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To decipher the interactions between various components of the tumor microenvironment (TME) and tumor cells in a preserved spatial context, a multiparametric approach is essential. In this pursuit, imaging mass cytometry (IMC) emerges as a valuable tool, capable of concurrently analyzing up to 40 parameters at subcellular resolution. In this study, a set of antibodies was selected to spatially resolve multiple cell types and TME elements, including a comprehensive panel targeted at dissecting the heterogeneity of cancer-associated fibroblasts (CAF), a pivotal TME component. This antibody panel was standardized and optimized using formalin-fixed paraffin-embedded tissue (FFPE) samples from different organs/lesions known to express the markers of interest. The final composition of the antibody panel was determined based on the performance of conjugated antibodies in both immunohistochemistry (IHC) and IMC. Tissue images were segmented employing the Steinbock framework. Unsupervised clustering of single-cell data was carried out using a bioinformatics pipeline developed in R program. This paper provides a detailed description of the staining procedure and analysis workflow. Subsequently, the panel underwent validation on clinical FFPE samples from head and neck squamous cell carcinoma (HNSCC). The panel and bioinformatics pipeline established here proved to be robust in characterizing different TME components of HNSCC while maintaining a high degree of spatial detail. The platform we describe shows promise for understanding the clinical implications of TMA heterogeneity in large patient cohorts with FFPE tissues available in diagnostic biobanks worldwide.
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Breast cancer (BC) patients aged <40 years at diagnosis experience aggressive disease and poorer survival compared with women diagnosed with BC at 40 to 49 years, but the age-related biology is described to little extent. Here, we explored transcriptional alterations in BC to gain better understanding of age-related tumor biology. We studied a subset of the Bergen in-house cohort (n = 127; age range, 26-49 years) and used the NanoString Breast Cancer 360 expression panel on formalin-fixed paraffin-embedded BC tissue, and publicly available global BC messenger RNA expression data (n = 204; age range, 22-49 years), to explore differentially expressed genes between the young (age <40 years) and older (age 40-49 years) patients. Unsupervised hierarchical clustering was applied to identify gene expression-based patient clusters. We applied established computational approaches to define the PAM50 subtypes, risk of recurrence scores (ROR), and risk groups and to infer the proportions of 22 immune cell types from bulk gene expression profiles of patients aged <50 years at BC diagnosis. Differentially expressed genes and gene sets were investigated using OncoEnrichR and g:Profiler to describe functional profiles and pathway enrichment. We identified 4 age-related patient clusters presenting distinct characteristics of PAM50 subtypes and ROR profiles, which demonstrated independent prognostic value when adjusted for traditional clinicopathologic variables and the known molecular subtypes. Our findings showed better survival than expected in the basal-enriched cluster 2 and in triple-negative and basal-like BC. Deconvolution analyses of immunophenotypes indicated higher levels of M0 and M1 macrophages than M2 macrophages in subsets of young BC. Our approach identifies age-based patient clusters with distinct clinicopathologic profiles, to a large extent overlapping with the PAM50 subtypes, although with independent prognostic values in multivariate survival analyses. The patient clusters provided new insight in the immune cell distribution across tumor subtypes, potentially contributing to survival differences between the clusters and the molecular subtypes and indicating age-related mechanisms improving outcome. Our study confirms the applicability of ROR as a valid prognosticator also in a young BC cohort.
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Neoplasias de la Mama , Fenotipo , Humanos , Femenino , Neoplasias de la Mama/inmunología , Neoplasias de la Mama/patología , Neoplasias de la Mama/genética , Neoplasias de la Mama/mortalidad , Adulto , Persona de Mediana Edad , Factores de Edad , Adulto Joven , Biomarcadores de Tumor/genética , Perfilación de la Expresión Génica , Pronóstico , Transcriptoma , Análisis por ConglomeradosRESUMEN
The prognosis of high-grade serous ovarian carcinoma (HGSOC) is poor, and treatment selection is challenging. A heterogeneous tumor microenvironment (TME) characterizes HGSOC and influences tumor growth, progression, and therapy response. Better characterization with multidimensional approaches for simultaneous identification and categorization of the various cell populations is needed to map the TME complexity. While mass cytometry allows the simultaneous detection of around 40 proteins, the CyTOFmerge MATLAB algorithm integrates data sets and extends the phenotyping. This pilot study explored the potential of combining two datasets for improved TME phenotyping by profiling single-cell suspensions from ten chemo-naïve HGSOC tumors by mass cytometry. A 35-marker pan-tumor dataset and a 34-marker pan-immune dataset were analyzed separately and combined with the CyTOFmerge, merging 18 shared markers. While the merged analysis confirmed heterogeneity across patients, it also identified a main tumor cell subset, additionally to the nine identified by the pan-tumor panel. Furthermore, the expression of traditional immune cell markers on tumor and stromal cells was revealed, as were marker combinations that have rarely been examined on individual cells. This study demonstrates the potential of merging mass cytometry data to generate new hypotheses on tumor biology and predictive biomarker research in HGSOC that could improve treatment effectiveness.
