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1.
Sci Rep ; 11(1): 16866, 2021 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-34654856

RESUMO

Prognosis of patients with HER2+ breast-to-brain-metastasis (BBM) is dismal even after current standard-of-care treatments, including surgical resection, whole-brain radiation, and systemic chemotherapy. Radiation and systemic chemotherapies can also induce cytotoxicity, leading to significant side effects. Studies indicate that donor-derived platelets can serve as immune-compatible drug carriers that interact with and deliver drugs to cancer cells with fewer side effects, making them a promising therapeutic option with enhanced antitumor activity. Moreover, human induced pluripotent stem cells (hiPSCs) provide a potentially renewable source of clinical-grade transfusable platelets that can be drug-loaded to complement the supply of donor-derived platelets. Here, we describe methods for ex vivo generation of megakaryocytes (MKs) and functional platelets from hiPSCs (hiPSC-platelets) in a scalable fashion. We then loaded hiPSC-platelets with lapatinib and infused them into BBM tumor-bearing NOD/SCID mouse models. Such treatment significantly increased intracellular lapatinib accumulation in BBMs in vivo, potentially via tumor cell-induced activation/aggregation. Lapatinib-loaded hiPSC-platelets exhibited normal morphology and function and released lapatinib pH-dependently. Importantly, lapatinib delivery to BBM cells via hiPSC-platelets inhibited tumor growth and prolonged survival of tumor-bearing mice. Overall, use of lapatinib-loaded hiPSC-platelets effectively reduced adverse effects of free lapatinib and enhanced its therapeutic efficacy, suggesting that they represent a novel means to deliver chemotherapeutic drugs as treatment for BBM.


Assuntos
Neoplasias Encefálicas/tratamento farmacológico , Neoplasias da Mama/tratamento farmacológico , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Lapatinib/farmacologia , Metástase Neoplásica/patologia , Receptor ErbB-2/efeitos dos fármacos , Animais , Antineoplásicos/farmacologia , Neoplasias Encefálicas/secundário , Neoplasias da Mama/patologia , Portadores de Fármacos/farmacologia , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Camundongos Endogâmicos NOD , Camundongos SCID , Quinazolinas/farmacologia , Receptor ErbB-2/metabolismo
2.
Breast Cancer Res Treat ; 189(3): 665-676, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34553296

RESUMO

PURPOSE: Neratinib, an irreversible pan-HER tyrosine kinase inhibitor, has demonstrated systemic efficacy and intracranial activity in various stages of HER2+breast cancer. NALA was a phase III randomized trial that assessed the efficacy and safety of neratinib+capecitabine (N+C) against lapatinib+capecitabine (L+C) in HER2+ metastatic breast cancer (mBC) patients who had received ≥ 2 HER2-directed regimens. Descriptive analysis results of the Asian subgroup in the NALA study are reported herein. METHODS: 621 centrally assessed HER2+ mBC patients were enrolled, 202 of whom were Asian. Those with stable, asymptomatic brain metastases (BM) were eligible for study entry. Patients were randomized 1:1 to N (240 mg qd) + C (750 mg/m2 bid, day 1-14) with loperamide prophylaxis or to L (1250 mg qd) + C (1000 mg/m2 bid, day 1-14) in 21-day cycles. Co-primary endpoints were centrally assessed progression-free survival (PFS) and overall survival (OS). Secondary endpoints included time to intervention for central nervous system (CNS) disease, objective response rate, duration of response (DoR), clinical benefit rate, and safety. RESULTS: 104 and 98 Asian patients were randomly assigned to receive N+C or L+C, respectively. Median PFS of N+C and L+C was 7.0 and 5.4 months (P = 0.0011), respectively. Overall cumulative incidence of intervention for CNS disease was lower with N+C (27.9 versus 33.8%; P = 0.039). Both median OS (23.8 versus 18.7 months; P = 0.185) and DoR (11.1 versus 4.2 months; P < 0.0001) were extended with N+C, compared to L+C. The incidences of grade 3/4 treatment emergent adverse events (TEAEs) and TEAEs leading to treatment discontinuation were mostly comparable between the two arms. Diarrhea and palmar-plantar erythrodysesthesia were the most frequent TEAEs in both arms, similar to the overall population in incidence and severity. CONCLUSION: Consistent with the efficacy profile observed in the overall study population, Asian patients with HER2+ mBC, who had received ≥ 2 HER2-directed regimens, may also benefit from N+C. No new safety signals were noted. CLINICAL TRIAL REGISTRATION: NCT01808573.


Assuntos
Neoplasias da Mama , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Capecitabina/uso terapêutico , Feminino , Humanos , Lapatinib/uso terapêutico , Quinolinas , Receptor ErbB-2/genética , Resultado do Tratamento
3.
PLoS One ; 16(8): e0254205, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34347777

