RESUMEN
AIMS: The diagnosis of breast cancer (BC) is based on clinical examination in combination with imaging, and confirmed by pathological assessment of core needle biopsy or fine-needle aspiration biopsy (FNAB). The biological profile of the lesion is needed to define the prognosis and establish therapy. Given the importance of an early and minimally invasive diagnosis, we aimed to verify whether the biological features detected in FNAB-derived cytological material reflect the biological characteristics of surgical samples. METHODS AND RESULTS: We used immunohistochemistry and fluorescence in-situ hybridisation to study a panel of conventional biomarkers [oestrogen receptor (ER), progesterone receptor (PgR), Ki67, and human epidermal growth factor receptor 2 (HER2)] in FNAB-derived cytological samples included in cell blocks of 93 BC patients, and compared the results with those obtained from histological evaluation of the same parameters in surgical samples. Median immunopositive values of ER, PgR and Ki67 were similar in cell blocks and surgical samples. The concordance rates of ER and PgR between FNAB-derived cell blocks and histological samples were 98% and 84%, respectively. The concordance rates of Ki67 and HER2 between the two sample types were 90% and 96%, respectively. Tumour subtype classification for triple-negative and HER2-positive BCs in FNAB-derived cell blocks was always concordant with the subtype determined in surgical material. CONCLUSIONS: We demonstrated that biological marker determination in FNAB-derived cell blocks is feasible, and provides useful information and comparable results to those obtained with histological evaluation. Given the low cost of the procedure and its minimal impact on patients, we believe that cytological samples could be used as an alternative to tissue samples for early BC biomarker evaluation.
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Biomarcadores de Tumor/análisis , Biopsia con Aguja Fina , Neoplasias de la Mama/diagnóstico , Adulto , Anciano , Anciano de 80 o más Años , Biopsia con Aguja Gruesa , Citodiagnóstico/métodos , Femenino , Humanos , Persona de Mediana EdadRESUMEN
Eribulin mesylate is a novel, non-taxane, synthetic microtubule inhibitor showing antitumor activity in a wide range of tumors including soft tissue sarcomas (STS). Eribulin has been recently approved for the treatment of metastatic liposarcoma (LPS) patients previously treated with anthracyclines. This work investigated the mechanism of action of this innovative antitubulin agent in well-differentiated/dedifferentiated LPS (ALT/DDLPS) which represents one of the most common adipocytic sarcoma histotypes. A primary culture of ALT/DDLPS from a 54-year-old patient was established. The anticancer activity of eribulin on the patient-derived primary culture was assessed by MTT and tunel assays. Eribulin efficacy was compared to other drugs approved for the treatment of STS. Cell migration and morphology were examined after exposure to eribulin to better understand the drug mechanism of action. Finally, Western blot analysis of apoptosis and migration proteins was performed. The results showed that eribulin exerts its antiproliferative effect by the arrest of cell motility and induction of apoptosis. Our results highlighted the activity of eribulin in the treatment of ALT/DDLPS patients.
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Antineoplásicos/farmacología , Furanos/farmacología , Cetonas/farmacología , Liposarcoma/tratamiento farmacológico , Moduladores de Tubulina/farmacología , Transformación Celular Neoplásica , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Liposarcoma/patología , Persona de Mediana Edad , Cultivo Primario de Células , Células Tumorales CultivadasRESUMEN
The present study deals with the preparation of albumin nanocapsules containing fenretinide and their evaluation in experimental models of human non-small cell lung cancer. These nanocapsules showed enhanced antitumor activity with respect to free fenretinide due to the solubilization effect of albumin on the hydrophobic drug, known to improve bioavailability. The high expression of caveolin-1 on the A549 cell surface further enhanced the antitumor activity of the nanoencapsulated fenretinide. Caveolin-1 favored albumin uptake and improved the efficacy of the fenretinide-loaded albumin nanocapsules, especially in 3-D cultures where the densely packed 3-D structures impaired drug diffusibility and severely reduced the activity of the free drug. The efficacy of the fenretinide albumin nanocapsules was further confirmed in tumor xenograft models of A549 by the significant delay in tumor progression observed with respect to control after intravenous administration of the novel formulation. FROM THE CLINICAL EDITOR: This study describes the preparation of fenretinide containing albumin nanocapsules and their evaluation in experimental models of non-small cell lung cancer, showing enhanced antitumor activity compared to free fenretinide.