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Metastatic castration-resistant prostate cancer (mCRPC) is an immunologically cold disease with dismal outcomes. Cryoablation destroys cancer tissue, releases tumor-associated antigens and creates a pro-inflammatory microenvironment, while dendritic cells (DCs) activate immune responses through processing of antigens. Immunotherapy combinations could enhance the anti-tumor efficacy. This open-label, single-arm, single-center phase I trial determined the safety and tolerability of combining cryoablation and autologous immature DC, without and with checkpoint inhibitors. Immune responses and clinical outcomes were evaluated. Patients with mCRPC, confirmed metastases and intact prostate gland were included. The first participants underwent prostate cryoablation with intratumoral injection of autologous DCs in a 3 + 3 design. In the second part, patients received cryoablation, the highest acceptable DC dose, and checkpoint inhibition with either ipilimumab or pembrolizumab. Sequentially collected information on adverse events, quality of life, blood values and images were analyzed by standard descriptive statistics. Neither dose-limiting toxicities nor adverse events > grade 3 were observed in the 18 participants. Results indicate antitumor activity through altered T cell receptor repertoires, and 33% durable (> 46 weeks) clinical benefit with median 40.7 months overall survival. Post-treatment pain and fatigue were associated with circulating tumor cell (CTC) presence at inclusion, while CTC responses correlated with clinical outcomes. This trial demonstrates that cryoimmunotherapy in mCRPC is safe and well tolerated, also for the highest DC dose (2.0 × 108) combined with checkpoint inhibitors. Further studies focusing on the biologic indications of antitumor activity and immune system activation could be considered through a phase II trial focusing on treatment responses and immunologic biomarkers.
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Neoplasias de la Próstata Resistentes a la Castración , Humanos , Masculino , Células Dendríticas , Ipilimumab/uso terapéutico , Estudios Prospectivos , Neoplasias de la Próstata Resistentes a la Castración/terapia , Calidad de Vida , Microambiente TumoralRESUMEN
PURPOSE: Predictive biomarkers are needed to aid the individualization of radiotherapy (RT) in breast cancer. Cancer-associated fibroblasts have been implicated in tumor radioresistance and can be identified by platelet-derived growth factor receptor-beta (PDGFRb). This study aims to analyze how PDGFRb expression affects RT benefit in a large randomized RT trial. METHODS: PDGFRb was assessed by immunohistochemistry on tissue microarrays from 989 tumors of the SweBCG91RT trial, which enrolled lymph node-negative, stage I/IIA breast cancer patients randomized to RT after breast-conserving surgery. Outcomes were analyzed at 10 years for ipsilateral breast tumor recurrence (IBTR) and any recurrence and 15 years for breast cancer specific death (BCSD). RESULTS: PDGFRb expression correlated with estrogen receptor negativity and younger age. An increased risk for any recurrence was noted in univariable analysis for the medium (HR 1.58, CI 95% 1.11-2.23, p = 0.011) or PDGFRb high group (1.49, 1.06-2.10, p = 0.021) compared to the low group. No differences in IBTR or BCSD risk were detected. RT benefit regarding IBTR risk was significant in the PDGFRb low (0.29, 0.12-0.67, p = 0.004) and medium (0.31, 0.16-0.59, p < 0.001) groups but not the PDGFRb high group (0.64, 0.36-1.11, p = 0.110) in multivariable analysis. Likewise, risk reduction for any recurrence was less pronounced in the PDGFRb high group. No significant interaction between RT and PDGFRb-score could be detected. CONCLUSION: A higher PDGFRb-score conferred an increased risk of any recurrence, which partly can be explained by its association with estrogen receptor negativity and young age. Reduced RT benefit was noted among patients with high PDGFRb, however without significant interaction.