RESUMO

Conventional frontline treatment for ovarian cancer consists of successive chemotherapy cycles of paclitaxel and platinum. Despite the initial favorable responses for most patients, chemotherapy resistance frequently leads to recurrent or refractory disease. New treatment strategies that circumvent or prevent mechanisms of resistance are needed to improve ovarian cancer therapy. We established in vitro paclitaxel-resistant ovarian cancer cell line and organoid models. Gene expression differences in resistant and sensitive lines were analyzed by RNA sequencing. We manipulated candidate genes associated with paclitaxel resistance using siRNA or small molecule inhibitors, and then screened the cells for paclitaxel sensitivity using cell viability assays. We used the Bliss independence model to evaluate the anti-proliferative synergy for drug combinations. ABCB1 expression was upregulated in paclitaxel-resistant TOV-21G (q < 1x10-300), OVCAR3 (q = 7.4x10-156) and novel ovarian tumor organoid (p = 2.4x10-4) models. Previous reports have shown some tyrosine kinase inhibitors can inhibit ABCB1 function. We tested a panel of tyrosine kinase inhibitors for the ability to sensitize resistant ABCB1-overexpressing ovarian cancer cell lines to paclitaxel. We observed synergy when we combined poziotinib or lapatinib with paclitaxel in resistant TOV-21G and OVCAR3 cells. Silencing ABCB1 expression in paclitaxel-resistant TOV-21G and OVCAR3 cells reduced paclitaxel IC50 by 20.7 and 6.2-fold, respectively. Furthermore, we demonstrated direct inhibition of paclitaxel-induced ABCB1 transporter activity by both lapatinib and poziotinib. In conclusion, lapatinib and poziotinib combined with paclitaxel synergizes to inhibit the proliferation of ABCB1-overexpressing ovarian cancer cells in vitro. The addition of FDA-approved lapatinib to second-line paclitaxel therapy is a promising strategy for patients with recurrent ovarian cancer.


Assuntos
Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Lapatinib/farmacologia , Proteínas de Neoplasias/metabolismo , Neoplasias Ovarianas , Quinazolinas/farmacologia , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/genética , Feminino , Humanos , Proteínas de Neoplasias/genética , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/metabolismo
4.
Aging (Albany NY) ; 13(16): 20793-20807, 2021 08 30.
Artigo em Inglês | MEDLINE | ID: mdl-34459788

RESUMO

PURPOSE: Apatinib resistance is the main obstacle to the effective treatment of advanced head and neck squamous cell carcinoma (HNSCC). This study aimed to evaluate the function of Erb-B2 receptor tyrosine kinase 2 (ERBB2) and stimulator of interferon response cGAMP interactor (STING) in apatinib resistance in HNSCC. METHOD: The Cancer Genome Atlas database of HNSCC was used to analyze the relationship between vascular endothelial growth factor receptor 2 (VEGFR2) expression and prognosis. An apatinib resistant (AR) HNSCC cell line was constructed based on the CAL27 cell line. RNA sequencing was performed to explore the differentially expressed mRNAs. Quantitative real-time reverse transcription PCR (qRT-PCR) and western blotting were used to evaluate the expression and phosphorylation level VEGFR2, ERBB2, STING, and related proteins. Apatinib resistance was evaluated by colony formation and cell viability assays. A mouse subcutaneous tumor formation model was established to evaluate the efficiency of combination treatment and vascularization was evaluated by assessing CD31 immunofluorescence. RESULT: The expression of VEGFR2 was high in tumor of patients with HNSCC. Western blotting and qRT-PCR revealed that in AR cells, ERBB2 expression was high, whereas the expression of STING was low. Targeted treatment of ERBB2 using lapatinib could attenuate apatinib resistance. Further research confirmed that overexpressing STING could decrease ERBB2 expression. CONCLUSION: STING could sensitize AR cells to apatinib by decreasing ERBB2 expression. The combination of lapatinib or a STING agonist with apatinib ameliorated acquired apatinib resistance in a synergistic manner.


Assuntos
Antineoplásicos/uso terapêutico , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Neoplasias de Cabeça e Pescoço/metabolismo , Proteínas de Membrana/metabolismo , Piridinas/uso terapêutico , Receptor ErbB-2/metabolismo , Carcinoma de Células Escamosas de Cabeça e Pescoço/metabolismo , Animais , Linhagem Celular Tumoral , Neoplasias de Cabeça e Pescoço/tratamento farmacológico , Humanos , Interferons , Lapatinib/uso terapêutico , Masculino , Camundongos Endogâmicos BALB C , Nucleotídeos Cíclicos , Carcinoma de Células Escamosas de Cabeça e Pescoço/tratamento farmacológico , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Xantonas/uso terapêutico , Ensaios Antitumorais Modelo de Xenoenxerto
5.
Colloids Surf B Biointerfaces ; 207: 112012, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34352656

RESUMO

Lapatinib, a dual tyrosine kinase inhibitor, has poor water solubility, which results in poor and incomplete absorption from the gastrointestinal tract. To overcome this obstacle, we designed a stable and high-loaded liposomal formulation encapsulating lapatinib and examined its therapeutic efficacy in vitro and in vivo on TUBO and 4T1 cell lines. We also assessed the impact of liposomal lapatinib on the extent of the tumor and spleen-infiltrating lymphocytes and the autophagy and apoptosis gene expression within the tumor site. Our results showed that liposomal lapatinib inhibits cell proliferation and significantly induces autophagy and apoptosis compared to control groups. Moreover, when it used in combination with liposomal doxorubicin, it extended the time to end from 22.4 ± 3.5 in the control group to 40 days in the TUBO cell line and from 29.2 ± 1.7 to 38.6 ± 2.2 days in 4T1 triple-negative breast cancer cell line, which reveals its promising effects on the survival of tumor-bearing mice. Our results indicated the need for further evaluations to understand liposomal lapatinib's potential effects on autophagy, apoptosis, and particularly on immune system cells.