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Antineoplásicos/química , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Fenretinida/administración & dosificación , Nanocápsulas/administración & dosificación , Albúminas/administración & dosificación , Albúminas/química , Animales , Antineoplásicos/administración & dosificación , Disponibilidad Biológica , Línea Celular Tumoral , Humanos , Ratones , Nanocápsulas/química , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Although combination chemotherapy and radiotherapy have become the standard of care in numerous tumors, the mechanisms of interaction are often still unclear. The purpose of this study was to analyze the efficacy of radiation treatment and cisplatin sequences and to investigate their mechanisms of interaction. Three melanoma cell lines were used to evaluate in vitro radiation-induced cytotoxicity before and after cisplatin treatment. Expression levels of a panel of genes were determined by real-time RT-PCR. Cytotoxic effect was evaluated by flow cytometry analysis and Comet assay. We also used normal human dermal fibroblasts (HUDE) to evaluate the cytotoxicity of the two treatments by clonogenic assay. Radiation and cisplatin used singly were not particularly effective in reducing proliferation in melanoma cells. Conversely, radiation treatment followed by cisplatin showed a strong synergistic interaction in all cell lines, with a ratio index ranging from 16 to >100. The synergistic effect was accompanied by apoptosis induction (up to 40%) and an increase in the percentage of comet-shaped nucleoids from 85% to 99%. In parallel, our results also showed that radiation treatment of HUDE fibroblasts followed by cisplatin only induced weak cytotoxicity. Our findings highlight the efficacy of the sequence radiation â cisplatin in reducing cell proliferation and in inducing apoptosis in melanoma cell lines. This sequence also modulated a network of proteins involved in DNA damage repair.
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Antineoplásicos/farmacología , Quimioradioterapia/métodos , Cisplatino/farmacología , Melanoma/patología , Neoplasias Cutáneas/patología , Antineoplásicos/toxicidad , Apoptosis/efectos de los fármacos , Apoptosis/efectos de la radiación , Ciclo Celular/efectos de los fármacos , Ciclo Celular/efectos de la radiación , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Proliferación Celular/efectos de la radiación , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/efectos de la radiación , Quimioradioterapia/efectos adversos , Cisplatino/toxicidad , Ensayo Cometa , Daño del ADN , Relación Dosis-Respuesta a Droga , Relación Dosis-Respuesta en la Radiación , Esquema de Medicación , Citometría de Flujo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de la radiación , Humanos , Concentración 50 Inhibidora , Melanoma/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Neoplasias Cutáneas/genética , Factores de TiempoRESUMEN
There are no validated predictors of benefit from anthracyclines. We compared cyclophosphamide, methotrexate, 5-fluorouracil (CMF), and epirubicin in different sequences with CMF alone in a phase III trial on operable breast cancers. Outcomes were analyzed in relation to tumor biological profiles to identify potential predictors of the efficacy of different treatments/drug combinations. Patients with N- or 1-3N+ tumors, were randomized to receive (a) epirubicin (4 cycles) followed by CMF (4 cycles); (b) CMF (4 cycles) followed by epirubicin (4 cycles), or (c) CMF (6 cycles) alone. Immunohistochemical assessments of estrogen (ER) and progesterone (PgR) receptors, HER2 and Ki67 were available for 705 patients (arm A/B/C: 276/269/160). Prognostic and predictive relevance was analyzed by log-rank tests and Cox models. Ki67 > 20 % and absent/low expression of ER and PgR were associated with worsen disease-free (DFS) and overall survival (OS). In patients with triple negative tumors (ER-, PgR-, HER2-), epirubicin-containing regimens yielded better DFS (HR 0.33, 95 % CI 0.17-0.62, P = 0.0007) and OS (HR 0.24, 95 % CI 0.10-0.57, P = 0.001) compared with CMF alone, whereas no differences were found in patients with HER2-positive (HER2+, ER-, PgR-) subtype. Treatment by subtype interaction (HER2-positive vs. others) was significant for DFS (χ (2) = 6.72, P = 0.009). In triple unfavorable (ER-, PgR-, Ki67 > 20 %) tumors, the use of epirubicin yielded better DFS (HR 0.45,95 % CI 0.26-0.78, P = 0.005) and OS (HR 0.30, 95 % CI 0.15-0.63, P = 0.001). Epirubicin-containing regimens seem to be superior to CMF alone in patients with highly proliferating, triple negative or triple unfavorable tumors.
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Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Biomarcadores de Tumor/biosíntesis , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Neoplasias de la Mama Triple Negativas/metabolismo , Adulto , Protocolos de Quimioterapia Combinada Antineoplásica/administración & dosificación , Ciclofosfamida/administración & dosificación , Supervivencia sin Enfermedad , Epirrubicina/administración & dosificación , Femenino , Fluorouracilo/administración & dosificación , Humanos , Inmunohistoquímica , Antígeno Ki-67/biosíntesis , Metotrexato/administración & dosificación , Persona de Mediana Edad , Receptor ErbB-2/biosíntesis , Receptores de Estrógenos/biosíntesis , Receptores de Progesterona/biosíntesisRESUMEN
OBJECTIVE: As hormone receptor and human epidermal growth factor receptor-2 (HER-2) expression in primary breast tumors frequently differs from that of paired metastases, we first evaluated the discordance rate (DR) of estrogen receptor (ER), progesterone receptor (PgR), HER-2, and Ki67 in breast cancer patients and then assessed the discordance effect on prognosis. METHODS: Of 145 cases reviewed, 120 with samples available from both primary tumors and paired metastases were included in the study. For each receptor, the DR was calculated as the proportion of discordant cases with respect to the total number of patients. RESULTS: A change in ER status was observed in 19 cases (DR 16.4%), while PgR status was modified in 48 cases (DR 41.7%). HER-2 was altered in 21 cases (DR 17.5%), and Ki67 in 33 patients (DR 38.8%). Patients with Ki67 <20% had a significantly higher postrecurrence survival (PRS) compared to patients with Ki67 ≥20% (p = 0.0006). Patients with ER-positive tumors showed a trend towards higher PRS (p = 0.064) compared to ER-negative patients. No differences in PRS were seen among patients with altered PgR or HER2 status. CONCLUSIONS: Changes in the cell biology of breast cancer metastasis would seem to occur and biopsy could potentially guide the choice of treatment and provide useful information on prognosis.
