Safety and immunogenicity of neoadjuvant treatment using WT1-immunotherapeutic in combination with standard therapy in patients with WT1-positive Stage II/III breast cancer: a randomized Phase I study.

PURPOSE: This Phase I, multicenter, randomized study (ClinicalTrials.gov NCT01220128) evaluated the safety and immunogenicity of recombinant Wilms' tumor 1 (WT1) protein combined with the immunostimulant AS15 (WT1-immunotherapeutic) as neoadjuvant therapy administered concurrently with standard treatments in WT1-positive breast cancer patients. METHODS: Patients were treated in 4 cohorts according to neoadjuvant treatment (A: post-menopausal, hormone receptor [HR]-positive patients receiving aromatase inhibitors; B: patients receiving chemotherapy; C: HER2-overexpressing patients on trastuzumab-chemotherapy combination; D: HR-positive/HER2-negative patients on chemotherapy). Patients (cohorts A-C) were randomized (2:1) to receive 6 or 8 doses of WT1-immunotherapeutic or placebo together with standard neoadjuvant treatment in a double-blind manner; cohort D patients received WT1-immunotherapeutic in an open manner. Safety was assessed throughout the study. WT1-specific antibodies were assessed pre- and post-vaccination. RESULTS: Sixty-two patients were randomized; 60 received ≥ one dose of WT1-immunotherapeutic. Two severe toxicities were reported: diarrhea (cohort C; also reported as a grade 3 serious adverse event) and decreased left ventricular ejection fraction (cohort B; also reported as a grade 2 adverse event). Post-dose 4 of WT1-immunotherapeutic, 10/10 patients from cohort A, 0/8 patients from cohort B, 6/11 patients from cohort C, and 2/3 patients from cohort D were humoral responders. The sponsor elected to close the trial prematurely. CONCLUSIONS: Concurrent administration of WT1-immunotherapeutic and standard neoadjuvant therapy was well tolerated and induced WT1-specific antibodies in patients receiving neoadjuvant aromatase inhibitors. In patients on neoadjuvant chemotherapy or trastuzumab-chemotherapy combination, the humoral response was impaired or blunted, likely due to either co-administration of corticosteroids and/or the chemotherapies themselves.

Safety and immunogenicity of the PRAME cancer immunotherapeutic in metastatic melanoma: results of a phase I dose escalation study.

PURPOSE: The PRAME tumour antigen is expressed in several tumour types but in few normal adult tissues. A dose-escalation phase I/II study (NCT01149343) assessed the safety, immunogenicity and clinical activity of the PRAME immunotherapeutic (recombinant PRAME protein (recPRAME) with the AS15 immunostimulant) in patients with advanced melanoma. Here, we report the phase I dose-escalation study segment. PATIENTS AND METHODS: Patients with stage IV PRAME-positive melanoma were enrolled to 3 consecutive cohorts to receive up to 24 intramuscular injections of the PRAME immunotherapeutic. The RecPRAME dose was 20, 100 or 500 µg in cohorts 1, 2 and 3, respectively, with a fixed dose of AS15. Adverse events (AEs), including predefined dose-limiting toxicity (DLT) and the anti-PRAME humoral response (ELISA), were coprimary end points. Cellular immune responses were evaluated using in vitro assays. RESULTS: 66 patients were treated (20, 24 and 22 in the respective cohorts). AEs considered by the investigator to be causally related were mostly grade 1 or 2 injection site symptoms, fatigue, chills, fever and headache. Two DLTs (grade 3 brain oedema and proteinuria) were recorded in two patients in two cohorts (cohorts 2 and 3). All patients had detectable anti-PRAME antibodies after four immunisations. Percentages of patients with predefined PRAME-specific-CD4+T-cell responses after four immunisations were similar in each cohort. No CD8+ T-cell responses were detected. CONCLUSIONS: The PRAME immunotherapeutic had an acceptable safety profile and induced similar anti-PRAME-specific humoral and cellular immune responses in all cohorts. As per protocol, the phase II study segment was initiated to further evaluate the 500 µg PRAME immunotherapeutic dose. TRIAL REGISTRATION NUMBER: NCT01149343, Results.

