Effects of Lapatinib on HER2-Positive and HER2-Negative Canine Mammary Carcinoma Cells Cultured In Vitro.
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ABSTRACT:
SUBMITTER: Leis-Filho AF
PROVIDER: S-EPMC8235449 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:IntroductionHuman epidermal growth factor receptor 2 (HER2) amplification is frequent in ductal carcinoma in situ (DCIS) of the breast and is associated with poorly differentiated tumors and adverse prognosis features. This study aimed to determine the molecular effects of the HER2 inhibitor lapatinib in patients with HER2 positive DCIS.MethodsPatients with HER2 positive DCIS received 1,500 mg daily of lapatinib for four consecutive weeks prior to surgical resection. Magnetic resonance imaging (MRI) was used to determine changes in tumor volume. The molecular effects of lapatinib on HER2 signaling (PI3K/AKT and RAS/MAPK pathways), cell proliferation (Ki67 and p27) and apoptosis (TUNEL) were determined in pre and post-lapatinib treatment samples.ResultsA total of 20 patients were included. Lapatinib was well tolerated with only minor and transient side effects. The agent effectively modulated HER2 signaling decreasing significantly pHER2 and pERK1 expression, together with a decrease in tumor size evaluated by MRI. There was no evidence of changes in Ki67.ConclusionsFour weeks of neoadjuvant lapatinib in patients with HER2-positive DCIS resulted in inhibition of HER2 and RAS/MAPK signaling pathway.Trial registration2008-004492-21 (Registered June 25th 2008).
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Project description:HER2-positive breast cancers may lose HER2 expression in recurrences and metastases. In this work, we studied cell lines derived from two transgenic mammary tumors driven by human HER2 that showed different dynamics of HER2 status. MamBo89HER2stable cell line displayed high and stable HER2 expression, which was maintained upon in vivo passages, whereas MamBo43HER2labile cell line gave rise to HER2-negative tumors from which MamBo38HER2loss cell line was derived. Both low-density seeding and in vitro trastuzumab treatment of MamBo43HER2labile cells induced the loss of HER2 expression. MamBo38HER2loss cells showed a spindle-like morphology, high stemness and acquired in vivo malignancy. A comprehensive molecular profile confirmed the loss of addiction to HER2 signaling and acquisition of an EMT signature, together with increased angiogenesis and migration ability. We identified PDGFR-B among the newly expressed determinants of MamBo38HER2loss cell tumorigenic ability. Sunitinib inhibited MamBo38HER2loss tumor growth in vivo and reduced stemness and IL6 production in vitro. In conclusion, HER2-positive mammary tumors can evolve into tumors that display distinctive traits of claudin-low tumors. Our dynamic model of HER2 status can lead to the identification of new druggable targets, such as PDGFR-B, in order to counteract the resistance to HER2-targeted therapy that is caused by HER2 loss.
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Project description:Human epidermal growth factor receptor 2 (HER2) is a tyrosine kinase receptor that promotes tumor cell growth and is implicated in the pathogenesis of human breast cancer. The role of HER2 in canine mammary carcinomas (CMCs) is not clear. Therefore, this study aimed to examine the protein expression and cytogenetic changes of HER2 and their correlation with other clinical-pathological parameters in CMC. We retrospectively selected 112 CMCs. HER2, ER, and Ki67 were assessed by immunohistochemistry. HER2 antibody validation was investigated by immunoblot on mammary tumor cell lines. Fluorescence in situ hybridization (FISH) was performed with probes for HER2 and CRYBA1 (control gene present on CFA9). HER2 protein overexpression was detected in 15 carcinomas (13.5%). A total of 90 carcinomas were considered technically adequate by FISH, and 8 out of 90 CMC (10%) were HER2 amplified, 3 of which showed a cluster-type pattern. HER2 overexpression was correlated with an increased number of HER2 gene copies (p = 0.01; R = 0.24) and overall survival (p = 0.03), but no correlation with ER, Ki67, grade, metastases, and tumor-specific survival was found. Surprisingly, co-amplification or polysomy was identified in three tumors, characterized by an increased copy number of both HER2 and CRYBA1. A morphological translocation-fusion pattern was recognized in 20 carcinomas (22%), with a co-localized signal of HER2 and CRYBA1. HER2 is not associated with clinical-pathological parameters of increased malignancy in canine mammary tumors, but it is suitable for studying different amplification patterns.
