Wainer Zoli - Academia.edu (original) (raw)

Papers by Wainer Zoli

Cancer Letters, 2013

The characterization of circulating tumor cells (CTCs) could substantially improve the management... more The characterization of circulating tumor cells (CTCs) could substantially improve the management of cancer patients. However, their study is still a matter of debate, often due to lymphocyte contamination. In the present paper, an investigation of CTCs was carried out for the first time using DEPArray, a dielectrophoresis-based platform able to detect and sort pure CTCs. Analyses were conducted on peripheral blood (PB) samples from patients with metastatic colon cancer. After 100% pure cell recovery and whole genome amplification, KRAS gene mutation of CTCs was screened and compared to gene status in the primary tumor tissue. CTCs were found in 21 colon cancer patients (52.5%), with more than three tumor cells per 7.5 ml. KRAS gene mutation analysis, showed a mutational concordance between CTCs and primary tumor in 50% of matched cases. The present study demonstrates for the first time the feasibility of analyzing at the molecular level pure CTCs avoiding lymphocyte contamination using an innovative instrumentation, and a KRAS discordance between CTCs and primary tissue. Our results present dielectrophoresis-based procedures as a new standard in single cell analysis and recovery and invite careful reflection on the value of CTCs characterization.

Journal of Cellular and Molecular Medicine, 2011

Gastric and oesophageal cancers are among the most common tumours in the world, representing the ... more Gastric and oesophageal cancers are among the most common tumours in the world, representing the second leading cause of cancer mortality. Furthermore, gastric tumours are more frequently diagnosed at an advanced, unresectable stage . Surgical resection is currently the mainstay of treatment and can cure patients with early-stage cancer. Conversely, very few improvements in treatment efficacy have been made over time for those with advanced disease [2]. However, multimodal strategies integrating pre-and post-operative treatments have clearly improved gastric cancer prognosis when used in association with curative intent surgery. In particular, the benefit of perioperative chemotherapy for patients with gastric and gastro-oesophageal cancers was confirmed in the two large phase III . The identification of more effective chemotherapy regimens and of the optimal treatment schedule in a neoadjuvant setting, aimed at obtaining a volumetric reduction and thus increasing the number of patients with locally advanced resectable disease, remain important goals. Interesting results in this area have been obtained from the use of multiple targeted therapies in gastric cancer. Different classes of new agents directed against specific targets, including monoclonal antibodies [7][8][9][10][11][12], tyrosine kinase inhibitors [13], anti-angiogenic compounds [14, 15] and the proteasome inhibitor bortezomib [16] have shown promising activity in clinical studies on advanced gastric cancer. In particular, preliminary phase II trials have produced encouraging Low-dose taxotere enhances the ability of sorafenib to induce apoptosis in gastric cancer models

Tumori

Preclinical studies have shown that several chemotherapeutic agents at low doses may affect the v... more Preclinical studies have shown that several chemotherapeutic agents at low doses may affect the vascular system. Here we report the case of a patient with long-term cancer control by metronomic chemotherapy. A 62-year-old woman with breast cancer underwent a left mastectomy in July 2007. For a liver metastasis she was given first-line chemotherapy with doxorubicin plus paclitaxel every 21 days. A CT scan after the sixth cycle showed a partial response. It was decided to stop the treatment with doxorubicin and paclitaxel, and start metronomic therapy with cyclophosphamide 50 mg daily orally and methotrexate 2.5 mg twice daily, 2 days a week. After 6 months of this maintenance treatment, CT scan showed a complete response. We examined the expression of vascular endothelial growth factor receptor 2 (VEGFR2) in histological sections of the primary tumor of our patient, finding evidence of overexpression of the receptor. The metronomic treatment is still ongoing, and after 60 months the ...

Oncology letters, 2015

Targeted therapy of metastatic colorectal cancer (mCRC) with monoclonal antibody anti-epidermal g... more Targeted therapy of metastatic colorectal cancer (mCRC) with monoclonal antibody anti-epidermal growth factor receptor (EGFR) agents, such as cetuximab (CTX) or panitumumab, is the treatment strategy of choice in patients characterised by a wild-type (wt) RAS gene status. However, despite selection based on RAS status, a high proportion of patients do not respond to therapy. EGFR methylation has been reported to have a role in predicting the response to anti-EGFR agents. The present study aimed to evaluate the role of EGFR methylation in association with the clinical outcome of patients with mCRC treated with CTX. In total, 64 patients with mCRC were assessed in the present study. Genomic DNA was extracted from tumoral tissue and EGFR methylation and mutation of the KRAS, BRAF and PIK3CA genes were analysed by pyrosequencing. EGFR expression was assessed by immunohistochemistry. The various alterations were analysed by assessing the objective response rate (ORR), progression free su...

