Nikolaos Tsavaris

Combination treatment with metronomic temozolomide, bevacizumab and long-acting octreotide for malignant neuroendocrine tumours.

Neuroendocrine tumours (NETs) are highly vascularised tumours that express high levels of the vascular endothelial growth factor (VEGF) ligand together with its receptor VEGFR (Modlin et al. 2008). Although advanced NETs may exhibit a 30–40% response rate to combination chemotherapeutic approaches, the response to single-agent chemotherapy is only 10% (Modlin et al. 2008). Bevacizumab (BVZ; Avastin, Roche, Basle, Switzerland), an anti-VEGF humanised monoclonal antibody, has been shown to exert objective tumour responses and improvement in median time to progression (TTP) in advanced carcinoid tumours (Yao et al. 2008). Additionally, a recently published study reported that temozolomide (TMZ), an oral chemotherapy derivative of dacarbazine, at a dose of 200 mg/m on first 5 days of each 28-day cycle may exert a significant effect on NETs (Ekeblad et al. 2007). A previous report that examined a variety of NETs suggested that the combination of BVZ and TMZ can be safely administered and shows promising activity in patients who had failed to prior treatments (Kulke et al. 2006a). Additionally, there is growing interest on regimens that introduce continuous low-dose TMZ administration as protracted low-dose TMZ regimens may deplete O6-methylguanine DNA methyltransferase (MGMT), an important factor in cases of TMZ resistance and/or inhibit endothelial cell proliferation and formation of tumour vasculature via the so-called metronomic effect (Tolcher et al. 2003, Lam et al. 2007). Although the exact mechanism of action of somatostatin analogues is not well understood, a long-standing hypothesis based on preclinical experiments suggests that they exert an antiangiogenic effect (Grozinsky-Glasberg et al. 2008). Given the high degree of endothelial proliferation, high vascular permeability and high expression of proangiogenic growth factors such as VEGF in NETs, angiogenesis inhibition by multiple pathways may be a rational treatment strategy for these tumours. We

Identification of a Novel Immunogenic HLA-A*0201-Binding Epitope of HER-2/neu with Potent Antitumor Properties

HER-2/neu oncoprotein is overexpressed in a variety of human tumors and is associated with aggressive disease. Immunogenic HER-2/neu CTL epitopes have been used as vaccines for the treatment of HER-2/neu positive malignancies with limited success. By applying prediction algorithms for MHC class I ligands and proteosomal cleavages, in this study, we describe the identification of HER-2/neu decamer LIAHNQVRQV spanning residues 85–94 (HER-2(1085)). HER-2(1085) proved to bind with high affinity to HLA-A2.1 and was stable for 4 h in an off-kinetics assay. This peptide was immunogenic in HLA-A2.1 transgenic (HHD) mice inducing peptide-specific CTL, which responded to tumor cell lines of various origin coexpressing human HER-2/neu and HLA-A2.1. This demonstrates that HER-2(1085) is naturally processed from endogenous HER-2/neu. Five of sixteen HER-2/neu+ HLA-A2.1+ breast cancer patients analyzed had HER-2(1085)-reactive T cells ranging from 0.35–0.70% of CD8+ T cells. Depletion of T regulatory cells from PBMC enabled the rapid expansion of HLA-A2.1/HER-2(1085)pentamer+/CD8+ cells (PENT+/CD8+), whereas significantly lower numbers of CTL could be generated from unfractionated PBMC. HER-2(1085)-specific human CTL recognized the HER-2/neu+ HLA-A2.1+ tumor cell line SKBR3.A2, as determined by IFN-γ intracellular staining and in the high sensitivity CD107α degranulation assay. Finally, HER-2(1085) significantly prolonged the survival of HHD mice inoculated with the transplantable ALC.A2.1.HER tumor both in prophylactic and therapeutic settings. These data demonstrate that HER-2(1085) is an immunogenic peptide, capable of eliciting CD8-mediated responses in vitro and in vivo, providing the platform for further exploitation of HER-2(1085) as a possible target for anticancer immunotherapy.