Assuntos
Antineoplásicos , Neoplasias da Mama , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Linhagem Celular Tumoral , Doxorrubicina/análogos & derivados , Feminino , Humanos , Lapatinib , Camundongos , Polietilenoglicóis , Quinazolinas/farmacologia , Quinazolinas/uso terapêutico
6.
Cancer Sci ; 112(10): 4234-4245, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34382727

RESUMO

Development of acquired resistance to lapatinib, a dual epidermal growth factor receptor (EGFR)/human epidermal growth factor receptor 2 (HER2) tyrosine kinase inhibitor, severely limits the duration of clinical response in advanced HER2-driven breast cancer patients. Although the compensatory activation of the PI3K/Akt survival signal has been proposed to cause acquired lapatinib resistance, comprehensive molecular mechanisms remain required to develop more efficient strategies to circumvent this therapeutic difficulty. In this study, we found that suppression of HER2 by lapatinib still led to Akt inactivation and elevation of FOX3a protein levels, but failed to induce the expression of their downstream pro-apoptotic effector p27kip1 , a cyclin-dependent kinase inhibitor. Elevation of miR-221 was found to contribute to the development of acquired lapatinib resistance by targeting p27kip1 expression. Furthermore, upregulation of miR-221 was mediated by the lapatinib-induced Src family tyrosine kinase and subsequent NF-κB activation. The reversal of miR-221 upregulation and p27kip1 downregulation by a Src inhibitor, dasatinib, can overcome lapatinib resistance. Our study not only identified miRNA-221 as a pivotal factor conferring the acquired resistance of HER2-positive breast cancer cells to lapatinib through negatively regulating p27kip1 expression, but also suggested Src inhibition as a potential strategy to overcome lapatinib resistance.


Assuntos
Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Resistencia a Medicamentos Antineoplásicos/fisiologia , Lapatinib/farmacologia , MicroRNAs/metabolismo , Receptor ErbB-2/antagonistas & inibidores , Animais , Neoplasias da Mama/química , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Inibidor de Quinase Dependente de Ciclina p27/efeitos dos fármacos , Dasatinibe/farmacologia , Regulação para Baixo/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Feminino , Proteína Forkhead Box O3/metabolismo , Fator 3-gama Nuclear de Hepatócito/metabolismo , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/efeitos dos fármacos , Análise em Microsséries , Subunidade p50 de NF-kappa B/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Regulação para Cima/efeitos dos fármacos , Quinases da Família src/antagonistas & inibidores , Quinases da Família src/metabolismo
7.
BMC Cancer ; 21(1): 923, 2021 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-34399705

RESUMO

BACKGROUND: Gastric cancer (GC) is one of the leading causes of cancer-related deaths worldwide. Human epidermal growth factor receptor 2 (HER2) amplification occurs in approximately 13-23% of all GC cases and patients with HER2 overexpression exhibit a poor prognosis. Lapatinib, a dual EGFR/HER2 tyrosine kinase inhibitor, is an effective agent to treat HER2-amplified breast cancer but it failed in gastric cancer (GC) clinical trials. However, the molecular mechanism of lapatinib resistance in HER2-amplified GC is not well studied. METHODS: We employed an unbiased, genome-scale screening with pooled CRISPR library on HER2-amplified GC cell lines to identify genes that are associated with resistance to lapatinib. To validate the candidate genes, we applied in vitro and in vivo pharmacological tests to confirm the function of the target genes. RESULTS: We found that loss of function of CSK or PTEN conferred lapatinib resistance in HER2-amplified GC cell lines NCI-N87 and OE19, respectively. Moreover, PI3K and MAPK signaling was significantly increased in CSK or PTEN null cells. Furthermore, in vitro and in vivo pharmacological study has shown that lapatinib resistance by the loss of function of CSK or PTEN, could be overcome by lapatinib combined with the PI3K inhibitor copanlisib and MEK inhibitor trametinib. CONCLUSIONS: Our study suggests that loss-of-function mutations of CSK and PTEN cause lapatinib resistance by re-activating MAPK and PI3K pathways, and further proved these two pathways are druggable targets. Inhibiting the two pathways synergistically are effective to overcome lapatinib resistance in HER2-amplified GC. This study provides insights for understanding the resistant mechanism of HER2 targeted therapy and novel strategies that may ultimately overcome resistance or limited efficacy of lapatinib treatment for subset of HER2 amplified GC.


Assuntos
Biomarcadores Tumorais/metabolismo , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Receptor ErbB-2/metabolismo , Neoplasias Gástricas/tratamento farmacológico , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Apoptose , Biomarcadores Tumorais/genética , Proliferação de Células , Feminino , Perfilação da Expressão Gênica , Humanos , Lapatinib/administração & dosagem , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Piridonas/administração & dosagem , Pirimidinas/administração & dosagem , Pirimidinonas/administração & dosagem , Quinazolinas/administração & dosagem , Receptor ErbB-2/genética , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
8.
Methods Mol Biol ; 2342: 809-823, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34272718

RESUMO

Often it may be convenient and efficient to address multiple research questions with a single experiment. In many instances, however, the best approach is to design the experiment to address one question at a time. The design of enzyme mapping experiments is discussed in this chapter, focusing on considerations pertinent to the study of aldehyde oxidase (AO) vs. cytochrome P450 metabolism. Specifically, a case is presented in which reduced glutathione (GSH) was included in an experiment with human liver S9 fraction to trap reactive metabolites generated from cytochrome P450-mediated metabolism of lapatinib and its O-dealkylated metabolite, M1 (question 1). The AO inhibitor hydralazine was included in this experiment to investigate the involvement of AO-mediated metabolism of M1 (question 2). The presence of GSH was found to interfere with the inhibitory activity of hydralazine. Consideration of the time-dependent nature of hydralazine inhibitory activity toward AO when designing this experiment could have predicted the potential for GSH to interfere with hydralazine. This case underscores the importance of clearly identifying the research question, tailoring the experimental protocol to answer that question, and then meticulously considering how the experimental conditions could influence the results, particularly if attempting to address multiple questions with a single experiment.


