Vincenzo Canzonieri

Targeted intraoperative radiotherapy impairs the stimulation of breast cancer cell proliferation and invasion caused by surgical wounding

Purpose: After apparently successful excision of breast cancer, risk of local recurrence remains high mainly in the area surrounding the original tumor, indicating that wound healing processes may be implicated. The proportional reduction of this risk by radiotherapy does not depend on the extent of surgery, suggesting that radiotherapy, in addition to killing tumor cells, may influence the tumor microenvironment. Experimental Design: We studied how normal and mammary carcinoma cell growth and motility are affected by surgical wound fluids (WF), collected over 24 h following breast-conserving surgery in 45 patients, 20 of whom had received additional TARGeted Intraoperative radioTherapy (TARGIT), immediately after the surgical excision. The proteomic profile of the WF and their effects on the activation of intracellular signal transduction pathways of breast cancer cells were also analyzed. Results: WF stimulated proliferation, migration, and invasion of breast cancer cell lines. The stimulatory effect was almost completely abrogated when fluids from TARGIT-treated patients were used. These fluids displayed altered expression of several cytokines and failed to properly stimulate the activation of some intracellular signal transduction pathways, when compared with fluids harvested from untreated patients. Conclusions: Delivery of TARGIT to the tumor bed alters the molecular composition and biological activity of surgical WF. This novel antitumoral effect could, at least partially, explain the very low recurrence rates found in a large pilot study using TARGIT. It also opens a novel avenue for identifying new molecular targets and testing novel therapeutic agents.

Stereotyped B-cell receptor is an independent risk factor of chronic lymphocytic leukemia transformation to Richter syndrome.

Purpose: Few biological prognosticators are useful for prediction of Richter syndrome (RS), representing the transformation of chronic lymphocytic leukemia (CLL) to aggressive lymphoma. Stereotyped B-cell receptors (BCR) may have prognostic effect in CLL progression. We tested the prognostic effect of stereotyped BCR for predicting RS transformation. Experimental Design: The prevalence of stereotyped BCR was compared in RS (n = 69) versus nontransformed CLL (n = 714) by a case-control analysis. Subsequently, the effect of stereotyped BCR at CLL diagnosis on risk of RS transformation was actuarially assessed in a consecutive CLL series (n = 753). Results: RS (n = 69) displayed a higher prevalence of stereotyped BCR (P < 0.001) compared with nontransformed CLL. The actuarial risk of RS transformation was significantly higher in CLL carrying stereotyped BCR (P < 0.001). Among BCR subsets most represented in CLL, subset 8 using IGHV4-39/IGHD6-13/IGHJ5 carried the highest risk of RS transformation [hazard ratio (HR), 24.50; P < 0.001]. Multivariate analysis selected stereotyped BCR (HR, 3.33; P = 0.001) and IGHV4-39 usage (HR, 4.03; P = 0.004) as independent predictors of RS transformation. The combination of IGHV4-39 usage and stereotyped BCR in the same patient identified CLL with a very high risk of RS transformation (5-year risk, 68.7%). The risk carried by stereotyped BCR and IGHV4-39 usage was specific for RS transformation and had no effect on CLL progression without transformation. Conclusions: Analysis of BCR features may help identify CLL patients at risk of RS. A close monitoring and a careful biopsy policy may help early recognition of RS in CLL patients using stereotyped BCR, particularly if combined with IGHV4-39.

Genome wide DNA-profiling of marginal zone lymphomas identifies subtype-specific lesions with an impact on the clinical outcome

