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In vivo diagnostic imaging of bacterial infections is currently reliant on targeting their metabolic pathways, an ineffective method to identify microbial species with low metabolic activity. Here, we establish HS-198 as a small-molecule fluorescent conjugate that selectively targets the highly conserved bacterial protein HtpG (high-temperature protein G), within Borrelia burgdorferi, the bacterium responsible for Lyme disease. We describe the use of HS-198 to target morphologic forms of B. burgdorferi in both the logarithmic growth phase and the metabolically dormant stationary phase as well as in inactivated spirochetes. Furthermore, in a murine infection model, systemically injected HS-198 identified B. burgdorferi as revealed by imaging in postnecropsy tissue sections. These findings demonstrate how small-molecule probes directed at conserved bacterial protein targets can function to identify the microbe using noninvasive imaging and potentially as scaffolds to deliver antimicrobial agents to the pathogen.
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Borrelia burgdorferi , Enfermedad de Lyme , Animales , Proteínas Bacterianas/genética , Diagnóstico por Imagen , Humanos , Enfermedad de Lyme/diagnóstico , RatonesRESUMEN
Hematopoietic regeneration following chemotherapy may be distinct from regeneration following radiation. While we have shown that epidermal growth factor (EGF) accelerates regeneration following radiation, its role following chemotherapy is currently unknown. We sought to identify EGF as a hematopoietic growth factor for chemotherapy-induced myelosuppression. Following 5-fluorouracil (5-FU), EGF accelerated hematopoietic stem cell regeneration and prolonged survival compared with saline-treated mice. To mitigate chemotherapy-induced injury to endothelial cells in vivo, we deleted Bax in VEcadherin+ cells (VEcadherinCre;BaxFL/FL mice). Following 5-FU, VEcadherinCre;BaxFL/FL mice displayed preserved hematopoietic stem/progenitor content compared with littermate controls. 5-FU and EGF treatment resulted in increased cellular proliferation, decreased apoptosis, and increased DNA double-strand break repair by non-homologous end-joining recombination compared with saline-treated control mice. When granulocyte colony stimulating factor (G-CSF) is given with EGF, this combination was synergistic for regeneration compared with either G-CSF or EGF alone. EGF increased G-CSF receptor (G-CSFR) expression following 5-FU. Conversely, G-CSF treatment increased both EGF receptor (EGFR) and phosphorylation of EGFR in hematopoietic stem/progenitor cells. In humans, the expression of EGFR is increased in patients with colorectal cancer treated with 5-FU compared with cancer patients not on 5-FU. Similarly, EGFR signaling is responsive to G-CSF in humans in vivo with both increased EGFR and phospho-EGFR in healthy human donors following G-CSF treatment compared with donors who did not receive G-CSF. These data identify EGF as a hematopoietic growth factor following myelosuppressive chemotherapy and that dual therapy with EGF and G-CSF may be an effective method to accelerate hematopoietic regeneration. Stem Cells 2018;36:252-264.
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Factor de Crecimiento Epidérmico/farmacología , Factor Estimulante de Colonias de Granulocitos/farmacología , Animales , Apoptosis/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Receptores ErbB/metabolismo , Fluorouracilo/farmacología , Hematopoyesis/efectos de los fármacos , Células Madre Hematopoyéticas/citología , Células Madre Hematopoyéticas/efectos de los fármacos , Ratones , Ratones Endogámicos C57BL , Transducción de Señal/efectos de los fármacosRESUMEN
CHAMBER was a regional educational initiative for providers of care to patients with HER2+ breast cancer. The study goals were to (1) enhance testing for HER2/neu overexpression in patients with invasive breast cancer; (2) increase the appropriate use of targeted therapy for patients with HER2+ breast cancer; and (3) enhance patients' coping ability. This Performance Improvement Continuing Medical Education (PI-CME) initiative included clinical practice assessment, educational activities, and reassessment. Chart review revealed a high rate of HER2 testing (98%) before and after education. Targeted therapy for patients with HER2+ breast cancer declined after the program (from 96% to 61%), perhaps attributable to an increase in awareness of medical reasons to avoid use of targeted therapy. Assessment for patients' emotional coping ability increased after education (from 55% to 76%; P=.01). Rates of testing for HER2 amplification and assessment of emotional well-being after education were consistent with ASCO Quality Oncology Practice Initiative benchmark values. Documentation of actions to address emotional problems remained an area for improvement.
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Neoplasias de la Mama/diagnóstico , Neoplasias de la Mama/terapia , Educación Médica Continua , Personal de Salud , Mejoramiento de la Calidad , Adaptación Psicológica , Neoplasias de la Mama/metabolismo , Femenino , Adhesión a Directriz , Personal de Salud/educación , Personal de Salud/normas , Humanos , Cumplimiento de la Medicación , Receptor ErbB-2/metabolismoRESUMEN
Cell migration driven by the epidermal growth factor receptor (EGFR) propels morphogenesis and involves reorganization of the actin cytoskeleton. Although de novo transcription precedes migration, transcript identity remains largely unknown. Through their actin-binding domains, tensins link the cytoskeleton to integrin-based adhesion sites. Here we report that EGF downregulates tensin-3 expression, and concomitantly upregulates cten, a tensin family member that lacks the actin-binding domain. Knockdown of cten or tensin-3, respectively, impairs or enhances mammary cell migration. Furthermore, cten displaces tensin-3 from the cytoplasmic tail of integrin beta1, thereby instigating actin fibre disassembly. In invasive breast cancer, cten expression correlates not only with high EGFR and HER2, but also with metastasis to lymph nodes. Moreover, treatment of inflammatory breast cancer patients with an EGFR/HER2 dual-specificity kinase inhibitor significantly downregulated cten expression. In conclusion, a transcriptional tensin-3-cten switch may contribute to the metastasis of mammary cancer.