Anti-PSMA antibody-coupled gold nanorods detection by optical and electron microscopies.

While cancer is one of the greatest challenges to public health care, prostate cancer was chosen as cancer model to develop a more accurate imaging assessment than those currently available. Indeed, an efficient imaging technique which considerably improves the sensitivity and specificity of the diagnostic and predicting the cancer behavior would be extremely valuable. The concept of optoacoustic imaging using home-made functionalized gold nanoparticles coupled to an antibody targeting PSMA (prostate specific membrane antigen) was evaluated on different cancer cell lines to demonstrate the specificity of the designed platform. Two commonly used microscopy techniques (indirect fluorescence and scanning electron microscopy) showed their straightforwardness and versatility for the nanoparticle binding investigations regardless the composition of the investigated nanoobjects. Moreover most of the research laboratories and centers are equipped with fluorescence microscopes, so indirect fluorescence using Quantum dots can be used for any active targeting nanocarriers (polymers, ceramics, metals, etc.). The second technique based on backscattered electron is not only limited to gold nanoparticles but also suits for any study of metallic nanoparticles as the electronic density difference between the nanoparticles and binding surface stays high enough. Optoacoustic imaging was finally performed on a 3D cellular model to assess and prove the concept of the developed platform.

Novel post-digest isotope coded protein labeling method for phospho- and glycoproteome analysis.

In the field of proteomics there is an apparent lack of reliable methodology for quantification of posttranslational modifications. Present study offers a novel post-digest ICPL quantification strategy directed towards characterization of phosphorylated and glycosylated proteins. The value of the method is demonstrated based on the comparison of two prostate related metastatic cell lines originating from two distinct metastasis sites (PC3 and LNCaP). The method consists of protein digestion, ICPL labeling, mixing of the samples, PTM enrichment and MS-analysis. Phosphorylated peptides were isolated using TiO(2), whereas the enrichment of glycosylated peptides was performed using hydrazide based chemistry. Isolated PTM peptides were analyzed along with non enriched sample using 2D-(SCX-RP)-Nano-HPLC-MS/MS instrumentation. Taken together the novel ICPL labeling method offered a significant improvement of the number of identified (∼600 individual proteins) and quantified proteins (>95%) in comparison to the classical ICPL method. The results were validated using alternative protein quantification strategies as well as label-free MS quantification method. On the biological level, the comparison of PC3 and LNCaP cells has shown specific modulation of proteins implicated in the fundamental process related to metastasis dissemination. Finally, a preliminary study involving clinically relevant autopsy cases reiterated the potential biological value of the discovered proteins.

Intra-arterial hepatic fotemustine for the treatment of liver metastases from uveal melanoma: experience in 101 patients.

BACKGROUND: Exclusive liver metastases occur in up to 40% of patients with uveal melanoma associated with a median survival of 2-7 months. Single agent response rates with commonly available chemotherapy are below 10%. We have investigated the use of fotemustine via direct intra-arterial hepatic (i.a.h.) administration in patients with uveal melanoma metastases. PATIENTS AND METHODS: A total of 101 patients from seven centers were treated with i.a.h. fotemustine, administered intra-arterially weekly for a 4-week induction period, and then as a maintenance treatment every 3 weeks until disease progression, unacceptable toxicity or patient refusal. RESULTS: A median of eight fotemustine infusions per patient were delivered (range 1-26). Catheter related complications occurred in 23% of patients; however, this required treatment discontinuation in only 10% of the patients. The overall response rate was 36% with a median overall survival of 15 months and a 2-year survival rate of 29%. LDH, time between diagnosis and treatment start and gender were significant predictors of survival. CONCLUSIONS: Locoregional treatment with fotemustine is well tolerated and seems to improve outcome of this poor prognosis patient population. Median survival rates are among the longest reported and one-third of the patients are still alive at 2 years.