Assuntos
Aldeído Oxidase/metabolismo , Sistema Enzimático do Citocromo P-450/metabolismo , Glutationa/metabolismo , Hidralazina/farmacocinética , Lapatinib/farmacocinética , Ativação Metabólica , Interações Medicamentosas , Hepatócitos/citologia , Humanos , Microssomos Hepáticos/enzimologia , Oxirredução , Projetos de Pesquisa , Fatores de Tempo
9.
Cell Death Dis ; 12(7): 684, 2021 07 08.
Artigo em Inglês | MEDLINE | ID: mdl-34238922

RESUMO

Exosomes are carriers of intercellular information that regulate the tumor microenvironment, and they have an essential role in drug resistance through various mechanisms such as transporting RNA molecules and proteins. Nevertheless, their effects on gemcitabine resistance in triple-negative breast cancer (TNBC) are unclear. In the present study, we examined the effects of exosomes on TNBC cell viability, colony formation, apoptosis, and annexin A6 (ANXA6)/EGFR expression. We addressed their roles in gemcitabine resistance and the underlying mechanism. Our results revealed that exosomes derived from resistant cancer cells improved cell viability and colony formation and inhibited apoptosis in sensitive cancer cells. The underlying mechanism included the transfer of exosomal ANXA6 from resistant cancer cells to sensitive cancer cells. Isobaric peptide labeling-liquid chromatography-tandem mass spectrometry and western blotting revealed that ANXA6 was upregulated in resistant cancer cells and their derived exosomes. Sensitive cancer cells exhibited resistance with increased viability and colony formation and decreased apoptosis when ANXA6 was stably overexpressed. On the contrary, knockdown ANXA6 restored the sensitivity of cells to gemcitabine. Co-immunoprecipitation expression and GST pulldown assay demonstrated that exosomal ANXA6 and EGFR could interact with each other and exosomal ANXA6 was associated with the suppression of EGFR ubiquitination and downregulation. While adding lapatinib reversed gemcitabine resistance induced by exosomal ANXA6. Moreover, ANXA6 and EGFR protein expression was correlated in TNBC tissues, and exosomal ANXA6 levels at baseline were lower in patients with highly sensitive TNBC than those with resistant TNBC when treated with first-line gemcitabine-based chemotherapy. In conclusion, resistant cancer cell-derived exosomes induced gemcitabine resistance via exosomal ANXA6, which was associated with the inhibition of EGFR ubiquitination and degradation. Exosomal ANXA6 levels in the serum of patients with TNBC might be predictive of the response to gemcitabine-based chemotherapy.


Assuntos
Anexina A6/metabolismo , Antimetabólitos Antineoplásicos/farmacologia , Desoxicitidina/análogos & derivados , Resistencia a Medicamentos Antineoplásicos , Exossomos/metabolismo , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Anexina A6/genética , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Desoxicitidina/farmacologia , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/metabolismo , Exossomos/genética , Humanos , Lapatinib/farmacologia , Proteólise , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/metabolismo , Neoplasias de Mama Triplo Negativas/patologia , Ubiquitinação
10.
Biochem Biophys Res Commun ; 567: 154-160, 2021 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-34157442

RESUMO

Epidermal growth factor receptor (EGFR) is a receptor tyrosine kinase and mutations in EGFR is a major driver force of lung cancer. EGFR tyrosine kinase inhibitors (TKIs) are group of promising agents to treat cancer patients with EGFR mutations. However, the application of TKIs is often hampered by the development of drug-resistance. In the present study, we studied the role of Glutathione peroxidase 4 (GPX4) and mammalian target of rapamycin (mTOR) in regulation of lung cancer cells response to Lapatinib (Lap). Lap resistant NSCLC cells A549/Lap and H1944/Lap were created and GPX4 was knockdown by lentivirus shGPX4. Change of cell viabilities and cell death were measured by MTT and flow cytometry, respectively. ROS, MDA, GSH and Fe2+ were detected by commercial kits. Xenograft mice was used to assay the in vivo effects of GPX4 on the sensitivity of Lap. We found that GPX4 and mTORC1 signalling was upregulated in Lap resistant NSCLC cells when compared to Lap sensitive NSCLC cells. Mechanistically, upregulation of GPX4 was due to enhanced activation of mTORC1 in Lap resistant NSCLC cells. Inhibition of mTORC1 led to the downregulation of GPX4 which promoted Lap induced ferroptosis as evidenced by increase of ROS, MDA, Fe 2+ and decrease of GSH. Rescue experiments confirmed the role of GPX4 in regulation of Lap induced ferroptosis. In vivo experiments also indicated that silencing of GPX4 enhanced the anticancer effect of Lap via promoting ferroptosis. Overall, targeting GPX4 might be a potential strategy to enhance antitumor effects of Lap.