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Neoplasias de la Mama/metabolismo , Movimiento Celular/fisiología , Factor de Crecimiento Epidérmico/metabolismo , Proteínas de Microfilamentos/metabolismo , Neoplasias de la Mama/patología , Línea Celular Tumoral , Inhibidores Enzimáticos/metabolismo , Receptores ErbB , Femenino , Humanos , Proteínas de Microfilamentos/genética , Análisis de Secuencia por Matrices de Oligonucleótidos , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , TensinasRESUMEN
INTRODUCTION: The human epidermal growth factor receptor 2 (HER2) receptor tyrosine kinase (RTK) oncogene is an attractive therapeutic target for the treatment of HER2-addicted tumors. Although lapatinib, an FDA-approved small-molecule HER2 and epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI), represents a significant therapeutic advancement in the treatment of HER2+ breast cancers, responses to lapatinib have not been durable. Consequently, elucidation of mechanisms of acquired therapeutic resistance to HER-directed therapies is of critical importance. METHODS: Using a functional protein-pathway activation mapping strategy, along with targeted genomic knockdowns applied to a series of isogenic-matched pairs of lapatinib-sensitive and resistant cell lines, we now report an unexpected mechanism of acquired resistance to lapatinib and similar TKIs. RESULTS: The signaling analysis revealed that whereas HER2 was appropriately inhibited in lapatinib-resistant cells, EGFR tyrosine phosphorylation was incompletely inhibited. Using a targeted molecular knockdown approach to interrogate the causal molecular underpinnings of EGFR-persistent activation, we found that lapatinib-resistant cells were no longer oncogene addicted to HER2-HER3-PI3K signaling, as seen in the parental lapatinib-sensitive cell lines, but instead were dependent on a heregulin (HRG)-driven HER3-EGFR-PI3K-PDK1 signaling axis. Two FDA-approved EGFR TKIs could not overcome HRG-HER3-mediated activation of EGFR, or reverse lapatinib resistance. The ability to overcome EGFR-mediated acquired therapeutic resistance to lapatinib was demonstrated through molecular knockdown of EGFR and treatment with the irreversible pan-HER TKI neratinib, which blocked HRG-dependent phosphorylation of HER3 and EGFR, resulting in apoptosis of resistant cells. In addition, whereas HRG reversed lapatinib-mediated antitumor effects in parental HER2+ breast cancer cells, neratinib was comparatively resistant to the effects of HRG in parental cells. Finally, we showed that HRG expression is an independent negative predictor of clinical outcome in HER2+ breast cancers, providing potential clinical relevance to our findings. CONCLUSIONS: Molecular analysis of acquired therapeutic resistance to lapatinib identified a new resistance mechanism based on incomplete and "leaky" inhibition of EGFR by lapatinib. The selective pressure applied by incomplete inhibition of the EGFR drug target resulted in selection of ligand-driven feedback that sustained EGFR activation in the face of constant exposure to the drug. Inadequate target inhibition driven by a ligand-mediated autocrine feedback loop may represent a broader mechanism of therapeutic resistance to HER TKIs and suggests adopting a different strategy for selecting more effective TKIs to advance into the clinic.
