The use of neoplastic donors to increase the donor pool.

The aim of the study was to evaluate the experience of the Centre-Sud Transplant Organization (OCST) area using cadaveric donor with neoplastic diseases to evaluate the possibility of transmission to recipients. From January 1, 2003, to December 31, 2010, the neoplastic risk has been reported to be 5.4% (377/4654 referred donors). In 2003, the number of donors with a tumor and their mean age were respectively: 60 (10.3%) and 59.6 ± 19.9; 2004: 33 (5.2%) and 61.4 ± 15.9; 2005: 32 (6%) and 62.8 ± 15.5; 2006: 46 (7%) and 60.7 ± 19.1; 2007: 51 (7%) and 58.9 ± 16; in 2008: 58 (7%) and 59.7 ± 19.6; 2009: 47 (7%) and 57 ± 26; 2010: 49 (7%) and 64 ± 16. The organ most affected by tumor has been the central nervous system (18%). The tumor was diagnosed before in 325 (86%) cases, versus during organ retrieval in 48 (12.7%) donor operations but before, which four cases (1%) occured after transplantation. According to the histological types and grades, 28 evaluated donors (8.2%) were suitable for transplantation. The histological types were: thyroid carcinoma (n = 3); prostate carcinoma (n = 8), renal clear cell carcinoma (n = 7), oncocytoma (n = 1), meningiomas (n = 2), dermofibrosarcoma (n = 1); verrucous carcinoma of the vulva (n = 1), colon adenocarcinoma (n = 1), grade II astrocytoma (n = 1), adrenal gland tumor (n = 1), gastric GIST (n = 1), oligodendroglioma (n = 1). Forty-five organs were retrieved (22 livers, 19 kidneys, 3 hearts, and 1 pancreas) and transplanted into 44 recipients with 1 liver-kidney combined transplantation. Four recipients died due to causes not related to the tumor. No donor-transmitted tumor was detected among the recipients. Donation is absolutely not indicated in cases of tumors with high metastatic potential and high grades. Performing an accurate evaluation of the donor, taking into account the histological grade, currently can allow, organ retrieval and transplantation with an acceptable risk.

Expanding the criteria of organ procurement from donors with prostate cancer: the application of the new Italian guidelines.

Prostate cancer (CaP) represents the most prevalent malignancy in men more than 60-year-old, posing a problem in organ procurement from elderly subjects. However, most of the currently diagnosed CaP are low-grade and intraprostatic, with low metastatic risk, and there is recent evidence that most patients are overdiagnosed. The Italian National guidelines about organ acceptance from neoplastic donors changed in March 2005, extending the pool of potential candidates with CaP and introducing the function of a second opinion expert. Between 2001 and February 2005, 40 candidate donors with total PSA>/=10 and/or positive digital rectal examination underwent histopathological analysis of the prostate: 15 (37.5%) donors harboured CaP, and 25 (62%) were judged at 'standard risk'. After the introduction of the new guidelines in 2005, the second opinion expert judged at 'standard risk' 48 of 65 donors, while 17 of 65 needed histopathological analysis. Four (6.2%) donors harboured CaP, and 61 (94%) where judged at 'standard risk', with a significant increase of donated and actually transplanted organs. The application of the new guidelines and the introduction of a second opinion expert allowed a significant extension of the 'standard risk' category also to CaP patients, decreasing the histopathological examinations and expanding the donor pool.

Two years' experience of acute rejection monitoring of intestinal transplant recipients by real-time PCR assessment of granzyme B and perforin up-regulation: considerations on diagnostic accuracy.

Granzyme B (GrB) and perforin are promising immunological markers to predict acute rejection of transplanted organs. Based on 2 years of experience with molecular monitoring on peripheral blood samples, we investigated the diagnostic accuracy of GrB/perforin gene up-regulation using real-time polymerase chain reaction (PCR) for prediction of acute cellular rejection (ACR) in intestinal transplantation recipients. Histology used as the reference standard. According to our definition of disease positivity (anything other than ACR score 0), GrB/perforin up-regulation showed 84% specificity but only 49% sensitivity. However, among the 26 false-negatives, 12 (46%) had an ACR score 1, which is indeterminate for rejection and no associated clinical manifestations; a further 10 (39%) had a score of 2 following rejection therapy (a confounder for GrB/perforin analysis). Thus only 4 (15%) false-negatives were actually associated with the onset of robust acute rejection. These data suggest that real-time PCR analysis for GrB/perforin up-regulation might play a role along with clinical criteria for detection of presymptomatic acute rejection episodes in intestinal recipients who require immediate endoscopy and pathological examination, especially during long-term follow-up.

