth of cells overexpressing NTS and NTSR1. All cellular effects, such as adherence, migration, invasion, altered by NTS/NTSR1 were abolished by a specific NTSR1 antagonist. A strong statistical correlation between NTS-NTSR1-and HER3 (p< 0.0001) as well as NTS-NTSR1-and HER3- HER2 (p< 0.001) expression was found in human breast tumors.Expression of NTS/NTSR1 on breast tumoral cells creates a cellular context associated with cancer aggressiveness by enhancing epidermal growth factor receptor activity. We propose the use of labeled NTS/NTSR1 complexes to enlarge the population eligible for therapy targeting HERs tyrosine kinase inhibitor or HER2 overexpression.     

EGFR / HER2 / HER3 expression in tumour and gefitinib treatment in Chinese patients with advanced non-small cell lung cancer

Objective: Biological markers performable in routine practice and able to predict the clinical outcome of advanced non-small cell lung cancer (NSCLC) treated with gefitinib are urgently needed.Methods: We analyzed EGFR / HER2 / HER3 primary tumour immunohistochemical expression in a prospective and consecutive series of 90 Chinese patients.Platinumpretreated patients received a 250 mg oral dose of gefitinib once daily until disease progression; EGFR / HER2 / HER3 tumour status was related with the clinical outcome in terms of response rate (RR), time to disease progression (TTP), and overall survival (OS).Results: A high expression (scores 2-3) of EGFR, HER2 and HER3 was verified in 16.7%, 43.3% and 21.1% of tumors, respectively.EGFR and HER3 status were not significantly related with response, while the HER2 overexpression result was significantly associated with a higher RR (35.9% vs.15.7%, P = 0.027).The RR in the 13 patients with both HER2 and HER3 expression was also significantly higher than in the other 77 patients (53.8% vs.22.1%, P = 0.036).EGFR / HER2 / HER3 status was not significantly correlated with TrP or OS.Conclusion: The HER2 immunohistochemical expression can play a role in the clinical management of Chinese patients with advanced NSCLC who are candidates for gefitinib therapy     

AZD8931, an equipotent,reversible inhibitor of signaling by epidermal growth factor receptor (EGFR), HER2, and HER3: preclinical activity in HER2 non-amplified inflammatory breast cancer models

Introduction

Epidermal growth factor receptor (EGFR) overexpression has been associated with prognostic and predictive value in inflammatory breast cancer (IBC). Epidermal growth factor receptor 2 (HER2) overexpression is observed at a higher rate in IBC compared with noninflammatory breast cancer. Current clinically available anti-HER2 therapies are effective only in patients with HER2 amplified breast cancer, including IBC. AZD8931 is a novel small-molecule equipotent inhibitor of EGFR, HER2, and HER3 signaling. In this study, we investigated the antitumor activity of AZD8931 alone or in combination with paclitaxel using preclinical models of EGFR-overexpressed and HER2 non-amplified IBC cells.

Methods

Two IBC cell lines SUM149 and FC-IBC-02 derived from pleural effusion of an IBC patient were used in this study. Cell growth and apoptotic cell death were examined in vitro. For the in vivo tumor growth studies, IBC cells were orthotopically transplanted into the mammary fat pads of immunodeficient mice. AZD8931 was given by daily oral gavage at doses of 25 mg/kg, 5 days/week for 4 weeks. Paclitaxel was subcutaneously injected twice weekly.

Results

AZD8931 significantly suppressed cell growth of IBC cells and induced apoptosis of human IBC cells in vitro. Significantly, we showed that AZD8931 monotherapy inhibited xenograft growth and the combination of paclitaxel + AZD8931 was demonstrably more effective than paclitaxel or AZD8931 alone treatment at delaying tumor growth in vivo in orthotopic IBC models.

Conclusion

AZD8931 single agent and in combination with paclitaxel demonstrated signal inhibition and antitumor activity in EGFR-overexpressed and HER2 non-amplified IBC models. These results suggest that AZD8931 may provide a novel therapeutic strategy for the treatment of IBC patients with HER2 non-amplified tumors.     

Antitumor activity of HER1/EGFR tyrosine kinase inhibitor erlotinib, alone and in combination with CPT-11 (irinotecan) in human colorectal cancer xenograft models