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Comunicación Autocrina , Neoplasias de la Mama/metabolismo , Receptores ErbB/metabolismo , Neurregulina-1/metabolismo , Receptor ErbB-2/metabolismo , Receptor ErbB-3/metabolismo , Transducción de Señal , Neoplasias de la Mama/genética , Neoplasias de la Mama/mortalidad , Proteína Tirosina Quinasa CSK , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Resistencia a Antineoplásicos , Receptores ErbB/genética , Femenino , Expresión Génica , Técnicas de Silenciamiento del Gen , Humanos , Lapatinib , Neurregulina-1/genética , Fosfatidilinositol 3-Quinasas , Fosfoproteínas Fosfatasas/metabolismo , Fosforilación , Pronóstico , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas c-met/antagonistas & inhibidores , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora , Quinazolinas/farmacología , Receptor ErbB-3/genética , Transducción de Señal/efectos de los fármacos , Serina-Treonina Quinasas TOR/metabolismo , Familia-src Quinasas/antagonistas & inhibidoresRESUMEN
INTRODUCTION: Although human epidermal growth factor receptor 2 (HER2) positive or estrogen receptor (ER) positive breast cancers are treated with clinically validated anti-HER2 or anti-estrogen therapies, intrinsic and acquired resistance to these therapies appears in a substantial proportion of breast cancer patients and new therapies are needed. Identification of additional molecular factors, especially those characterized by aggressive behavior and poor prognosis, could prioritize interventional opportunities to improve the diagnosis and treatment of breast cancer. METHODS: We compiled a collection of 4,010 breast tumor gene expression data derived from 23 datasets that have been posted on the National Center for Biotechnology Information (NCBI) Gene Expression Omnibus (GEO) database. We performed a genome-scale survival analysis using Cox-regression survival analyses, and validated using Kaplan-Meier Estimates survival and Cox Proportional-Hazards Regression survival analyses. We conducted a genome-scale analysis of chromosome alteration using 481 breast cancer samples obtained from The Cancer Genome Atlas (TCGA), from which combined expression and copy number data were available. We assessed the correlation between somatic copy number alterations and gene expression using analysis of variance (ANOVA). RESULTS: Increased expression of each of the heat shock protein (HSP) 90 isoforms, as well as HSP transcriptional factor 1 (HSF1), was correlated with poor prognosis in different subtypes of breast cancer. High-level expression of HSP90AA1 and HSP90AB1, two cytoplasmic HSP90 isoforms, was driven by chromosome coding region amplifications and were independent factors that led to death from breast cancer among patients with triple-negative (TNBC) and HER2-/ER+ subtypes, respectively. Furthermore, amplification of HSF1 was correlated with higher HSP90AA1 and HSP90AB1 mRNA expression among the breast cancer cells without amplifications of these two genes. A collection of HSP90AA1, HSP90AB1 and HSF1 amplifications defined a subpopulation of breast cancer with up-regulated HSP90 gene expression, and up-regulated HSP90 expression independently elevated the risk of recurrence of TNBC and poor prognosis of HER2-/ER+ breast cancer. CONCLUSIONS: Up-regulated HSP90 mRNA expression represents a confluence of genomic vulnerability that renders HER2 negative breast cancers more aggressive, resulting in poor prognosis. Targeting breast cancer with up-regulated HSP90 may potentially improve the effectiveness of clinical intervention in this disease.
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Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Regulación Neoplásica de la Expresión Génica , Proteínas HSP90 de Choque Térmico/genética , Receptor ErbB-2/metabolismo , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/mortalidad , Proteínas de Unión al ADN/genética , Femenino , Amplificación de Genes , Factores de Transcripción del Choque Térmico , Humanos , Estimación de Kaplan-Meier , Fenotipo , Pronóstico , Modelos de Riesgos Proporcionales , Análisis de Supervivencia , Factores de Transcripción/genética , Regulación hacia ArribaRESUMEN
The ERBB2 proto-oncogene is associated with an aggressive phenotype in breast cancer. Its role in hematologic malignancies is incompletely defined, in part because ERBB2 is not readily detected on the surface of cancer cells. We demonstrate that truncated ERBB2, which lacks the extracellular domain, is overexpressed on primary CD34+ myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) cells compared with healthy hematopoietic cells. This overexpression of ERBB2 is associated with aberrant, oncogenic signaling with autophosphorylation of multiple tyrosine sites. Like in breast cancers, ERBB2 can exist as truncated isoforms p95ERBB2 and p110ERBB2 in MDS and AML. Neutralization of ERBB2 signaling with ERBB2 tyrosine kinase inhibitors (i.e., lapatinib, afatinib, and neratinib) increases apoptotic cell death and reduces human engraftment of MDS cells in mice at 21 weeks posttransplantation. Inhibition of ERBB2 modulates the expression of multiple pro- and anti-apoptotic mitochondrial proteins, including B-cell lymphoma 2 (BCL2). Dual blockade with ERBB2 and BCL2 inhibitors triggers additional reductions of BCL2 phosphorylation and myeloid cell leukemia-1 (MCL1) expression compared with single drug treatment. Dual therapy was synergistic at all tested doses, with a dose reduction index of up to 29 for lapatinib + venetoclax compared with venetoclax alone. Notably, these agents operated together and shifted cancer cells to a pro-apoptotic phenotype, resulting in increased mitochondrial cytochrome c release and activated caspase-3-mediated cell death. IMPLICATIONS: These findings warrant study of ERBB2 and BCL2 combination therapy in patients with MDS and AML. VISUAL OVERVIEW: http://mcr.aacrjournals.org/content/molcanres/19/5/886/F1.large.jpg.