[Rupture of hepatic focal nodular hyperplasia. About two cases]. / Rupture d'une hyperplasie nodulaire focale. A propos de deux cas.

The diagnostics of focal nodular hyperplasia is reached through the use of imaging. When the diagnostic is certain, surgical abstention is the rule. Nevertheless, we were confronted with two cases of a rare complication; that of intraperitoneal rupture. In this situation, we suggest to first do an arteriography to control the bleeding, then to perform surgery when the patient has reached hemodynamic stability. Spontaneous rupture as a complication of benign nodular hyperplasia remains a rare event and only five cases were reported in litterature.

Induction of apoptosis in human promyelocytic leukemia cells by a natural trachylobane diterpene.

BACKGROUND: Trachylobane diterpenes are secondary metabolites, quite rare in nature, and their bioactivities are poorly understood. Recently, we have described the cytotoxic activity of ent-trachyloban-3beta-ol isolated from the leaves of Croton zambesicus, a plant used in African folk medicine. MATERIALS AND METHODS: Cell viability on several cell lines, cell morphology, DNA laddering, annexin Vand caspase-3 activation experiments were undertaken in order to analyse the cytotoxicty of trachylobane diterpene and to determine if this compound is able to induce apoptosis. RESULTS: ent-Trachyloban-3beta-ol exerts a dose-dependent cytotoxic effect, which varies between cell lines. Induction of apoptosis in HL-60 cells could be detected at a concentration of 50 microM after 24-h treatment. CONCLUSION: We show here, for the first time, that a trachylobane diterpene is able to induce apoptosis in human promyelocytic leukemia cells via caspase-3 activation in a concentration-dependent manner.

Cystic pneumatosis of the colon after liver transplantation.

Cystic pneumatosis (CP) is an uncommon but significant condition in adults in which gas is found in a linear or cystic form in the submucosa or the subserosa of the bowel wall. The diagnosis was made by conventional X-ray and confirmed by abdominal computed tomography. Benign pneumoperitoneum due to CP should be considered in the differential diagnosis of free intra-abdominal air after chemotherapeutic or immunosuppressive therapy. As such, pneumatosis intestinalis is only a sign and must be interpreted in light of the clinical findings because it may be found in various scenarios: in patients who are otherwise healthy, and associated with pyloric stenosis, jejunoileal bypass, progressive systemic sclerosis, transplantation, chemotherapy, immunosuppression (including AIDS), obstructive pulmonary disease and finally, as in our case, after liver transplantation. Since there were no signs of secondary complications such as peritonitis, ischemia, or perforation, conservative treatment with broad-spectrum antibiotics and parenteral nutrition was initiated.

Impaired insulin response after oral but not intravenous glucose in heart- and liver-transplant recipients.

BACKGROUND: The prevalence of diabetes is high after transplantation. We hypothesized that liver transplantation induces additional alterations of glucose homeostasis because of liver denervation. METHODS: Nondiabetic patients with a heart (n=9) or liver (n=9) transplant and healthy subjects (n=8) were assessed using a two-step hyperglycemic clamp (7.5 and 10 mmol/L). Thereafter, an oral glucose load (0.65 g/kg fat free mass) was administered while glucose was clamped at 10 mmol/L. Glucose appearance from the gut was calculated as the difference between glucose appearance (6,6 2H2 glucose) and exogenous glucose infusion. Plasma insulin, glucagon-like peptide (GLP)-1 and gastric inhibitory polypeptide(GIP) concentrations were compared after intravenous and oral glucose. RESULTS: After oral glucose, the glucose appearance from the gut was increased 52% and 81% in liver- and heart-transplant recipients (P<0.05). First-pass splanchnic glucose uptake was reduced by 39% in liver-transplant and 64% in heart-transplant patients (P<0.05). After oral but not intravenous glucose, there was an impairment of insulin secretion in both transplant groups relative to the controls. Plasma concentrations of GIP and GLP-1 increased similarly in all three groups after oral glucose. CONCLUSIONS: First-pass hepatic glucose extraction is decreased after heart and liver transplant. Insulin secretion elicited by oral, but not intravenous glucose, is significantly reduced in both groups of patients. There was no difference between liver- and heart-transplant recipients, indicating that hepatic denervation was not involved. These data suggest an impairment in the beta-cell response to neural factors or incretin hormones secondary to immunosuppressive treatment.