The prognostic significance of immune changes in patients with renal cancer, melanoma and colorectal cancer, treated with interferon α2b

Abstract In the present study we evaluated the response rate and the immunorestorative properties of interferon α2b (IFnα2b) administered to patients with advanced renal cell carcinoma (RCC), melanoma (MEL) or colorectal cancer (CC). We studied the immune status and correlated it with clinical responses. Thirty-five patients with advanced RCC, and 14 with MEL were treated with recombinant INFα2b. The dose was increased progressively from 5×106 IU/day in the first week (three times every week) to 10×106 IU/day in the second week and thereafter to 15×106 IU/day subcutaneously. In patients with CC INFα2b was given at 5×106 IU/day every other day (three times every week); these patients also received (together with INF) leucovorin 200 mg m–2 day–1 in a 1-h i. v. infusion every week, and mid-infusion 400 mg/m2 5-FU was administered as an intravenous bolus every week. The response rate was as follows: for RCC, 6 patients achieved partial response (PR), 10 stable disease (SD), and 21 progressed (PD); for MEL, 5 patients achieved PR and 9 PD; for CC, 6 achieved PR, 5 SD, and 9 PD. In all patients blood was withdrawn prior to INFα2b treatment and then monthly. T lymphocytes, after isolation from peripheral blood, were tested for proliferation in the autologous mixed-lymphocyte reaction and allogeneic mixed-lymphocyte reaction, interleukin-2 (IL-2) production, expression of IL-2 receptors during the allogeneic-mixed-lymphocyte reaction, and the production of IL-1 by peripheral blood monocytes. Striking increases were demonstrated in all parameters 2 months after treatment with INFα2b. In comparison to normal controls, all patients with the malignant neoplasms presented decreased (>45%) mean values of the immunological parameters under investigation (P 0.0001). Responders (patients with RCC, MEL, and PR) presented lower mean values of all the parameters studied than did non-responders (P 0.0001). Patients with CC presented the lowest mean values of the parameters than did the other patients (RCC, MEL) (P 0.0001). After therapy with INFα2b, patients with RCC experiencing PR showed a mean increase of more than 30% (P 0.0001). Patients with SD showed a mean increase of about 20% (P 0.0001), and those with PD showed a 6% increase in the immunological parameters under investigation. Patients with MEL experiencing PR showed a mean increase of more than 30% and patients with PD a decrease of more than 10% (P 0.0001). All patients, regardless of the clinical response, achieved an increase of more than 60% (P 0.0001). Administration of IFNα2b resulted in a marked potentiation of a deficient cellular immune response in vitro in those patients with RCC and MEL who responded to the treatment. On the other hand, non-responders demonstrated a decrease in the examined parameters and, in some, deterioration of the already depressed immunological functions was observed. This observation can have prognostic significance regarding clinical response of INF. In contrast, our findings show that the immune stimulation associated with INFα treatment in all our CC patients did not predict an improved clinical outcome. There are several theoretical explanations for this discrepancy.

Polycystin-1 and polycystin-2 are involved in the acquisition of aggressive phenotypes in colorectal cancer.

The polycystins PC1 and PC2 are emerging as major players in mechanotransduction, a process that influences all steps of the invasion/metastasis cascade. We hypothesized that PC1 and PC2 facilitate cancer aggressiveness. Immunoblotting, RT‐PCR, semi‐quantitative and quantitative real‐time PCR and FACS analyses were employed to investigate the effect of polycystin overexpression in colorectal cancer (CRC) cells. The impact of PC1 inhibition on cancer‐cell proliferation was evaluated through an MTT assay. In vitro data were analyzed by Students t‐test. HT29 human xenografts were treated with anti‐PC1 (extracellular domain) inhibitory antibody and analyzed via immunohistochemistry to determine the in vivo role of PC1 in CRC. Clinical significance was assessed by examining PC1 and PC2 protein expression in CRC patients (immunohistochemistry). In vivo and clinical data were analyzed by non‐parametric tests, Kaplan‐Meier curves, log‐rank test and Cox model. All statistical tests were two‐sided. PC1 overexpression promotes epithelial‐to‐mesenchymal transition (EMT) in HCT116 cells, while PC2 overexpression results in upregulation of the mTOR pathway in SW480 cells. PC1 inhibition causes reduced cell proliferation in CRC cells inducing tumor necrosis and suppressing EMT in HT29 tumor xenografts. In clinical study, PC1 and PC2 overexpression associates with adverse pathological parameters, including invasiveness and mucinous carcinomas. Moreover, PC1 overexpression appears as an independent prognostic factor of reduced recurrence‐free survival (HR = 1.016, p = 0.03) and lowers overall survival probability, while aberrant PC2 expression predicts poor overall survival (p = 0.0468). These results support, for the first time, a direct link between mechanosensing polycystins (PC1 and PC2) and CRC progression.