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Leucemia Mieloide Aguda/genética , Síndromes Mielodisplásicos/genética , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Receptor ErbB-2/antagonistas & inhibidores , Animales , Apoptosis , Humanos , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Receptor ErbB-2/metabolismoRESUMEN
PURPOSE: Next-generation sequencing (NGS) multigene panel testing has become widespread, including the Veterans Affairs (VA), through the VA National Precision Oncology Program (NPOP). The interpretation of genomic alterations remains a bottleneck for realizing precision medicine. We sought to examine the concordance for pathogenicity determination and clinical actionability of annotation services in NPOP. METHODS: Unique gene variants were generated from NGS gene panel results using two sequencing services. For each unique gene variant, annotations were provided through N-of-One (NoO), IBM Watson for Genomics (WfG), and OncoKB. Annotations for pathogenicity (all three sources) and actionability (WfG and OncoKB) were examined for concordance. Cohen's kappa statistic was calculated to measure agreement between annotation services. RESULTS: Among 1,227 NGS results obtained between 2015 and 2017, 1,388 unique variants were identified in 117 genes. The genes with the largest number of variants included TP53 (270), STK11 (92), and CDKN2A (81). The most common cancer type was lung adenocarcinoma (440), followed by colon adenocarcinoma (113). For pathogenic and likely pathogenic variants, there was 30% agreement between WfG and NoO (kappa, -0.26), 76% agreement between WfG and OncoKB (kappa, 0.22), and 42% agreement between NoO and OncoKB (kappa, -0.07). For level 1 drug actionability of gene variant-diagnosis combinations, there was moderate agreement between WfG and OncoKB (96.9%; kappa, 0.44), with 27 combinations identified as level 1 by both services, 58 by WfG alone, and 6 variants by OncoKB alone. CONCLUSION: There is substantial variability in pathogenicity assessment of NGS variants in solid tumors by annotation services. In addition, there was only moderate agreement in level 1 therapeutic actionability recommendations between WfG and OncoKB. Improvement in the precision of NGS multigene panel annotation is needed.
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BACKGROUND: To support the rising need for testing and to standardize tumor DNA sequencing practices within the U.S. Department of Veterans Affairs (VA)'s Veterans Health Administration (VHA), the National Precision Oncology Program (NPOP) was launched in 2016. We sought to assess oncologists' practices, concerns, and perceptions regarding Next-Generation Sequencing (NGS) and the NPOP. MATERIALS AND METHODS: Using a purposive total sampling approach, oncologists who had previously ordered NGS for at least one tumor sample through the NPOP were invited to participate in semi-structured interviews. Questions assessed the following: expectations for the NPOP, procedural requirements, applicability of testing results, and the summative utility of the NPOP. Interviews were assessed using an open coding approach. Thematic analysis was conducted to evaluate the completed codebook. Themes were defined deductively by reviewing the direct responses to interview questions as well as inductively by identifying emerging patterns of data. RESULTS: Of the 105 medical oncologists who were invited to participate, 20 (19%) were interviewed from 19 different VA medical centers in 14 states. Five recurrent themes were observed: (1) Educational Efforts Regarding Tumor DNA Sequencing Should be Undertaken, (2) Pathology Departments Share a Critical Role in Facilitating Test Completion, (3) Tumor DNA Sequencing via NGS Serves as the Most Comprehensive Testing Modality within Precision Oncology, (4) The Availability of the NPOP Has Expanded Options for Select Patients, and (5) The Completion of Tumor DNA Sequencing through the NPOP Could Help Improve Research Efforts within VHA Oncology Practices. CONCLUSION: Medical oncologists believe that the availability of tumor DNA sequencing through the NPOP could potentially lead to an improvement in outcomes for veterans with metastatic solid tumors. Efforts should be directed toward improving oncologists' understanding of sequencing, strengthening collaborative relationships between oncologists and pathologists, and assessing the role of comprehensive NGS panels within the battery of precision tests.
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Conocimientos, Actitudes y Práctica en Salud , Secuenciación de Nucleótidos de Alto Rendimiento/normas , Neoplasias/genética , Oncólogos/psicología , Análisis de Secuencia de ADN/normas , United States Department of Veterans Affairs , Adulto , Detección Precoz del Cáncer/normas , Femenino , Pruebas Genéticas/normas , Humanos , Masculino , Persona de Mediana Edad , Neoplasias/diagnóstico , Medicina de Precisión/normas , Planes Estatales de Salud , Encuestas y Cuestionarios , Estados UnidosRESUMEN
Breast cancers overexpressing the ErbB2 (HER2) receptor tyrosine kinase oncogene are treated with targeted therapies such as trastuzumab (Herceptin), an anti-ErbB2 antibody, and lapatinib (GW572016/Tykerb), a selective small molecule inhibitor of ErbB2 and epidermal growth factor receptor tyrosine kinases that was recently approved for ErbB2+ breast cancers that progressed on trastuzumab-based therapy. The efficacy of lapatinib as a monotherapy or in combination with chemotherapy, however, is limited by the development of therapeutic resistance that typically occurs within 12 months of starting therapy. In contrast to small molecule inhibitors targeting other receptor tyrosine kinases where resistance has been attributed to mutations within the targeted receptor, ErbB2 mutations have not been commonly found in breast tumors. Instead, acquired resistance to lapatinib seems to be mediated by redundant survival pathways that are activated as a consequence of marked inhibition of ErbB2 kinase activity. For example, inhibition of phosphatidylinositol3 kinase-Akt in lapatinib-treated cells leads to derepression of FOXO3A, a transcription factor that up-regulates estrogen receptor (ER) signaling, resulting in a switch in the regulation of survival factors (e.g., survivin) and cell survival from ErbB2 alone to ER and ErbB2 in resistant cells. In this review, we discuss the effects of lapatinib on signaling networks in ErbB2+ breast cancer cells to elucidate potential mechanisms of therapeutic resistance and strategies to overcome or prevent its development.