11C-choline positron emission tomography/computerized tomography for tumor localization of primary prostate cancer in comparison with 12-core biopsy.

PURPOSE: (11)C-choline positron emission tomography is an innovative imaging technique for prostate cancer. We assessed the sensitivity of positron emission tomography used together with computerized tomography for intraprostatic localization of primary prostate cancer on a nodule-by-nodule basis, and compared its performance with 12-core transrectal biopsy. MATERIALS AND METHODS: In 43 patients with known prostate cancer who had received positron emission tomography/computerized tomography before initial biopsy, we assessed sensitivity of positron emission tomography/computerized tomography for localization of nodules 5 mm or greater (those theoretically large enough for visualization) using radical prostatectomy histopathology as the reference standard. Comparison with transrectal ultrasound guided biopsy was based on sextant assessment of all cancer foci following sextant-by-sextant matching and reconstruction. Sensitivity/specificity of positron emission tomography/computerized tomography and magnetic resonance imaging for prediction of extraprostatic extension was also assessed. RESULTS: Positron emission tomography/computerized tomography showed 83% sensitivity for localization of nodules 5 mm or greater. At logistic regression analysis only nodule size appeared to influence sensitivity. At sextant assessment positron emission tomography/computerized tomography had slightly better sensitivity than transrectal ultrasound guided biopsy (66% vs 61%, p = 0.434) but was less specific (84% vs 97%, p = 0.008). For assessment of extraprostatic extension, sensitivity of PET/CT was low in comparison with magnetic resonance imaging (22% vs 63%, p <0.001). CONCLUSIONS: Positron emission tomography/computerized tomography has good sensitivity for intraprostatic localization of primary prostate cancer nodules 5 mm or greater. Positron emission tomography/computerized tomography and transrectal ultrasound guided biopsy show similar sensitivity for localization of any cancer focus. Positron emission tomography/computerized tomography does not seem to have any role in extraprostatic extension detection. Studies of diagnostic accuracy (as opposed to tumor localization) are needed in patients with suspected prostate cancer to see whether positron emission tomography/computerized tomography could have a role in not selected patients.

Toward the definition of immunosuppressive regimens with antitumor activity.

Immunosuppressive therapies associated with organ transplantation produce an increased risk of cancer development. Malignancies are increased in transplant recipients because of the impaired immune system. Moreover, experimental data point to a tumor-promoting activity of various immunosuppressive agents. In this study, we compared the effects of 4 immunosuppressive agents with different mechanisms of action (cyclosporine, rapamycin, mycophenolic acid, and leflunomide) on the in vitro growth of various tumor cell lines and umbilical vein endothelial cells. To varying degrees rapamycin (10 ng/mL), mycophenolic acid (300 nmol/L), and leflunomide (30 micromol/L) highly inhibited the growth of human rhabdomyosarcoma, hepatocellular carcinoma, colorectal carcinoma, and endothelial cells. In contrast, cyclosporine (100 ng/mL) did not affect their growth. Our data suggest that regimens containing rapamycin, mycophenolic acid, or leflunomide, which have both immunosuppressive and antitumor activities, should be preferred in transplant recipients to minimize the risk of tumors.

Expression of connective tissue growth factor is a prognostic marker for patients with intrahepatic cholangiocarcinoma.

BACKGROUND AND AIMS: Connective tissue growth factor is a member of the 'CCN' protein family. Consistent with its profibrotic properties, it is over-expressed in several human epithelial malignancies. PATIENTS AND METHODS: We have retrospectively evaluated by immunohistochemistry the presence of connective tissue growth factor in archival tissues from 55 resected intrahepatic cholangiocarcinomas and compared its expression to the main pathological parameters, disease free and overall survival. RESULTS: Tumours were scored as high and low/absent expressers (> or =50%, 0-50% cells, respectively). Thirty-three of 55 cholangiocarcinomas (60%) were high and 22 (40%) low expressers. No significant correlation was found between connective tissue growth factor and tumour grade, tumour location, vascular and perineural invasion. Eighteen of 22 (82%) low/absent expressers and 12/33 (36%) high expressers had recurrence of disease (P=0.001). Low/absent expressers showed a poor disease free and overall survival compared with the higher expressers (P<0.001). Vascular invasion was related to tumour recurrence (P=0.025) and to decreased disease free survival (P<0.05). During proportional hazard regression analysis, only connective tissue growth factor was found to influence disease free survival (P=0.01). CONCLUSIONS: Expression of connective tissue growth factor is an independent prognostic indicator of both tumour recurrence and overall survival for intrahepatic cholangiocarcinoma patients regardless of tumour location, tumour grade, vascular and perineural invasion.