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Neoplasias de la Mama , Neoplasias de la Mama/cirugía , Neoplasias de la Mama/terapia , Femenino , Humanos , Inmunohistoquímica , Mastectomía Segmentaria , Recurrencia Local de Neoplasia , Pronóstico , Receptor beta de Factor de Crecimiento Derivado de Plaquetas/genéticaRESUMEN
BACKGROUND: Preoperative predictors for the need of prophylactic lymph node dissection in the lateral neck have been studied in patients with medullary thyroid carcinoma (MTC). OBJECTIVES: To evaluate the ability of serum calcitonin to predict the extent of surgery needed in the lateral neck. METHODS: This retrospective population-based cohort study includes data from 94 of 139 patients with MTC surgically treated in Norway from 2003 to 2016. Patients were identified in the 4 regional centers treating MTC and by the Cancer Registry of Norway, and grouped according to calcitonin levels. In 58 patients without distant metastases or disease progression to the next tumor level (NPNL), data were compared in prognostic groups (N0-NPNL), (N1a-NPNL), and (N1b-NPNL). RESULTS: At calcitonin levels ≤500, 501-1,000, and >1,000 pmol/L, metastatic lymph nodes in the lateral neck were found in 16, 50, and 71% of the patients, respectively. In the prognostic groups, 19% of N0-NPNL patients had calcitonin >500 pmol/L and 17% of N1b-NPNL patients had calcitonin ≤500 pmol/L. In multivariate analysis, factors predicting biochemical cure and calcitonin level ≤500 pmol/L were no metastatic lymph nodes in the lateral neck (p = 0.030) and tumor diameter ≤20 mm (p < 0.001), respectively. Factors related to metastatic lymph nodes in the lateral neck were extrathyroidal extension (p = 0.007) and no biochemical cure (p = 0.028). CONCLUSIONS: Basal calcitonin cannot predict the need for prophylactic lateral lymph node dissection in patients with MTC. Further prospective, randomized studies are warranted.
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BACKGROUND: Medullary thyroid carcinoma (MTC) is rare. Nationwide population-based studies are important to evaluate its clinical course. OBJECTIVES: To describe all patients with MTC in Norway during 1994-2016 and compare time-related trends in diagnostics and surgical treatment, including prognostic factors for biochemical cure and disease-specific survival (DSS). METHODS: This retrospective population-based cohort study includes data for 228 out of 237 patients (96%) with MTC; 201 patients were surgically treated. Patients were identified in the 4 regional centers treating MTC and by the Cancer Registry of Norway. Data were collected from patients' files. Trends were compared over 2 study periods. RESULTS: MTC accounted for 4.2% of thyroid carcinomas. During the study periods, the incidence increased from 0.18 to 0.25: 100,000 per year, preoperative diagnostics improved with increased use of calcitonin, ultrasound, and fine-needle cytology (p = 0.010, p < 0,001, and p = 0.001), patients were diagnosed at an earlier tumor stage (p = 0.004), and more patients were cured (p = 0.002). Via multivariate analysis of patients with metastatic lymph nodes, independent prognostic factors for cure were: a low ratio of metastatic and total number of dissected lymph nodes (p = 0.021) and no extrathyroidal extension (p = 0.030). Independent prognostic factors for DSS were: no distant metastasis, a younger age, and a low ratio of metastatic and dissected lymph nodes (p = 0.005, p = 0.020, p = 0.022). CONCLUSIONS: Preoperative diagnostics have improved over time with increased therapeutic control. A low ratio of metastatic and dissected lymph nodes predicts better outcomes in patients with metastatic lymph nodes.
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OBJECTIVES: To explore performance measures among non-immigrants and immigrants attending BreastScreen Norway. METHODS: We analysed data from 2,951,375 screening examinations among non-immigrants and 153,026 among immigrants from 1996 to 2015. Immigrants were categorised into high- and low-incidence countries according to the incidence of breast cancer in their birth country. Performance measures, including attendance and recall rates, rates of screen-detected cancer (SDC) and interval breast cancer (IBC), positive predictive value (PPV) and histopathological tumour characteristics, were analysed. We used Fisher's exact model and t tests for descriptive statistics, and Poisson regression, adjusting for age and screening history, comparing results for non-immigrants versus immigrants. RESULTS: Attendance rates were 78% for non-immigrants and 56% for immigrants (p < 0.001). Rates of prevalent screens were 24% for non-immigrants and 32% for immigrants (p < 0.001). Immigrants from low-incidence countries were younger at diagnosis than non-immigrants (57 years versus 60 years, p < 0.001). Recall rates were 3.1% for non-immigrants and 3.8% for immigrants (p < 0.001), while PPVs were 17% and 14% (p < 0.001), respectively. IBCs in immigrants from low-incidence countries were more often triple negative (RRadj 1.81, 95% CI 1.11-2.94) than those in non-immigrants. Both SDC and IBC in immigrants from low-incidence countries tended more often to be histological grade 3 than those in non-immigrants. CONCLUSION: Immigrants had lower attendance rates, higher recall rates and lower PPV than non-immigrants. The optimal age range and screening interval for immigrant women from low-incidence countries need to be further investigated. KEY POINTS: ⢠Immigrants from countries with a low incidence of breast cancer had their breast cancer diagnosed at a younger age than non-immigrants. ⢠Interval breast cancers detected in immigrants from countries with a low incidence of breast cancers were more often triple negative than those in non-immigrants. ⢠The optimal age range and screening interval for immigrant women from low-incidence countries and non-immigrants might differ.