Annals of translational medicine, 2014

Until now detection and numeration of circulating tumor cells (CTCs) were essentially used as a p... more Until now detection and numeration of circulating tumor cells (CTCs) were essentially used as a prognostic factor in cancer progression. To extend the role of these kinds of analysis, it seems necessary to improve analytical methods related to isolation and characterization of CTCs. Discrepancies between published results corroborates this requirement. In this review we suggest a combination of markers able to reach the goal. Moreover to improve the clinical utility of CTC analysis, particularly in the therapeutic follow up of the disease, epithelial mesenchymal transition (EMT) level of a global CTC population should be studied.

Cancer Epidemiology Biomarkers & Prevention, 2014

Background: DNA integrity analysis could represent an alternative approach to the early detection... more Background: DNA integrity analysis could represent an alternative approach to the early detection of colorectal cancer. Previously, fluorescence long DNA (FL-DNA) in stools was extracted using a manual approach and analyzed by capillary electrophoresis assay (CE FL-DNA). We aimed to improve diagnostic accuracy using a simpler and more standardized method [Real Time PCR FL-DNA (RT FL-DNA)] for the detection of early malignant lesions in a population undergoing colorectal cancer screening.

Journal of Nucleic Acids Investigation, 2010

... In addition, miR-128b seems to be directly implicated in EGFR regulation. In particular,miR-1... more ... In addition, miR-128b seems to be directly implicated in EGFR regulation. In particular,miR-128b loss of heterozygosity is frequently found in tumors and correlates significantly with clinical response and survival following gefitinib ...

Two human cancer cell lines were established from metastatic lesions of an adenocarcinoma (RAL) a... more Two human cancer cell lines were established from metastatic lesions of an adenocarcinoma (RAL) and a squamous cell (CAEP) carcinoma of the lung. The clinical histories of the patients from whom the cell lines were derived are reported. The lines were maintained in continuous culture with doubling times of 65 (RAL) and 50 (CAEP) hours. The RAL and CAEP cell lines, whose morphology and ultrastructural features are presented, showed extensively rearranged karyotypes with modal number of 85 (RAL) and 98 (CAEP). In particular, chromosome 2 pentasomy and several clonal markers were evident in the RAL cells, whereas a telomeric deletion of chromosome 1, del (1)(q32), was observed in the CAEP cells. The morphologic data were confirmed by high expression of specific antigens for each histotype. A marked positivity of the neuron-specific enolase (NSE) levels was evident by immunoenzymatic assays in the cell lines cytosol with respect to those present in the respective patient's sera. No amplification or rearrangements were evident in the CMYC, LMYC, NMYC, INT-2, ERBB2, HRAS, KRAS, MOS, HST-1 genes by Southern blotting analysis in each cell line. Point mutations in exon 1 of KRAS and in exon 7 of TP53 were evident by polymerase chain reaction (PCR)-DNA sequencing in the RAL cell line, whereas no alterations were present in the HRAS and RB genes. The four genes studied did not show point mutations in the CAEP cell line. The RAL cell line was resistant to all the drugs tested, whereas the CAEP cells were sensitive to vinblastine. These cell lines may represent useful experimental models to investigate lung cancer biology and anticancer drug response. © Elsevier Science Inc.

BMC cancer, 2006

Patients with metastatic breast cancer are frequently treated with anthracyclines and taxanes, wh... more Patients with metastatic breast cancer are frequently treated with anthracyclines and taxanes, which are among the most active agents in this disease. Gemcitabine is an interesting candidate for a three-drug combination because of its different mechanism of action and non-overlapping toxicity with respect to the other two drugs. We aimed to evaluate the activity and toxicity of the GAT (gemcitabine, doxorubicin and paclitaxel) regimen, derived from experimental preclinical studies, as first-line chemotherapy in patients with stage IIIB-IV breast cancer. Patients with locally advanced or metastatic breast cancer and at least one bidimensionally measurable lesion were included in the present study. Adequate bone marrow reserve, normal cardiac, hepatic and renal function, and an ECOG performance status of 0 to 2 were required. Only prior adjuvant non anthracycline-based chemotherapy was permitted. Treatment consisted of doxorubicin 50 mg/m2 on day 1, paclitaxel 160 mg/m2 on day 2 and g...

BioMed Research International, 2014

The aim of the present study was to identify sensitive and noninvasive biomarkers of early carcin... more The aim of the present study was to identify sensitive and noninvasive biomarkers of early carcinogenic effect at target organ to use in biomonitoring studies of workers at risk for previous occupational exposure to potential carcinogens. Standard urine cytology (Papanicolaou staining test), comet assay, and quantitative telomerase repeat amplification protocol (TRAP) assay were performed in 159 ex-rubber workers employed in tyres production and 97 unexposed subjects. In TRAP positive cases, a second level analysis using FISH (Urovysion) was done. Cystoscopy results were available for 11 individuals whose 6 FISH/TRAP/comet positive showed in 3 cases a dysplastic condition confirmed by biopsy, 1 comet positive resulted in infiltrating UBC to the biopsy and with hyperplasia and slight dysplasia to the urinary cytology, 1 comet positive resulted in papillary superficial UBC to the biopsy, 1 FISH/TRAP positive showed a normal condition, and 2 TRAP positive showed in one case a phlogosis condition. The results evidenced good concordance of TRAP, comet, and FISH assays as early biomarkers of procarcinogenic effect confirmed by the dysplastic condition and UBC found by cystoscopy-biopsy analysis. The analysis of these markers in urine cells could be potentially more accurate than conventional cytology in monitoring workers exposed to mixture of bladder potential carcinogens.

