Marco Lucchi

Bcl-2 protein: a prognostic factor inversely correlated to p53 in non-small-cell lung cancer.

Non-small-cell lung cancer (NSCLC) prognosis is strictly related to well-established clinicopathological parameters which have unfortunately become insufficient in the prognostic evaluation of this type of cancer. As p53 and bcl-2 gene deregulations are frequently involved in several types of epithelial malignancies, we investigated the Bcl-2 and p53 protein expression in 91 and 101 cases of NSCLC respectively. The expression was then compared with established indicators of prognosis and biological behaviour of the tumours. No relationship was observed between Bcl-2 and either clinicopathological or biological parameters such as histology, grading, tumour status, nodal metastasis and proliferative activity evaluated by scoring proliferating cell nuclear antigen expression and Ki-67 immunoreactivity. However, the mean Bcl-2 expression was significantly lower in patients who developed metastasis during follow-up or died of metastatic disease (P = 0.006 and P = 0.01 respectively). Moreover, survival probability was higher in patients who expressed the Bcl-2 protein (P = 0.0002). In contrast with this, p53 protein accumulation was observed in tumours with metastatic nodal involvement (P = 0.02) or in patients who developed metastasis during follow-up (P = 0.01), although no correlation was found between p53 expression and overall survival. An inverse relationship was also found between Bcl-2 and the anti-oncogene protein product p53 (P = 0.01). Thus, a high proportion of NSCLCs express p53 and Bcl-2 proteins and their expression may have prognostic importance.

Neoangiogenesis and p53 protein in lung cancer: their prognostic role and their relation with vascular endothelial growth factor (VEGF) expression.

Following up-regulation of an angiogenesis inhibitor by the wild-type p53 protein proven recently, we have analysed on the one hand the prognostic impact of microvessel count (MC) and p53 protein overexpression in non-small-cell lung carcinoma (NSCLC) progression and, on the other hand, the inter-relation between the microvascular pattern and the p53 protein expression. Moreover, we assessed the expression of vascular endothelial growth factor (VEGF), one of the pivotal mediators of tumour angiogenesis, in order to investigate its relation to p53 protein expression and MC. Tumours from 73 patients resected for NSCLC between March 1991 and April 1992 (median follow-up 47 months, range 32-51 months) were analysed using an immunohistochemical method. In univariate analysis, MC and p53 accumulation were shown to affect metastatic nodal involvement, recurrence and death significantly. Multiple logistic regression analysis showed an important prognostic influence of MC and nodal status on overall (P = 0.0009; P = 0.01) and disease-free survival (P = 0.0001; P = 0.03). Interestingly, a strong statistical association was observed between p53 nuclear accumulation and MC (P = 0.0003). The same inter-relationship was found in non-squamous histotype (P = 0.002). When we analysed the concomitant influence of MC and p53 expression on overall survival, we were able to confirm a real predominant role of MC in comparison with p53. With regard to VEGF expression, p53-negative and lowly vascularized tumours showed a mean VEGF expression significantly lower than p53-positive and highly vascularized cancers (P = 0.02). These results underline the prognostic impact of MC and p53 protein accumulation in NSCLC and their reciprocal inter-relationship, supporting the hypothesis of a wild-type p53 regulation on the angiogenetic process through a VEGF up-regulation.

A high vascular count and overexpression of vascular endothelial growth factor are associated with unfavourable prognosis in operated small cell lung carcinoma

It has been widely demonstrated that neo-angiogenesis and its mediators (i.e. vascular endothelial growth factor), represent useful indicators of poor prognosis in non small cell lung carcinoma. In order to verify whether neovascularization and vascular endothelial growth factor may be considered useful markers of clinical outcome also in the small cell lung cancer subgroup, we retrospectively investigated a series of 75 patients with small cell lung carcinoma treated by surgery between 1980 and 1990. Immunohistochemically-detected microvessels and vascular endothelial growth factor expressing cells were significantly associated with poor prognosis, as well as with nodal status and pathological stage. In fact, patients whose tumours had vascular count and vascular endothelial growth factor expression higher than median value of the entire series (59 vessels per 0.74 mm2 and 50% of positive cells, respectively), showed a shorter overall and disease-free survival (P=0.001, P=0.001; P=0.008, P=0.03). Moreover, the presence of hilar and/or mediastinal nodal metastasis and advanced stage significantly affected overall and disease-free interval (P=0.00009, P=0.00001; P=0.0001, P=0.00001). At multivariate analysis, only vascular endothelial growth factor expression retained its influence on overall survival (P=0.001), suggesting that angiogenic phenomenon may have an important role in the clinical behaviour of this lung cancer subgroup.

