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Cancer stem cells (CSCs) are a subpopulation of cancer cells endowed with high tumorigenic, chemoresistant and metastatic potential. Nongenetic mechanisms of acquired resistance are increasingly being discovered, but molecular insights into the evolutionary process of CSCs are limited. Here, we show that type I interferons (IFNs-I) function as molecular hubs of resistance during immunogenic chemotherapy, triggering the epigenetic regulator demethylase 1B (KDM1B) to promote an adaptive, yet reversible, transcriptional rewiring of cancer cells towards stemness and immune escape. Accordingly, KDM1B inhibition prevents the appearance of IFN-I-induced CSCs, both in vitro and in vivo. Notably, IFN-I-induced CSCs are heterogeneous in terms of multidrug resistance, plasticity, invasiveness and immunogenicity. Moreover, in breast cancer (BC) patients receiving anthracycline-based chemotherapy, KDM1B positively correlated with CSC signatures. Our study identifies an IFN-I â KDM1B axis as a potent engine of cancer cell reprogramming, supporting KDM1B targeting as an attractive adjunctive to immunogenic drugs to prevent CSC expansion and increase the long-term benefit of therapy.
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Neoplasias de la Mama , Epigénesis Genética , Histona Demetilasas , Interferón Tipo I , Antraciclinas/metabolismo , Antraciclinas/uso terapéutico , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Femenino , Histona Demetilasas/metabolismo , Humanos , Interferón Tipo I/metabolismo , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patologíaRESUMEN
Male breast cancer (MBC) is an uncommon malignancy. We have previously reported that the expression of the Hippo transducers TAZ/YAP and their target CTGF was associated with inferior survival in MBC patients. Preclinical evidence demonstrated that Axl is a transcriptional target of TAZ/YAP. Thus, we herein assessed AXL expression to further investigate the significance of active TAZ/YAP-driven transcription in MBC. For this study, 255 MBC samples represented in tissue microarrays were screened for AXL expression, and 116 patients were included. The association between categorical variables was verified by the Pearson's Chi-squared test of independence (2-tailed) or the Fisher Exact test. The relationship between continuous variables was tested with the Pearson's correlation coefficient. The Kaplan-Meier method was used for estimating survival curves, which were compared by log-rank test. Factors potentially impacting 10-year and overall survival were verified in Cox proportional regression models. AXL was positively associated with the TAZ/CTGF and YAP/CTGF phenotypes (P = 0.001 and P = 0.002, respectively). Patients with TAZ/CTGF/AXL- or YAP/CTGF/AXL-expressing tumors had inferior survival compared with non-triple-positive patients (log rank P = 0.042 and P = 0.048, respectively). The variables TAZ/CTGF/AXL and YAP/CTGF/AXL were adverse factors for 10-year survival in the multivariate Cox models (HR 2.31, 95%CI:1.02-5.22, P = 0.045, and HR 2.27, 95%CI:1.00-5.13, P = 0.050). Nearly comparable results were obtained from multivariate analyses of overall survival. The expression pattern of AXL corroborates the idea of the detrimental role of TAZ/YAP activation in MBC. Overall, Hippo-linked biomarkers deserve increased attention in this rare disease. J. Cell. Physiol. 232: 2246-2252, 2017. © 2016 Wiley Periodicals, Inc.
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Proteínas Adaptadoras Transductoras de Señales/análisis , Biomarcadores de Tumor/análisis , Neoplasias de la Mama Masculina/química , Fosfoproteínas/análisis , Proteínas Proto-Oncogénicas/análisis , Proteínas Tirosina Quinasas Receptoras/análisis , Factores de Transcripción/análisis , Aciltransferasas , Anciano , Neoplasias de la Mama Masculina/mortalidad , Neoplasias de la Mama Masculina/patología , Distribución de Chi-Cuadrado , Factor de Crecimiento del Tejido Conjuntivo/análisis , Humanos , Inmunohistoquímica , Estimación de Kaplan-Meier , Masculino , Persona de Mediana Edad , Análisis Multivariante , Valor Predictivo de las Pruebas , Pronóstico , Modelos de Riesgos Proporcionales , Estudios Retrospectivos , Factores de Riesgo , Transducción de Señal , Factores de Tiempo , Análisis de Matrices Tisulares , Proteínas Señalizadoras YAP , Tirosina Quinasa del Receptor AxlRESUMEN