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Neoplasias de la Mama/prevención & control , Carcinoma Intraductal no Infiltrante/prevención & control , Emigrantes e Inmigrantes/estadística & datos numéricos , Mama/patología , Neoplasias de la Mama/etnología , Neoplasias de la Mama/patología , Carcinoma Intraductal no Infiltrante/etnología , Carcinoma Intraductal no Infiltrante/patología , Detección Precoz del Cáncer/métodos , Femenino , Humanos , Incidencia , Mamografía/métodos , Tamizaje Masivo/métodos , Persona de Mediana Edad , Invasividad Neoplásica , Noruega/epidemiología , Aceptación de la Atención de Salud/estadística & datos numéricos , Prevalencia , Estudios ProspectivosRESUMEN
PURPOSE: Proliferation is a hallmark of cancer. Using a combined genomic approach, FGD5 amplification has been identified as a driver of proliferation in Luminal breast cancer. We aimed to describe FGD5 copy number change in breast cancer, and to assess a possible association with tumour proliferation and prognosis. METHODS: We used fluorescence in situ hybridization targeting FGD5 and chromosome 3 centromere (CEP3) on formalin-fixed, paraffin-embedded tissue from 430 primary breast cancers and 108 lymph node metastases, from a cohort of Norwegian breast cancer patients. We tested the association between FGD5 copy number status and proliferation (assessed by Ki67 levels and mitotic count) using Pearson's Chi square test, and assessed the prognostic impact of FGD5 copy number change by estimating cumulative risks of death and hazard ratios. RESULTS: We identified FGD5 amplification (defined as FGD5/CEP3 ratio ≥2 or mean FGD5/tumour cell ≥4) in 9.5% of tumours. Mitotic count and Ki67 levels were higher in tumours with FGD5 copy number increase, compared to tumours with no copy number change. After 10 years of follow-up, cumulative risk of death from breast cancer was higher among cases with FGD5 amplification [48.1% (95% CI 33.8-64.7)], compared to non-amplified cases [27.7% (95% CI 23.4-32.6)]. CONCLUSIONS: FGD5 is a new prognostic marker in breast cancer, and increased copy number is associated with higher tumour proliferation and poorer long-term prognosis.
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Neoplasias de la Mama/genética , Neoplasias de la Mama/mortalidad , Amplificación de Genes , Factores de Intercambio de Guanina Nucleótido/genética , Anciano , Anciano de 80 o más Años , Neoplasias de la Mama/patología , Variaciones en el Número de Copia de ADN , Femenino , Estudios de Seguimiento , Reguladores de Proteínas de Unión al GTP/genética , Humanos , Hibridación Fluorescente in Situ , Clasificación del Tumor , Metástasis de la Neoplasia , Pronóstico , Modelos de Riesgos ProporcionalesRESUMEN
BACKGROUND: Secular trends in incidence and prognosis of molecular breast cancer subtypes are poorly described. We studied long-term trends in a population of Norwegian women born 1886-1977. METHODS: A total of 52,949 women were followed for breast cancer incidence, and 1,423 tumors were reclassified into molecular subtypes using IHC and in situ hybridization. We compared incidence rates among women born 1886-1928 and 1929-1977, estimated age-specific incidence rate ratios (IRR), and performed multiple imputations to account for unknown subtype. Prognosis was compared for women diagnosed before 1995 and in 1995 or later, estimating cumulative risk of death and HRs. RESULTS: Between 50 and 69 years of age, incidence rates of Luminal A and Luminal B (HER2-) were higher among women born in 1929 or later, compared with before 1929 [IRRs 50-54 years; after imputations: 3.5; 95% confidence interval (CI), 1.8-6.9 and 2.5; 95% CI, 1.2-5.2, respectively], with no clear differences for other subtypes. Rates of death were lower in women diagnosed in 1995 or later, compared to before 1995, for Luminal A (HR 0.4; 95% CI, 0.3-0.5), Luminal B (HER2-; HR 0.5; 95% CI, 0.3-0.7), and Basal phenotype (HR 0.4; 95% CI, 0.2-0.9). CONCLUSIONS: We found a strong secular incidence increase restricted to Luminal A and Luminal B (HER2-) subtypes, combined with a markedly improved prognosis for these subtypes and for the Basal phenotype. IMPACT: This study documents a clear secular increase in incidence and a concomitant improved prognosis for specific molecular breast cancer subtypes. Cancer Epidemiol Biomarkers Prev; 25(12); 1625-34. ©2016 AACR.