Erlotinib (Tarceva, OSI-774) is a potent, orally available, small-molecule inhibitor of HER1/EGFR tyrosine-kinase activity. In this study, the antitumor activity of erlotinib was evaluated in two human colorectal tumor xenograft models (LoVo and HCT116) in athymic mice. When erlotinib was administered as monotherapy, significant tumor growth inhibition (TGI) was seen in the LoVo model at both 100 mg/kg [TGI > 100%, P < 0.001; 6/10 partial regressions (PRs)] and 25 mg/kg (TGI = 79%, P < 0.001) doses. However, the HCT116 xenograft model was not responsive to any dose of erlotinib tested. The differential response to erlotinib of these two tumor models was not a result of differences in HER1/EGFR expression levels since these were similar in both cell lines. However, it was demonstrated that resistance to erlotinib in the HCT116 model may be a result of persistent activation of ERK in these tumors. Based on the single agent activity of erlotinib in LoVo tumors, a combination study with CPT-11 (Camptosar, irinotecan) was performed. CPT-11 at the optimal dose of 60 mg/kg or a lower dose of 15 mg/kg resulted in significant TGI (TGI > 100%, P < 0.001, and TGI = 93%, P < 0.001, respectively) in LoVo-bearing mice. Combination treatment with erlotinib (25 mg/kg) and CPT-11 (15 mg/kg) produced significantly greater antitumor activity (TGI > 100%, P < 0.001; 10/10 PRs) than either agent alone (P < 0.05), with no increase in toxicity. These data indicate that erlotinib can enhance the antitumor activity of CPT-11, without enhanced toxicity, in the LoVo human colorectal tumor xenograft model.     

Increased signalling of EGFR and IGF1R, and deregulation of PTEN/PI3K/Akt pathway are related with trastuzumab resistance in HER2 breast carcinomas

Background:

Trastuzumab resistance hampers its well-known efficacy to control HER2-positive breast cancer. The involvement of PI3K/Akt pathway in this mechanism is still not definitively confirmed.

Methods:

We selected 155 patients treated with trastuzumab after development of metastasis or as adjuvant/neoadjuvant therapy. We performed immunohistochemistry for HER2, ER/PR, epidermal growth factor 1-receptor (EGFR), α-insulin-like growth factor 1-receptor (IGF1R), phosphatase and tensin homologue (PTEN), p110α, pAkt, pBad, pmTOR, pMAPK, MUC1, Ki67, p53 and p27; mutational analysis of PIK3CA and PTEN, and PTEN promoter hypermethylation.

Results:

We found 46% ER/PR-positive tumours, overexpression of EGFR (15%), α-IGF1R (25%), p110α (19%), pAkt (28%), pBad (22%), pmTOR (23%), pMAPK (24%), MUC1 (80%), PTEN loss (20%), and PTEN promoter hypermethylation (20%). PIK3CA and PTEN mutations were detected in 17% and 26% tumours, respectively. Patients receiving adjuvant trastuzumab with α-IGF1R or pBad overexpressing tumours presented shorter progression-free survival (PFS) (all P⩽0.043). Also, p110α and mTOR overexpression, liver and brain relapses implied poor overall survival (OS) (all P⩽0.041). In patients with metastatic disease, decreased PFS correlated with p110α expression (P=0.024), whereas for OS were the presence of vascular invasion and EGFR expression (P⩽0.019; Cox analysis).

Conclusion:

Our results support that trastuzumab resistance mechanisms are related with deregulation of PTEN/PI3K/Akt/mTOR pathway, and/or EGFR and IGF1R overexpression in a subset of HER2-positive breast carcinomas.     

UBE2C promotes the progression of pancreatic cancer and glycolytic activity via EGFR stabilization-mediated PI3K-Akt pathway activation

BackgroundPancreatic cancer (PC) is among the most prevalent and deadliest endocrine tumors, yet the mechanisms governing its pathogenesis remain to be fully clarified. While ubiquitin-conjugating enzyme E2C (UBE2C) has been identified as an important oncogene in several cancers, its importance in PC has yet to be established.MethodsUBE2C expression in PC tumor samples and cell lines was examined via quantitative real-time polymerase chain reaction (qRT-PCR), while appropriate commercial kits were used to assess lactate production, ATP generation, and the uptake of glucose.ResultsUBE2C was found to be upregulated in PC patient tumors and correlated with poorer survival outcomes. In PC cell lines, the silencing of this gene suppressed the malignant activity of cells, thus supporting its identification as an oncogene in this cancer type. Mechanistically, UBE2C was found to promote enhanced matrix metalloproteinase (MMP) protein expression via activating the PI3K-Akt pathway. Moreover, it was found to bind to the epidermal growth factor receptor (EGFR), stabilizing it and driving additional PI3K-Akt pathway activation. UBE2C knockdown in PC cells impaired their uptake of glucose and their ability to produce lactate and ATP.ConclusionsIn conclusion, the results of this study support a role for UBE2C as a driver of metastatic PC progression owing to its ability to bind to EGFR and to induce signaling via the PI3K-Akt pathway.     