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Neoplasias de la Mama/patología , Antígeno Ki-67/metabolismo , Recurrencia Local de Neoplasia/diagnóstico , Receptor ErbB-2/metabolismo , Receptores de Estrógenos/metabolismo , Receptores de Progesterona/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/metabolismo , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/mortalidad , Carcinoma Ductal de Mama/metabolismo , Carcinoma Ductal de Mama/mortalidad , Carcinoma Ductal de Mama/secundario , Carcinoma Lobular/metabolismo , Carcinoma Lobular/mortalidad , Carcinoma Lobular/secundario , Femenino , Estudios de Seguimiento , Humanos , Técnicas para Inmunoenzimas , Incidencia , Persona de Mediana Edad , Metástasis de la Neoplasia , Recurrencia Local de Neoplasia/metabolismo , Recurrencia Local de Neoplasia/mortalidad , Estadificación de Neoplasias , Pronóstico , Estudios Retrospectivos , Tasa de SupervivenciaRESUMEN
BACKGROUND: Despite numerous studies aimed at verifying the antitumor activity of nitric oxide-releasing nonsteroidal antiflammatory drugs (NO-NSAIDs), little is known about the molecular targets responsible for their antineoplastic properties. In the present study, we investigated the mechanisms underlying the cytotoxicity of NCX 4040, a novel NO-aspirin with promising antineoplastic action, in in vitro human colon cancer models. METHODS: The effect on tumor growth was evaluated in four human colon cancer cell lines (LoVo, LRWZ, WiDr and LoVo Dx) by sulforhodamine B assay, oxidative stress by immunohistochemistry, apoptosis by laddering assay, mitochondrial membrane potential (DeltaPsim) by flow cytometry, and apoptosis- and chemoresistance-related markers by western-blot and real-time method, respectively. Prostaglandin E2 levels were determined by ELISA. RESULTS: NCX 4040 produced a higher cytotoxic effect in all the cell lines than that produced by other NO donors tested. In particular, in LoVo and LRWZ cells, NCX 4040 induced a cytocidal effect and apoptosis through p53 and NAG-1 expression, an early DeltaPsim collapse, and a sequential release of cytoplasmatic cytochrome c and caspase -9 and -3 active forms. 8-hydroxyguanine lesions, indicative of oxidative stress, were also observed. Conversely, in WiDr line, the drug caused a cytocidal effect, albeit not through apoptosis, and a concomitant increase in COX-2 activity. In LoVo Dx line, characterized by high levels drug resistance and DNA repair-related markers, only a cytostatic effect was observed, again in concomitance with the increase in COX-2 enzyme activity. CONCLUSION: This study highlights the multiplicity of mechanisms involved in sensitivity or resistance to NCX 4040 and could provide useful indications for tailored therapy by identifying potentially drug-responsive tumors.
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Apoptosis/efectos de los fármacos , Aspirina/análogos & derivados , Neoplasias del Colon/patología , Nitrocompuestos/farmacología , Antiinflamatorios no Esteroideos/farmacología , Aspirina/química , Aspirina/farmacología , Línea Celular Tumoral , Humanos , Potenciales de la Membrana/efectos de los fármacos , Membranas Mitocondriales/efectos de los fármacos , Membranas Mitocondriales/fisiología , Conformación Molecular , Óxido Nítrico/metabolismo , Nitrocompuestos/química , Nitroprusiato/farmacologíaRESUMEN
We previously showed that NCX 4040 inhibits in vitro and in vivo tumor growth and induces apoptosis in human colon cancer cell lines. On the basis of these results, NCX 4040 antitumor activity in combination with 5-fluorouracil (5-FU) or oxaliplatin was evaluated in vitro and in vivo in human colon cancer models. The cytotoxicity of different NCX 4040 and 5-FU or oxaliplatin combination schemes was evaluated on a panel of colon cancer lines (LoVo, LoVo Dx, WiDr, and LRWZ) by the sulforhodamine B assay, and apoptosis was assessed by flow cytometry. NCX 4040 and 5-FU combination was always additive in vitro regardless of the scheme used. Sequential NCX 4040-->oxaliplatin treatment produced a strong synergism in three cell lines, with a ratio index ranging from 3.7 to 4. The synergistic effect was accompanied by apoptosis induction (up to 40%). In the in vivo experiments, xenografted mice were treated with the sequential combination of NCX 4040 and oxaliplatin, and apoptosis was evaluated immunohistochemically in excised tumors. Furthermore, in WiDr xenografts, this sequence caused a significantly higher reduction ( approximately 60%) in tumor growth compared with single-drug treatments and produced extensive apoptotic cell death (15.3%), significantly higher (P < 0.01) than that observed in untreated tumors (2.7%) or in tumors treated with NCX 4040 (5.1%) or oxaliplatin (5.7%) alone. These data show that NCX 4040 sensitizes colon cancer cell lines to the effect of antitumor drugs and suggests that their combination could be useful for the clinical management of colon cancer.