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Neoplasias de la Mama/epidemiología , Receptor ErbB-2/análisis , Anciano , Biomarcadores de Tumor , Neoplasias de la Mama/clasificación , Neoplasias de la Mama/diagnóstico , Neoplasias de la Mama/mortalidad , Femenino , Humanos , Incidencia , Persona de Mediana Edad , Noruega/epidemiología , Pronóstico , Receptor ErbB-2/genéticaRESUMEN
BACKGROUND: Most studies of cutaneous head and neck melanomas (CHNM) have reported poorer survival in CHNM compared with other sites, especially on the scalp/neck. OBJECTIVE: We sought to compare patient and tumor characteristics between CHNM and cutaneous trunk and extremity melanomas and between CHNM locations (face/ear vs scalp/neck, anterior vs posterior), and to study prognostic factors in patients with CHNM. METHODS: We studied all CHNM (n = 1074) from 8120 cases of cutaneous melanomas diagnosed in Norway in 2008 to 2012. RESULTS: Compared with cutaneous trunk and extremity melanomas, CHNM were more frequently found in men, more often nodular and lentigo maligna cutaneous melanomas, and diagnosed at higher T stage (P ≤ .01). CHNM located on posterior sites were diagnosed at significantly higher T stage, and were significantly more often diagnosed with ulceration and at more advanced stage compared with CHNM located on anterior sites (P < .001). T stage and clinical stage were the only significant prognostic factors for melanoma-specific and overall death in the multivariable analysis (P < .001). LIMITATIONS: Low number of cases and the relatively high frequency of missing values are limitations. CONCLUSION: More advanced CHNM were diagnosed on posterior compared with anterior locations, but location was not a significant prognostic factor for cutaneous melanoma-specific or overall death in the multivariable models.
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Melanoma/mortalidad , Neoplasias Cutáneas/microbiología , Adulto , Anciano , Extremidades , Femenino , Neoplasias de Cabeza y Cuello/mortalidad , Humanos , Peca Melanótica de Hutchinson/mortalidad , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Noruega/epidemiología , Especificidad de Órganos , Pronóstico , Sistema de Registros , Torso , Melanoma Cutáneo MalignoRESUMEN
BACKGROUND: Multiple endocrine neoplasia type 2A (MEN 2A) is an autosomal dominant syndrome caused by activating germline mutations in the RET (REarranged during Transfection) proto-oncogene. MEN 2A has a strong (>95%) and age-dependent (5-25 years) clinical penetrance of medullary thyroid carcinoma (MTC). Several major studies have analyzed the predictive and prognostic factors for MEN 2A to find indicators that predict the optimal timing of prophylactic thyroidectomy. The aims of this study were to describe all known RET positive MEN 2A patients diagnosed in Norway and to evaluate the clinical course of MTC, as well as its predictive and prognostic factors. METHODS: This nationwide retrospective cohort study included data for 65 (14 index and 51 screening patients) out of a total of 67 MEN 2A patients with the RET gene mutation who were diagnosed in Norway since 1974. Data were collected by reviewing patient files. The variables analyzed were genotype, phenotype, preoperative basal calcitonin, age at thyroid surgery, central lymph node dissection and nodal status at primary surgery, number of surgical procedures, and biochemical cure. Of the 65 patients, 60 had undergone thyroid surgery. The median follow-up period was 9.9 years. The patients were divided into pre-RET-and RET-era, which included patients who had thyroid surgery before January 1, 1994, and after, respectively. RESULTS: In index and screening patients, MTC was found, respectively, in 100% and 45% of cases, central lymph node dissection at primary surgery was done for 64% and 52% of patients, and the median total number of surgical procedures was two (range 1-6) and one (range 1-4). At primary surgery, all patients (n = 13) with lymph node metastases had preoperative basal calcitonin levels ≥68 pg/mL, and all patients (n = 17) without central lymph node dissection and preoperative basal calcitonin <40 pg/mL were biochemically cured. Multivariate analysis showed that preoperative basal calcitonin was a significant predictive factor for MTC superior to age at thyroid surgery when analyzing the entire period (p = 0.009) and the RET-era separately (p = 0.021). Prognostic factors for biochemical cure were preoperative basal calcitonin, central lymph node dissection, and nodal status at primary surgery (p = 0.037, p = 0.002, and p = 0.005) when analyzing the entire period, but only nodal status at primary surgery when the RET-era was considered separately (p = 0.006). CONCLUSIONS: Preoperative basal calcitonin alone can serve as an indicator for optimal timing and the extent of thyroid surgery for MEN 2A patients that could be considered safe. The results are consistent with previously reported data.