EGFR基因突变及HER2/HER3蛋白表达水平与吉非替尼治疗晚期非小细胞肺癌疗效的关系

背景与目的:吉非替尼治疗非小细胞肺癌(NSCLC)的疗效不一,如何选择对吉非替尼敏感的患者,提高药物的疗效是临床中的难点。本研究探讨了中国人群中表皮生长因子受体(epidermal growth factor receptor,EGFR)的突变及HER2/HER3蛋白表达与吉非替尼治疗局部晚期或转移性NSCLC疗效的关系。方法:2002年5月至2005年2月,符合入组条件的106例患者每日口服250mg吉非替尼一次,直至疾病进展或出现不可耐受的毒副反应。收集吉非替尼治疗前的肿瘤组织。提取基因组DNA后,采用nest PCR技术扩增EGFR基因的18～24外显子,并从正反两个方向进行DNA测序和分析。同时采用免疫组化法检测84例肿瘤组织中的HER2/HER3蛋白的表达。结果:106例肿瘤标本中32例(30.2%)发生了突变。HER2高表达患者的有效率显著高于低表达的患者(36.8%vs.17.4%,P=0.044)。HER2/HER3的表达水平与疾病进展时间(TTP)及总生存期(OS)无关,但HER2/HER3高表达的患者生存期较低表达组略长(9.1个月vs.6.1个月,P=0.725;9.0个月vs.6.1个...     

Curcumin and its Analogues (PGV-0 and PGV-1) Enhance Sensitivity of Resistant MCF-7 Cells to Doxorubicin through Inhibition of HER2 and NF-kB Activation

Chemoresistance of breast cancer to doxorubicin is mediated mainly through activation of NF-kB and overexpression of HER2. Curcumin and its analogues (PGV-0 and PGV-1) exert cytotoxic effects on T47D breastcancer cells. Suppression of NF-kB activation is suggested to contribute to this activity. The present studyaimed to explore the effects of curcumin, PGV-0, and PGV-1 singly and in combination with doxorubicin onMCF-7/Dox cells featuring over-expression of HER2. In MTT assays, curcumin, PGV-0, and PGV-1 showedcytotoxicity effects against MCF-7/Dox with IC50 values of 80 μM, 21 μM, and 82 μM respectively. Thesecompounds increased MCF-7/Dox sensitivity to doxorubicin. Cell cycle distribution analysis exhibited that thecombination of curcumin and its analogues with Dox increased sub G-1 cell populations. Curcumin and PGV-1but not PGV-0 decreased localization of p65 into the nucleus induced by Dox, indicating that activation of NFkBwas inhibited. Molecular docking of curcumin, PGV-0, and PGV-1 demonstrated high affinity to HER2 atATP binding site. This interaction were directly comparable with those of ATP and lapatinib. These findingssuggested that curcumin, PGV-0 and PGV-1 enhance the Dox cytotoxicity to MCF-7 cells through inhibition ofHER2 activity and NF-kB activation.     

A phase I dose escalation study of BIBW 2992, an irreversible dual inhibitor of epidermal growth factor receptor 1 (EGFR) and 2 (HER2) tyrosine kinase in a 2-week on, 2-week off schedule in patients with advanced solid tumours

To assess tolerability, pharmacokinetics (PK), pharmacodynamics (PD) and clinical activity of the dual epidermal growth factor receptor (EGFR) 1 and 2 (HER2) tyrosine kinase inhibitor BIBW 2992. An escalating schedule of once-daily (OD) BIBW 2992 for 14 days followed by 14 days off medication was explored. Thirty-eight patients were enrolled. Dose levels were 10, 20, 30, 45, 70, 85, and 100 mg. At 100 mg dose-limiting toxicity (DLT) (common toxicity criteria grade 3 skin rash and grade 3 diarrhoea despite treatment with loperamide) occurred in two patients. In the next-lower dose of 70 mg, DLT (grade 3 fatigue and ALAT elevation) occurred in one of six patients. An intermediate dose level of 85 mg was studied. Here DLT occurred in two patients (grade 3 diarrhoea despite treatment and grade 2 diarrhoea lasting more than 7 days despite treatment). An additional 12 patients were treated at 70 mg. BIBW 2992 PK after single and multiple doses revealed moderately fast absorption, and no deviation from dose proportionality. Pharmacodynamics analysis in skin biopsies did not show significant changes in EGFR-associated biomarkers. However, a significant inhibitory effect on the proliferation index of epidermal keratinocytes was observed. No partial or complete responses were observed, stable disease lasting more than four cycles was seen in seven patients. The recommended dose for studies with BIBW 2992 for 14 days followed by 14 days off medication is 70 mg OD.     