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Neoplasias de la Mama/tratamiento farmacológico , Resistencia a Antineoplásicos , Inhibidores de Proteínas Quinasas/uso terapéutico , Quinazolinas/uso terapéutico , Receptor ErbB-2/metabolismo , Antineoplásicos/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Neoplasias de la Mama/metabolismo , Humanos , Lapatinib , Modelos Biológicos , Receptor ErbB-2/antagonistas & inhibidores , Transducción de Señal/efectos de los fármacosRESUMEN
Trastuzumab antitumor activity in ErbB2-overexpressing breast cancers seems to be dependent upon the presence of phosphatase and tensin homologue deleted on chromosome 10 (PTEN), a phosphatase that dampens phosphatidylinositol 3-kinase-Akt signaling. Consequently, PTEN deficiency, which occurs in 50% of breast cancers, predicts for resistance to trastuzumab monotherapy. Here, we show that lapatinib, a small-molecule inhibitor of ErbB1 and ErbB2 tyrosine kinases, exerts its antitumor activity in a PTEN-independent manner. Steady-state phosphorylated ErbB2 (p-ErbB2) and p-Akt (S473) protein levels were inhibited within 30 min following lapatinib but not in response to trastuzumab in BT474 and Au565 cells (two ErbB2-overexpressing breast cancer cell lines that are sensitive to the proapoptotic effects of lapatinib). Whereas trastuzumab reportedly inhibits SRC phosphorylation (Y416), which in turn reduced SRC-ErbB2 protein interactions, lapatinib had no effect on either variable. To assess the potential functional role that PTEN might play in lapatinib antitumor activity, we selectively knocked down PTEN in BT474 and Au565 cells using small interfering RNA transfection. Loss of PTEN did not affect induction of tumor cell apoptosis by lapatinib in either cell line. In addition, lapatinib inhibited Akt phosphorylation in MDA-MB-468 cells, an ErbB1-expressing/ErbB2 non-overexpressing breast cancer line, despite their PTEN-null status. Moreover, patients with ErbB2-overexpressing inflammatory breast cancers responded to lapatinib monotherapy regardless of PTEN status. Thus, lapatinib seems to exert its antitumor activity in ErbB2-overexpressing breast cancers in a PTEN-independent manner. These data emphasize the importance of assessing PTEN status in tumors when selecting ErbB2-targeted therapies in patients with breast cancer.
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Neoplasias de la Mama/tratamiento farmacológico , Fosfohidrolasa PTEN/deficiencia , Quinazolinas/farmacología , Receptor ErbB-2/biosíntesis , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/fisiología , Neoplasias de la Mama/enzimología , Neoplasias de la Mama/genética , Línea Celular Tumoral , Humanos , Lapatinib , Fosfohidrolasa PTEN/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Fosforilación , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptor ErbB-2/antagonistas & inhibidores , Receptor ErbB-2/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
PURPOSE: The Veterans Health Administration (VHA) is the largest cancer care provider in the United States, with the added challenge of serving more than twice the percentage of patients with cancer in rural areas than the national average. The VHA established the National Precision Oncology Program in 2016 to implement and standardize the practice of precision oncology across the diverse VHA system. METHODS: Tumor or peripheral blood specimens from veterans with advanced solid tumors who were eligible for treatment were submitted for next-generation sequencing (NGS) at two commercial laboratories. Annotated results were generated by the laboratories and independently using IBM Watson for Genomics. Levels-of-evidence treatment recommendations were based on OncoKB criteria. RESULTS: From July 2016 to June 2018, 3,698 samples from 72 VHA facilities were submitted for NGS testing, of which 3,182 samples (86%) were successfully sequenced. Most samples came from men with lung, prostate, and colorectal cancers. Thirty-four percent of samples were from patients who lived in a rural area. TP53, ATM, and KRAS were among the most commonly mutated genes. Approximately 70% of samples had at least one actionable mutation, with clinical trials identified as the recommended option in more than 50%. Mutations in genes associated with a neuroendocrine prostate cancer phenotype were expressed at increased frequency among veterans than in the general population. The most frequent therapies prescribed in response to NGS testing were immune checkpoint inhibitors, EGFR kinase inhibitors, and PARP inhibitors. CONCLUSION: Clinical implementation of precision oncology is feasible across the VHA health care system, including rural sites. Veterans have unique occupational exposures that might inform the nature of the mutational signatures identified here. Importantly, these results underscore the importance of increasing clinical trial availability to veterans.
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In breast cancer, overexpression of ErbB2 or aberrant regulation of survivin, a member of the inhibitor of apoptosis family, is associated with resistance to chemo/hormone therapy and predicts for a poor clinical outcome. A functional link between the two predictive factors has not been previously shown. Here, using genetic and pharmacologic approaches to block ErbB2 signaling, we show that ErbB2 regulates survivin protein expression in ErbB2-overexpressing breast cancer cells. Selective knockdown of ErbB2 using small interfering RNA markedly reduced survivin protein, resulting in apoptosis of ErbB2-overexpressing breast cancer cell lines such as BT474. Alternatively, inhibition of ErbB2 signaling using lapatinib (GW572016), a reversible small-molecule inhibitor of ErbB1/ErbB2 tyrosine kinases, at pharmacologically relevant concentrations, leads to marked inhibition of survivin protein with subsequent apoptosis. The effect of lapatinib on survivin seems to be predominantly posttranslational, mediated by ubiquitin-proteosome degradation as lactacystin, a proteosome inhibitor, reverses these effects. Furthermore, lapatinib down-regulated the expression of His-tagged survivin, which was under the transcriptional control of a heterologous promoter, providing additional evidence supporting a posttranslational mechanism of regulation. In contrast, trastuzumab and gefitinib failed to down-regulate survivin in ErbB2-overexpressing breast cancer cells. Importantly, the clinical relevance of these findings was illustrated in patients with ErbB2-overexpressing breast cancer whose clinical response to lapatinib was associated with marked inhibition of survivin in their tumors. These findings shed new light on the mechanism by which ErbB2 overexpression protects against apoptotic stimuli in breast cancer and identifies therapeutic interventions to improve clinical outcomes in these aggressive tumors.