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Antiinflamatorios no Esteroideos/farmacología , Antineoplásicos/farmacología , Aspirina/análogos & derivados , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/patología , Óxido Nítrico/metabolismo , Nitrocompuestos/farmacología , Animales , Antiinflamatorios no Esteroideos/uso terapéutico , Antineoplásicos/uso terapéutico , Aspirina/farmacología , Supervivencia Celular/efectos de los fármacos , Citometría de Flujo , Fluorouracilo/farmacología , Fluorouracilo/uso terapéutico , Humanos , Masculino , Ratones , Ratones Desnudos , Compuestos Organoplatinos/farmacología , Compuestos Organoplatinos/uso terapéutico , Oxaliplatino , Trasplante HeterólogoRESUMEN
BACKGROUND: Epidermal growth factor receptor (EGFR) and v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) mutations, and echinoderm microtubule-associated protein-like 4 (EML4) anaplastic lymphoma kinase (ALK) translocation are generally considered to be mutually exclusive. However, concomitant mutations are found in a small number of patients and the effect of these on response to targeted therapy is still unknown. PATIENTS AND METHODS: We considered 380 non-small-cell lung cancer (NSCLC) patients who underwent nonsequential testing for EGFR and EML4-ALK translocation. KRAS mutation analysis was also performed on 282 patients. RESULTS: We found 1.6%, 1.1%, and 2.5% of patients who showed a double mutation comprising EGFR and EML4-ALK, EGFR and KRAS, and EML4-ALK and KRAS, respectively. Twenty-eight patients with EGFR mutation underwent first-line therapy with a tyrosine kinase receptor; a clinical benefit was observed in 81.8% of patients with EGFR mutations only and in 67% of those who also showed an EML4-ALK translocation. Twelve patients with an EML4-ALK translocation received crizotinib and 7 of these had disease progression within 3 months (2 had a concomitant KRAS mutation and 1 had a concomitant EGFR mutation). Two patients showed stable disease, 1 of whom also had a KRAS mutation. Two patients obtained a partial response and 1 had a complete response; all harbored an EML4-ALK translocation only. The median overall survival of patients who carried an EML4-ALK translocation alone or concomitant with a KRAS mutation was 57.1 (range, 10.7-not reached) and 10.7 (range, 4.6-not reached) months, respectively. CONCLUSION: Concomitant EGFR, EML4-ALK, or KRAS mutations can occur in NSCLC. Concomitant KRAS mutation and EML4-ALK translocation represents the most common double alteration and confers a poor prognosis.
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Carcinoma de Pulmón de Células no Pequeñas/genética , Receptores ErbB/genética , Neoplasias Pulmonares/genética , Proteínas de Fusión Oncogénica/genética , Proteínas Proto-Oncogénicas p21(ras)/genética , Adulto , Anciano , Anciano de 80 o más Años , Antineoplásicos/uso terapéutico , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/patología , Crizotinib , Progresión de la Enfermedad , Femenino , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/patología , Masculino , Persona de Mediana Edad , Terapia Molecular Dirigida , Mutación , Pronóstico , Pirazoles/uso terapéutico , Piridinas/uso terapéutico , Estudios Retrospectivos , Tasa de Supervivencia , Resultado del TratamientoRESUMEN
OBJECTIVE: Recent years have seen a considerable wealth of studies conducted on the potential usefulness of telomerase determination in diagnosis, prognosis and targeted cancer therapy. The frequently used Telomeric Repeat Amplification Protocol assay suffers from some drawbacks, the most important being the rate of false positives. In situ analysis using well characterised antibodies directed against the human telomerase reverse transcriptase (hTERT) would therefore appear to be important to morphologically identify the nature of telomerase positive cells. METHODS: We performed immunostaining in a series of cultured cells and in normal, preneoplastic and tumour tissues from different organs using a monoclonal antibody directed against the catalytic subunit of telomerase. RESULTS: Immunoreactivity was not observed in perennial cells of terminally differentiated cardiac and skeletal muscular tissues or in small pyramidal cells of the cerebral cortex. Conversely, it was found in other normal somatic tissues as well as in precancerous lesions and in all tumour histotypes. CONCLUSIONS: Immunohistochemistry with a well characterised hTERT-specific monoclonal antibody permitted the identification of hTERT immunopositive cells in normal somatic tissues. Whether hTERT protein detected by immunostaining with hTERT-specific Tel 3 36-10 antibody is actually the degraded form of the protein that retains hTERT antigenicity but not enzymatic function, or whether it represents the real, potentially functional catalytic subunit of the enzyme, immunohistochemistry would not seem to represent a useful tool to investigate the role of telomerase and the mechanisms involved in its regulation.