BACKGROUND: Body mass index (BMI) is largely investigated as a prognostic and predictive factor in triple-negative breast cancer (TNBC). Overweight and obesity are linked to a variety of pathways regulating tumor-promoting functions, including the DNA damage response (DDR). The DDR physiologically safeguards genome integrity but, in a neoplastic background, it is aberrantly engaged and protects cancer cells from chemotherapy. We herein verified the role of BMI on a previously assessed association between DDR biomarkers and pathological complete response (pCR) in TNBC patients treated with neoadjuvant chemotherapy (NACT). METHODS: In this retrospective analysis 54 TNBC patients treated with NACT were included. The relationship between DDR biomarkers, namely phosphorylated H2A Histone Family Member X (γ-H2AX) and phosphorylated checkpoint kinase 1 (pChk1), and pCR was reconsidered in light of BMI data. The Pearson's Chi-squared test of independence (2-tailed) and the Fisher Exact test were employed to assess the relationship between clinical-molecular variables and pCR. Uni- and multivariate logistic regression models were used to identify variables impacting pCR. Internal validation was carried out. RESULTS: We observed a significant association between elevated levels of the two DDR biomarkers and pCR in patients with BMI < 25 (p = 0.009 and p = 0.022 for γ-H2AX and pChk1, respectively), but not in their heavier counterpart. Results regarding γ-H2AX were confirmed in uni- and multivariate models and, again, for leaner patients only (γ-H2AXhigh vs γ-H2AXlow: OR 10.83, 95% CI: 1.79-65.55, p = 0.009). The consistency of this finding was confirmed upon internal validation. CONCLUSIONS: The predictive significance of γ-H2AX varies according to BMI status. Indeed, elevated levels of γ-H2AX seemed associated with lower pCR rate only in leaner patients, whereas differences in pCR rate according to γ-H2AX levels were not appreciable in heavier patients. Larger investigations are warranted concerning the potential role of BMI as effect modifier of the relationship between DDR-related biomarkers and clinical outcomes in TNBC.
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Índice de Masa Corporal , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1)/análisis , Daño del ADN , Histonas/análisis , Terapia Neoadyuvante , Neoplasias de la Mama Triple Negativas/patología , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1)/química , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1)/metabolismo , Femenino , Histonas/metabolismo , Humanos , Persona de Mediana Edad , Obesidad/complicaciones , Fosforilación , Estudios Retrospectivos , Neoplasias de la Mama Triple Negativas/complicaciones , Neoplasias de la Mama Triple Negativas/diagnóstico , Neoplasias de la Mama Triple Negativas/terapiaAsunto(s)
Ado-Trastuzumab Emtansina/farmacología , Biomarcadores de Tumor , Neoplasias de la Mama/diagnóstico , Neoplasias de la Mama/etiología , Resistencia a Antineoplásicos , Biopsia Líquida , Receptor ErbB-2 , Ado-Trastuzumab Emtansina/uso terapéutico , Adulto , Anciano , Anciano de 80 o más Años , Neoplasias de la Mama/tratamiento farmacológico , Progresión de la Enfermedad , Resistencia a Antineoplásicos/genética , Femenino , Amplificación de Genes , Humanos , Biopsia Líquida/métodos , Persona de Mediana Edad , Clasificación del Tumor , Estadificación de Neoplasias , Pronóstico , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismoRESUMEN
BACKGROUND: The primary objectives of this study on carcinomas with equivocal HER2 expression were to assess the impact of distinct recommendations with regard to identifying patients eligible for anti-HER2 agents by fluorescence in situ hybridization (FISH) and to elucidate whether multiplex ligation-dependent probe amplification (MLPA) may be of support in assessing HER2 gene status. METHODS: A cohort of 957 immunohistochemistry-evaluated HER2-equivocal cases was analyzed by dual-color FISH. The results were assessed according to U.S. Food and Drug Administration (FDA) and European Medicines Agency (EMA) guidelines and American Society of Clinical Oncology (ASCO) and College of American Pathologists (CAP) 2007 and 2013 guidelines for dual- and single-signal in situ hybridization (ISH) assays. A subgroup of 112 cases was subjected to MLPA. RESULTS: HER2 amplification varied from 15% (ASCO/CAP 2007 HER2/CEP17 ratio) to 29.5% (FDA/EMA HER2 copy number). According to the ASCO/CAP 2013 interpretation of the dual-signal HER2 assay, ISH-positive carcinomas accounted for 19.7%. In contrast with the ASCO/CAP 2007 ratio, this approach labeled as positive all 32 cases (3.34%) with a HER2/CEP17 ratio <2 and an average HER2 copy number ≥6.0 signals per cell. In contrast, only one case showing a HER2 copy number <4 but a ratio ≥2 was diagnosed as positive. MLPA data correlated poorly with FISH results because of the presence of heterogeneous HER2 amplification in 33.9% of all amplified carcinomas; however, MLPA ruled out HER2 amplification in 75% of ISH-evaluated HER2-equivocal carcinomas. CONCLUSION: The ASCO/CAP 2013 guidelines seem to improve the identification of HER2-positive carcinomas. Polymerase chain reaction-based methods such as MLPA can be of help, provided that heterogeneous amplification has been ruled out by ISH.