[Agenesis of the gallbladder]. / Agénésie de la vésicule biliaire.

Isolated agenesis of the gallbladder (AG) is a rare anomaly. Twenty-three percent of the patients are symptomatic and present with right upper abdominal pain, nausea and fatty food intolerance. The condition is frequently mistaken with excluded or sclero-atrophic gallbladder, regardless of the imaging modality used. Consequently, AG leads often to unnecessary and potentially dangerous surgery. During laparoscopy, the absence of normal anatomical structures and the impossibility of pulling on the gallbladder to dissect the triangle of Callot represent an increased risk of iatrogenic injury to biliary or portal structures. The aim of this study is to discuss the pitfalls of the available radiological exams and the management of this rare condition. We describe two cases of AG, with a review of the literature. A high index of suspicion is necessary when interpreting the radiological images. In case of doubt, a MRI-cholangiography is mandatory. Because of possible inherited transmission, relatives with a history of biliary symptoms should be investigated, even when asymptomatic.

Cytotoxic aporphine alkaloids from Cassytha filiformis.

Purification of a cytotoxic crude alkaloid extract of Cassytha filiformis led to the isolation of four known aporphine alkaloids: neolitsine, dicentrine, cassythine (= cassyfiline) and actinodaphnine. Their structures were determined by analysis of spectroscopic data. All isolated alkaloids were tested for their cytotoxic activities on cancer and non-cancer cell lines in vitro. Neolitsine was the most active against HeLa and 3T3 cells (IC 50 :21.6 microM, and 21.4 microM, respectively). Cassythine and actinodaphnine showed the highest activity against Mel-5 (IC 50 : 24.3 microM and 25.7 microM, respectively) and HL-60 (IC 50 : 19.9 microM and 15.4 microM, respectively). This is the first report on the cytotoxic activity of C. filiformis extract and of neolitsine and cassythine. Furthermore, the complete NMR data of cassythine and actinodaphnine are given here for the first time.

ent-trachyloban-3beta-ol, a new cytotoxic diterpene from Croton zambesicus.

The dichloromethane extract of leaves of Croton zambesicus (Euphorbiaceae) showing in vitro cytotoxicity against human cervix carcinoma cells was investigated in order to identify its active compounds. A bio-guided fractionation by HSCCC followed by MPLC led us to isolate a trachylobane diterpene, ent-trachyloban-3beta-ol, with cytotoxic properties (IC50 on HeLa cells = 7.3 microg/ml). This is the first report on the cytotoxicity of a trachylobane diterpene.

[Should benign tumors of the liver be operated?]. / Faut-il opérer les tumeurs benignes du foie?

OBJECTIVES: To define a strategy for benign liver tumours treatment and to report our experience. DESIGN: A retrospective study. PATIENTS: Sixty-eight patients operated for benign liver tumours among 424 operated of liver resection. RESULTS: Benign liver tumours were discovered during radiologic exploration for upper abdominal pain in 69%, incidentally during radiologic examination in 23% or at laparotomy in 8% of patients. Preoperative assessment of solid benign liver tumours was based on 4 radiologic examinations in average whereas cystic lesions needed only 2. The surgical treatment ranged from a simple tumorectomy to liver transplantation depending on the type, size and location of the tumour. There were no postoperative deaths and minor complications occurred in 7 patients (10%). Twenty hemangiomas, 20 hepatic cysts, 22 focal nodular hyperplasias (FNH), 4 hepatic adenomas (HA) and 2 cystadenomas were found on histology. The final diagnosis differed from the preoperative one in 36 patients (53%). CONCLUSIONS: Differential diagnosis for cystic lesions are the hepatic cyst, the hydatid cyst and the cystadenoma. Hydatid cysts and cystadenomas are resected whereas the asymptomatic hepatic cyst is left untreated. Hemangiomas must be resected only when symptomatic or pedunculated. The differentiation between FNH, HA and hepatocellular carcinoma is difficult and final diagnosis is most often achieved only by surgery.