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Carcinoma Medular/patología , Neoplasia Endocrina Múltiple Tipo 2a/patología , Proteínas Proto-Oncogénicas c-ret/genética , Neoplasias de la Tiroides/patología , Tiroidectomía , Adolescente , Adulto , Anciano , Biomarcadores de Tumor/sangre , Calcitonina/sangre , Carcinoma Medular/sangre , Carcinoma Medular/genética , Carcinoma Medular/cirugía , Niño , Preescolar , Progresión de la Enfermedad , Femenino , Humanos , Escisión del Ganglio Linfático , Masculino , Persona de Mediana Edad , Neoplasia Endocrina Múltiple Tipo 2a/sangre , Neoplasia Endocrina Múltiple Tipo 2a/genética , Neoplasia Endocrina Múltiple Tipo 2a/cirugía , Noruega , Pronóstico , Proto-Oncogenes Mas , Neoplasias de la Tiroides/sangre , Neoplasias de la Tiroides/genética , Neoplasias de la Tiroides/cirugía , Adulto JovenAsunto(s)
Subunidades alfa de la Proteína de Unión al GTP/genética , Melanoma/genética , Proteínas Proto-Oncogénicas c-kit/genética , Neoplasias Cutáneas/genética , Población Negra , Análisis Mutacional de ADN , Femenino , Pie/patología , GTP Fosfohidrolasas/genética , Subunidades alfa de la Proteína de Unión al GTP Gq-G11 , Mano/patología , Humanos , Masculino , Melanoma/etnología , Proteínas de la Membrana/genética , Reacción en Cadena de la Polimerasa Multiplex , Mutación , Neoplasias Cutáneas/etnología , Población BlancaRESUMEN
The significance of tumor necrosis in cutaneous melanoma has not been well elucidated. The purpose of this study was to explore the prognostic impact of necrosis in comparison with other known clinicopathologic factors in these tumors. Initially, 457 consecutive cases of nodular cutaneous melanoma (1981 to 2008) were included in this series. Tumor necrosis was assessed on hematoxylin and eosin-stained sections and was recorded as significant when an area of at least a quarter of a high-power field (×400; 0.07 mm) was occupied by necrotic cells and as sparse when clusters of at least 5 necrotic cells were observed. Tumor necrosis (26% of the cases) was associated with increased tumor thickness, high mitotic count, presence of tumor ulceration, and decreased survival. Stratified analyses (univariate and multivariate) with standard microscopic variables indicated the strongest prognostic influence of necrosis in tumors thicker than 4 mm. Notably, in the stratum of pT4 tumors, presence of necrosis was a stronger prognostic predictor than was ulceration. Tumor necrosis was a significant prognostic indicator providing additional information to established predictors of patient outcome in this series of nodular cutaneous melanoma, predominantly among thick tumors (>4 mm). Presence of necrosis was a stronger indicator for worse outcome compared with ulceration in pT4 tumors.
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Melanoma/diagnóstico , Neoplasias Cutáneas/diagnóstico , Piel/patología , Anciano , Femenino , Humanos , Masculino , Melanoma/mortalidad , Necrosis , Estadificación de Neoplasias , Noruega/epidemiología , Pronóstico , Neoplasias Cutáneas/mortalidadRESUMEN
Tumor hypoxia is relevant for tumor growth, metabolism and epithelial-to-mesenchymal transition (EMT). We report that hyperbaric oxygen (HBO) treatment induced mesenchymal-to-epithelial transition (MET) in a dimethyl-alpha-benzantracene induced mammary rat adenocarcinoma model, and the MET was associated with extensive coordinated gene expression changes and less aggressive tumors. One group of tumor bearing rats was exposed to HBO (2 bar, pO(2) = 2 bar, 4 exposures à 90 minutes), whereas the control group was housed under normal atmosphere (1 bar, pO(2) = 0.2 bar). Treatment effects were determined by assessment of tumor growth, tumor vascularisation, tumor cell proliferation, cell death, collagen fibrils and gene expression profile. Tumor growth was significantly reduced (approximately 16%) after HBO treatment compared to day 1 levels, whereas control tumors increased almost 100% in volume. Significant decreases in tumor cell proliferation, tumor blood vessels and collagen fibrils, together with an increase in cell death, are consistent with tumor growth reduction and tumor stroma influence after hyperoxic treatment. Gene expression profiling showed that HBO induced MET. In conclusion, hyperoxia induced MET with coordinated expression of gene modules involved in cell junctions and attachments together with a shift towards non-tumorigenic metabolism. This leads to more differentiated and less aggressive tumors, and indicates that oxygen per se might be an important factor in the "switches" of EMT and MET in vivo. HBO treatment also attenuated tumor growth and changed tumor stroma, by targeting the vascular system, having anti-proliferative and pro-apoptotic effects.