Disease-specific expression of VEGF and its receptors in AML cells: possible autocrine pathway of VEGF/type1 receptor of VEGF in t(15;17) AML and VEGF/type2 receptor of VEGF in t(8;21) AML

Various angiogenic factors, such as vascular endothelial growth factor (VEGF) and an associated molecule, placenta growth factor (PlGF), are thought to be important for normal and malignant hematopoiesis. This study examined mRNA expression of VEGF, PlGF and receptors for these molecules in AML cells and identified the disease-specific patterns of expression. AML M₃ having t(15;17) abnormality showed highest expression of VEGF and VEGF receptor type 1 (VEGFR1), suggesting the autocrine pathway of VEGF-VEGFR1. Then, t(8;21) AML demonstrated augmented expression of VEGF and VEGF receptor type 2 (VEGFR2), suggesting VEGF-VEGFR2 autocrine pathway. Then, addition of VEGFR2 kinase inhibitor in Kasumi-1, a t(8;21) AML cell line, resulted in marked inhibition of cell growth, although growth inhibitory effect of R2 kinase inhibitor to HL-60 was marginal. In addition, cell cycle analysis study showed S-phase cell population reduction by R2 kinase inhibitor in Kasumi-1, but not in HL-60. This observation is thought to be the rationale for novel molecular target therapy directed to angiogenic molecules.     

Human epidermal growth factor receptor-2 and estrogen receptor expression, a demonstration project using the residual tissue repository of the Surveillance, Epidemiology, and End Results (SEER) program

Background In 2001, the National Cancer Institute’s Surveillance, Epidemiology, and End Results (SEER) program established Residual Tissue Repositories (RTR) in the Hawaii, Iowa, and Los Angeles Tumor Registries to collect discarded tissue blocks from pathologic laboratories within their catchment areas. To validate the utility of the RTR for supplementing SEER’s central database, we assessed human epidermal growth factor receptor-2 (HER2) and estrogen receptor expression (ER) in a demonstration project. Materials Using a prepared set of tissue microarrays (TMAs) residing in the Hawaii Tumor Registry (HTR), we performed standard immunohistochemistry. Breast cancers in the TMA were diagnosed in 1995, followed through 2006, and linked to SEER’s main database. Results The TMA included 354 cases, representing 51% of 687 breast cancers in the HTR (1995). The HTR and TMA cases were similar with respect to patient demographics and tumor characteristics. Seventy-six percent (76%, 268 of 354) of TMA cases were HER2+ and/or ER+, i.e., 28 HER2+ER−, 12 HER2+ER+, and 228 HER2−ER+. There were 67 HER2−ER− cases and 19 were unclassified. Age distributions at diagnosis were bimodal with dominant early-onset modes for HER2+ER− tumors and dominant late-onset modes for HER2−ER+ breast cancers. Epidemiologic patterns for concordant HER2+ER+ (double-positive) and HER2−ER− (double-negative) were intermediate to discordant HER2+ER− and HER2−ER+. Conclusion Results showed contrasting incidence patterns for HER2+ (HER2+ER−) and ER+ (HER2−ER+) breast cancers, diagnosed in 1995. Though sample sizes were small, this demonstration project validates the potential utility of the RTR for supplementing the SEER program. The U.S. Government's right to retain a non-exclusive, royalty-free license in and to any copyright is acknowledged.     

Overexpression of NLRP3, NLRC4 and AIM2 inflammasomes and their priming‐associated molecules (TLR2, TLR4, Dectin‐1, Dectin‐2 and NFκB) in Malassezia folliculitis

The activation of NLRP3, NLRC4 and AIM2 inflammasomes is pivotal for innate immunity against some pathogenic fungi, but their role in the pathogenesis of Malassezia folliculitis (MF) remains unclear. The objective of the study was to determine the expression of 4 canonical inflammasomes (NLRP1, NLRP3, NLRC4 and AIM2) and their priming‐associated molecules (TLR2, TLR4, Dectin‐1, Dectin‐2 and NFκB) in MF lesion. Expression of NLRP1, NLRP3, NLRC4, AIM2, caspase‐1, IL‐1β, TLR2, TLR4, Dectin‐1, Dectin‐2 and NFκB was detected by immunohistochemistry in skin lesion of 23 MF patients and normal skin of 12 healthy subjects. Furthermore, NLRP1, NLRP3, NLRC4, AIM2, caspase‐1 and IL‐1β mRNA was measured by quantitative real‐time PCR (qRT‐PCR) in 12 MF cases and 10 controls. Immunohistochemical analysis revealed that NLRP3, NLRC4, AIM2, Casp‐1, IL‐1β, TLR2, TLR4, Dectin‐1, Dectin‐2 and NFκB expression was up‐regulated in the epidermis and dermal inflammatory cells of MF lesion compared with control skin (P < .01‐.05), but NLRP1 expression was not different between both groups (P > .05). qRT‐PCR showed that levels of NLRP3, Casp‐1 and IL‐1β mRNA were significantly increased (P < .01‐.05), whereas those of NLRP1, NLRC4 and AIM2 mRNA were slightly augmented compared to control skin (P > .05). Our observation suggests that simultaneous activation of NLRP3, NLRC4 and AIM2 inflammasomes may play an important role in the pathogenesis of MF.