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Neoplasias de la Mama/metabolismo , Proteínas Asociadas a Microtúbulos/biosíntesis , Proteínas de Neoplasias/biosíntesis , Receptor ErbB-2/fisiología , Apoptosis/efectos de los fármacos , Apoptosis/fisiología , Neoplasias de la Mama/genética , Línea Celular Tumoral , Regulación hacia Abajo , Receptores ErbB/antagonistas & inhibidores , Receptores ErbB/metabolismo , Humanos , Proteínas Inhibidoras de la Apoptosis , Lapatinib , Proteínas Asociadas a Microtúbulos/antagonistas & inhibidores , Proteínas Asociadas a Microtúbulos/genética , Proteínas de Neoplasias/antagonistas & inhibidores , Proteínas de Neoplasias/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Quinazolinas/farmacología , Interferencia de ARN , Receptor ErbB-2/antagonistas & inhibidores , Receptor ErbB-2/biosíntesis , Receptor ErbB-2/genética , Receptor ErbB-3/fisiología , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , SurvivinRESUMEN
Extracellular expression of heat shock protein 90 (eHsp90) by tumor cells is correlated with malignancy. Development of small molecule probes that can detect eHsp90 in vivo may therefore have utility in the early detection of malignancy. We synthesized a cell impermeable far-red fluorophore-tagged Hsp90 inhibitor to target eHsp90 in vivo. High resolution confocal and lattice light sheet microscopy show that probe-bound eHsp90 accumulates in punctate structures on the plasma membrane of breast tumor cells and is actively internalized. The extent of internalization correlates with tumor cell aggressiveness, and this process can be induced in benign cells by overexpressing p110HER2. Whole body cryoslicing, imaging, and histology of flank and spontaneous tumor-bearing mice strongly suggests that eHsp90 expression and internalization is a phenomenon unique to tumor cells in vivo and may provide an "Achilles heel" for the early diagnosis of metastatic disease and targeted drug delivery.
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Neoplasias de la Mama/patología , Colorantes Fluorescentes/metabolismo , Proteínas HSP90 de Choque Térmico/metabolismo , Animales , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Endocitosis , Espacio Extracelular/metabolismo , Genes erbB-2 , Xenoinjertos , Humanos , RatonesRESUMEN
Antibodies and small molecule tyrosine kinase inhibitors targeting ErbB2 exhibit distinct, noncross resistant mechanisms of action. Here, apoptosis of ErbB2-overexpressing breast cancer cells was enhanced by combining lapatinib, an inhibitor of ErbB1 and ErbB2 tyrosine kinases, with anti-ErbB2 antibodies, including (i) trastuzumab, a humanized monoclonal antibody, and (ii) pAb, rabbit polyclonal antisera generated by vaccination with a human ErbB2 fusion protein. Treating ErbB2-overexpressing breast cancer cell lines with a relatively low concentration of lapatinib alone resulted in a minimal increase in tumor cell apoptosis with an associated decrease in steady-state protein levels of p-ErbB2, p-Akt, p-Erk1/2, and notably survivin, compared to baseline. Exposure to pAb alone reduced total ErbB2 protein, disrupting ErbB3 transactivation, leading to a marked inhibition of p-Akt; however, survivin protein levels remained unchanged and apoptosis only increased slightly. Treatment with trastuzumab alone had relatively little effect on survivin and apoptosis was unaffected. Combining lapatinib with either pAb or trastuzumab markedly downregulated survivin protein and enhanced tumor cell apoptosis. The association between the inhibition of survivin and enhanced apoptosis following the combination of ErbB2-targeted therapies provides a biological effect in order to identify therapeutic strategies that promote tumor cell apoptosis and might improve clinical response.