Laparoscopic treatment of acute small bowel obstruction: a multicentre retrospective study.

BACKGROUND: Laparoscopic surgery is thought to promote early recovery and quicker return to bowel function. The objective was to evaluate the rate and predictive factors of success, the causes of failure, the morbidity, and mortality during and after hospitalization, as well as to determine whether laparoscopic treatment of acute small bowel obstruction offers the same benefits as for other laparoscopic procedures. METHODS: The records of 308 patients with acute small bowel obstruction treated laparoscopically in 35 centres between 1 October 1988 and 30 September 1996 were retrospectively reviewed. RESULTS: Treatment was implemented completely by laparoscopy ('success' group) in 168 patients (54.6%). Conversion to laparotomy ('failure' group) was required in 140 patients (45.4%; during the same operation in 126 patients and after a median delay of 4 days (range: 1-12 days) in 14 patients). There were significantly more successes in patients with a history of one or two surgical interventions than in those with more than two (56% vs 37%; P < 0.05). There were significantly more successes in patients who had undergone appendectomy only (67/94; 71%) than in patients who (i) had no antecedent surgery (52%; P < 0.05), or (ii) underwent other surgery (33%; P < 0.001). The rate of success was significantly higher (P < 0.001) in patients operated on early (< 24 h) and in patients with bands (54%), than in those with adhesions (31%) or with other causes of obstruction (15%). The median duration of postoperative ileus was significantly shorter in the 'success' group than in the 'failure' group (2 days vs 4 days; P < 0.001). The median duration of postoperative hospital stay was shorter in the 'success' group than in the 'failure' group (4 days vs 10 days; P < 0.001). Fewer immediate wound complications were sustained in the 'success' group than in the 'failure' group (1.2% vs 10.7%; P < 0.001). The total number of immediate or delayed complications and particularly the number of recurrent obstructions after hospitalization as well as the number of deaths did not differ significantly between the two groups. CONCLUSIONS: Successful laparoscopic treatment of small bowel obstruction can be expected in patients who are seen early, and who have had one or two previous interventions (particularly appendectomy, especially if bands are found).

Malacoplakia simulating organ invasion in a rectosigmoid adenocarcinoma: report of a case.

Malacoplakia is a histiocytic inflammatory response that may be associated with colorectal tumors. We report the case of a 65-year-old male taking steroids for a severe pulmonary disease. He presented with a rectosigmoid tumor that seemed to infiltrate the urinary bladder and the sacrum on the preoperative CT scan and echography and at laparotomy. A low anterior resection en bloc with a partial cystectomy was performed. The pathologic analysis showed a pT3pN0 adenocarcinoma with an extensive malacoplakia infiltrating the bladder and the pericolic and perirectal tissues. This case report emphasizes the overstaging that malacoplakia may induce and underlines a situation the surgeon may possibly confront. Our observation confirms the association of malacoplakia, colorectal carcinoma, and steroid treatment.

Disappearance rate of catecholamines, total metanephrines, and neuropeptide Y from the plasma of patients after resection of pheochromocytoma.