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Núcleo Celular/metabolismo , Proteínas de Unión al ADN/biosíntesis , Proteínas de Unión al ADN/genética , Regulación Neoplásica de la Expresión Génica , Técnicas Inmunológicas , Neoplasias/metabolismo , Telomerasa/metabolismo , Encéfalo/patología , Dominio Catalítico , Línea Celular Tumoral , ADN Complementario/metabolismo , Fibroblastos/metabolismo , Humanos , Inmunohistoquímica , Pronóstico , Estructura Terciaria de Proteína , ARN Mensajero/metabolismo , ARN Neoplásico/metabolismo , Telomerasa/biosíntesis , Telomerasa/genética , Rayos Ultravioleta , Neoplasias de la Vejiga Urinaria/metabolismoRESUMEN
Telomerase activity plays an important role in the two complementary processes of cellular immortalization and senescence. This enzyme is active in almost all tumors, but also in inflammatory and many normal proliferating cells. Therefore, the main limits of molecular determinations, such as telomeric repeat amplification protocol assay is that they are not able to discriminate between the enzymatic activity of tumor and normal cells. The most appropriate technique for this would be immunohistochemical determination using monoclonal antibodies. Very few monoclonal antibodies (Mabs) directed against the human telomerase reverse transcriptase (hTERT) are commercially available and in the present study, we developed a new Mab directed against this protein (TERT-3 36-10) to investigate the possibility of detecting immunoreactivity to this Mab by immunohistochemical and flow cytometric approaches. Immunohistochemical determination showed a lack of reactivity to the Mab in highly differentiated striated muscle tissue, a variable reactivity in dysplastic cervical epithelial tissue and similar and widespread immunoreactivity in cell lines and clinical tumors. Furthermore, we demonstrated the ability of this Mab to inhibit enzyme activity in cell extract from MCR bladder tumor cell line.
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Anticuerpos Monoclonales/inmunología , Proteínas de Unión al ADN/inmunología , Telomerasa/inmunología , Animales , Anticuerpos Monoclonales/aislamiento & purificación , Anticuerpos Monoclonales/farmacología , Células COS , Proliferación Celular , Senescencia Celular/fisiología , Chlorocebus aethiops , Proteínas de Unión al ADN/antagonistas & inhibidores , Proteínas de Unión al ADN/genética , Femenino , Células HL-60 , Humanos , Hibridomas , Ratones , Ratones Endogámicos BALB C , Músculo Esquelético/citología , Músculo Esquelético/enzimología , Neoplasias/enzimología , Telomerasa/antagonistas & inhibidores , Telomerasa/genéticaRESUMEN
AIMS AND BACKGROUND: The aim of this retrospective multicenter study was to evaluate the impact of progesterone receptor (PgR) loss on locoregional recurrence in patients with estrogen receptor (ER)-positive primary breast cancer and ER-positive locoregional recurrence. PATIENTS AND METHODS: Eight Italian oncology centers collected data from consecutive patients with ER-positive breast cancer and a subsequent ER-positive locoregional recurrence. RESULTS: Data were available for 265 patients diagnosed with breast cancer between 1990 and 2009. Median metastasis-free survival was 111 months in patients with PgR-positive primary tumors and locoregional recurrence (PgRpos), 38 months in patients with PgR-negative primary tumors and locoregional recurrence (PgRneg), and 63 months in patients with PgR-positive primary tumors and PgR-negative locoregional recurrence (PgRloss). In multivariate analysis, PgR status was independently associated with metastasis-free survival, with a hazard ratio of 2.84 (95% CI 1.34-6.00) for PgRneg compared with PgRpos, and 2.93 (95% CI: 1.51-5.70) for PgRloss compared with PgRpos. CONCLUSIONS: PgR absence was found to be a negative prognostic factor in breast cancer patients with ER-positive locoregional recurrence. Thus, PgR status could be a biological marker in ER-positive recurrent breast cancer.