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Project description:Feline mammary carcinoma (FMC) is a common neoplasia in cat, being HER2-positive the most prevalent subtype. In woman's breast cancer, tyrosine kinase inhibitors (TKi) are used as a therapeutic option, by blocking the phosphorylation of the HER2 tyrosine kinase domain. Moreover, clinical trials demonstrated that TKi produce synergistic antiproliferative effects in combination with mTOR inhibitors, overcoming resistance to therapy. Thus, to uncover new chemotherapeutic strategies for cats, the antiproliferative effects of two TKi (lapatinib and neratinib), and their combination with a mTOR inhibitor (rapamycin), were evaluated in FMC cell lines (CAT-M, FMCp and FMCm) and compared with a human breast cancer cell line (SkBR-3). Results revealed that both TKi induced antiproliferative effects in all feline cell lines, by blocking the phosphorylation of EGFR members and its downstream effectors. Furthermore, combined treatments with rapamycin presented synergetic antiproliferative effects. Additionally, the DNA sequence of the her2 TK domain (exons 18 to 20) was determined in 40 FMC tissue samples, and despite several mutations were found none of them were described as inducing resistance to therapy. Altogether, our results demonstrated that TKi and combined protocols may be useful in the treatment of cats with mammary carcinomas, and that TKi-resistant FMC are rare.
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Project description:Salinomycin (SAL), a polyether ionophore antibiotic, has been demonstrated to selectively kill cancer stem cells (CSCs) in various types of human tumor. The aim of the present study was to investigate the effects of SAL on canine mammary CSCs. CSCs in canine mammary carcinoma cell lines (CMT7364 and CIPp) were identified using a sphere formation assay and flow cytometry. The chemoresistance, invasive potential and expression of stem cell-associated proteins of these spheres was then analyzed. This demonstrated that the spheres exhibited characteristics of CSCs, including a cluster of differentiation (CD)44+/CD24-/low phenotype, upregulation of Wnt/β-catenin signaling pathway-associated proteins and chemoresistance. The viability of the spheres was decreased in a concentration- and time-dependent manner following treatment with SAL, and the spheres did not exhibit increased resistance to SAL compared with their parental cells. In addition, exposure to SAL inhibited sphere-formation and invasive potential in canine mammary CSCs in a dose-dependent manner. Furthermore, SAL decreased the CD44+/CD24-/low population and downregulated the expression of Wnt/β-catenin signaling-associated proteins (β-catenin, Cyclin D1 and octamer-binding transcription factor 4) in the spheres. In conclusion, the present study demonstrated that SAL is an effective inhibitor of canine mammary CSCs in vitro, indicating that SAL is a promising chemotherapeutic agent for the treatment of canine mammary carcinoma.
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Project description:Molecular mechanisms of lapatinib resistance in breast cancer are not well understood. The aim of this study was to correlate expression of selected proteins involved in ErbB family signaling pathways with clinical efficacy of lapatinib. Study group included 270 HER2-positive advanced breast cancer patients treated with lapatinib and capecitabine. Immunohistochemical expression of phosphorylated adenosine monophosphate-activated protein (p-AMPK), mitogen-activated protein kinase (p-MAPK), phospho (p)-p70S6K, cyclin E, phosphatase and tensin homolog were analyzed in primary breast cancer samples. The best discriminative value for progression-free survival (PFS) was established for each biomarker and subjected to multivariate analysis. At least one biomarker was determined in 199 patients. Expression of p-p70S6K was independently associated with longer (HR 0.45; 95% CI: 0.25-0.81; p = 0.009), and cyclin E with shorter PFS (HR 1.83; 95% CI: 1.06-3.14; p = 0.029). Expression of p-MAPK (HR 1.61; 95% CI 1.13-2.29; p = 0.009) and cyclin E (HR 2.99; 95% CI: 1.29-6.94; p = 0.011) was correlated with shorter, and expression of estrogen receptor (HR 0.65; 95% CI 0.43-0.98; p = 0.041) with longer overall survival. Expression of p-AMPK negatively impacted response to treatment (HR 3.31; 95% CI 1.48-7.44; p = 0.004) and disease control (HR 3.07; 95% CI 1.25-7.58; p = 0.015).the efficacy of lapatinib seems to be associated with the activity of downstream signaling pathways - AMPK/mTOR and Ras/Raf/MAPK. Further research is warranted to assess the clinical utility of these data and to determine a potential role of combining lapatinib with MAPK pathway inhibitors.