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Neoplasias de la Mama/genética , Reacción en Cadena de la Polimerasa/métodos , Receptor ErbB-2/genética , Autoantígenos/genética , Biomarcadores de Tumor/genética , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Proteínas de Ciclo Celular/genética , Estudios de Cohortes , Femenino , Amplificación de Genes , Dosificación de Gen , Guías como Asunto , Humanos , Inmunohistoquímica , Hibridación Fluorescente in Situ , Receptor ErbB-2/metabolismoRESUMEN
BACKGROUND: During targeted treatment, HER2-positive breast cancers invariably lose HER2 DNA amplification. In contrast, and interestingly, HER2 proteins may be either lost or gained. To longitudinally and systematically appreciate complex/discordant changes in HER2 DNA/protein stoichiometry, HER2 DNA copy numbers and soluble blood proteins (aHER2/sHER2) were tested in parallel, non-invasively (by liquid biopsy), and in two-dimensions, hence HER2-2D. METHODS: aHER2 and sHER2 were assessed by digital PCR and ELISA before and after standard-of-care treatment of advanced HER2-positive breast cancer patients (n=37) with the antibody-drug conjugate (ADC) Trastuzumab-emtansine (T-DM1). RESULTS: As expected, aHER2 was invariably suppressed by T-DM1, but this loss was surprisingly mirrored by sHER2 gain, sometimes of considerable entity, in most (30/37; 81%) patients. This unorthodox split in HER2 oncogenic dosage was supported by reciprocal aHER2/sHER2 kinetics in two representative cases, and an immunohistochemistry-high status despite copy-number-neutrality in 4/5 available post-T-DM1 tumor re-biopsies from sHER2-gain patients. Moreover, sHER2 was preferentially released by dying breast cancer cell lines treated in vitro by T-DM1. Finally, sHER2 gain was associated with a longer PFS than sHER2 loss (mean PFS 282 vs 133 days, 95% CI [210-354] vs [56-209], log-rank test p=0.047), particularly when cases (n=11) developing circulating HER2-bypass alterations during T-DM1 treatment were excluded (mean PFS 349 vs 139 days, 95% CI [255-444] vs [45-232], log-rank test p=0.009). CONCLUSIONS: HER2 gain is adaptively selected in tumor tissues and recapitulated in blood by sHER2 gain. Possibly, an increased oncogenic dosage is beneficial to the tumor during anti-HER2 treatment with naked antibodies, but favorable to the host during treatment with a strongly cytotoxic ADC such as T-DM1. In the latter case, HER2-gain tumors may be kept transiently in check until alternative oncogenic drivers, revealed by liquid biopsy, bypass HER2. Whichever the interpretation, HER2-2D might help to tailor/prioritize anti-HER2 treatments, particularly ADCs active on aHER2-low/sHER2-low tumors. TRIAL REGISTRATION: NCT05735392 retrospectively registered on January 31, 2023 https://www. CLINICALTRIALS: gov/search?term=NCT05735392.
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Neoplasias de la Mama , Receptor ErbB-2 , Humanos , Femenino , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/patología , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Receptor ErbB-2/metabolismo , Biopsia Líquida/métodos , Persona de Mediana Edad , Ado-Trastuzumab Emtansina/uso terapéutico , Anciano , Trastuzumab/uso terapéutico , Trastuzumab/farmacología , Adulto , Biomarcadores de TumorRESUMEN
We assessed the impact of DNA damage response and repair (DDR) biomarker expressions in 222 node-positive early breast cancer (BC) patients from a previous Phase III GOIM 9902 trial of adjuvant taxanes. At a median follow-up of 64 months, the original study showed no disease-free survival (DFS) or overall survival (OS) differences with the addition of docetaxel (D) to epirubicine-cyclophosphamide (EC). Immunohistochemistry was employed to assess the expression of DDR phosphoproteins (pATM, pATR, pCHK1, γH2AX, pRPA32, and pWEE1) in tumor tissue, and their association with clinical outcomes was evaluated through the Cox elastic net model. Over an extended follow-up of 234 months, we confirmed no significant differences in DFS or OS between patients treated with EC and those receiving D â EC. A DDR risk score, inversely driven by ATM and ATR expression, emerged as an independent prognostic factor for both DFS (HR = 0.41, p < 0.0001) and OS (HR = 0.61, p = 0.046). Further validation in a public adjuvant BC cohort was possible only for ATM, confirming its protective role. Overall, our findings confirm the potential role of the DDR pathway in BC prognostication and in shaping treatment strategies advocating for an integrated approach, combining molecular markers with clinical-pathological factors.