Array comparative genomic hybridization-based characterization of genetic alterations in pulmonary neuroendocrine tumors

The goal of this study was to characterize and classify pulmonary neuroendocrine tumors based on array comparative genomic hybridization (aCGH). Using aCGH, we performed karyotype analysis of 33 small cell lung cancer (SCLC) tumors, 13 SCLC cell lines, 19 bronchial carcinoids, and 9 gastrointestinal carcinoids. In contrast to the relatively conserved karyotypes of carcinoid tumors, the karyotypes of SCLC tumors and cell lines were highly aberrant. High copy number (CN) gains were detected in SCLC tumors and cell lines in cytogenetic bands encoding JAK2, FGFR1, and MYC family members. In some of those samples, the CN of these genes exceeded 100, suggesting that they could represent driver alterations and potential drug targets in subgroups of SCLC patients. In SCLC tumors, as well as bronchial carcinoids and carcinoids of gastrointestinal origin, recurrent CN alterations were observed in 203 genes, including the RB1 gene and 59 microRNAs of which 51 locate in the DLK1-DIO3 domain. These findings suggest the existence of partially shared CN alterations in these tumor types. In contrast, CN alterations of the TP53 gene and the MYC family members were predominantly observed in SCLC. Furthermore, we demonstrated that the aCGH profile of SCLC cell lines highly resembles that of clinical SCLC specimens. Finally, by analyzing potential drug targets, we provide a genomics-based rationale for targeting the AKT-mTOR and apoptosis pathways in SCLC.

Small cell lung carcinoma (SCLC): the angiogenic phenomenon

OBJECTIVES Tumor angiogenesis, expressed by the microvessel count (MVC), and its mediators (i.e. vascular endothelial growth factor) significantly correlate with metastases in surgically treated non-small cell lung carcinoma/cancer (NSCLC). SCLC is rarely treated by surgery, as a consequence, few specimens are available to perform a biological characterization. We reviewed our experience in the surgical treatment of SCLC with particular reference to the angiogenetic expression and its correlation to the stage of disease and prognosis. METHODS We retrospectively investigated a homogenous cohort of 87 patients with SCLC, who were primarily operated on and then underwent adjuvant chemotherapy between 1980 and 1998. Their median age was 62 years (range 34-73). All the patients were completely staged. The surgical procedures included: 32 pneumonectomies and 55 lobectomies. There were 46 N0, 17 N1 and 24 N2-disease. The adjuvant chemotherapy consisted of four to six courses of cyclophosphamide, epidoxorubicine and etoposide. The MVC was determined highlighting the microvessels with anti-CD34 monoclonal antibodies. Immunostaining for VEGF was performed using the ABC method with anti-VEGF monoclonal antibodies. The p53 protein expression was assessed by NCL-DO7 anti-p53 monoclonal antibody. RESULTS With a median follow-up of 109.6 months (range 25-238), 37 patients are alive and well, two are alive with systemic metastases. Forty-four patients died of local (n=5) or systemic (n=39) relapse, while four patients died from other causes. The median MVC was 59 (range 18-145). Among the clinico-pathological parameters, metastatic nodal-involvement (P=0.002) and advanced stage of disease (P=0.005) were associated with a worse overall survival (OS). MVC and VEGF protein expression significantly affected the survival (P<0.001 and P=0.0008, respectively). No statistical association was found between p53 alterations and OS as well as no association was found among p53 alterations, MVC and VEGF expression. On multivariate analysis only the VEGF expression (P=0.003) was an independent prognostic factor. CONCLUSIONS Angiogenesis plays a role in the metastatic process of the SCLC as well as NSCLC. SCLC has a higher vascularization than NSCLC as results from the higher number of microvessels; however, tumor angiogenesis tested by the MVC and the VEGF protein expression correlates with the prognosis also in SCLC. SCLC may be an ideal field to test new antiangiogenic drugs associated to chemotherapy.