Potential of real-time PCR assessment of granzyme B and perforin up-regulation for rejection monitoring in intestinal transplant recipients.

Granzyme B (GrB) and perforin are promising markers to predict acute rejection episodes of transplanted organs. Having recently reported that immunohistochemical expression of GrB/perforin correlates with histologically assessed acute cellular rejection (ACR) episodes in intestinal transplantation recipients, herein we have additionally explored the potential of real-time polymerase chain reaction (PCR) assessment of GrB/perforin gene up-regulation in peripheral blood mononuclear cells. Both immunohistochemical evaluation of GrB/perforin expression and real-time PCR assessment of up-regulation, which was defined as a 2-fold increase with respect to "basal" levels during maintenance immunosuppressive protocols, were performed among a population of 23 intestinal transplant recipients under routine surveillance, in addition to histological analysis of ACR. The ACR scores showed direct relationships both with GrB/perforin immunohistochemistry (IHC) scores (P < .001) and with gene up-regulation by real-time PCR (P = .004). Furthermore, real-time PCR upregulation was associated with the IHC score (P < .001). A preliminary analysis of diagnostic accuracy-performed to gain information to plan future studies-indicated that when using histological assessment as the reference technique, our current definition of PCR up-regulation provided good specificity (84%) but insufficient sensitivity (44%) for a noninvasive prediction of ACR. The results of this pilot study suggested that real-time PCR analysis of GrB/perforin upregulation may help therapeutic decision making, and have the potential for detection of presymptomatic rejection. More extensive studies must investigate strategies to improve the sensitivity of the analyses of GrB/perforin up-regulation.

Molecular monitoring of organ recipients from cancer-affected donors by detection of circulating tumor cells.

We have initiated regular molecular monitoring based on nested RT-PCR detection of circulating tumor cells for monitoring recipients of organs from cancer-affected donors in Italy (in the context of a "Donation Safety and Donated Organ Quality" project organized by the Centro Nazionale Trapianti). Five patients are being monitored. For two patients who each received a kidney from a single donor with prostate adenocarcinoma, RT-PCR was performed using PSA mRNA. For three recipients of organs (two livers and one kidney) from donors with renal cell carcinoma, RT-PCR was performed using cytokeratine 18 and 19 mRNA. Blood samples from five healthy subjects were used as negative controls. After a median monitoring time of 26 months (range 8 to 32), none of the regular 3-month assays has tested positive. This pilot study suggests that detection of circulating tumor cells by nested RT-PCR may provide a feasible molecular monitoring, which might assist decision making regarding other forms of clinical surveillance.

Endoscopic finding of granular cell tumour associated with leiomyomas in the oesophagus.

Following a single report in the literature of granular cell tumour associated with diffuse leiomyomatosis in the oesophagus, we describe the case of a 39-year-old man in whom a granular cell tumour and two leiomyomas were endoscopically removed from this site. This previously unreported association of granular cell tumour with isolated leiomyomas suggests the need to bear in mind the possibility of other mesenchymal lesions, including leiomyomas or leiomyomatosis, when a granular cell tumour is found in the oesophagus.

Granzyme B and perforin as predictive markers for acute rejection in human intestinal transplantation.

In human heart and kidney transplantations, granzyme B (GrB) and perforin have both been shown to be predictive markers for acute cellular rejection (ACR). We investigated the tissue expression and possible relationship of GrB and perforin to the clinical outcome, histopathology, and function of intestinal transplants. In 13 consecutive patients undergoing small intestine transplantation, histologic/immunohistochemical rejection monitoring was performed together with GrB and perforin immunostaining (score "0", 0%-10% positive lymphocytes; "1", 10%-25%; "2", 25%-50%; "3", >50%). Eleven patients are currently alive and well. All 11 had at least one episode of ACR: one patient had 6 episodes of severe ACR requiring retransplantation; the remaining 10 experienced only mild or moderate rejection. Both GrB and perforin were always co-expressed. A highly significant correlation was observed between GrB/perforin scores and histological severity of ACR (Pearson's coefficient, R < 0.0009). Interestingly, score 3 GrB/perforin immunostaining was recorded only in the context of severe ACR; all the histologically negative or "indeterminate" biopsies (n = 6) taken from a single affected patient showed GrB/perforin scores of 1 or 2. By contrast, none of the other tested histologically negative/"indeterminate" biopsies (n = 350), including those performed during graft stabilization, had raised GrB or perforin scores. We conclude that in intestinal transplantation recipients, a direct correlation seems to exist between histologically confirmed ACR and raised GrB/perforin immunohistochemical scores. Our findings suggest the need to investigate the possibility of predicting ACR by routine serum polymerase chain reaction (PCR) monitoring, which would reduce discomfort to patients.