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Biomarcadores de Tumor/análisis , Neoplasias de la Mama/química , Neoplasias de la Mama/patología , Recurrencia Local de Neoplasia/química , Recurrencia Local de Neoplasia/patología , Receptores de Estrógenos/análisis , Receptores de Progesterona/análisis , Adulto , Anciano , Anciano de 80 o más Años , Proliferación Celular , Supervivencia sin Enfermedad , Femenino , Humanos , Italia , Metástasis Linfática , Persona de Mediana Edad , Clasificación del Tumor , Estadificación de Neoplasias , Oportunidad Relativa , Valor Predictivo de las Pruebas , Receptor ErbB-2/análisis , Estudios Retrospectivos , Factores de TiempoRESUMEN
Telomerase activity is present in most human malignant tumors, whereas it is generally not detectable, with some exceptions, in normal cells. Therefore, it represents a potential tool for tumor detection. In the present study, telomerase activity was determined in urine from 79 healthy individuals and 121 previously untreated bladder cancer patients using a highly sensitive telomeric repeat amplification protocol (TRAP) assay and the results were expressed as arbitrary enzymatic units (AEU). This approach enabled us to identify cutoff values characterized by high sensitivity (from 75% to 93%) and specificity (from 72% to 92%). Moreover, analysis as a function of gender showed a higher accuracy of TRAP assay in males (93% sensitivity and 90% specificity at the cutoff of 50 AEU) than in females. This sensitivity was confirmed in patients with nonassessable or negative cytology. In women, morphological and immunocytochemical determinations using a monoclonal antibody (anti-hTERT) recently developed in our laboratory showed a large fraction of immunoreactive inflammatory or nonbladder cells, which may justify the false-positive TRAP results. In conclusion, this assay represents an important noninvasive diagnostic tool to detect bladder cancer also in patients with negative or nonassessable urine cytology and with low-grade and early-stage lesions.
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Biomarcadores de Tumor/orina , Telomerasa/orina , Neoplasias de la Vejiga Urinaria/diagnóstico , Neoplasias de la Vejiga Urinaria/orina , Proteínas de Unión al ADN , Femenino , Humanos , Masculino , Sensibilidad y Especificidad , Telomerasa/metabolismo , Neoplasias de la Vejiga Urinaria/enzimología , Neoplasias de la Vejiga Urinaria/patologíaRESUMEN
INTRODUCTION: The aim of the present study was to analyze the relationship between the expression of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) in breast cancer cells and the corresponding serum levels in individual patients. The study also evaluated the potential of serum levels of the two growth factors as diagnostic markers in a case-control study. METHODS: VEGF expression and bFGF expression were determined in 62 and 63 tumor samples, respectively. Serum VEGF and bFGF levels were determined in 54 and 65 healthy women and in 69 and 73 breast cancer patients, respectively, using a quantitative sandwich enzyme immunoassay technique. RESULTS: A direct correlation was observed between VEGF expression and bFGF expression in individual tumors (P = 0.001) and between serum levels (P = 0.038) in individual patients, but not between tumor cell expression and the corresponding serum level for either growth factor. Median values of serum levels in healthy women and breast cancer patients were not different for VEGF (P = 0.055), but were significantly different for bFGF (P < 0.001). The receiver operating characteristic curve identified a serum bFGF concentration of 1.0 pg/ml, with 84.9% sensitivity and 63.1% specificity, as the best cut-off value to discriminate between healthy women and breast cancer patients. An age-based subgroup analysis showed that serum values of patients older than 70 years of age mainly contributed to the high accuracy. CONCLUSIONS: Our data repropose bFGF as a noninvasive diagnostic tool for breast cancer.
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Biomarcadores de Tumor/sangre , Neoplasias de la Mama/diagnóstico , Factor 2 de Crecimiento de Fibroblastos/sangre , Factor A de Crecimiento Endotelial Vascular/sangre , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/biosíntesis , Neoplasias de la Mama/sangre , Neoplasias de la Mama/metabolismo , Estudios de Casos y Controles , Femenino , Factor 2 de Crecimiento de Fibroblastos/biosíntesis , Humanos , Técnicas para Inmunoenzimas/métodos , Inmunohistoquímica , Persona de Mediana Edad , Sensibilidad y Especificidad , Factor A de Crecimiento Endotelial Vascular/biosíntesisRESUMEN
Nitric oxide-releasing non-steroidal antiinflammatory drugs (NO-NSAIDs) are safer than NSAIDs due to their ability to reduce gastric toxicity. We assessed the cytotoxic activity of a new aspirin derivative, NCX 4016, after different exposure schedules, in three human colon adenocarcinoma cell lines. All the lines were positive for COX-1 protein and mRNA, as evaluated by Western blot and RT-PCR, respectively, while only one was positive for COX-2. The cytostatic and cytocidal activity was determined by sulforhodamine B assay and evaluated according to Monks' model. Cytostatic activity was observed after a 24-h drug exposure and 50% growth inhibition was reached at concentrations ranging from 165 to 250 micro M in all cell lines, whereas with aspirin the IC50 was never reached, even at the maximum concentration tested (500 micro M), and was independent of COX-1 or COX-2 status. Cytocidal activity was observed only at the highest concentrations and persisted for a long time after drug removal. Flow cytometric analysis showed that the NO-aspirin compound induced a persistent accumulation of cells in G2-M phase in all the cell lines after at least 48 h exposure. Specifically, the block pertained mainly to G2 phase, whereas mitotic index was not affected at all. Our results indicate that NCX 4016 has an in vitro cytostatic activity superior to that of its parental aspirin compound, which makes it a potentially important tumor preventive agent. Furthermore, the cytocidal effect observed at the highest concentrations and the induction of a specific block in G2 phase renders it a promising candidate for drug combination treatments.