Combined treatment with Bevacizumab and standard chemotherapy restores abnormal immune parameters in advanced colorectal cancer patients

SummaryBackground Bevacizumab, a monoclonal antibody (mAb) targeting vascular endothelial growth factor (VEGF), has produced promising results when combined with chemotherapy in the treatment of advanced colorectal cancer (CRC). The aim of the present study was to define the immunological profile of metastatic CRC patients at baseline and following chemotherapy with either irinotecan/5-fluorouracil/leucovorin (IFL) alone or IFL in combination with.bevacizumab (B-IFL). Methods Peripheral blood mononuclear cells (PBMCs) obtained from healthy donors (HD) (n = 20) and patients (n = 40) were tested for T-cell proliferation in the autologous mixed lymphocyte reaction (auto-MLR), and cytokine production following stimulation with anti-CD3 mAb. Results,PBMCs obtained from CRC patients prior to treatment exhibited lower auto-MLR responses and low production of IL-2, IFN-γ, IL-12 and IL-18 cytokines, whereas IL-4 and IL-10 cytokines were increased as compared to HD (p < 0.001, for all parameters) following in vitro stimulation with anti-CD3 mAb. During treatment, and in particular in week 12 of evaluation, IL-2 (p < 0.001 for both IFL and B-IFL groups), IFN-γ (p < 0.001 for IFL and p = 0.001 for B-IFL), IL-12 (p < 0.001 for both IFL and B-IFL) and IL-18 (p < 0.001 for both IFL and B-IFL) production, as well as auto-MLR responses increased (p < 0.001 for both IFL and B-IFL), whereas IL-4 (p < 0.001 for IFL and p = 0.001 for B-IFL) and IL-10 [p < 0.001 for IFL and p = 0.067 (non-significant) for B-IFL] production decreased over baseline in the two treatment groups, yet their respective values never reached those of HD. Moreover, IL-2, IFN-γ production, and auto-MLR were higher in the B-IFL over the IFL treatment group (p < 0.001, p < 0.04, p < 0.001, respectively). Conclusion Our study demonstrates that the abnormal immune parameters observed in metastatic CRC patients at presentation can substantially improve during treatment with either IFL or B-IFL. The immune parameters examined can provide a sensitive and valuable tool for monitoring immune function in CRC patients, and could be applied as surrogate markers predicting treatment-related outcome.

Comparison of two oral regimens for the outpatient treatment of low-risk cancer patients with chemotherapy-induced neutropenia and fever: Ciprofloxacin plus cefuroxime axetil versus ciprofloxacin plus amoxicillin/clavulanate

The objective of this investigation was to assess retrospectively the safety and the efficacy of oral ciprofloxacin plus cefuroxime axetil compared to the combination of oral ciprofloxacin plus amoxicillin/clavulanate, as initial outpatient treatment, in low-risk cancer patients with fever and neutropenia. We analysed retrospectively 120 episodes of febrile neutropenia, treated on an outpatient basis at 2 different oncology units; 63 episodes were treated with the oral regimen of ciprofloxacin plus amoxicillin/clavulanate and 57 were treated with the combination of oral ciprofloxacin plus cefuroxime. 20 treatment failures were recorded–2 of them among patients receiving ciprofloxacin plus amoxicillin/clavulanate and 18 in the ciprofloxacin plus cefuroxime group. Univariate analysis showed that the administration of ciprofloxacin plus cefuroxime was associated with a worse outcome compared to the regimen ciprofloxacin plus amoxicillin/clavulanate (OR 11, CI 2.42–49.9, p =0.002). In the multivariate model, after adjusting for the absolute number of neutrophils and the duration of neutropenia, the effect of the antibiotic regimen on the outcome disappeared, and no significant differences between the 2 regimens were noted, although the regimen of ciprofloxacin plus cefuroxime was associated with a trend to a worse outcome (OR 4.74, CI 0.72–31.1, p =0.10). In conclusion, the 2 regimens appeared equally safe and effective but prospective studies are needed to confirm these results.

Cytochemical Distribution of 5-Nucleotidase Activity in Human Leukemic Cells

5-Nucleotidase activity was cytochemically determined in normal and leukemic cells by the application of a modified cytochemical reaction developed in our laboratory. Blood smears from 10 normal donors, 54 patients with acute or chronic leukemias, and 5 patients with nodular poorly differentiated lymphocytic lymphoma in leukemic phase, were stained for this reaction. 5-Nucleotidase (5N) positivity was found in 20.6 ± 2.9% of the normal lymphocytes. 5N activity was highly decreased in the leukemic cells of acute myeloblastic leukemia, B chronic lymphocytic leukemia (B CLL) (2.73 ± 3.7%), B prolymphocytic leukemia (1.5 ± 1.7%), and hairy cell leukemia (HCL) (2.4 ± 2.4%). In two cases of T chronic lymphocytic leukemia (T CLL), one case of T prolymphocytic leukemia, and one case of T acute lymphoblastic leukemia (T ALL), a very low 5N activity was also observed.