Investigation of ErbB1 and ErbB2 expression for therapeutic targeting in primary liver tumours.

BACKGROUND: Molecular targets are needed for primary liver tumours. AIMS: ErbB1 and ErbB2 expression was analysed in neoplastic and surrounding tissue in surgical specimens from 52 hepatocellular carcinomas and 48 intrahepatic cholangiocarcinomas, randomly chosen from cases surgically treated in this institution. METHODS: ErbB1 and ErbB2 expression were evaluated immunohistochemically, the latter by Herceptest. Gene amplification of ErbB2 was tested by chromogenic in situ hybridisation. RESULTS: In normal/cirrhotic non-neoplastic tissue, the ErbB1 (but not ErbB2) antibody commonly stained normal hepatocytes and mature intrahepatic ducts. In neoplastic tissue, moderate/strong ErbB1 immunostaining occurred in 43/52 (85%) hepatocellular carcinomas and 39/48 (81%) intra-hepatic cholangiocarcinomas. With ErbB2 Herceptest, 0/52 (0%) hepatocellular carcinomas and 2/48 (4%) intra-hepatic cholangiocarcinomas had treatable scores of 2+/3+ (chromogenic in situ hybridisation confirmed gene amplification in the latter two cases only). Neither ErbB1 nor ErbB2 expression correlated with any of the main clinical-pathologic features or survival. CONCLUSIONS: Although not related to prognosis, ErbB1 could be a molecular target in a large percentage of patients with hepatocellular carcinoma or intrahepatic cholangiocarcinoma. Inclusion of anti-ErbB1 drugs such as ZD 1839 and c225 (and possibly also anti-ErbB2 drugs like Trastuzumab for a small subset of patients) in clinical trials is suggested.

Ovarian tissue banking and fertility preservation in cancer patients: histological and immunohistochemical evaluation.

OBJECTIVE: A combination of chemotherapy and radiotherapy in young females with cancer has greatly enhanced the life expectancy of these patients, even if these treatments have a highly deleterious effect on the ovary and cause a severe depletion of the follicular store. Cryopreservation of ovarian tissue before chemotherapy and/or radiotherapy, followed by autograft after remission or in in vitro maturation, could restore gonadal function and fertility. The aim of this study is to verify the efficiency of the ovarian tissue cryopreservation procedure by histological and immunohistochemical analyses. METHODS: Ovarian tissue was obtained by laparoscopy from 22 patients affected with different malignant diseases. Tissue specimens were frozen using a combination of PROH (1,2-propanediol) and sucrose as cryoprotectants, and the cryopreservation protocol used consisted of a slow freezing-rapid thawing program. Both fresh and frozen/thawed tissues were embedded in paraffin blocks for histological and immunohistochemical analyses. RESULTS: Good stromal and follicular morphology was found in fresh and frozen/thawed tissue. No significant differences were found in follicular density, distribution, and diameters in fresh and frozen/thawed tissue. Follicle immunohistochemical analysis showed a high percentage of negative staining for both estrogen receptor (ER) (100% both in fresh and frozen/thawed specimens) and progesterone receptor (PR) (97% versus 91%, respectively). Regarding the Ki67 protein, positive staining was found in both the granulosa cells and/or the oocytes (36% in fresh and 56% in frozen/thawed). For the Bcl2 protein, positive staining was observed in the follicle granulosa cells but not in the oocytes in 74% of the fresh and in 79% of the frozen/thawed specimens. For the stromal cells, ER showed a negative staining distribution in 97% of the fresh and 100% of the frozen/thawed specimens. The stroma staining distribution was diffuse/focal in fresh versus frozen/thawed specimens (50% versus 74% respectively) for PR, patch/focal (70% versus 80%, respectively) for Ki67 protein, and diffuse (55% versus 54%, respectively) for Bcl2. CONCLUSIONS: These results suggest that human ovarian tissue morphology, antigenicity, cellular proliferation, and anti-apoptotic index were well preserved by cryopreservation in PROH and sucrose.

Low p27 expression is an independent predictor of survival for patients with either hilar or peripheral intrahepatic cholangiocarcinoma.