Assuntos
Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Lapatinib/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Fosfolipídeo Hidroperóxido Glutationa Peroxidase/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Células A549 , Animais , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Resistencia a Medicamentos Antineoplásicos , Ferroptose/efeitos dos fármacos , Inativação Gênica , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Masculino , Camundongos Endogâmicos BALB C , Camundongos Nus , Fosfolipídeo Hidroperóxido Glutationa Peroxidase/genética , Serina-Treonina Quinases TOR/genética
11.
BMC Cancer ; 21(1): 652, 2021 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-34074257

RESUMO

BACKGROUND: Even though targeted therapies are available for cancers expressing oncogenic epidermal growth receptor (EGFR) and (or) human EGFR2 (HER2), acquired or intrinsic resistance often confounds therapy success. Common mechanisms of therapy resistance involve activating receptor point mutations and (or) upregulation of signaling downstream of EGFR/HER2 to Akt and (or) mitogen activated protein kinase (MAPK) pathways. However, additional pathways of resistance may exist thus, confounding successful therapy. METHODS: To determine novel mechanisms of EGFR/HER2 therapy resistance in breast cancer, gefitinib or lapatinib resistant variants were created from SKBR3 breast cancer cells. Syngenic therapy sensitive and resistant SKBR3 variants were characterized for mechanisms of resistance by mammosphere assays, viability assays, and western blotting for total and phospho proteins. RESULTS: Gefitinib and lapatinib treatments reduced mammosphere formation in the sensitive cells, but not in the therapy resistant variants, indicating enhanced mesenchymal and cancer stem cell-like characteristics in therapy resistant cells. The therapy resistant variants did not show significant changes in known therapy resistant pathways of AKT and MAPK activities downstream of EGFR/HER2. However, these cells exhibited elevated expression and activation of the small GTPase Rac, which is a pivotal intermediate of GFR signaling in EMT and metastasis. Therefore, the potential of the Rac inhibitors EHop-016 and MBQ-167 to overcome therapy resistance was tested, and found to inhibit viability and induce apoptosis of therapy resistant cells. CONCLUSIONS: Rac inhibition may represent a viable strategy for treatment of EGFR/HER2 targeted therapy resistant breast cancer.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Neoplasias da Mama/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Proteínas rac de Ligação ao GTP/antagonistas & inibidores , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Apoptose , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Carbazóis/farmacologia , Carbazóis/uso terapêutico , Linhagem Celular Tumoral , Ensaios de Seleção de Medicamentos Antitumorais , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/genética , Feminino , Mutação com Ganho de Função , Gefitinibe/farmacologia , Gefitinibe/uso terapêutico , Regulação Neoplásica da Expressão Gênica , Humanos , Lapatinib , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/genética , Mutação Puntual , Inibidores de Proteínas Quinases/uso terapêutico , Pirimidinas/farmacologia , Pirimidinas/uso terapêutico , Receptor ErbB-2/antagonistas & inibidores , Receptor ErbB-2/genética , Esferoides Celulares , Regulação para Cima
12.
Biochem Pharmacol ; 190: 114635, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34058187

RESUMO

Since trastuzumab-resistance remains a major obstacle to the successful treatment of HER2-positive breast cancer, a detailed understanding of the mechanisms responsible is required to direct future pharmacotherapeutic strategies. Recently, several studies have indicated that the quiescent natures of cancer stem cells contribute to treatment resistance and tumor recurrence. Thus, in this study, we investigated the mechanism underlying trastuzumab resistance in a quiescent cell population using tumorsphere cultures and explored better therapeutic strategies to overcome trastuzumab resistance in HER2-positive breast cancer patients. We observed that most cells in SK-BR-3 tumorspheres were quiescent, showing the accumulation of cells at the G0/G1 phase as compared to cells in monolayer culture. Furthermore, SK-BR-3 tumorspheres exhibited enhanced EGFR/HER2 signaling, which was incompletely inhibited by trastuzumab, and subsequently led to trastuzumab-resistance. Interestingly, cytoplasmic estrogen receptor α (ERα) expression was markedly elevated in tumorspheres and was associated with enhanced EGFR/HER2 signaling. Accordingly, inhibition of ERα with tamoxifen selectively targeted tumorspheres rather than cells in monolayer culture and overcame trastuzumab resistance in tumorspheres. Taken together, our findings indicate that crosstalk between cytoplasmic ERα and the HER2/EGFR signaling pathway can be considered a novel therapeutic target for quiescent cell populations within HER2-positive breast cancer and that simultaneous inhibition of ER and the EGFR/HER2 pathway may prevent trastuzumab resistance. We hope that these results provide a basis for the use of combinations of tamoxifen and trastuzumab in HER2-positive breast cancer patients.


Assuntos
Resistencia a Medicamentos Antineoplásicos , Receptores ErbB/metabolismo , Receptor alfa de Estrogênio/metabolismo , Receptor ErbB-2/metabolismo , Tamoxifeno/farmacologia , Trastuzumab/farmacologia , Antineoplásicos Hormonais/farmacologia , Antineoplásicos Imunológicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Linhagem Celular Tumoral , Citoplasma , Receptores ErbB/genética , Estradiol/farmacologia , Receptor alfa de Estrogênio/genética , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Lapatinib/farmacologia , Receptor ErbB-2/genética , Transdução de Sinais
13.
J Transl Med ; 19(1): 184, 2021 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-33933113