| S-EPMC4808017 | biostudies-literature
Project description:The incidence and prevalence of paraneoplastic glomerulopathy, especially associated with carcinoma, are a matter of debate and the causal link between cancer and glomerular diseases remains unclear. The aim of this study was to evaluate renal biopsies of selected bitches with spontaneous mammary gland carcinoma. We hypothesized that dogs with mammary carcinomas would show histologic evidence of glomerular pathology. A prospective study was performed in dogs with naturally occurring mammary carcinoma that were undergoing tumor resection and ovariohysterectomy. We evaluated renal biopsies of 32 bitches with spontaneous mammary gland carcinoma and 11 control dogs without mammary gland neoplasia. Samples were obtained from the left kidney and the biopsy material was divided for light microscopy (LM), immunofluorescence (IF) and transmission electron microscopy (TEM). Light microscopy abnormalities were identified in 78.1% of dogs with mammary carcinoma (n = 25) and in none of the dogs in the control group. Focal glomerular mesangial matrix expansion was the most common alteration (n = 15, 60.0%), but mesangial cell proliferation (n = 9, 36.0%) and focal segmental glomerulosclerosis (n = 9, 36.0%), synechiae (n = 7, 28.0%), and globally sclerotic glomeruli (n = 6, 24.0%) were also frequent in dogs with malignancy. Immunofluorescence microscopy revealed strong IgM staining was demonstrated in 64.3% (n = 18) of carcinoma dogs. Transmission electron microscopy from dogs with carcinoma revealed slight changes, the most frequent of which was faint sub-endothelial and mesangial deposits of electron-dense material (78%). Mesangial cell interpositioning and segmental effacement of podocyte foot processes were identified in some specimens (45%). Changes in the glomerulus and proteinuria are common in dogs with naturally occurring mammary carcinoma and this condition appears to provide an excellent large animal model for cancer-associated glomerulopathy in humans.
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Project description:Dogs can be excellent models for spontaneous studies about breast cancers, presenting similarities in clinical behavior and molecular pathways of the disease. Thus, analyses of the canine transcriptome can identify deregulated genes and pathways, contributing to the identification of biomarkers and new therapeutic targets, benefiting humans and animals. In this context, this study aimed to determine the transcriptional profile of canine mammary ductal carcinoma and contribute to the clarification of the importance of deregulated molecules in the molecular pathways involved in the disease. Therefore, we used mammary ductal carcinoma tissue samples and non-tumor mammary tissue from the radical mastectomy of six female dogs. Sequencing was performed on the NextSeq-500 System platform. A comparison of carcinoma tissue and normal tissue revealed 633 downregulated and 573 upregulated genes, which were able to differentiate the groups by principal component analysis. Gene ontology analysis indicated that inflammatory, cell differentiation and adhesion, and extracellular matrix maintenance pathways were mainly deregulated in this series. The main differentially expressed genes observed in this research can indicate greater disease aggressiveness and worse prognosis. Finally, the study of the canine transcriptome indicates that it is an excellent model to generate information relevant to oncology in both species.
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Project description:Lapatinib is an FDA-approved EGFR and HER2 tyrosine kinase inhibitor for the treatment of HER2-positive breast cancer patients. However, its therapeutic efficacy is limited by primary or acquired resistance. In the present study, we established breast cancers cells with acquired lapatinib resistance and investigated the antitumor activity of the second-generation HSP90 inhibitor ganetespib in association with lapatinib in lapatinib-sensitive and -resistant cells. The combination treatment showed synergistic inhibition of HER and the downstream PI3K/Akt and Ras/MEK/ERK pathways, in addition to enhancing induction of early apoptotic cell death and G1 arrest in both parent and lapatinib-resistant cells in vitro. The joint administration of ganetespib and lapatinib depleted the aberrant nuclear transcription factor STAT3, a mediator of the cell cycle and apoptosis-related pathways that is probably involved in the lapatinib resistance of HER2-positive breast cancer cells. In conjunctive with the augmented inhibition of tumor growth observed in both SKBR3 and SKBR3-L xenografts compared to monotherapy, our data provide a sound preclinical basis for combination treatment with lapatinib and ganetespib for refractory HER2-positive breast cancer.
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Project description:Tumor microenvironment has gained great relevance due to its ability to regulate distinct checkpoints mediators, orchestrating tumor progression. Serum programmed cell death protein-1 (PD-1) and programmed death ligand-1 (PD-L1) levels were compared with healthy controls and with serum cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) and tumor necrosis factor-alpha (TNF-?) levels in order to understand the role of PD-1/PD-L1 axis in cats with mammary carcinoma. PD-1 and PD-L1 expression was evaluated in tumor-infiltrating lymphocytes (TILs) and cancer cells, as the presence of somatic mutations. Results showed that serum PD-1 and PD-L1 levels were significantly higher in cats with HER2-positive (p = 0.017; p = 0.032) and triple negative (TN) normal-like mammary carcinomas (p = 0.004; p = 0.015), showing a strong positive correlation between serum CTLA-4 and TNF-? levels. In tumors, PD-L1 expression in cancer cells was significantly higher in HER2-positive samples than in TN normal-like tumors (p = 0.010), as the percentage of PD-L1-positive TILs (p = 0.037). PD-L1 gene sequencing identified two heterozygous mutations in exon 4 (A245T; V252M) and one in exon 5 (T267S). In summary, results support the use of spontaneous feline mammary carcinoma as a model for human breast cancer and suggest that the development of monoclonal antibodies may be a therapeutic strategy.
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