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Phospholipase Cγ1 (PLCγ1) is highly expressed in several tumors. We have previously reported that both stable and inducible PLCγ1 down-regulation resulted in an almost complete inhibition of breast cancer-derived experimental lung metastasis formation. The aim of our study is to evaluate the association between the expression of PLCγ1 and of PLCγ1 phosphorylated at Tyr1253 (PLCγ1-pY1253) and at Tyr783 (PLCγ1-pY783) with the clinical outcome of patients with node negative, T1/T2 breast cancers. The study groups consisted of 292 (training set) and 122 (validation set) patients presenting with primary unilateral breast carcinoma (T1-T2), with no evidence of nodal involvement and distant metastases. PLCγ1, PLCγ1-pY1253 and PLCγ1-pY783 protein expression were assessed by immunohistochemistry on tissue microarrays and the results correlated with the clinical data using Kaplan-Meier curves and multivariate Cox regression analysis. Tumor cells while expressing variable proportions of cytoplasmic PLCγ1, express PLCγ1-pY1253 and PLCγ1-pY783 predominantly in the nucleus. High expression of PLCγ1, and of its activated forms, is associated with a worse clinical outcome in terms of incidence of distant metastases, and not of local relapse in T1-T2, N0 breast cancer patients undergone adjuvant chemotherapy. PLCγ1 over-expression appears to be a reliable predictive surrogate marker of development of metastases. Thus, targeting PLCγ1 pathways might represent a potential therapeutic approach for the prevention of metastatic disease in breast cancer.
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Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/enzimología , Fosfolipasa C gamma/biosíntesis , Anciano , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Quimioterapia Adyuvante , Supervivencia sin Enfermedad , Femenino , Humanos , Inmunohistoquímica , Persona de Mediana Edad , Metástasis de la Neoplasia , Estadificación de Neoplasias , Fosfolipasa C gamma/genética , Fosfolipasa C gamma/metabolismo , Factores de RiesgoRESUMEN
KRAS mutations are the most common oncogenic event in colorectal cancer (CRC) progression and their occurrence is associated with lack of response to anti epidermal growth factor receptor (EGFR) targeted therapies. Using preclinical models and patients' samples we recently reported that the emergence of KRAS mutations but also KRAS amplification is associated with acquired resistance to the EGFR inhibitors cetuximab or panitumumab. We reasoned that KRAS amplification may also be responsible for primary resistance to these agents. Furthermore, while the prevalence of KRAS mutations has been well established in CRC, little is known about the frequency of KRAS amplification in large CRC series. We performed a screening of 1,039 CRC samples to assess the prevalence of KRAS amplification in this tumor type and further evaluated the role of this genetic alteration on the sensitivity to anti EGFR therapies. We detected KRAS amplification in 7/1,039 (0.67%) and 1/102 evaluable CRC specimens and cell lines, respectively. KRAS amplification was mutually exclusive with KRAS mutations. Tumors or cell lines harboring this genetic lesion are not responsive to anti-EGFR inhibitors. Although KRAS amplification is an infrequent event in CRC, it might be responsible for precluding response to anti-EGFR treatment in a small proportion of patients.
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Neoplasias Colorrectales/genética , Receptores ErbB/antagonistas & inhibidores , Amplificación de Genes , Proteínas Proto-Oncogénicas/genética , Proteínas ras/genética , Línea Celular Tumoral , Neoplasias Colorrectales/tratamiento farmacológico , Humanos , Proteínas Proto-Oncogénicas p21(ras) , Estudios RetrospectivosRESUMEN
Under therapeutic pressure aggressive tumors evolve rapidly. Herein, a luminal B/HER2-low breast cancer was tracked for >3 years during a total of 6 largely unsuccessful therapy lines, from adjuvant to advanced settings. Targeted next generation sequencing (NGS) of the primary lesion, two metastases and 14 blood drawings suggested a striking, unprecedented coexistence of three evolution modes: punctuated, branched and convergent. Punctuated evolution of the trunk was supported by en bloc inheritance of a large set (19 distinct genes) of copy number alterations. Branched evolution was supported by the distribution of site-specific SNVs. Convergent evolution was characterized by a unique asynchronous expansion of three actionable (OncoKB level 3A) mutations at two consecutive ESR1 codons. Low or undetectable in all the sampled tumor tissues, ESR1 mutations expanded rapidly in blood during HER2/hormone double-blockade, and predicted life-threatening local progression at lung and liver metastatic foci. Dramatic clinical response to Fulvestrant (assigned off-label exclusively based on liquid biopsy) was associated with clearance of all 3 subclones and was in stark contrast to the poor therapeutic efficacy reported in large liquid biopsy-informed interventional trials. Altogether, deconvolution of the tumor phylogenetic tree, as shown herein, may help to customize treatment in breast cancers that rapidly develop refractoriness to multiple drugs.