RESUMO

BACKGROUND: Aberrant PI3K signalling is implicated in trastuzumab resistance in HER2-positive gastric cancer (GC). The role of PI3K or MEK inhibitors in sensitising HER2-positive GCs to trastuzumab or in overcoming trastuzumab resistance is unclear. METHODS: Using mass spectrometry-based genotyping we analysed 105 hotspot, non-synonymous somatic mutations in PIK3CA and ERBB-family (EGFR, ERBB2, ERBB3 and ERBB4) genes in gastric tumour samples from 69 patients. A panel of gastric cell lines (N87, OE19, ESO26, SNU16, KATOIII) were profiled for anti-proliferative response to the PI3K inhibitor copanlisib and the MEK1/2 inhibitor refametinib alone and in combination with anti-HER2 therapies. RESULTS: Patients with HER2-positive GC had significantly poorer overall survival compared to HER2-negative patients (15.9 months vs. 35.7 months). Mutations in PIK3CA were only identified in HER2-negative tumours, while ERBB-family mutations were identified in HER2-positive and HER2-negative tumours. Copanlisib had anti-proliferative effects in 4/5 cell lines, with IC50s ranging from 23.4 (N87) to 93.8 nM (SNU16). All HER2-positive cell lines except SNU16 were sensitive to lapatinib (IC50s 0.04 µM-1.5 µM). OE19 cells were resistant to trastuzumab. The combination of lapatinib and copanlisib was synergistic in ESO-26 and OE-19 cells (ED50: 0.83 ± 0.19 and 0.88 ± 0.13, respectively) and additive in NCI-N87 cells (ED50:1.01 ± 0.55). The combination of copanlisib and trastuzumab significantly improved growth inhibition compared to either therapy alone in NCI-N87, ESO26 and OE19 cells (p < 0.05). CONCLUSIONS: PI3K or MEK inhibition alone or in combination with anti-HER2 therapy may represent an improved treatment strategy for some patients with HER2-positive GC, and warrants further investigation in a clinical trial setting.


Assuntos
Neoplasias Gástricas , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/genética , Humanos , Lapatinib , Fosfatidilinositol 3-Quinases , Receptor ErbB-2/genética , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/genética , Trastuzumab/farmacologia , Trastuzumab/uso terapêutico
14.
PLoS One ; 16(5): e0251163, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33951110

RESUMO

Our previous pre-clinical work defined BCL-2 induction as a critical component of the adaptive response to lapatinib-mediated inhibition of HER2. To determine whether a similar BCL-2 upregulation occurs in lapatinib-treated patients, we evaluated gene expression within tumor biopsies, collected before and after lapatinib or trastuzumab treatment, from the TRIO-B-07 clinical trial (NCT#00769470). We detected BCL2 mRNA upregulation in both HER2+/ER- as well as HER2+/ER+ patient tumors treated with lapatinib or trastuzumab. To address whether mRNA expression correlated with protein expression, we evaluated pre- and post-treatment tumors for BCL-2 via immunohistochemistry. Despite BCL2 mRNA upregulation within HER2+/ER- tumors, BCL-2 protein levels were undetectable in most of the lapatinib- or trastuzumab-treated HER2+/ER- tumors. BCL-2 upregulation was evident within the majority of lapatinib-treated HER2+/ER+ tumors and was often coupled with increased ER expression and decreased proliferation. Comparable BCL-2 upregulation was not observed within the trastuzumab-treated HER2+/ER+ tumors. Together, these results provide clinical validation of the BCL-2 induction associated with the adaptive response to lapatinib and support evaluation of BCL-2 inhibitors within the context of lapatinib and other HER2-targeted receptor tyrosine kinase inhibitors.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Receptor ErbB-2/metabolismo , Adulto , Idoso , Anticorpos Monoclonais Humanizados/uso terapêutico , Antineoplásicos/uso terapêutico , Biomarcadores Tumorais/metabolismo , Proliferação de Células/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Lapatinib/uso terapêutico , Pessoa de Meia-Idade , Terapia Neoadjuvante/métodos , Inibidores de Proteínas Quinases/uso terapêutico , Quinazolinas/uso terapêutico , RNA Mensageiro/metabolismo , Trastuzumab/uso terapêutico , Regulação para Cima/efeitos dos fármacos
15.
Sci Rep ; 11(1): 10893, 2021 05 25.
Artigo em Inglês | MEDLINE | ID: mdl-34035375

RESUMO

HER2-positive (HER2 +) breast cancer patients that do not respond to targeted treatment have a poor prognosis. The effects of targeted treatment on endogenous microRNA (miRNA) expression levels are unclear. We report that responsive HER2 + breast cancer cell lines had a higher number of miRNAs with altered expression after treatment with trastuzumab and lapatinib compared to poorly responsive cell lines. To evaluate whether miRNAs can sensitize HER2 + cells to treatment, we performed a high-throughput screen of 1626 miRNA mimics and inhibitors in combination with trastuzumab and lapatinib in HER2 + breast cancer cells. We identified eight miRNA mimics sensitizing cells to targeted treatment, miR-101-5p, mir-518a-5p, miR-19b-2-5p, miR-1237-3p, miR-29a-3p, miR-29c-3p, miR-106a-5p, and miR-744-3p. A higher expression of miR-101-5p predicted better prognosis in patients with HER2 + breast cancer (OS: p = 0.039; BCSS: p = 0.012), supporting the tumor-suppressing role of this miRNA. In conclusion, we have identified miRNAs that sensitize HER2 + breast cancer cells to targeted therapy. This indicates the potential of combining targeted drugs with miRNAs to improve current treatments for HER2 + breast cancers.