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Adenocarcinoma/patología , Células Epiteliales/citología , Hiperoxia/tratamiento farmacológico , Mesodermo/citología , Modelos Biológicos , Animales , Femenino , Inmunohistoquímica , Ratas , Ratas Sprague-DawleyRESUMEN
The aim of this work was to compare DNA microarray results using either total RNA or affinity-purified poly(A) RNA from the same biological sample for target preparation. The high-density oligonucleotide microarrays of both Agilent Technologies (based on two-color detection) and Applied Biosystems (based on single-color detection) were evaluated. Real-time quantitative PCR was used to quantify messenger RNA (mRNA) and ribosomal RNA (rRNA) at different stages of target preparations. Poly(A) RNA versus total RNA target hybridizations exhibited slightly lower correlation coefficients than did self versus self hybridizations (i.e., poly(A) RNA targets vs. poly(A) RNA targets or total RNA targets vs. total RNA targets). Only a small fraction of all transcripts appeared to be significantly over- or underrepresented when total RNA targets or poly(A) RNA targets from the same biological sample were compared. Therefore, the conclusion is that poly(A) affinity purification from total RNA can be omitted during target preparation for routine mRNA expression analysis using high-density oligonucleotide microarrays. Among consistently overrepresented transcripts in total RNA targets were histone mRNAs known to lack poly(A) tails. Therefore, structurally exceptional RNA species can be identified by comparing targets derived from either poly(A) RNA or total RNA using microarray hybridization.
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Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , ARN Mensajero/genética , ARN/genética , Secuencia de Bases , Línea Celular , Cromatografía de Afinidad , Cartilla de ADN/genética , Desoxirribonucleasas , Neoplasias Endometriales/genética , Femenino , Perfilación de la Expresión Génica , Humanos , Leucemia Mieloide Aguda/genética , Masculino , Oligodesoxirribonucleótidos , Reacción en Cadena de la Polimerasa , Hiperplasia Prostática/genética , ARN/aislamiento & purificación , ARN Mensajero/aislamiento & purificación , ARN Neoplásico/genética , ARN Neoplásico/aislamiento & purificación , ARN Ribosómico/genética , ARN Ribosómico/aislamiento & purificaciónRESUMEN
PURPOSE: The human SIM2 gene is located within the Down's syndrome critical region of chromosome 21 and encodes transcription factors involved in brain development and neuronal differentiation. SIM2 has been assigned a possible role in the pathogenesis of solid tumors, and the SIM2-short isoform (SIM2-s) was recently proposed as a molecular target for cancer therapy. We previously reported SIM2 among the highly up-regulated genes in 29 prostate cancers, and the purpose of our present study was to examine the expression status of SIM2 at the transcriptional and protein level as related to outcome in prostate cancer. EXPERIMENTAL DESIGN: By quantitative PCR, mRNA in situ hybridization, and immunohistochemistry, we evaluated the expression and significance of SIM2 isoforms in 39 patients with clinically localized prostate cancer and validated the expression of SIM2-s protein in an independent cohort of 103 radical prostatectomies from patients with long and complete follow-up. RESULTS: The SIM2 isoforms (SIM2-s and SIM2-l) were significantly coexpressed and increased in prostate cancer. Tumor cell expression of SIM2-s protein was associated with adverse clinicopathologic factors like increased preoperative serum prostate-specific antigen, high histologic grade, invasive tumor growth with extra-prostatic extension, and increased tumor cell proliferation by Ki-67 expression. SIM2-s protein expression was significantly associated with reduced cancer-specific survival in multivariate analyses. CONCLUSIONS: These novel findings indicate for the first time that SIM2 expression might be important for clinical progress of human cancer and support the recent proposal of SIM2-s as a candidate for targeted therapy in prostate cancer.
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Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/biosíntesis , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/química , Regulación Neoplásica de la Expresión Génica , Neoplasias de la Próstata/metabolismo , Anciano , Biomarcadores de Tumor , Progresión de la Enfermedad , Humanos , Inmunohistoquímica , Hibridación in Situ , Antígeno Ki-67/biosíntesis , Masculino , Persona de Mediana Edad , Análisis Multivariante , Invasividad Neoplásica , Análisis de Secuencia por Matrices de Oligonucleótidos , Neoplasias de la Próstata/mortalidad , Neoplasias de la Próstata/cirugíaRESUMEN
The aim of this study was to identify and validate differentially expressed genes in matched pairs of benign and malignant prostate tissue. Samples included 29 histologically verified primary tumors and 23 benign controls. Microarray analysis was initially performed using a sequence verified set of 40,000 human cDNA clones. Among the genes most consistently and highly upregulated in prostate cancer was the ETS family transcription factor ERG (ETS related gene). This finding was validated in an expanded patient series (37 tumors and 38 benign samples) using DNA oligonucleotide microarray and real-time quantitative PCR assays. ERG was 20- to more than 100-fold overexpressed in prostate cancer compared with benign prostate tissue in more than 50% of patients according to quantitative PCR. Surprisingly, ERG mRNA levels were found to be significantly higher in the endothelial cell line, HUVEC, than in the prostate cell lines PC3, DU145 and LNCaP. In situ hybridization of prostate cancer tissue revealed that ERG was abundantly expressed in both prostate cancer cells and associated endothelial cells. The consistency and magnitude of ERG overexpression in prostate cancer appeared unique, but several related ETS transcription factors were also overexpressed in matched pairs of tumor and benign samples, whereas ETS2 was significantly underexpressed. Our findings support the hypothesis that ERG overexpression and related ETS transcription factors are important for early prostate carcinogenesis.