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AIMS: The minimally invasive procedures used in the diagnostic workup of patients with advanced non-small cell lung cancer (NSCLC) often provide poor yields of pathological material suitable for molecular analyses. Not infrequently, the DNA yield from small biopsies/cytological samples is insufficient for the assessment of genomic biomarkers that inform personalised therapies. The Idylla EGFR mutation test (IEMT) has been specifically designed to process formalin-fixed paraffin-embedded sections without requiring preliminary DNA extraction.This study aims to evaluate the diagnostic accuracy of IEMT when used to analyse archival histopathology material. More specifically, our objective was to establish whether or not different staining procedures could affect assay performance. METHODS: Twenty NSCLC samples were selected accordingly to EGFR mutational status. To mimic archived stained material, sections were subjected to H&E staining, fluorescent in situ hybridisation analyses or immunodetection by immunohistochemistry before being processed for IEMT. RESULTS: Parallel assessment of EGFR mutational status by IEMT on stained sections and next-generation sequencing on DNA yielded a concordant result in 50 out of 60 tests (83.3%). The discoloration of H&E of the archived sample was found to be the optimal procedure to highlight all the actionable alterations of EGFR. CONCLUSIONS: IEMT can provide remarkable diagnostic accuracy for the assessment of EGFR mutational status also when the only source of pathological material available for molecular analyses is represented by H&E stained sections. Ad hoc supervision by a qualified molecular biologist is in any case recommended.
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Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Humanos , Carcinoma de Pulmón de Células no Pequeñas/diagnóstico , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/patología , Análisis Mutacional de ADN/métodos , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Estudios de Factibilidad , Receptores ErbB/genética , MutaciónRESUMEN
The approval of osimertinib for adjuvant treatment of stage I-II-III EGFR-mutated NSCLC (early stage) represents a paradigm shift, raising the question of whether other genotype-matched therapeutics approved for advanced-stage NSCLC can also provide clinical benefit in the adjuvant setting. However, there is a paucity of real-world data on the prevalence of actionable genomic alterations (GAs) in early-stage NSCLC. We used next-generation sequencing, complemented by immunohistochemistry and fluorescence in situ hybridization, to screen our single-institution cohort of 1961 NSCLC consecutive cases for actionable molecular targets. The prevalence of actionable GAs was comparable in early versus advanced-stage NSCLC, the only exception being KRAS mutations (more frequent in early-stage cases). Consistent with advanced-stage tumors being more aggressive, co-occurrence of TP53 and EGFR GAs as well as copy number gains were less frequent in early-stage tumors. EGFR mutations and high expression of PD-L1 were inversely associated, whereas KRAS mutations and high PD-L1 reactivity showed positive association. Recapitulating advanced-stage tumors, early-stage NSCLC had the highest share of EGFR mutations in lepidic and acinar subtypes. Resected lepidic tumors contained the highest proportion of the KRAS G12C actionable variant. These results, obtained with routine diagnostic technologies in an unselected clinical setting, provide a significant addition of real-world data in early-stage NSCLC.
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Targeted therapy in lung cancer requires the assessment of multiple oncogenic driver alterations, including fusion genes. This retrospective study evaluated the Idylla GeneFusion prototype, an automated and ease-of-use (<2 minutes) test, with a short turnaround time (3 hours) to detect fusions involving ALK, ROS1, RET, and NTRK1/2/3 genes and MET exon 14 skipping. This multicenter study (18 centers) included 313 tissue samples from lung cancer patients with 97 ALK, 44 ROS1, 20 RET, and 5 NTRKs fusions, 32 MET exon 14 skipping, and 115 wild-type samples, previously identified with reference methods (RNA-based next-generation sequencing/fluorescence in situ hybridization/quantitative PCR). Valid results were obtained for 306 cases (98%), overall concordance between Idylla and the reference methods was 89% (273/306); overall sensitivity and specificity were 85% (165/193) and 96% (108/113), respectively. Discordances were observed in 28 samples, where Idylla did not detect the alteration identified by the reference methods; and 5 samples where Idylla identified an alteration not detected by the reference methods. All of the ALK-, ROS1-, and RET-specific fusions and MET exon 14 skipping identified by Idylla GeneFusion were confirmed by reference method. To conclude, Idylla GeneFusion is a clinically valuable test that does not require a specific infrastructure, allowing a rapid result. The absence of alteration or the detection of expression imbalance only requires additional testing by orthogonal methods.