International Journal of Molecular Sciences, 2013

Patients with solid cancer frequently develop bone metastases (BM). Zoledronic acid (Zometa®, ZA)... more Patients with solid cancer frequently develop bone metastases (BM). Zoledronic acid (Zometa®, ZA), routinely used to treat patients with BM, acts on osteoclasts and also has antitumor properties. We aimed to assess the effect of ZA over time in novel bone turnover markers (RANK/receptor activator of nuclear factor-k B ligand (RANK-L)/ Osteoprotegerin (OPG)) and to correlate these with serum N-terminal telopeptide (NTX). The study prospectively evaluated levels of RANK, RANK-L and OPG transcripts by real-time PCR and NTX expression by ELISA in the peripheral blood of 49 consecutive patients with advanced breast, lung or prostate cancer. All patients received the standard ZA schedule and were monitored for 12 months. Median baseline values of RANK, RANK-L and OPG were 78.28 (range 7.34-620.64), 319.06 (21.42-1884.41) and 1.52 (0.10-58.02), respectively. At 12 months, the median RANK-L value had decreased by 22% with respect to the baseline, whereas median OPG levels had increased by about 96%. Consequently, the RANK-L/OPG ratio decreased by 56% from the baseline. Median serum NTX levels decreased over the 12-month period, reaching statistical significance (p < 0.0001). Our results would seem to indicate that ZA modulates RANK, RANK-L and OPG expression, thus decreasing osteoclast activity.

Bone, 2014

Metastatic bone disease has a major impact on the morbidity and mortality of breast cancer patien... more Metastatic bone disease has a major impact on the morbidity and mortality of breast cancer patients, and studies on bone metastasis biology have led to the development of the most widely used drugs for bone metastases treatment: zoledronate (Zol) and denosumab (Den). The aim of the present study was to assess the effect of soluble mediators produced by breast cancer cells on human osteoclast maturation in a co-culture model. We also tested the ability of zoledronate, denosumab and 5H4, an antibody directed against CSF-1, to interfere with the osteoclastogenic potential of breast cancer. The study was performed on the triple negative cell line MDA-MB-231 and on human osteoclasts obtained from the differentiation of peripheral blood monocytes of a healthy volunteer. Osteoclastogenesis was evaluated by TRAP assay after 14days of differentiation with 10% MDA-MB-231-conditioned media or with CSF-1 and RANKL. Den, Zol and 5H4 were administered after 7days of differentiation. MDA-MB-231-conditioned media doubled the differentiation of monocytes into osteoclasts. MDA-MB-231 secreted CSF-1, especially when cells were cultured to confluence. Induced osteoclasts were sensitive to bone-targeted drugs: Den and 5H4 blocked osteoclast differentiation and survival, while Zol induced osteoclast apoptosis. Osteoclasts differentiated by breast cancer cells were less sensitive to Zol than those induced by differentiation factors, whereas sensitivity to Den was similar. Conversely, breast cancer-induced osteoclast activation resulted in a higher sensitivity to 5H4. A significant increase in CSF-1 secretion was observed in osteoclast precursors after treatment with the highest concentration of Den. Further research is ongoing to evaluate the efficacy of 5H4 combination with Den.

Nanomedicine: Nanotechnology, Biology and Medicine, 2015

The present study deals with the preparation of albumin nanocapsules containing fenretinide and t... more 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 fenretinideloaded 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.

Molecular and Clinical Oncology, 2013

In recent years, a number of new agents that target specific molecular pathways in non-small cell... more In recent years, a number of new agents that target specific molecular pathways in non-small cell lung cancer (NSCLC) have been investigated. Much effort has been focused on identifying specific markers that are predictive of treatment response, given that a tailored approach would maximise the therapeutic index and cost-effectiveness. Gefitinib and erlotinib are selective epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (EGFR-TKIs) and have produced good results in selected cases in terms of objective response rate and overall survival. At present, EGFR gene mutations are considered the most important predictors of clinical response to TKI therapy and tumour characterisation for these alterations is mandatory prior to any decision making. Echinoderm microtubule-like protein 4-anaplastic lymphoma kinase (EML4-ALK) translocation is another alteration capable of predicting the efficacy of anti-ALK agents, such as crizotinib. Moreover, emerging target agents, such as MET inhibitors, are likely to increase the amount of molecular characterisation required before a decision is made on treatment. The main limiting factor for adequate characterisation of metastatic NSCLC patients is the small quantity of tumour cells available for molecular analysis. In this study, we provided an overview of the most important and clinically relevant target agents in NSCLC patients as well as the most important mechanisms of resistance. The issue of the scant amount of biological samples available for analysis as well as alternative sampling approaches such as plasma-or serum-derived DNA were also examined.