Asunto(s)
Proteínas de Unión al ADN/genética , Regulación Neoplásica de la Expresión Génica , Neoplasias de la Próstata/genética , Proteínas Proto-Oncogénicas c-ets/genética , Transactivadores/genética , Cartilla de ADN , Humanos , Hibridación in Situ , Masculino , Análisis de Secuencia por Matrices de Oligonucleótidos , Reacción en Cadena de la Polimerasa , Próstata/fisiología , Prostatectomía , Neoplasias de la Próstata/cirugía , Regulador Transcripcional ERGRESUMEN
We previously reported that high tumour cell proliferation evaluated by Ki-67 expression, high mitotic frequency and high histological grade were associated with resistance to primary doxorubicin monotherapy in locally advanced breast cancer harbouring wild-type (wt) TP53. The aim of our present study was to evaluate the predictive and prognostic impact of proliferation parameters assessed in tumour tissue obtained after chemotherapy, and alterations induced in tumour cell proliferation. While we found a significant reduction in Ki-67 expression and mitotic frequency in tumours with wtTP53 (p=0.001 and p=0.008, respectively), no significant change was recorded in tumours expressing mutant TP53. For histological grade there was no significant change in either group. There was a direct correlation between pre- and post-treatment values for Ki-67 and mitotic frequency in tumours harbouring wtTP53 (p=0.0001 for both), but no correlation in tumours harbouring mutated TP53. High post-treatment Ki-67 expression and mitotic frequency were found to predict doxorubicin resistance only in patients with wtTP53 (p=0.04 and p=0.03, respectively). The prognostic importance of proliferation markers and histological grade was found to be similar whether they were determined in the pre- or post-treatment samples (Ki-67; pre: p=0.02; post: p=0.03; mitotic frequency; p=0.002 and p=0.01, respectively; histological grade; p=0.0001 and p=0.002, respectively). While the reduction in mitotic frequency was associated with improved survival (p=0.03), no significant associations between changes in other parameters and outcome were recorded.
Asunto(s)
Antibióticos Antineoplásicos/uso terapéutico , Biomarcadores de Tumor/análisis , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Proliferación Celular , Doxorrubicina/uso terapéutico , Antígeno Ki-67/biosíntesis , Neoplasias de la Mama/patología , Femenino , Humanos , Inmunohistoquímica , Antígeno Ki-67/análisis , Mitosis , Valor Predictivo de las Pruebas , Pronóstico , Análisis de Supervivencia , Resultado del TratamientoRESUMEN
PURPOSE: To our knowledge we introduce the ROC partial area under the curve (AUC) index as a method of evaluating the discriminative power of different prostate cancer predictors. Peripheral zone volume and peripheral zone prostate specific antigen (PSA) density are introduced as potential predictors and compared with other known predictors of prostate cancer. MATERIALS AND METHODS: During 1999, 220 consecutive patients with suspected early prostate cancer were examined using total PSA, free PSA, total prostate volume, transition zone volume and transrectal ultrasonography guided sextant biopsy of the prostate. The free-to-total PSA ratio, PSA density, transition zone PSA density, peripheral zone volume and peripheral zone PSA density were calculated. Usually total AUC is used to evaluate the discriminative power of different parameters. In this study parameters were evaluated by the ROC partial area index, which includes only the AUC in highly sensitivity parts of the ROC curve. Explorative analysis using logistic regression analysis was performed to investigate the ability of combinations of parameters to predict cancer. RESULTS: Of the 220 patients 75 were diagnosed with cancer. In the subgroup of 160 patients with PSA less than 10 microg/l 44 had cancer. Transition zone PSA density and PSA density had significant discriminative power in the total group, while none of the parameters were discriminative in the subgroup of patients. CONCLUSIONS: When high sensitivity is demanded, the ROC partial area index seems to be meaningful for evaluating the discriminative power of potential predictors. In our study none of the evaluated parameters had discriminative power in patients with PSA less than 10 microg/l, while transition zone PSA density and PSA density showed discriminative power in the total group of patients.