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Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Carcinoma de Pulmón de Células no Pequeñas/diagnóstico , Carcinoma de Pulmón de Células no Pequeñas/genética , Humanos , Hibridación Fluorescente in Situ , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genética , Mutación , Proteínas Tirosina Quinasas/genética , Proteínas Tirosina Quinasas/metabolismo , Proteínas Proto-Oncogénicas/genética , Proteínas Tirosina Quinasas Receptoras/genética , Estudios RetrospectivosRESUMEN
The Hippo pathway and its two key effectors, Yes-associated protein (YAP) and transcriptional coactivator with PDZ-binding motif (TAZ), are consistently altered in breast cancer. Pivotal regulators of cell metabolism such as the AMP-activated protein kinase (AMPK), Stearoyl-CoA-desaturase 1 (SCD1), and HMG-CoA reductase (HMGCR) are relevant modulators of TAZ/YAP activity. In this prospective study, we measured the tumor expression of TAZ, YAP, AMPK, SCD1, and HMGCR by immunohistochemistry in 65 Her2+ breast cancer patients who underwent trastuzumab-based neoadjuvant treatment. The aim of the study was to assess the impact of the immunohistochemical expression of the Hippo pathway transducers and cell metabolism regulators on pathological complete response. Low expression of cytoplasmic TAZ, both alone and in the context of a composite signature identified by machine learning including also low nuclear levels of YAP and HMGCR and high cytoplasmic levels of SCD1, was a predictor of residual disease in the univariate logistic regression. This finding was not confirmed in the multivariate model including estrogen receptor > 70% and body mass index > 20. However, our findings were concordant with overall survival data from the TCGA cohort. Our results, possibly affected by the relatively small sample size of this study population, deserve further investigation in adequately sized, ad hoc prospective studies.
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BACKGROUND: Human Leukocyte Antigen (HLA)-E is a non-classical class I HLA molecule that can be stabilized by ligands donated by other classical (HLA-A, -B, -C) and non-classical (HLA-G) family members. HLA-E engages a variety of immune receptors expressed by cytotoxic T lymphocytes (CTLs), Natural killer (NK) cells and NK-CTLs. In view of the opposing outcomes (activation or inhibition) of the different HLA-E receptors, the preferred role (if any) of HLA-E expressed in vivo on tumor cells remains to be established. METHODS: Taking advantage of MEM-E/02, a recently characterized antibody to denatured HLA-E molecules, HLA-E expression was assessed by immunohistochemistry on an archival collection (formalin-fixed paraffin-embedded) of 149 colorectal primary carcinoma lesions paired with their morphologically normal mucosae. Lymphoid infiltrates were assessed for the expression of the HLA-E-specific, inhibitory, non-rearranging receptor NKG2A. RESULTS: High HLA-E expression did not significantly correlate with the expression of classical HLA-B and HLA-C molecules, but it did correlate with high expression of its preferential ligand donor HLA-A. In addition, it correlated with lymphoid cell infiltrates expressing the inhibitory NKG2A receptor, and was an independent predictor of good prognosis, particularly in a subset of patients whose tumors express HLA-A levels resembling those of their paired normal counterparts (HLA-A). Thus, combination phenotypes (HLA-Elo-int/HLA-AE and HLA-Ehi/HLA-AE) of classical and non-classical class I HLA molecules mark two graded levels of good prognosis. CONCLUSIONS: These results suggest that HLA-E favors activating immune responses to colorectal carcinoma. They also provide evidence in humans that tumor cells entertain extensive negotiation with the immune system until a compromise between recognition and escape is reached. It is implied that this process occurs stepwise, as predicted by the widely accepted 'immunoediting' model.
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Neoplasias Colorrectales/diagnóstico , Neoplasias Colorrectales/inmunología , Antígenos de Histocompatibilidad Clase I/inmunología , Adulto , Anciano , Anciano de 80 o más Años , Alelos , Anticuerpos/inmunología , Antígenos CD8/inmunología , Neoplasias Colorrectales/patología , Femenino , Antígenos de Histocompatibilidad Clase I/genética , Humanos , Inmunohistoquímica , Mucosa Intestinal/inmunología , Mucosa Intestinal/patología , Estimación de Kaplan-Meier , Masculino , Persona de Mediana Edad , Modelos Inmunológicos , Análisis Multivariante , Subfamília C de Receptores Similares a Lectina de Células NK/inmunología , Estadificación de Neoplasias , Adhesión en Parafina , Pronóstico , Fijación del Tejido , Antígenos HLA-ERESUMEN
BACKGROUND: Understanding the signalling pathways involved in angiogenesis, and developing anti-angiogenic drugs are one of the major focuses on cancer research. Herein, we assessed the effect of CPTH6, a lysine acetyltransferase inhibitor and anti-tumoral compound, on angiogenesis-related properties of both endothelial and cancer cells. METHODS: The in vitro effect of CPTH6 on protein acetylation and anti-angiogenic properties on endothelial and lung cancer cells was evaluated via wound healing, trans-well invasion and migration, tube formation, immunoblotting and immunofluorescence. Matrigel plug assay, zebrafish embryo and mouse xenograft models were used to evaluate in vivo anti-angiogenic effect of CPTH6. RESULTS: CPTH6 impaired in vitro endothelial angiogenesis-related functions, and decreased the in vivo vascularization both in mice xenografts and zebrafish embryos. Mechanistically, CPTH6 reduced α-tubulin acetylation and induced accumulation of acetylated microtubules in the perinuclear region of endothelial cells. Interestingly, CPTH6 also affected the angiogenesis-related properties of lung cancer cells, and conditioned media derived from CPTH6-treated lung cancer cells impaired endothelial cells morphogenesis. CPTH6 also modulated the VEGF/VEGFR2 pathway, and reshaped cytoskeletal organization of lung cancer cells. Finally, anti-migratory effect of CPTH6, dependent on α-tubulin acetylation, was also demonstrated by genetic approaches in lung cancer cells. CONCLUSION: Overall, this study indicates that α-tubulin acetylation could play a role in the anti-angiogenic effect of CPTH6 and, more in general, it adds information to the role of histone acetyltransferases in tumor angiogenesis, and proposes the inhibition of these enzymes as an antiangiogenic therapy of cancer.