Platinum and Other Heavy Metal Compounds in Cancer Chemotherapy, 2009

Lipoplatin is a liposome encapsulated form of cisplatin. Phase I studies on Lipoplatin showed an ... more Lipoplatin is a liposome encapsulated form of cisplatin. Phase I studies on Lipoplatin showed an excellent toxicity profile of the compound. Therefore we performed a phase II trial in heavily pre-treated patients with advanced non–small cell lung cancer (NSCLC). This was an open label single-arm trial in patients with NSCLC with stage IV disease already pre-treated with first line chemotherapy.

Molecules, 2014

Lung cancer is a leading cause of cancer death and late diagnosis is one of the most important re... more Lung cancer is a leading cause of cancer death and late diagnosis is one of the most important reasons for the high mortality rate. Circulating microRNAs (miRNAs) represent stable and reproducible markers for numerous solid tumors, including lung cancer, and have been hypothesized as non-invasive diagnostic markers. Serum, plasma or whole peripheral blood can be used as starting material, and several methodological approaches have been proposed to evaluate miRNA expression. The present review provides an in depth summary of current knowledge on circulating miRNAs in different types of biological samples used as diagnostic markers of lung cancer. We also evaluate the diagnostic accuracy of each miRNA or group of miRNAs in relation to the different housekeeping miRNAs used. Finally, the limitations and potential of miRNA analysis are discussed.

World Journal of Gastroenterology, 2014

Core tip: Although the importance of circulating free DNA is widely recognized, numerous studies ... more Core tip: Although the importance of circulating free DNA is widely recognized, numerous studies evaluating its presence in blood and stool samples have reported analytic variability and non conforming approaches. Nonetheless, circulating free DNA has shown high potential as a biomarker for the early non-invasive detection of cancer and for monitoring disease progression. Population studies are now needed to confirm its usefulness for colorectal cancer diagnosis.

The Prostate, 2009

BACKGROUND. The efficacy of current therapy for hormone-refractory prostate cancer is still unsat... more BACKGROUND. The efficacy of current therapy for hormone-refractory prostate cancer is still unsatisfactory and new agents and therapeutic modalities are needed. The aims of the present work were to examine the in vitro activity and mechanisms of action of different antitumor drug combinations in hormone-resistant prostate cancer (HRPC) cell lines. METHODS. The activity of docetaxel (Doc), cisplatin (Cis), oxaliplatin (Oxa), SN-38 and ST1481, singly or in combination, was assessed in different HRPC cell lines (PC3, parental DU145 and taxane-resistant DU145-R) by SRB test. Apoptosis was evaluated by TUNEL and ANN-V assays. Extrusion pump activity was studied by Hoechst 33342 assay, while gene expression related to drug efflux mechanisms and DNA damage repair was analyzed by RT-PCR. RESULTS. Doc induced a high cytocidal effect in the HRPC cells, whereas Cis, Oxa, SN-38 and ST1481 exerted prevalently cytostatic activity. Doc followed by ST1481 proved to be the most effective drug sequence among those investigated, producing an important synergistic effect (R.I. from 2.0 to 5.2) in all the tested cell lines. Moreover, this sequence induced a significant downregulation of xenobiotic extrusion pump and DNA damage repair gene expression. ST1481 synergistically increased the cytocidal effect of Doc, probably through a downregulation of extrusion pump activity and DNA damage repair-related genes. CONCLUSIONS. Our results show that the Doc ! ST1481 sequence effectively reduces the cancer cell population and restores Doc activity in taxane-resistant HRPC, indicating its potential usefulness as first-or second-line treatment of hormone-refractory prostate cancer.

PLoS ONE, 2014

Anti-EGFR therapy appears to be a potential treatment option for squamous cell anal carcinoma (SC... more Anti-EGFR therapy appears to be a potential treatment option for squamous cell anal carcinoma (SCAC). KRAS mutation is a rare event in SCAC, indicating the absence of the principal mechanism of resistance to this type of therapy. However, no information is available from the literature regarding the status of BRAF or PIK3CA in this cancer type. We analysed KRAS, BRAF and PIK3CA status in SCAC patients in relation to the clinical-pathological characteristics of patients and to the presence of the human papilloma virus (HPV). One hundred and three patients were treated with the Nigro scheme for anal cancer from March 2001 to August 2012. Fifty patients were considered for the study as there was insufficient paraffin-embedded tumour tissue to perform molecular analysis the remaining 53. DNA was extracted from paraffin-embedded sections. KRAS, BRAF and PIK3CA gene status and HPV genotype were evaluated by pyrosequencing. KRAS and BRAF genes were wild-type in all cases. Conversely, PIK3CA gene was found to be mutated in 11 (22%) cases. In particular, 8 mutations occurred in exon 9 and 3 in exon 20 of the PIK3CA gene. These findings suggest that SCAC could potentially respond to an anti-EGFR drug. PIK3CA mutation may be involved in the process of carcinogenesis in some cases of SCAC.