Expression of vascular endothelial growth factor mRNA in non-small-cell lung carcinomas

SummaryThe vascular endothelial growth factor (VEGF) has been shown to be strictly related to vascular permeability and endothelial cell growth under physiological and pathological conditions. In tumour development and progression, VEGF plays a pivotal role in the development of the tumoral vascular network, and useful information in the progression of human cancer can be obtained by analysing the vascular endothelial growth factor expression of the tumours. In this study, we investigated the vascular endothelial growth factor transcript expression in non-small-cell lung carcinomas to evaluate the significance of this factor in a group of cancers in which the vascular pattern has been shown to significantly affect progression. Surgical samples of 42 patients with NSCLC were studied using reverse transcription polymerase chain reaction (PCR) analysis and in situ hybridization. Thirty-three out of 42 cases (78.6%) showed VEGF transcript expression predominantly as transcripts for the secretory forms of VEGF (isoforms 121 and 165). In situ hybridization, performed on 24 out of 42 samples, showed that the VEGF transcript expression was in several cases present in the cytoplasm both of neoplastic and normal cells, even if the VEGF mRNA was less expressed in the corresponding non-tumoral part. The VEGF 121 expression was associated with hilar and/or mediastinal nodal involvement (P = 0.02), and, taken together, the VEGF isoforms were shown to significantly influence overall (P = 0.02) and disease-free survival (P = 0.03). As a regulator of tumour angiogenesis, VEGF may represent a useful indicator of progression and poor prognosis in non-small-cell lung carcinomas.

Osteopontin Expression and Prognostic Significance in Non–Small Cell Lung Cancer

Purpose: The survival rate of non–small cell lung cancer patients is very low, and knowledge of predictors of outcome is inadequate. To improve the curability of lung cancer, we need to identify new specific molecules involved in tumorigenesis and progression. The purpose of this study was to better define the role of osteopontin in non–small cell lung cancer biology by determining its prognostic significance. Experimental Design: Osteopontin expression was evaluated by immunohistochemistry, as percentage of neoplastic cells with cytoplasmic immunoreactivity, in a wide series of patients with stage I-IIIA non–small cell lung cancer (207 cases). The median value of this series (20% of positive cells) was used as the cutoff value to distinguish tumors with low (<20%) from tumors with high (≥20%) osteopontin expression. Results: Taking the series of patients as a whole (207 cases), osteopontin expression was associated with neither overall survival (P = 0.14) nor disease-free survival (P = 0.074). However, among patients with at least 6 years of follow-up (163 cases), 6-year overall survival and disease-free survival were significantly reduced if osteopontin expression was high (P = 0.0085 for overall survival, P = 0.0023 for disease-free survival). Moreover, a statistically significant correlation between high levels of osteopontin and shorter overall survival (P = 0.034) and disease-free survival (P = 0.011) in patients with stage I tumors (136 cases) was shown. Conclusions: Our results support the hypothesis of an association between high osteopontin expression and poor survival of patients with stage I non–small cell lung cancer, suggesting that osteopontin could be a candidate target for cancer therapy.

Bcl2 and p53 regulate vascular endothelial growth factor (VEGF)-mediated angiogenesis in non-small cell lung carcinoma.

The aim of this study was to investigate the expression of p53 and bcl2 proteins in a series of 107 non-small cell lung cancers (NSCLC), and to relate such protein expression to neovascularisation and the expression of vascular endothelial growth factor (VEGF). Moreover, we analysed the prognostic impact of these biological parameters on overall survival, both in univariate and multivariate analyses. An inverse association was found between bcl2 expression and microvessel count (MVC; P = 0.0004) and bcl2 and VEGF (P = 0.007). In contrast, a significant association was found between p53 expression and MVC (P = 0.03) and p53 and VEGF expression (P = 0.04). In univariate analysis, nodal status (P < 0.000001), MVC (P < 0.000001), bcl2 (P = 0.002), p53 (P = 0.03) and VEGF expression (P < 0.000001) significantly affected overall survival, but in multivariate analysis only MVC and VEGF expression retained their prognostic influence. Our results suggest that bcl2 and p53 possibly control the development of tumour angiogenesis in NSCLC, with putative mediation by VEGF. Moreover, the important influence of angiogenesis in the progression of NSCLC is further highlighted.