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Inhibidores de la Angiogénesis/farmacología , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Neoplasias Pulmonares/irrigación sanguínea , Lisina Acetiltransferasas/antagonistas & inhibidores , Neovascularización Patológica/tratamiento farmacológico , Tiazoles/farmacología , Animales , Apoptosis , Diferenciación Celular , Movimiento Celular , Proliferación Celular , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Ratones , Ratones Endogámicos NOD , Ratones SCID , Neovascularización Patológica/metabolismo , Neovascularización Patológica/patología , Transducción de Señal , Células Tumorales Cultivadas , Factor A de Crecimiento Endotelial Vascular/genética , Factor A de Crecimiento Endotelial Vascular/metabolismo , Receptor 2 de Factores de Crecimiento Endotelial Vascular/genética , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
INTRODUCTION: p53 and antiapoptotic B-cell leukemia/lymphoma 2 (BLC2) have been proposed as prognostic markers for early breast cancer (BC), although their relationship with conventional parameters and patient prognosis, as well as their distribution within the molecular BC subtypes remains uncertain. PATIENTS AND METHODS: In this observational study, we analyzed the immunohistochemical expression of p53 and BLC2 in 1099 early BC patients surgically treated between 2000 and 2006 and followed for at least 5 years, also considering their association with pathologic factors and molecular subtypes, as well as their influence on disease-free survival. RESULTS: p53 and BLC2 are distributed differently across molecular subtypes (P < .0001); in particular, p53 positivity and BLC2 negativity seems to be associated with more aggressive conventional tumor phenotypes. Moreover, BLC2 negativity seems to be a significant discriminating factor for disease-free survival (P = .003) according to Kaplan-Meier analysis, while p53 seems to have no discriminating effect. Among patients with discordant p53/BLC2 phenotype, the combination p53+BLC2- seems to be associated with the worst outcomes (P = .007) and significantly influenced the clinical course of node-negative patients treated only with hormone therapy (P = .004). CONCLUSION: These two biomarkers, in addition to conventional pathologic factors and molecular subtype, could help define the risk and outcome of BC.
Asunto(s)
Biomarcadores de Tumor/metabolismo , Neoplasias de la Mama/mortalidad , Recurrencia Local de Neoplasia/epidemiología , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Antineoplásicos Hormonales/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Biomarcadores de Tumor/análisis , Mama/patología , Mama/cirugía , Neoplasias de la Mama/diagnóstico , Neoplasias de la Mama/patología , Neoplasias de la Mama/terapia , Quimioterapia Adyuvante/métodos , Supervivencia sin Enfermedad , Femenino , Estudios de Seguimiento , Humanos , Inmunohistoquímica , Estimación de Kaplan-Meier , Mastectomía , Persona de Mediana Edad , Recurrencia Local de Neoplasia/patología , Pronóstico , Proteínas Proto-Oncogénicas c-bcl-2/análisis , Receptores de Progesterona , Proteína p53 Supresora de Tumor/análisis , Adulto JovenRESUMEN
Evaluating the expression levels of miR-378a-5p both in a large melanoma patient cohort from The Cancer Genome Atlas database and in melanoma patients from our Institute, we found that miR-378a-5p is upregulated in metastatic melanoma specimens. miR-378a-5p expression was also increased in melanoma cells resistant to target therapy, and decreased in response to drug treatment. We also demonstrated that overexpression of miR-378a-5p enhances in vitro cell invasion and migration, and facilitates the ability of melanoma cells to form de novo vasculogenic structures. While performing downstream targeting studies, we confirmed the ability of miR-378a-5p to modulate the expression of known target genes, such as SUFU, FUS-1, and KLF9. Luciferase-3'UTR experiments also identified STAMBP and HOXD10 as new miR-378a-5p target genes. MMP2 and uPAR, two HOXD10 target genes, were positively regulated by miR-378a-5p. Genetic and pharmacologic approaches inhibiting uPAR expression and activity evidenced that the in vitro tumor-promoting functions of miR-378a-5p, were in part mediated by uPAR. Of note miR-378a-5p was also able to increase VEGF, as well as in vitro and in vivo angiogenesis. Finally, genetic and pharmacologic modulation of Bcl-2 evidenced Bcl-2 ability to regulate miR-378a-5p expression. In conclusion, to the best of our knowledge, this is the first study demonstrating that miR-378a-5p acts as an oncogenic microRNA in melanoma.