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Apoptosis/efectos de los fármacos , Neoplasias de la Mama/inmunología , Neoplasias de la Mama/terapia , Inhibidores Enzimáticos/farmacología , Receptores ErbB/antagonistas & inhibidores , Quinazolinas/farmacología , Receptor ErbB-2/antagonistas & inhibidores , Anticuerpos Monoclonales/inmunología , Neoplasias de la Mama/patología , Línea Celular Tumoral , Receptores ErbB/inmunología , Humanos , Lapatinib , Receptor ErbB-2/inmunologíaRESUMEN
PURPOSE: This study (EGF10004) assessed the safety/tolerability, pharmacokinetics, and clinical activity of daily oral dosing with lapatinib (GW572016) in patients with ErbB1-expressing and/or ErbB2-overexpressing advanced-stage refractory solid tumors. PATIENTS AND METHODS: Heavily pretreated patients with ErbB1-expressing and/or ErbB2-overexpressing metastatic cancers were randomly assigned to one of five dose cohorts of lapatinib administered once daily. Pharmacokinetic samples were obtained on days 1 and 20. Clinical response was assessed every 8 weeks. RESULTS: Sixty-seven patients with metastatic solid tumors were treated with lapatinib. The most frequently reported drug-related adverse events were diarrhea (42%) and rash (31%). No grade 4 drug-related adverse events were reported. Five grade 3 drug-related toxicities (gastrointestinal events and rash) were experienced by four patients. Drug-related interstitial pneumonitis or cardiac dysfunction associated with other ErbB-targeted therapies was not reported. Four patients with trastuzumab-resistant metastatic breast cancer-two of whom were classified as having inflammatory breast cancer-had partial responses (PRs). Twenty-four patients with various other carcinomas experienced stable disease, of whom 10 received lapatinib for > or = 6 months. The relationships between lapatinib dose or serum concentration and clinical response could not be adequately characterized due to the limited response data. The incidence of diarrhea increased with increasing dose, whereas the incidence of rash was not related to dose. CONCLUSION: Lapatinib was well tolerated at doses ranging from 500 to 1,600 mg once daily. Clinical activity was observed in heavily pretreated patients with ErbB1-expressing and/or ErbB2-overexpressing metastatic cancers, including four PRs in patients with trastuzumab-resistant breast cancers and prolonged stable disease in 10 patients.
Asunto(s)
Antineoplásicos/farmacocinética , Receptores ErbB/antagonistas & inhibidores , Neoplasias/tratamiento farmacológico , Quinazolinas/farmacocinética , Receptor ErbB-2/antagonistas & inhibidores , Adulto , Anciano , Anciano de 80 o más Años , Antineoplásicos/uso terapéutico , Biomarcadores de Tumor/análisis , Esquema de Medicación , Resistencia a Antineoplásicos , Receptores ErbB/metabolismo , Femenino , Humanos , Lapatinib , Masculino , Dosis Máxima Tolerada , Persona de Mediana Edad , Metástasis de la Neoplasia/tratamiento farmacológico , Recurrencia Local de Neoplasia/tratamiento farmacológico , Neoplasias/sangre , Quinazolinas/uso terapéutico , Receptor ErbB-2/metabolismo , Seguridad , Resultado del TratamientoRESUMEN
PURPOSE: This was a pilot study to assess the biologic effects of lapatinib on various tumor growth/survival pathways in patients with advanced ErbB1 and/or ErbB2-overexpressing solid malignancies. PATIENTS AND METHODS: Heavily pretreated patients with metastatic cancers overexpressing ErbB2 and/or expressing ErbB1 were randomly assigned to one of five dose cohorts of lapatinib (GW572016) administered orally once daily continuously. The biologic effects of lapatinib on tumor growth and survival pathways were assessed in tumor biopsies obtained before and after 21 days of therapy. Clinical response was determined at 8 weeks. RESULTS: Sequential tumor biopsies from 33 patients were examined. Partial responses occurred in four patients with breast cancer, and disease stabilization occurred in 11 others with various malignancies. Responders exhibited variable levels of inhibition of p-ErbB1, p-ErbB2, p-Erk1/2, p-Akt, cyclin D1, and transforming growth factor alpha. Even some nonresponders demonstrated varying degrees of biomarker inhibition. Increased tumor cell apoptosis (TUNEL) occurred in patients with evidence of tumor regression but not in nonresponders (progressive disease). Clinical response was associated with a pretreatment TUNEL score > 0 and increased pretreatment expression of ErbB2, p-ErbB2, Erk1/2, p-Erk1/2, insulin-like growth factor receptor-1, p70 S6 kinase, and transforming growth factor alpha compared with nonresponders. CONCLUSION: Lapatinib exhibited preliminary evidence of biologic and clinical activity in ErbB1 and/or ErbB2-overexpressing tumors. However, the limited sample size of this study and the variability of the biologic endpoints suggest that further work is needed to prioritize biomarkers for disease-directed studies, and underscores the need for improved trial design strategies in early clinical studies of targeted agents.
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Receptores ErbB/biosíntesis , Neoplasias/tratamiento farmacológico , Quinazolinas/farmacología , Quinazolinas/uso terapéutico , Receptor ErbB-2/biosíntesis , Adulto , Anciano , Anciano de 80 o más Años , Apoptosis , Biomarcadores de Tumor/análisis , Supervivencia Celular , Relación Dosis-Respuesta a Droga , Determinación de Punto Final , Femenino , Humanos , Inmunohistoquímica , Etiquetado Corte-Fin in Situ , Lapatinib , Masculino , Persona de Mediana Edad , Neoplasias/fisiopatología , Quinazolinas/administración & dosificación , Resultado del TratamientoRESUMEN
We report the case of a woman who conceived while being treated on a phase I clinical trial with lapatinib, a dual inhibitor of epidermal growth factor receptor (EGFR) and HER2/neu, for metastatic breast cancer. Despite approximately 11 weeks of exposure to lapatinib in the first and second trimesters, the pregnancy was uncomplicated and resulted in the delivery of a healthy baby. Although concomitant cancer and pregnancy is relatively rare, the increasing use of biologic agents among fertile women, sometimes for as long as a year in the adjuvant setting increases the probability that some women will conceive while taking a growth factor pathway inhibitor. As with systemic chemotherapy given during pregnancy, there exists the potential for teratogenicity or fetal demise from exposure of the developing embryo to inhibitors of EGFR and HER2/neu. Despite the positive outcome of this case, continued caution is warranted with the use of EGFR and HER2/neu inhibitors in pregnancy.