The IASLC Lung Cancer Staging Project: Proposals for Coding T Categories for Subsolid Nodules and Assessment of Tumor Size in Part-Solid Tumors in the Forthcoming Eighth Edition of the TNM Classification of Lung Cancer

ABSTRACT This article proposes codes for the primary tumor categories of adenocarcinoma in situ (AIS) and minimally invasive adenocarcinoma (MIA) and a uniform way to measure tumor size in part‐solid tumors for the eighth edition of the tumor, node, and metastasis classification of lung cancer. In 2011, new entities of AIS, MIA, and lepidic predominant adenocarcinoma were defined, and they were later incorporated into the 2015 World Health Organization classification of lung cancer. To fit these entities into the T component of the staging system, the Tis category is proposed for AIS, with Tis (AIS) specified if it is to be distinguished from squamous cell carcinoma in situ (SCIS), which is to be designated Tis (SCIS). We also propose that MIA be classified as T1mi. Furthermore, the use of the invasive size for T descriptor size follows a recommendation made in three editions of the Union for International Cancer Control tumor, node, and metastasis supplement since 2003. For tumor size, the greatest dimension should be reported both clinically and pathologically. In nonmucinous lung adenocarcinomas, the computed tomography (CT) findings of ground glass versus solid opacities tend to correspond respectively to lepidic versus invasive patterns seen pathologically. However, this correlation is not absolute; so when CT features suggest nonmucinous AIS, MIA, and lepidic predominant adenocarcinoma, the suspected diagnosis and clinical staging should be regarded as a preliminary assessment that is subject to revision after pathologic evaluation of resected specimens. The ability to predict invasive versus noninvasive size on the basis of solid versus ground glass components is not applicable to mucinous AIS, MIA, or invasive mucinous adenocarcinomas because they generally show solid nodules or consolidation on CT.

A pilot study of the role of TC-99 radionuclide in localization of pulmonary nodular lesions for thoracoscopic resection

OBJECTIVE Video-assisted thoracic surgery (VATS) is an interesting and emerging procedure for diagnosis and treatment of peripheral pulmonary nodules. However, thoracoscopy has limits in the detection of small nodules, below the pleural surface, deep in the lung parenchyma, which cannot be seen as much as palpated. Methods to localize such lesions, including the methylene blue injection or the introduction of a hooked-wire under the radiological vision, have some advantages but a lot of limitations. We are developing a new technique for the detection of pulmonary nodules smaller than 2 cm, deep in the lung parenchyma. METHODS The technique consisted of a intra-lesional injection of 0.3 ml of solution of 99m Tc-labelled human serum albumin microspheres (5-10 MBq) under the CT-scan guide, 2 h before surgery. During thoracoscopy a 11 mm diameter-collimated probe connected to a gamma ray detector (Scinti Probe MR 100 - Pol. hi.tech., Aquila - Italy), is introduced by a 11.5 mm trocar and the pleural surface of the suspected area was scanned. A hot-spot indicated the presence of the injected nodule and as a consequence, the area to be resected. RESULTS from June 1997 to June 1999 we treated 39 patients with small pulmonary nodules. The patients were 27 men and 12 women with a mean age of 60.8 years (range: 13-80). In 19 cases the anamnesis was positive for synchronous or metachronous malignant neoplasm. The mean surgical procedure length was 50 min (range 20-100 min). In all the cases the nodule was resected and the resection margins were pathologically free of tumour. The mean post-operative hospital stay was 3 days (range 2-6 days). Histological examination showed 21 benign lesions and 18 malignant lesions (seven metastases and 11 primary lung cancers). Nine pts with primary lung carcinoma underwent a completion lobectomy by open surgery. CONCLUSIONS Radiolocalization by gamma-probe allows the detection and exeresis of small nodules in a easy and safe way. Future and predictable advances in radio-marked monoclonal antibodies, as well as in the development of endoscopic beta-detector probe, will offer a more effective method for detection of primary and metastatic tumours, targets of thoracoscopic resections.