Cancer Letters, 2013

The characterization of circulating tumor cells (CTCs) could substantially improve the management... more The characterization of circulating tumor cells (CTCs) could substantially improve the management of cancer patients. However, their study is still a matter of debate, often due to lymphocyte contamination. In the present paper, an investigation of CTCs was carried out for the first time using DEPArray, a dielectrophoresis-based platform able to detect and sort pure CTCs. Analyses were conducted on peripheral blood (PB) samples from patients with metastatic colon cancer. After 100% pure cell recovery and whole genome amplification, KRAS gene mutation of CTCs was screened and compared to gene status in the primary tumor tissue. CTCs were found in 21 colon cancer patients (52.5%), with more than three tumor cells per 7.5 ml. KRAS gene mutation analysis, showed a mutational concordance between CTCs and primary tumor in 50% of matched cases. The present study demonstrates for the first time the feasibility of analyzing at the molecular level pure CTCs avoiding lymphocyte contamination using an innovative instrumentation, and a KRAS discordance between CTCs and primary tissue. Our results present dielectrophoresis-based procedures as a new standard in single cell analysis and recovery and invite careful reflection on the value of CTCs characterization.

Journal of Cellular and Molecular Medicine, 2011

Gastric and oesophageal cancers are among the most common tumours in the world, representing the ... more Gastric and oesophageal cancers are among the most common tumours in the world, representing the second leading cause of cancer mortality. Furthermore, gastric tumours are more frequently diagnosed at an advanced, unresectable stage . Surgical resection is currently the mainstay of treatment and can cure patients with early-stage cancer. Conversely, very few improvements in treatment efficacy have been made over time for those with advanced disease [2]. However, multimodal strategies integrating pre-and post-operative treatments have clearly improved gastric cancer prognosis when used in association with curative intent surgery. In particular, the benefit of perioperative chemotherapy for patients with gastric and gastro-oesophageal cancers was confirmed in the two large phase III . The identification of more effective chemotherapy regimens and of the optimal treatment schedule in a neoadjuvant setting, aimed at obtaining a volumetric reduction and thus increasing the number of patients with locally advanced resectable disease, remain important goals. Interesting results in this area have been obtained from the use of multiple targeted therapies in gastric cancer. Different classes of new agents directed against specific targets, including monoclonal antibodies [7][8][9][10][11][12], tyrosine kinase inhibitors [13], anti-angiogenic compounds [14, 15] and the proteasome inhibitor bortezomib [16] have shown promising activity in clinical studies on advanced gastric cancer. In particular, preliminary phase II trials have produced encouraging Low-dose taxotere enhances the ability of sorafenib to induce apoptosis in gastric cancer models

Tumori

Preclinical studies have shown that several chemotherapeutic agents at low doses may affect the v... more Preclinical studies have shown that several chemotherapeutic agents at low doses may affect the vascular system. Here we report the case of a patient with long-term cancer control by metronomic chemotherapy. A 62-year-old woman with breast cancer underwent a left mastectomy in July 2007. For a liver metastasis she was given first-line chemotherapy with doxorubicin plus paclitaxel every 21 days. A CT scan after the sixth cycle showed a partial response. It was decided to stop the treatment with doxorubicin and paclitaxel, and start metronomic therapy with cyclophosphamide 50 mg daily orally and methotrexate 2.5 mg twice daily, 2 days a week. After 6 months of this maintenance treatment, CT scan showed a complete response. We examined the expression of vascular endothelial growth factor receptor 2 (VEGFR2) in histological sections of the primary tumor of our patient, finding evidence of overexpression of the receptor. The metronomic treatment is still ongoing, and after 60 months the ...

Oncology letters, 2015

Targeted therapy of metastatic colorectal cancer (mCRC) with monoclonal antibody anti-epidermal g... more Targeted therapy of metastatic colorectal cancer (mCRC) with monoclonal antibody anti-epidermal growth factor receptor (EGFR) agents, such as cetuximab (CTX) or panitumumab, is the treatment strategy of choice in patients characterised by a wild-type (wt) RAS gene status. However, despite selection based on RAS status, a high proportion of patients do not respond to therapy. EGFR methylation has been reported to have a role in predicting the response to anti-EGFR agents. The present study aimed to evaluate the role of EGFR methylation in association with the clinical outcome of patients with mCRC treated with CTX. In total, 64 patients with mCRC were assessed in the present study. Genomic DNA was extracted from tumoral tissue and EGFR methylation and mutation of the KRAS, BRAF and PIK3CA genes were analysed by pyrosequencing. EGFR expression was assessed by immunohistochemistry. The various alterations were analysed by assessing the objective response rate (ORR), progression free su...