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Anticarcinógenos/farmacología , Aspirina/análogos & derivados , Aspirina/farmacología , Ciclo Celular/efectos de los fármacos , División Celular/efectos de los fármacos , Inhibidores de Agregación Plaquetaria/farmacología , Adenocarcinoma/patología , Supervivencia Celular/efectos de los fármacos , Neoplasias del Colon/patología , Ciclooxigenasa 1 , Ciclooxigenasa 2 , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Isoenzimas/genética , Isoenzimas/metabolismo , Cinética , Proteínas de la Membrana , Prostaglandina-Endoperóxido Sintasas/genética , Prostaglandina-Endoperóxido Sintasas/metabolismo , Células Tumorales CultivadasRESUMEN
BACKGROUND: Although two-dimensional (2-D) monolayer cell cultures provide important information on basic tumor biology and radiobiology, they are not representative of the complexity of three-dimensional (3-D) solid tumors. In particular, new models reproducing clinical conditions as closely as possible are needed for radiobiological studies to provide information that can be translated from bench to bedside. METHODS: We developed a novel system for the irradiation, under sterile conditions, of 3-D tumor spheroids, the in vitro model considered as a bridge between the complex architectural organization of in vivo tumors and the very simple one of in vitro monolayer cell cultures. The system exploits the same equipment as that used for patient treatments, without the need for dedicated and highly expensive instruments. To mimic the passage of radiation beams through human tissues before they reach the target tumor mass, 96-multiwell plates containing the multicellular tumor spheroids (MCTS) are inserted into a custom-built phantom made of plexiglass, the material most similar to water, the main component of human tissue. RESULTS: The system was used to irradiate CAEP- and A549-derived MCTS, pre-treated or not with 20 µM cisplatin, with a dose of 20 Gy delivered in one session. We also tested the same treatment schemes on monolayer CAEP and A549 cells. Our preliminary results indicated a significant increment in radiotoxicity 20 days after the end of irradiation in the CAEP spheroids pre-treated with cisplatin compared to those treated with cisplatin or irradiation alone. Conversely, the effect of the radio- chemotherapy combination in A549-derived MCTS was similar to that induced by cisplatin or irradiation alone. Finally, the 20 Gy dose did not affect cell survival in monolayer CAEP and A549 cells, whereas cisplatin or cisplatin plus radiation caused 100% cell death, regardless of the type of cell line used. CONCLUSIONS: We set up a system for the irradiation, under sterile conditions, of tumor cells grown in 3-D which allows for the use of the same dose intensities and schedules utilized in clinical practice. This irradiation system, coupled with 3-D cell cultures, has the potential to generate information that could be used to individually tailor radiotherapy.
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Técnicas de Cultivo de Célula/métodos , Radioterapia/métodos , Esferoides Celulares/patología , Esferoides Celulares/efectos de la radiación , Animales , Reactores Biológicos , Línea Celular Tumoral , Supervivencia Celular , Células Cultivadas , Cisplatino/administración & dosificación , Fraccionamiento de la Dosis de Radiación , Humanos , Inmunohistoquímica , Modelos Lineales , Microscopía Electrónica de Transmisión , Radiobiología/métodos , Radiometría/métodos , Proyectos de Investigación , Células Tumorales CultivadasRESUMEN
PURPOSE: The increasing use of breast-conserving surgery makes it essential to identify biofunctional profiles responsible for the progression of in situ to invasive carcinomas to facilitate the detection of lesions that are most likely to relapse or progress and, thus, to be able to offer patients tailored treatment options. Our objective was to analyse and compare biofunctional profiles in ductal carcinomas in situ (DCIS) and invasive ductal carcinomas (IDC). We also aimed to identify markers in tumor and normal surrounding tissues that may be predictive of locoregional recurrence in patients with DCIS. METHODS: Biofunctional parameters including mitotic activity, estrogen receptor, progesterone receptor, microvessel density (MVD), c-kit and p27 expression were evaluated in 829 in situ and invasive carcinomas. The impact of the biomarker profiles of DCIS, IDC and normal surrounding tissues on loco-regional recurrence was analyzed. RESULTS: A progressive increase in cell proliferation and a concomitant decrease in steroid hormone receptor-positive lesions was observed during the transition from in situ to invasive carcinomas, as also within each subgroup as grade increased. Conversely, p27 expression and MVD dramatically decreased during the transition from in situ to invasive carcinomas. Finally, we found that a low c-kit expression was indicative of IDC relapse. CONCLUSIONS: Cell proliferation, hormonal and differentiation characteristics differed in DCIS with respect to IDC, and the main variation in the transition between the two histologic lesions was the decrease in p27 expression and MVD.