BACKGROUND: Plasma free metanephrines are a more reliable analyte to measure than catecholamines for the biochemical diagnosis of pheochromocytomas. We hypothesized that the long persistence of total (sulfate-conjugated plus free) metanephrines in the blood might have a significant diagnostic value. METHODS: We measured plasma concentrations of catecholamines and total metanephrines (sulfate-conjugated plus free forms) by HPLC with amperometric detection, and neuropeptide Y (NPY) by an amplified ELISA in seven patients before and after removal of their pheochromocytomas. The results for catecholamine, total metanephrines, and NPY in each patient were analyzed for up to 120 min, starting from the time of tumor vessel clamping. The persistence of analytes was quantified as the area under the concentration-time curve over 120 min. RESULTS: On the basis of the upper reference limit for each variable, plasma free norepinephrine (NE) and epinephrine (E) concentrations were increased preoperatively in at least one sample in seven and six patients, respectively. Total normetanephrine (NMN) and metanephrine (MN) were increased in all samples in seven and six patients, respectively. NPY was increased 2- to 465-fold. After removal of the tumor, MN and NMN showed a higher average relative increase above the upper limit of the reference interval than NE and E (P = 0.05), whereas NPY was intermediate. The persistence of increased values was significantly shorter for catecholamines than for metanephrines. The half-life estimated by nonlinear regression was 12.3 +/- 7.8 min for NPY. Significant correlations were observed among NE, E, NMN, MN, and NPY concentrations, but parent markers (E and MN or NE and NMN) did not appear significantly intercorrelated. CONCLUSIONS: A larger increase and a longer persistence of total metanephrines (reflecting predominantly sulfo-conjugated metanephrines) than catecholamines and NPY in plasma may contribute to their greater diagnostic accuracy in pheochromocytoma.

Methylation silencing and mutations of the p14ARF and p16INK4a genes in colon cancer.

The INK4a-ARF locus encodes two tumor suppressor proteins involved in cell-cycle regulation, p16INK4a and p14ARF, whose functions are inactivated in many human cancers. The aim of this study was to evaluate p14ARF and p16INK4a gene inactivation and its association with some clinocopathological parameters in colon cancer. The mutational and methylation status of the p14ARF and p16INK4a genes was analyzed in 60 primary colon carcinomas and 8 colon cancer cell lines. We have identified the first two reported mutations affecting exon 1beta of p14ARF in the HCT116 cell line and in one of the primary colon carcinomas. Both mutations occur within the N-terminal region of p14ARF, documented as important for nucleolar localization and interaction with Mdm2. Tumor-specific methylation of the p14ARF and p16INK4a genes was found in 33% and 32% of primary colon carcinomas, respectively. Methylation of the p14ARF was inversely correlated with p53 overexpression (p = 0.02). p14ARF and p16INK4a gene methylation was significantly more frequent in right-sided than in left-sided tumors (p = 0.02). Methylation of the p14ARF gene occurred more frequently in well-differentiated adenocarcinomas (p = 0.005), whereas the p16INK4a gene was more often methylated in poorly differentiated adenocarcinomas (p = 0.002). The present results underline the role of p14ARF and p16INK4a gene inactivation in the development of colon carcinoma. They suggest that the methylation profile of specific genes, in particular p14ARF and p16INK4a, might be related to biologically distinct subsets of colon carcinomas and possibly to different tumorigenic pathways.

Effects of a glucose meal on energy metabolism in patients with cirrhosis before and after liver transplantation.

HYPOTHESIS: Liver transplantation results in hepatic denervation. This may produce alterations of liver energy and substrate metabolism, which may contribute to weight gain after liver transplantation. DESIGN: Prospective clinical study. SETTING: Liver transplantation clinics in a university hospital. PATIENTS: Seven nondiabetic patients with cirrhosis were recruited while on a waiting list for liver transplantation. Seven healthy subjects were recruited as controls. INTERVENTION: Orthotopic liver transplantation. MAIN OUTCOME MEASURES: Evaluation of energy and substrate metabolism after ingestion of a glucose load with indirect calorimetry was performed before, 2 to 6 weeks after, and 5 to 19 months after transplantation. Whole-body glucose oxidation and storage and glucose-induced thermogenesis were calculated. RESULTS: Patients with cirrhosis had modestly elevated resting energy expenditure and normal glucose-induced thermogenesis and postprandial glucose oxidation and storage. These measures remained unchanged after liver transplantation despite a significant increase in postprandial glycemia. Patients, however, gained an average of 3 kg of body weight after 5 to 19 months compared with their weight before transplantation. CONCLUSION: Liver denervation secondary to transplantation does not lead to alterations of energy metabolism after ingestion of a glucose load.