Marginal zone B-cell lymphomas (MZLs) have been divided into 3 distinct subtypes (extranodal MZLs of mucosa-associated lymphoid tissue [MALT] type, nodal MZLs, and splenic MZLs). Nevertheless, the relationship between the subtypes is still unclear. We performed a comprehensive analysis of genomic DNA copy number changes in a very large series of MZL cases with the aim of addressing this question. Samples from 218 MZL patients (25 nodal, 57 MALT, 134 splenic, and 2 not better specified MZLs) were analyzed with the Affymetrix Human Mapping 250K SNP arrays, and the data combined with matched gene expression in 33 of 218 cases. MALT lymphoma presented significantly more frequently gains at 3p, 6p, 18p, and del(6q23) (TNFAIP3/A20), whereas splenic MZLs was associated with del(7q31), del(8p). Nodal MZLs did not show statistically significant differences compared with MALT lymphoma while lacking the splenic MZLs-related 7q losses. Gains of 3q and 18q were common to all 3 subtypes. del(8p) was often present together with del(17p) (TP53). Although del(17p) did not determine a worse outcome and del(8p) was only of borderline significance, the presence of both deletions had a highly significant negative impact on the outcome of splenic MZLs.

Stathmin activity influences sarcoma cell shape, motility, and metastatic potential

The balanced activity of microtubule-stabilizing and -destabilizing proteins determines the extent of microtubule dynamics, which is implicated in many cellular processes, including adhesion, migration, and morphology. Among the destabilizing proteins, stathmin is overexpressed in different human malignancies and has been recently linked to the regulation of cell motility. The observation that stathmin was overexpressed in human recurrent and metastatic sarcomas prompted us to investigate stathmin contribution to tumor local invasiveness and distant dissemination. We found that stathmin stimulated cell motility in and through the extracellular matrix (ECM) in vitro and increased the metastatic potential of sarcoma cells in vivo. On contact with the ECM, stathmin was negatively regulated by phosphorylation. Accordingly, a less phosphorylable stathmin point mutant impaired ECM-induced microtubule stabilization and conferred a higher invasive potential, inducing a rounded cell shape coupled with amoeboid-like motility in three-dimensional matrices. Our results indicate that stathmin plays a significant role in tumor metastasis formation, a finding that could lead to exploitation of stathmin as a target of new antimetastatic drugs.

A microRNA signature defines chemoresistance in ovarian cancer through modulation of angiogenesis

Epithelial ovarian cancer is the most lethal gynecologic malignancy; it is highly aggressive and causes almost 125,000 deaths yearly. Despite advances in detection and cytotoxic therapies, a low percentage of patients with advanced stage disease survive 5 y after the initial diagnosis. The high mortality of this disease is mainly caused by resistance to the available therapies. Here, we profiled microRNA (miR) expression in serous epithelial ovarian carcinomas to assess the possibility of a miR signature associated with chemoresistance. We analyzed tumor samples from 198 patients (86 patients as a training set and 112 patients as a validation set) for human miRs. A signature of 23 miRs associated with chemoresistance was generated by array analysis in the training set. Quantitative RT-PCR in the validation set confirmed that three miRs (miR-484, -642, and -217) were able to predict chemoresistance of these tumors. Additional analysis of miR-484 revealed that the sensitive phenotype is caused by a modulation of tumor vasculature through the regulation of the VEGFB and VEGFR2 pathways. We present compelling evidence that three miRs can classify the response to chemotherapy of ovarian cancer patients in a large multicenter cohort and that one of these three miRs is involved in the control of tumor angiogenesis, indicating an option in the treatment of these patients. Our results suggest, in fact, that blockage of VEGF through the use of an anti-VEGFA antibody may not be sufficient to improve survival in ovarian cancer patients unless VEGFB signaling is also blocked.

Local excision after preoperative chemoradiotherapy for rectal cancer: results of a multicenter phase II clinical trial.