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Biomarcadores de Tumor/metabolismo , Neoplasias de la Mama/metabolismo , Carcinoma Ductal de Mama/metabolismo , Carcinoma Intraductal no Infiltrante/metabolismo , Neoplasias de la Mama/patología , Neoplasias de la Mama/terapia , Carcinoma Ductal de Mama/patología , Carcinoma Ductal de Mama/terapia , Carcinoma Intraductal no Infiltrante/patología , Carcinoma Intraductal no Infiltrante/terapia , Línea Celular Tumoral , Proliferación Celular , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/metabolismo , Progresión de la Enfermedad , Femenino , Humanos , Recurrencia Local de Neoplasia , Neovascularización Patológica/patología , Evaluación de Resultado en la Atención de Salud/estadística & datos numéricos , Pronóstico , Modelos de Riesgos Proporcionales , Proteínas Proto-Oncogénicas c-kit/metabolismo , Receptores de Estrógenos/metabolismo , Receptores de Progesterona/metabolismoRESUMEN
A significant proportion of breast cancers with HER2 amplification show simultaneous amplification or deletion of Topo 2. Amplification of Topo 2 may lead to the overexpression of the Topo 2 protein and ultimately to hypersensitivity to Topo 2 inhibitors. HER2 and Topo 2 gene status in breast cancer patients has been determined in several studies using immunohistochemistry, florescence in situ hybridisation (FISH) and multiplex ligation-dependent probe amplification (MLPA). Although comparisons of FISH and MLPA have been reported for HER2, it is believed that there are no similar studies for Topo 2. In this study, HER2 and Topo 2 were analysed by MLPA and FISH. There was a high agreement between the two approaches, although MLPA was easier to perform and cheaper than FISH. In conclusion, MLPA is a fast and accurate quantitative method to detect HER2 and Topo 2 amplification, and could be considered a good alternative to FISH.
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Neoplasias de la Mama/genética , ADN-Topoisomerasas de Tipo II/genética , Amplificación de Genes , Genes erbB-2 , Hibridación Fluorescente in Situ , Técnicas de Amplificación de Ácido Nucleico , Femenino , HumanosRESUMEN
Deregulation of signal transduction pathways frequently confers selective biological advantages to tumors. Phosphoinositides play an essential role in numerous cellular functions and, among the enzymes implicated in these processes, phosphoinositide-specific phospholipase C ß1 (PI-PLCß1) is one of the key regulators. In the present study, a fluorescence in situ hybridization (FISH) approach was used to investigate PI-PLCß1 gene copy number alterations in various types of breast cancer differing in their invasiveness and proliferative activity, according to their mitotic index. At the molecular level, we also performed both real-time PCR and immunohistochemical analyses on PI-PLCß1 to further investigate its expression in primary breast cancers. Finally, we analyzed the correlation between PI-PLCß1 gene copy number and clinicopathological parameters. Our results show that most of our cases had aneusomies on the PI-PLCß1 locus (20p12) and amplification of this specific region was the most frequent alteration observed. Our findings also indicate that the amplification of the region containing the PI-PLCß1 gene was mostly related to the mitotic index, rather than to the invasion status. Finally, even though our case series is limited, PI-PLCß1 gene amplification seems to be correlated to clinicopathological parameters.
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Neoplasias de la Mama/genética , Dosificación de Gen , Fosfolipasa C beta/genética , Adulto , Anciano , Anciano de 80 o más Años , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Persona de Mediana Edad , Clasificación del Tumor , Fenotipo , ARN Mensajero/análisisRESUMEN
Oligodendrogliomas are rare primary brain tumors with variable patient outcomes which are not always adequately accounted for by clinical or pathological variables. The present study evaluated the prognostic implications of chromosome 1p and 19q status in a set of 23 low grade oligodendrogliomas (OGD II), and correlated the results with patient outcome. Loss of heterozygosity (LOH) and fluorescent in situ hybridization (FISH) analyses, the most widely used standard procedures, were used. 1p and 19q deletions were found in 65 and 61% of cases, respectively, using FISH and in 78 and 72% of cases using LOH. Both deletions were found in 56 and 64% of patients using FISH and LOH, respectively. Concordance between the results from the two techniques, determined by the Kappa statistics, ranged from fair to substantial depending on whether single or combined deletions were considered. Our results showed that the molecular alterations are associated with age and tumor localization. With regard to the impact of chromosomal alterations on clinical outcome, chromosome 19q deletions detected by LOH would seem to indicate a subgroup of patients at a higher risk of relapse, although the small number of patients recruited does not permit any definitive conclusions to be drawn. Further studies are now ongoing to determine whether this methodological approach could be potentially useful in low grade oligodendrogliomas to better characterize chromosomal alterations of 1p/19q and identify subgroups of patients with a higher risk of disease recurrence.