Annals of translational medicine, 2014

Until now detection and numeration of circulating tumor cells (CTCs) were essentially used as a p... more Until now detection and numeration of circulating tumor cells (CTCs) were essentially used as a prognostic factor in cancer progression. To extend the role of these kinds of analysis, it seems necessary to improve analytical methods related to isolation and characterization of CTCs. Discrepancies between published results corroborates this requirement. In this review we suggest a combination of markers able to reach the goal. Moreover to improve the clinical utility of CTC analysis, particularly in the therapeutic follow up of the disease, epithelial mesenchymal transition (EMT) level of a global CTC population should be studied.

Cancer Epidemiology Biomarkers & Prevention, 2014

Background: DNA integrity analysis could represent an alternative approach to the early detection... more Background: DNA integrity analysis could represent an alternative approach to the early detection of colorectal cancer. Previously, fluorescence long DNA (FL-DNA) in stools was extracted using a manual approach and analyzed by capillary electrophoresis assay (CE FL-DNA). We aimed to improve diagnostic accuracy using a simpler and more standardized method [Real Time PCR FL-DNA (RT FL-DNA)] for the detection of early malignant lesions in a population undergoing colorectal cancer screening.

Journal of Nucleic Acids Investigation, 2010

... In addition, miR-128b seems to be directly implicated in EGFR regulation. In particular,miR-1... more ... In addition, miR-128b seems to be directly implicated in EGFR regulation. In particular,miR-128b loss of heterozygosity is frequently found in tumors and correlates significantly with clinical response and survival following gefitinib ...

Two human cancer cell lines were established from metastatic lesions of an adenocarcinoma (RAL) a... more Two human cancer cell lines were established from metastatic lesions of an adenocarcinoma (RAL) and a squamous cell (CAEP) carcinoma of the lung. The clinical histories of the patients from whom the cell lines were derived are reported. The lines were maintained in continuous culture with doubling times of 65 (RAL) and 50 (CAEP) hours. The RAL and CAEP cell lines, whose morphology and ultrastructural features are presented, showed extensively rearranged karyotypes with modal number of 85 (RAL) and 98 (CAEP). In particular, chromosome 2 pentasomy and several clonal markers were evident in the RAL cells, whereas a telomeric deletion of chromosome 1, del (1)(q32), was observed in the CAEP cells. The morphologic data were confirmed by high expression of specific antigens for each histotype. A marked positivity of the neuron-specific enolase (NSE) levels was evident by immunoenzymatic assays in the cell lines cytosol with respect to those present in the respective patient's sera. No amplification or rearrangements were evident in the CMYC, LMYC, NMYC, INT-2, ERBB2, HRAS, KRAS, MOS, HST-1 genes by Southern blotting analysis in each cell line. Point mutations in exon 1 of KRAS and in exon 7 of TP53 were evident by polymerase chain reaction (PCR)-DNA sequencing in the RAL cell line, whereas no alterations were present in the HRAS and RB genes. The four genes studied did not show point mutations in the CAEP cell line. The RAL cell line was resistant to all the drugs tested, whereas the CAEP cells were sensitive to vinblastine. These cell lines may represent useful experimental models to investigate lung cancer biology and anticancer drug response. © Elsevier Science Inc.

BMC cancer, 2006

Patients with metastatic breast cancer are frequently treated with anthracyclines and taxanes, wh... more Patients with metastatic breast cancer are frequently treated with anthracyclines and taxanes, which are among the most active agents in this disease. Gemcitabine is an interesting candidate for a three-drug combination because of its different mechanism of action and non-overlapping toxicity with respect to the other two drugs. We aimed to evaluate the activity and toxicity of the GAT (gemcitabine, doxorubicin and paclitaxel) regimen, derived from experimental preclinical studies, as first-line chemotherapy in patients with stage IIIB-IV breast cancer. Patients with locally advanced or metastatic breast cancer and at least one bidimensionally measurable lesion were included in the present study. Adequate bone marrow reserve, normal cardiac, hepatic and renal function, and an ECOG performance status of 0 to 2 were required. Only prior adjuvant non anthracycline-based chemotherapy was permitted. Treatment consisted of doxorubicin 50 mg/m2 on day 1, paclitaxel 160 mg/m2 on day 2 and g...

BioMed Research International, 2014

The aim of the present study was to identify sensitive and noninvasive biomarkers of early carcin... more The aim of the present study was to identify sensitive and noninvasive biomarkers of early carcinogenic effect at target organ to use in biomonitoring studies of workers at risk for previous occupational exposure to potential carcinogens. Standard urine cytology (Papanicolaou staining test), comet assay, and quantitative telomerase repeat amplification protocol (TRAP) assay were performed in 159 ex-rubber workers employed in tyres production and 97 unexposed subjects. In TRAP positive cases, a second level analysis using FISH (Urovysion) was done. Cystoscopy results were available for 11 individuals whose 6 FISH/TRAP/comet positive showed in 3 cases a dysplastic condition confirmed by biopsy, 1 comet positive resulted in infiltrating UBC to the biopsy and with hyperplasia and slight dysplasia to the urinary cytology, 1 comet positive resulted in papillary superficial UBC to the biopsy, 1 FISH/TRAP positive showed a normal condition, and 2 TRAP positive showed in one case a phlogosis condition. The results evidenced good concordance of TRAP, comet, and FISH assays as early biomarkers of procarcinogenic effect confirmed by the dysplastic condition and UBC found by cystoscopy-biopsy analysis. The analysis of these markers in urine cells could be potentially more accurate than conventional cytology in monitoring workers exposed to mixture of bladder potential carcinogens.