Assuntos
Neoplasias da Mama/genética , Perfilação da Expressão Gênica/métodos , Lapatinib/farmacologia , MicroRNAs/genética , Trastuzumab/farmacologia , Neoplasias da Mama/tratamento farmacológico , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Prognóstico , Análise de Sobrevida , Regulação para Cima
16.
Brief Bioinform ; 22(5)2021 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-33857297

RESUMO

Signalling transduction pathways (STPs) are commonly hijacked by many cancers for their growth and malignancy, but demystifying their underlying mechanisms is difficult. Here, we developed methodologies with a fully Bayesian approach in discovering novel driver bio-markers in aberrant STPs given high-throughput gene expression (GE) data. This project, namely 'PathTurbEr' (Pathway Perturbation Driver) uses the GE dataset derived from the lapatinib (an EGFR/HER dual inhibitor) sensitive and resistant samples from breast cancer cell lines (SKBR3). Differential expression analysis revealed 512 differentially expressed genes (DEGs) and their pathway enrichment revealed 13 highly perturbed singalling pathways in lapatinib resistance, including PI3K-AKT, Chemokine, Hippo and TGF-$\beta $ singalling pathways. Next, the aberration in TGF-$\beta $ STP was modelled as a causal Bayesian network (BN) using three MCMC sampling methods, i.e. Neighbourhood sampler (NS) and Hit-and-Run (HAR) sampler that potentially yield robust inference with lower chances of getting stuck at local optima and faster convergence compared to other state-of-art methods. Next, we examined the structural features of the optimal BN as a statistical process that generates the global structure using $p_1$-model, a special class of Exponential Random Graph Models (ERGMs), and MCMC methods for their hyper-parameter sampling. This step enabled key drivers identification that drive the aberration within the perturbed BN structure of STP, and yielded 34, 34 and 23 perturbation driver genes out of 80 constituent genes of three perturbed STP models of TGF-$\beta $ signalling inferred by NS, HAR and MH sampling methods, respectively. Functional-relevance and disease-relevance analyses suggested their significant associations with breast cancer progression/resistance.


Assuntos
Teorema de Bayes , Biomarcadores Tumorais/genética , Neoplasias da Mama/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Lapatinib/uso terapêutico , Algoritmos , Antineoplásicos/uso terapêutico , Neoplasias da Mama/genética , Biologia Computacional/métodos , Feminino , Perfilação da Expressão Gênica/métodos , Ontologia Genética , Redes Reguladoras de Genes/efeitos dos fármacos , Redes Reguladoras de Genes/genética , Humanos , Modelos Genéticos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética
17.
Breast Cancer Res Treat ; 188(2): 477-487, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33852121

RESUMO

PURPOSE: To compare efficacy and safety of capecitabine and lapatinib with or without IMC-A12 (cituxumumab) in patients with HER2-positive metastatic breast cancer (MBC) previously treated with trastuzumab. PATIENTS AND METHODS: Following an initial safety run-in cohort, patients were randomized 1:2 to Arm A (capecitabine and lapatinib) or to Arm B (capecitabine, lapatinib, and cituxumumab). Given the frequency of non-hematologic grade ≥ 3 adverse events in those receiving the three-drug combination in the safety cohort, lapatinib and capecitabine doses were reduced in Arm B only. The primary objective was to determine if the addition of cituxumumab to capecitabine and lapatinib improved progression-free survival (PFS) compared with capecitabine and lapatinib. Secondary objectives included a comparison between arms of other clinical endpoints, safety, change in overall quality of life (QOL) and self-assessed fatigue, rash, diarrhea, and hand-foot syndrome. RESULTS: From July 2008 to March 2012, 68 patients (out of 142 planned) were enrolled and 63 were evaluable, including 8 for the safety run-in and 55 for the randomized cohort. Study enrollment was stopped early due to slow accrual. The addition of cituxumumab to capecitabine and lapatinib did not improve PFS (HR 0.93, 95% CI: 0.52-1.64). Furthermore, no difference in objective response rate or overall survival (OS) was observed. No difference between arms was observed in grade ≥ 3 adverse events, overall QOL change from baseline after 4 cycles of treatment. CONCLUSION: The addition of cituxumumab to lapatinib and capecitabine did not improve PFS or OS compared with lapatinib and capecitabine in patients with HER2-positive MBC. CLINICAL TRIAL REGISTRY: ClinicalTrials.gov Identifier: NCT00684983.


Assuntos
Neoplasias da Mama , Qualidade de Vida , Anticorpos Monoclonais Humanizados , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Neoplasias da Mama/tratamento farmacológico , Capecitabina/uso terapêutico , Intervalo Livre de Doença , Feminino , Humanos , Lapatinib/uso terapêutico , Quinazolinas/efeitos adversos , Receptor ErbB-2/genética , Trastuzumab/uso terapêutico
18.
Sci Rep ; 11(1): 9091, 2021 04 27.
Artigo em Inglês | MEDLINE | ID: mdl-33907275