BACKGROUND: Transanal local excision has been suggested as an attractive approach for patients with rectal cancer who show a major clinical response after preoperative chemoradiotherapy. OBJECTIVE: To evaluate the impact of transanal local excision on the local recurrence of rectal cancer in patients who had a major clinical response after preoperative chemoradiotherapy. DESIGN: Sequential 2-stage phase II study for early efficacy. SETTING: Multicenter study. PATIENTS: Patients with clinical T3 or low-lying T2 rectal adenocarcinoma that showed a major clinical response after a preoperative chemoradiotherapy. Eligible patients underwent a full-thickness transanal local excision. According to their histopathology, the patients staged as ypT0-1 were observed, while the remaining patients were recommended to undergo a subsequent total mesorectal excision. MAIN OUTCOME MEASURES: A local recurrence rate of ⩽5% was set as a successful rate for stopping the trial early after the first stage. RESULTS: The study group included 63 patients. Before chemoradiotherapy, patients were staged as clinical T3 (n = 42) and T2 (n = 21). After the local excision, 43 patients fulfilled the criteria to be observed with no further treatment. Nine of the remaining 20 patients for whom a subsequent total mesorectal excision was recommended refused surgery. Two of these patients who refused surgery had intraluminal local recurrence; both had a ypT2 tumor and underwent salvage surgery. The estimated cumulative 3-year overall survival, disease-free survival and local disease-free survival were 91.5% (95% CI: 75.9–97.2), 91.0% (95% CI: 77.0–96.6) and 96.9% (95% CI: 80.3–99.5), respectively. LIMITATIONS: The time of follow-up is still short and the sample size is limited. CONCLUSIONS: Our data suggest that local excision is a good option for patients with a major clinical response after chemoradiotherapy. A longer period of follow-up is required to confirm these findings.

Establishment of HHV-8-positive and HHV-8-negative lymphoma cell lines from primary lymphomatous effusions.

Primary lymphomatous effusions are represented by cases of non‐Hodgkins lymphoma (NHL) which grow in liquid phase in the serous body cavities in the absence of solid tumour masses. Based on morphologic, immunophenotypic, virologic and genotypic features, primary lymphomatous effusions are distinguished into body cavity–based lymphoma (BCBL), Burkitts lymphoma (BL) and immunoblastic large‐cell lymphoma. The histogenesis and pathogenesis of primary lymphomatous effusions are virtually unclarified. In this study, we have established 2 cell lines (CRO‐AP/1 and CRO‐AP/2) representative of 2 distinct categories of primary lymphomatous effusion, BCBL (CRO‐AP/2) and BL (CRO‐AP/1). Both CRO‐AP/1 and CRO‐AP/2 carry monoclonal re‐arrangements of the immunoglobulin genes which are identical to those of the respective parental tumours. Consistent with its BCBL origin, CRO‐AP/2 is characterised by a non‐B, non‐T phenotype and harbours infection by HHV‐8 (approx. 100 viral copies/cell) and Epstein‐Barr virus. Conversely, CRO‐AP/1 expresses several B cell–associated antigens, lacks HHV‐8 infection and carries the genetic hallmark of BL, i.e., a chromosomal breakpoint of band 8q24. CRO‐AP/1 and CRO‐AP/2 may be valuable for the biologic characterization of primary lymphomatous effusions, particularly since the number of available cell lines derived from such lymphomatous effusions is extremely limited. Int. J. Cancer 73:562–569, 1997.

Solid pseudopapillary tumour of the pancreas.

Solid pseudopapillary tumour (Frantz’s tumour) is a rare benign or low-grade neoplasm of the pancreas with distinct clinicopathological features. The diagnosis may be difficult, but should be considered as a possibility in young women who present with a large abdominal mass involving the pancreas. In this report we describe the clinical and pathological characteristics of these lesions and discuss therapeutic options. A 27-year-old woman was admitted to our hospital in October 1999 with a 3-month history of mild asthenia, weight loss, and a feeling of heaviness localised to the left lumbar region. Routine laboratory analyses were all within the normal range; however, abdominal ultrasound revealed the presence of a mass in the superior abdomen, which was composed of high and low echoic areas. Computed tomography showed the mass to be solid, encapsulated, and about 9 cm in diameter (figure 1). It was composed of high and low density areas and was located close to the splenic hilum—above the pancreatic tail, behind the stomach, and between the spleen and the aorta (Albarran-Chatelin’s anatomosurgical quadrilateral). Nuclear magnetic resonance imaging revealed a posterior anastomosis between the mass drainage vessels and the diaphragmatic veins. The patient underwent a complete surgical resection of the tumour (enucleation) and a resection of the pancreatic tail. The spleen was removed because of vascular damage consisting of large patches of venous stasis and ischaemia which did not resolve over time or after application of warm, moist dressings. The pathological diagnosis was epithelial solid papillary cystic neoplasm with signs of incipient pseudocapsular invasion. Immunohistochemical studies established that the tumour cells were positive for cytokeratins and vimentin, and negative for S100 protein, CD34, chromogranin, neurone-specific enolase (NSE), and glial fibrillary acid protein. The postoperative course was uneventful and the patient was discharged from hospital on day 7 and referred to the Centro di Riferimento Oncologico di Aviano for counselling. No residual tumour or metastases were found with clinical imaging, so no adjuvant therapy was recommended. In November 2001, routine work-up identified no signs of tumour relapse. This case is a typical example of solid pseudopapillary tumour of the pancreas or Frantz’s tumour. The diagnostic imaging techniques used were effective in defining the location of the tumour, although a nonfunctioning neoplasm of the left adrenal gland was initially suspected. In 1959, Frantz 1