International Journal of Molecular Sciences, 2013

Patients with solid cancer frequently develop bone metastases (BM). Zoledronic acid (Zometa®, ZA)... more Patients with solid cancer frequently develop bone metastases (BM). Zoledronic acid (Zometa®, ZA), routinely used to treat patients with BM, acts on osteoclasts and also has antitumor properties. We aimed to assess the effect of ZA over time in novel bone turnover markers (RANK/receptor activator of nuclear factor-k B ligand (RANK-L)/ Osteoprotegerin (OPG)) and to correlate these with serum N-terminal telopeptide (NTX). The study prospectively evaluated levels of RANK, RANK-L and OPG transcripts by real-time PCR and NTX expression by ELISA in the peripheral blood of 49 consecutive patients with advanced breast, lung or prostate cancer. All patients received the standard ZA schedule and were monitored for 12 months. Median baseline values of RANK, RANK-L and OPG were 78.28 (range 7.34-620.64), 319.06 (21.42-1884.41) and 1.52 (0.10-58.02), respectively. At 12 months, the median RANK-L value had decreased by 22% with respect to the baseline, whereas median OPG levels had increased by about 96%. Consequently, the RANK-L/OPG ratio decreased by 56% from the baseline. Median serum NTX levels decreased over the 12-month period, reaching statistical significance (p < 0.0001). Our results would seem to indicate that ZA modulates RANK, RANK-L and OPG expression, thus decreasing osteoclast activity.

Bone, 2014

Metastatic bone disease has a major impact on the morbidity and mortality of breast cancer patien... more Metastatic bone disease has a major impact on the morbidity and mortality of breast cancer patients, and studies on bone metastasis biology have led to the development of the most widely used drugs for bone metastases treatment: zoledronate (Zol) and denosumab (Den). The aim of the present study was to assess the effect of soluble mediators produced by breast cancer cells on human osteoclast maturation in a co-culture model. We also tested the ability of zoledronate, denosumab and 5H4, an antibody directed against CSF-1, to interfere with the osteoclastogenic potential of breast cancer. The study was performed on the triple negative cell line MDA-MB-231 and on human osteoclasts obtained from the differentiation of peripheral blood monocytes of a healthy volunteer. Osteoclastogenesis was evaluated by TRAP assay after 14days of differentiation with 10% MDA-MB-231-conditioned media or with CSF-1 and RANKL. Den, Zol and 5H4 were administered after 7days of differentiation. MDA-MB-231-conditioned media doubled the differentiation of monocytes into osteoclasts. MDA-MB-231 secreted CSF-1, especially when cells were cultured to confluence. Induced osteoclasts were sensitive to bone-targeted drugs: Den and 5H4 blocked osteoclast differentiation and survival, while Zol induced osteoclast apoptosis. Osteoclasts differentiated by breast cancer cells were less sensitive to Zol than those induced by differentiation factors, whereas sensitivity to Den was similar. Conversely, breast cancer-induced osteoclast activation resulted in a higher sensitivity to 5H4. A significant increase in CSF-1 secretion was observed in osteoclast precursors after treatment with the highest concentration of Den. Further research is ongoing to evaluate the efficacy of 5H4 combination with Den.

Nanomedicine: Nanotechnology, Biology and Medicine, 2015

The present study deals with the preparation of albumin nanocapsules containing fenretinide and t... more 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 fenretinideloaded 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.

Molecular and Clinical Oncology, 2013

In recent years, a number of new agents that target specific molecular pathways in non-small cell... more In recent years, a number of new agents that target specific molecular pathways in non-small cell lung cancer (NSCLC) have been investigated. Much effort has been focused on identifying specific markers that are predictive of treatment response, given that a tailored approach would maximise the therapeutic index and cost-effectiveness. Gefitinib and erlotinib are selective epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (EGFR-TKIs) and have produced good results in selected cases in terms of objective response rate and overall survival. At present, EGFR gene mutations are considered the most important predictors of clinical response to TKI therapy and tumour characterisation for these alterations is mandatory prior to any decision making. Echinoderm microtubule-like protein 4-anaplastic lymphoma kinase (EML4-ALK) translocation is another alteration capable of predicting the efficacy of anti-ALK agents, such as crizotinib. Moreover, emerging target agents, such as MET inhibitors, are likely to increase the amount of molecular characterisation required before a decision is made on treatment. The main limiting factor for adequate characterisation of metastatic NSCLC patients is the small quantity of tumour cells available for molecular analysis. In this study, we provided an overview of the most important and clinically relevant target agents in NSCLC patients as well as the most important mechanisms of resistance. The issue of the scant amount of biological samples available for analysis as well as alternative sampling approaches such as plasma-or serum-derived DNA were also examined.