RESUMO

HER2 and HER3 play key driving functions in the pathophysiology of HER2-amplified breast cancers, but this function is less well characterized in other cancers driven by HER2 amplification. This study aimed to explore the role of HER2 and HER3 signaling in other types of HER2-amplified cancer. The expression and signaling activity of HER2, HER3, and downstream pathway proteins were studied in cell panels representing HER2-amplified cancers of the breast, bladder, colon and rectal, stomach, esophagus, lung, tongue, and endometrium along with controls lacking HER2 amplification. We report that HER2-amplified cancers are addicted to HER2 across different cancer types and the depth of addiction is best linked with the expression level of HER2, but not with HER3 expression. We report that the expression and constitutive phosphorylation of HER3 are ubiquitous in HER2-amplified breast cancer cell lines, but much more variable in HER2-amplified cancer cells from other tissues. We observed the lapatinib-induced compensatory upregulation of HER3 signaling in many types of HER2-amplified cancers, although with much variability. We find that HER3 expression is essential for in vivo tumorigenic growth in some HER2-amplified tumors but not others. Importantly HER3 expression level does not correlate well with its functional importance. More biomarkers will be needed to guide the optimal use of HER3 inhibitors in HER2-amplified cancers from non-breast origin. Unlike oncogenes activated through mutational events, the activation of HER2 through overexpression represents a gradient of activities and depth of addiction and the response to inhibitors follows a similar gradient.


Assuntos
Neoplasias/genética , Receptor ErbB-2/genética , Receptor ErbB-3/genética , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Lapatinib/farmacologia , Camundongos Endogâmicos NOD , Neoplasias/patologia , Fosforilação/efeitos dos fármacos , Receptor ErbB-2/metabolismo , Receptor ErbB-3/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
19.
J Mol Model ; 27(4): 105, 2021 Mar 08.
Artigo em Inglês | MEDLINE | ID: mdl-33686576

RESUMO

Experimental and theoretical studies have provided structural information regarding the shift from inactive to active EGFR, throughout which both conformations are linked via binding to specific tyrosine kinase inhibitors. For HER2, an intermediate active-inactive receptor conformation is present in the PDB, which has been co-crystallized with tak-285. The affinity of HER2 in monomeric state to tak-285 has been previously reported. However, the lack of structural knowledge of HER2 limits our capacity to understand whether tak-285, or other known HER2 inhibitors, selectively bind active, inactive, or intermediate forms of HER2. To elucidate mechanisms by which tak-285 binds to HER2, we first obtained information regarding the structural features of the active state of HER2 via microsecond MD simulations from the crystallized intermediate structure previously determined. Based on these HER2 conformers, together with the inactive HER2 conformer obtained in a previous study, we used docking and MD simulations coupled to MMGBSA approach to assess binding of tak-285 and lapatinib, known HER2/EGFR dual inhibitors, to HER2. Structural and energetic studies revealed that tak-285 binds with a greater affinity than lapatinib to active and intermediate active-inactive forms of HER2. This is in accordance with experimental findings that showed the tak-285 inhibitor has increased activity relative to lapatinib in breast cancer cell lines.


Assuntos
Compostos Bicíclicos Heterocíclicos com Pontes/química , Hidroxibutiratos/química , Lapatinib/química , Modelos Moleculares , Receptor ErbB-2/química , Antineoplásicos/química , Humanos , Conformação Molecular , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular
20.
Exp Neurol ; 341: 113697, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33727095

RESUMO

Epidermal growth factor receptor (EGFR) signaling plays a substantial role in learning and memory. The upregulation of EGFR has been embroiled in the pathophysiology of Alzheimer's disease (AD). Nevertheless, most of EGFR-tyrosine kinase inhibitors (EGFR-TKIs) have been extensively studied for non-CNS diseases such as cancer and rheumatoid arthritis. TKIs targeting-based research in neurodegenerative disorders sounds to be lagging behind those of other diseases. Hence, this study aims to explore the molecular signaling pathways and the efficacy of treatment with lapatinib ditosylate (LAP), as one of EGFR-TKIs that has not yet been investigated in AD, on cognitive decline induced by ovariectomy (OVX) with chronic administration of D-galactose (D-gal) in female Wistar albino rats. OVX rats were injected with 150 mg/kg/day D-gal ip for 8 weeks to induce AD. Administration of 100 mg/kg/day LAP p.o. for 3 weeks starting after the 8th week of D-gal administration improved memory and debilitated histopathological alterations. LAP decreased the expression of GFAP, p-tau, and Aß 1-42. Besides, it reduced EGFR, HER-2, TNF-α, NOX-1, GluR-II, p38 MAPK, and p-mTOR. LAP increased nitrite, and neuronal pro-survival transduction proteins; p-PI3K, p-AKT, and p-GSK-3ß levels. Taken together, these findings suggest the role of LAP in ameliorating D-gal-induced AD in OVX rats via activating the pro-survival pathway; PI3K-Akt-GSK-3ß, while inhibiting p-mTOR, NOX-1, and p38 MAPK pathways. Moreover, this research offered a significant opportunity to advance awareness of the repositioning of TKI anti-cancer drugs for the treatment of AD.


Assuntos
Doença de Alzheimer/tratamento farmacológico , Reposicionamento de Medicamentos/métodos , Galactose/toxicidade , Lapatinib/uso terapêutico , Transtornos da Memória/tratamento farmacológico , Ovariectomia/efeitos adversos , Doença de Alzheimer/induzido quimicamente , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/metabolismo , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Relação Dose-Resposta a Droga , Feminino , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Hipocampo/patologia , Lapatinib/farmacologia , Aprendizagem em Labirinto/efeitos dos fármacos , Aprendizagem em Labirinto/fisiologia , Transtornos da Memória/metabolismo , Transtornos da Memória/patologia , Fragmentos de Peptídeos/metabolismo , Ratos , Ratos Wistar , Resultado do Tratamento
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