Prostate cancer and body size at different ages: an Italian multicentre case–control study

We investigated the influence of anthropometric measures at diagnosis and at different ages on prostate cancer risk using an Italian multicentre case–control study conducted between 1991 and 2002 of 1294 histologically confirmed cases and 1451 controls admitted to the same network of hospitals for acute non-neoplastic conditions. Height, weight, body mass index (BMI), waist-to-hip ratio, lean body mass 1 year before diagnosis/interview were not significantly associated with risk. However, a positive association with high BMI at age 30 years was found (odds ratio=1.2 for BMI⩾24.7 vs <22.7) and: for less differentiated prostate cancer, with BMI 1 year before diagnosis/interview. This study supports possible relationships between high body mass in young adulthood, and a tendency to high weight throughout adult life, and the risk of prostate cancer.

Tumor response is predicted by patient genetic profile in rectal cancer patients treated with neo-adjuvant chemo-radiotherapy

The aim of the study was the identification of a pharmacogenetic profile predictive of the tumor regression grade (TRG), considered as tumor response parameter, after neo-adjuvant treatment in rectal cancer patients. A total of 238 rectal cancer patients treated in a neo-adjuvant setting by a fluoropyrimidines-based chemo-radiotherapy (RT) were genotyped for 25 genetic polymorphisms in 16 genes relevant for treatment-associated pathways. Two polymorphisms were associated with TRG in a multivariate analysis: hOGG1-1245C>G, which can affect radiosensitivity and MTHFR-677C>T, which is involved in fluoropyrimidines action. Patients bearing at least one variant allele had a lower chance to get TRG⩽2 (OR=0.46 95% CI 0.23–0.90, P=0.024; and OR=0.48 95% CI 0.24–0.96, P=0.034; respectively). An association trend was observed for ABCB1-3435C>T, which is responsible for the multi-drug resistance (odds ratio (OR)=1.96, 95% confidence interval (CI) 0.98–3.95, P=0.057). Exploratory classification and regression tree (CART) analysis highlighted high-order gene–gene and gene–environment interactions and a genetic signature associated with differential response, with hOGG1-1245C>G as the most predictive factor. Other significant variables were: ABCB1-3435C>T, MTHFR-677C>T, ERCC1-8092C>A, ABCC2-1249G>A, XRCC1-28152G>A, XRCC3-4541A>G and patients gender. On the basis of CART results, patients were categorized into three groups according to tumor response probability: intermediate and high profiles had a higher probability to get TRG⩽2 as compared with low profiles (OR=4.12 95% CI 1.46–11.65, P<0.001 and OR=12.44, 95% CI 5.52–28.04, P<0.0001, respectively). This study evidences a major role of hOGG1-1245C>G and MTHFR-677C>T polymorphisms in the tumor response of rectal cancer patients treated with chemo-RT in neo-adjuvant setting, and shows the relevance of gene–gene and gene–environment interactions for complex phenotypes as tumor response.