PURPOSE AND EXPERIMENTAL DESIGN: The prognosis for intrahepatic cholangiocarcinoma (ICC) depends mainly on the feasibility of complete surgical resection. In the absence of demonstrated biological predictors of survival, we evaluated the prognostic value of the cyclin-dependent kinase inhibitor p27 by immunohistochemistry in a series of routine specimens from 47 ICC patients, 22 with the hilar and 25 with the peripheral subtype. Proliferation rate was also evaluated in the same cases by the MIB1 index. Tumors were scored as high, low, and negative p27 expressers (> or =50%, <50%, and no positive nuclei, respectively). RESULTS: High, low, and negative p27 expression was recorded in 18 (38%), 17 (36%), and 12 (26%) cases, respectively. No significant correlation was found between p27 expression and gender, age, tumor grade, tumor location, vascular or perineural invasion, or proliferative index. Tumors with low or absent p27 expression were associated with poor survival compared with the high-expresser group. Kaplan-Meier curves comparing different p27 expression levels with survival showed highly significant separation (P < 0.0001, log-rank test). With univariate Cox proportional hazard regression, only p27 score among all of the parameters was found to influence survival (P = 0.0003). CONCLUSION: We conclude that in ICC, low or absent p27 expression can predict poor survival, regardless of tumor location, pathological features, and tumor proliferation. Immunohistochemical detection of p27 on routine sections may provide the first biological prognostic marker for ICC, thus influencing the therapeutic strategies for these patients.

Expression of Epstein-Barr virus-encoded RNA and biological markers in Italian nasopharingeal carcinomas.

Nasopharingeal carcinoma (NPC) is frequently associated with Epstein-Barr virus (EBV), but little is known about its pathogenesis in Western countries, where its incidence is low and EBV is not endemic. We studied 26 Italian cases of NPC: 24 of the non-keratinizing subtype, and 2 keratinizing. We used immunohistochemistry to evaluate p53, bcl-2 and Ki-67 protein expression, and in situ hybridization (ISH) to assess bcl-2 mRNA and the EBV transcript, EBER1, with bcl-2 protein and mRNA expression being compared in serial sections. As regards bcl-2, the protein or mRNA was expressed in 15/26 (58%) and 17 (65%) cases respectively, with co-expression being found only in 9 (35%) cases; 23 (88%) cases were positive for one or the other. EBER1 was detected in 17/26 (65%) cases, of which 80% and 71% coexpressed bcl-2 protein or mRNA, respectively. Only 9/26 (35%) cases had a high Ki67 proliferative index; 3/26 (12%) overexpressed p53. This study of NPC in Italy confirms the importance of EBV and bcl-2 in Western cases. The discrepancy between bcl-2 mRNA and protein expression invites investigation into a possible internal bcl-2 regulation system.

Surgical therapy for adenocarcinoma of the cardia: modalities of recurrence and extension of resection.

In order to define the optimal extent of resection for cancer of the cardia, we considered 116 patients operated upon with five different surgical techniques. The procedures were: transabdominal total gastrectomy associated with distal esophagectomy in 38 patients; transabdominal total gastrectomy and left thoracotomic esophageal resection at the inferior pulmonary vein level in 26 patients; transabdominal total gastrectomy and right thoracotomic esophageal resection at the azygos vein level in 27 patients; transabdominal total gastrectomy and transhiatal lower third esophagectomy in 18 patients; transhiatal total esophagectomy and upper third gastrectomy with cervical esophago-gastroplasty in seven patients. Grading, staging, neoplastic lymphangitis, satellite intramural metastases, infiltration of the resection margin, site of recurrence, and survival were analyzed. N+ was the single independent prognostic factor for survival. A poorly differentiated grading was related to T (P = 0.0009), N (P = 0.001), satellite growth (P = 0.05), and infiltration of the resection margin (P = 0.0001). Recurrence was local in 26% and distant in 74% of patients. The modalities of recurrence were not related to the aggressiveness parameters and the surgical technique. Infiltration of the esophageal resection margin was related to the type of operation (P = 0.005) and survival (P = 0.02), but it was not related to the site of recurrence. Transabdominal total gastrectomy and the right thoracotomic esophageal resection procedure achieved free margins and control of the lymph nodal metastatic spread. Transabdominal total gastrectomy and right thoracotomic esophageal resection at the azygos vein level provides a radical oncologic resection, particularly in poorly differentiated tumors. However, surgery alone cannot cure the majority of adenocarcinomas of the cardia.