RESUMEN
BACKGROUND: A bidirectional crosstalk between tumor cells and the surrounding microenvironment contributes to tumor progression and response to therapy. Our previous studies have demonstrated that bcl-2 affects melanoma progression and regulates the tumor microenvironment. The aim of this study was to evaluate whether bcl-2 expression in melanoma cells could influence tumor-promoting functions of tumor-associated macrophages, a major constituent of the tumor microenvironment that affects anticancer immunity favoring tumor progression. METHODS: THP-1 monocytic cells, monocyte-derived macrophages and melanoma cells expressing different levels of bcl-2 protein were used. ELISA, qRT-PCR and Western blot analyses were used to evaluate macrophage polarization markers and protein expression levels. Chromatin immunoprecipitation assay was performed to evaluate transcription factor recruitment at specific promoters. Boyden chamber was used for migration experiments. Cytofluorimetric and immunohistochemical analyses were carried out to evaluate infiltrating macrophages and T cells in melanoma specimens from patients or mice. RESULTS: Higher production of tumor-promoting and chemotactic factors, and M2-polarized activation was observed when macrophages were exposed to culture media from melanoma cells overexpressing bcl-2, while bcl-2 silencing in melanoma cells inhibited the M2 macrophage polarization. In agreement, the number of melanoma-infiltrating macrophages in vivo was increased, in parallel with a greater expression of bcl-2 in tumor cells. Tumor-derived interleukin-1ß has been identified as the effector cytokine of bcl-2-dependent macrophage reprogramming, according to reduced tumor growth, decreased number of M2-polarized tumor-associated macrophages and increased number of infiltrating CD4+IFNγ+ and CD8+IFNγ+ effector T lymphocytes, which we observed in response to in vivo treatment with the IL-1 receptor antagonist kineret. Finally, in tumor specimens from patients with melanoma, high bcl-2 expression correlated with increased infiltration of M2-polarized CD163+ macrophages, hence supporting the clinical relevance of the crosstalk between tumor cells and microenvironment. CONCLUSIONS: Taken together, our results show that melanoma-specific bcl-2 controls an IL-1ß-driven axis of macrophage diversion that establishes tumor microenvironmental conditions favoring melanoma development. Interfering with this pathway might provide novel therapeutic strategies.
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Melanoma/patología , Monocitos/inmunología , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Microambiente Tumoral/inmunología , Macrófagos Asociados a Tumores/patología , Animales , Apoptosis , Diferenciación Celular , Movimiento Celular , Proliferación Celular , Femenino , Humanos , Melanoma/inmunología , Melanoma/metabolismo , Ratones , Ratones Endogámicos C57BL , Fenotipo , Células Tumorales Cultivadas , Macrófagos Asociados a Tumores/inmunología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Cancer stem-like cells (CSC) induce aggressive tumor phenotypes such as metastasis formation, which is associated with poor prognosis in triple-negative breast cancer (TNBC). Repurposing of FDA-approved drugs that can eradicate the CSC subcompartment in primary tumors may prevent metastatic disease, thus representing an effective strategy to improve the prognosis of TNBC. Here, we investigated spheroid-forming cells in a metastatic TNBC model. This strategy enabled us to specifically study a population of long-lived tumor cells enriched in CSCs, which show stem-like characteristics and induce metastases. To repurpose FDA-approved drugs potentially toxic for CSCs, we focused on pyrvinium pamoate (PP), an anthelmintic drug with documented anticancer activity in preclinical models. PP induced cytotoxic effects in CSCs and prevented metastasis formation. Mechanistically, the cell killing effects of PP were a result of inhibition of lipid anabolism and, more specifically, the impairment of anabolic flux from glucose to cholesterol and fatty acids. CSCs were strongly dependent upon activation of lipid biosynthetic pathways; activation of these pathways exhibited an unfavorable prognostic value in a cohort of breast cancer patients, where it predicted high probability of metastatic dissemination and tumor relapse. Overall, this work describes a new approach to target aggressive CSCs that may substantially improve clinical outcomes for patients with TNBC, who currently lack effective targeted therapeutic options. SIGNIFICANCE: These findings provide preclinical evidence that a drug repurposing approach to prevent metastatic disease in TNBC exploits lipid anabolism as a metabolic vulnerability against CSCs in primary tumors.