Platinum and Other Heavy Metal Compounds in Cancer Chemotherapy, 2009

Lipoplatin is a liposome encapsulated form of cisplatin. Phase I studies on Lipoplatin showed an ... more Lipoplatin is a liposome encapsulated form of cisplatin. Phase I studies on Lipoplatin showed an excellent toxicity profile of the compound. Therefore we performed a phase II trial in heavily pre-treated patients with advanced non–small cell lung cancer (NSCLC). This was an open label single-arm trial in patients with NSCLC with stage IV disease already pre-treated with first line chemotherapy.

Molecules, 2014

Lung cancer is a leading cause of cancer death and late diagnosis is one of the most important re... more Lung cancer is a leading cause of cancer death and late diagnosis is one of the most important reasons for the high mortality rate. Circulating microRNAs (miRNAs) represent stable and reproducible markers for numerous solid tumors, including lung cancer, and have been hypothesized as non-invasive diagnostic markers. Serum, plasma or whole peripheral blood can be used as starting material, and several methodological approaches have been proposed to evaluate miRNA expression. The present review provides an in depth summary of current knowledge on circulating miRNAs in different types of biological samples used as diagnostic markers of lung cancer. We also evaluate the diagnostic accuracy of each miRNA or group of miRNAs in relation to the different housekeeping miRNAs used. Finally, the limitations and potential of miRNA analysis are discussed.

World Journal of Gastroenterology, 2014

Core tip: Although the importance of circulating free DNA is widely recognized, numerous studies ... more Core tip: Although the importance of circulating free DNA is widely recognized, numerous studies evaluating its presence in blood and stool samples have reported analytic variability and non conforming approaches. Nonetheless, circulating free DNA has shown high potential as a biomarker for the early non-invasive detection of cancer and for monitoring disease progression. Population studies are now needed to confirm its usefulness for colorectal cancer diagnosis.

The Prostate, 2009

BACKGROUND. The efficacy of current therapy for hormone-refractory prostate cancer is still unsat... more BACKGROUND. The efficacy of current therapy for hormone-refractory prostate cancer is still unsatisfactory and new agents and therapeutic modalities are needed. The aims of the present work were to examine the in vitro activity and mechanisms of action of different antitumor drug combinations in hormone-resistant prostate cancer (HRPC) cell lines. METHODS. The activity of docetaxel (Doc), cisplatin (Cis), oxaliplatin (Oxa), SN-38 and ST1481, singly or in combination, was assessed in different HRPC cell lines (PC3, parental DU145 and taxane-resistant DU145-R) by SRB test. Apoptosis was evaluated by TUNEL and ANN-V assays. Extrusion pump activity was studied by Hoechst 33342 assay, while gene expression related to drug efflux mechanisms and DNA damage repair was analyzed by RT-PCR. RESULTS. Doc induced a high cytocidal effect in the HRPC cells, whereas Cis, Oxa, SN-38 and ST1481 exerted prevalently cytostatic activity. Doc followed by ST1481 proved to be the most effective drug sequence among those investigated, producing an important synergistic effect (R.I. from 2.0 to 5.2) in all the tested cell lines. Moreover, this sequence induced a significant downregulation of xenobiotic extrusion pump and DNA damage repair gene expression. ST1481 synergistically increased the cytocidal effect of Doc, probably through a downregulation of extrusion pump activity and DNA damage repair-related genes. CONCLUSIONS. Our results show that the Doc ! ST1481 sequence effectively reduces the cancer cell population and restores Doc activity in taxane-resistant HRPC, indicating its potential usefulness as first-or second-line treatment of hormone-refractory prostate cancer.

PLoS ONE, 2014

Anti-EGFR therapy appears to be a potential treatment option for squamous cell anal carcinoma (SC... more Anti-EGFR therapy appears to be a potential treatment option for squamous cell anal carcinoma (SCAC). KRAS mutation is a rare event in SCAC, indicating the absence of the principal mechanism of resistance to this type of therapy. However, no information is available from the literature regarding the status of BRAF or PIK3CA in this cancer type. We analysed KRAS, BRAF and PIK3CA status in SCAC patients in relation to the clinical-pathological characteristics of patients and to the presence of the human papilloma virus (HPV). One hundred and three patients were treated with the Nigro scheme for anal cancer from March 2001 to August 2012. Fifty patients were considered for the study as there was insufficient paraffin-embedded tumour tissue to perform molecular analysis the remaining 53. DNA was extracted from paraffin-embedded sections. KRAS, BRAF and PIK3CA gene status and HPV genotype were evaluated by pyrosequencing. KRAS and BRAF genes were wild-type in all cases. Conversely, PIK3CA gene was found to be mutated in 11 (22%) cases. In particular, 8 mutations occurred in exon 9 and 3 in exon 20 of the PIK3CA gene. These findings suggest that SCAC could potentially respond to an anti-EGFR drug. PIK3CA mutation may be involved in the process of carcinogenesis in some cases of SCAC.