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Neoplasias de la Mama/tratamiento farmacológico , Nacimiento Vivo , Complicaciones Neoplásicas del Embarazo/tratamiento farmacológico , Primer Trimestre del Embarazo/efectos de los fármacos , Quinazolinas/uso terapéutico , Adulto , Antineoplásicos/uso terapéutico , Carcinoma Ductal de Mama/tratamiento farmacológico , Femenino , Humanos , Recién Nacido , Lapatinib , Metástasis de la Neoplasia/tratamiento farmacológico , EmbarazoRESUMEN
This work investigates X-PACT (X-ray Psoralen Activated Cancer Therapy): a new approach for the treatment of solid cancer. X-PACT utilizes psoralen, a potent anti-cancer therapeutic with current application to proliferative disease and extracorporeal photopheresis (ECP) of cutaneous T Cell Lymphoma. An immunogenic role for light-activated psoralen has been reported, contributing to long-term clinical responses. Psoralen therapies have to-date been limited to superficial or extracorporeal scenarios due to the requirement for psoralen activation by UVA light, which has limited penetration in tissue. X-PACT solves this challenge by activating psoralen with UV light emitted from novel non-tethered phosphors (co-incubated with psoralen) that absorb x-rays and re-radiate (phosphoresce) at UV wavelengths. The efficacy of X-PACT was evaluated in both in-vitro and in-vivo settings. In-vitro studies utilized breast (4T1), glioma (CT2A) and sarcoma (KP-B) cell lines. Cells were exposed to X-PACT treatments where the concentrations of drug (psoralen and phosphor) and radiation parameters (energy, dose, and dose rate) were varied. Efficacy was evaluated primarily using flow cell cytometry in combination with complimentary assays, and the in-vivo mouse study. In an in-vitro study, we show that X-PACT induces significant tumor cell apoptosis and cytotoxicity, unlike psoralen or phosphor alone (p<0.0001). We also show that apoptosis increases as doses of phosphor, psoralen, or radiation increase. Finally, in an in-vivo pilot study of BALBc mice with syngeneic 4T1 tumors, we show that the rate of tumor growth is slower with X-PACT than with saline or AMT + X-ray (p<0.0001). Overall these studies demonstrate a potential therapeutic effect for X-PACT, and provide a foundation and rationale for future studies. In summary, X-PACT represents a novel treatment approach in which well-tolerated low doses of x-ray radiation are delivered to a specific tumor site to generate UVA light which in-turn unleashes both short- and potentially long-term antitumor activity of photo-active therapeutics like psoralen.
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Ficusina/farmacología , Neoplasias/radioterapia , Terapia por Rayos X/métodos , Animales , Apoptosis/efectos de los fármacos , Apoptosis/efectos de la radiación , Línea Celular Tumoral , Transformación Celular Neoplásica , Relación Dosis-Respuesta en la Radiación , Ficusina/uso terapéutico , RatonesRESUMEN
The expression of the NH2 terminally truncated ErbB2 receptor (p95ErbB2) in breast cancer correlates with metastatic disease progression compared with the expression of full-length p185ErbB2. We now show that heregulin (HRG), but not EGF, stimulates p95ErbB2 phosphorylation in BT474 breast cancer cells. Furthermore, phospho-p95ErbB2 forms heterodimers with ErbB3, but not EGFR, while p185ErbB2 heterodimerizes with both EGFR and ErbB3. The predilection of p95ErbB2 to heterodimerize with ErbB3 provides an explanation for its regulation by HRG, an ErbB3 ligand. GW572016, a reversible small molecule inhibitor of EGFR and ErbB2 tyrosine kinases, inhibits baseline p95ErbB2 phosphorylation in BT474 cells and tumor xenografts. Inhibition of p95ErbB2, p185ErbB2, and EGFR phosphorylation by GW572016 resulted in the inhibition of downstream phospho-Erk1/2, phospho-AKT, and cyclin D steady-state protein levels. Increased phosphorylation of p95ErbB2 and AKT in response to HRG was abrogated to varying degrees by GW572016. In contrast, trastuzumab did not inhibit p95ErbB2 phosphorylation or the expression of downstream phospho-Erk1/2, phospho-AKT, or cyclin D. It is tempting to speculate that trastuzumab resistance may be mediated in part by the selection of p95ErbB2-expressing breast cancer cells capable of exerting potent growth and prosurvival signals through p95ErbB2-ErbB3 heterodimers. Thus, p95ErbB2 represents a target for therapeutic intervention, and one that is sensitive to GW572016 therapy.