Purpose: Tumor cell killing by anticancer drugs may be supported by their immuno- and pharmacologic effects. Chemotherapy is in fact able to (A) upregulate tumor-associated antigen expression, including carcinoembryonic antigen (CEA) or other target molecules such as thymidylate synthase (TS); and (B) downregulate tumor cell resistance to the death signals induced by tumor antigen–specific cytotoxic T lymphocytes. This provides the rationale for combining chemo- and immunotherapy. Materials and Methods: We describe the results of a translational phase II trial designed to evaluate the toxicity, antitumor activity and immunologic effects of gemcitabine + FOLFOX-4 (oxaliplatin, fluorouracil, and folinic acid) polychemotherapy followed by the subcutaneous administration of granulocyte macrophage colony-stimulating factor and low-dose interleukin-2 in colorectal carcinoma patients. The study involved 29 patients (16 males and 13 females with a mean age of 69 years), 21 of whom had received a previous line of treatment, and 19 had liver involvement. Results: The treatment was well tolerated and induced very high objective response (68.9%) and disease control rates (96.5%), with an average time to progression of 12.5 months. An immunologic study of peripheral blood mononuclear cells (PBMCs) taken from 20 patients showed an enhanced proliferative response to colon carcinoma antigen and a significant reduction in suppressive regulatory T lymphocytes (CD4⁺CD25_T-reg⁺). A cytofluorimetric study of the PBMCs of five HLA-A(^★)02.01⁺ patients who achieved an objective response showed an increased frequency of cytolytic T lymphocyte precursors specific for known CEA- and TS-derived epitopes. Conclusion: The results show that our regimen has strong immunologic and antitumor activity in colorectal cancer patients and deserves to be investigated in phase III trials.

Background: To investigate the cardiotoxic role of reactive oxygen species (ROS) and of products derived from catecholamines auto‐oxidation, we studied: (1) the response of antioxidant cardiac cellular defence systems to oxidative stress induced by norepinephrine (NE) administration, (2) the effect of NE administration on cardiac β₁‐adrenergic receptors by means of receptor binding assay, (3) the cellular morphological alterations related to the biologically cross‐talk between the NE administration and cytokines [tumor necrosis factor‐alpha (TNF‐α), monocyte chemotactic protein‐1 (MCP‐1), interleukins IL6, IL8, IL10]Methods and Results: A total of 195 male rats was used in the experiment. All animals underwent electrocardiogram (EKG) before being sacrificed. The results obtained show that NE administration influences the antioxidant cellular defence system significantly increasing glutathione peroxidase (GPx) activity, glutathione reductase (GR) and superoxide dismutase (SOD). The oxidized glutathione (GSH/GSSG) ratio significantly decreases and malondialdehyde (MDA) levels increase showing a state of lipoperoxidation of cardiac tissue. We describe a significant apoptotic process randomly sparse in the damaged myocardium and the effect of ROS on the NE‐mediated TNF‐α, MCP‐1, and IL6, IL8, IL10 production. Conclusions: Our results support the hypothesis that catecholamines may induce oxidative damage through reactive intermediates resulting from their auto‐oxidation, irrespective of their interaction with adrenergic receptors, thus representing an important factor in the pathogenesis of catecholamines‐induced cardiotoxicity. The rise of the cardioinhibitory cytokines may be interpreted as the adaptive response of jeopardized myocardium with respect to the cardiac dysfunction resulting from NE injection.

Antioxidant activity of fresh Allium sativum L. (garlic) is well known and is mainly due to unstable and irritating organosulphur compounds. Fresh garlic extracted over a prolonged period (up to 20 months) produces odourless aged garlic extract (AGE) containing stable and water soluble organosulphur compounds that prevent oxidative damage by scavenging free radicals. The aim of this study was to investigate the in vitro antioxidant activity of aged (up to 20 months) 15% hydroethanolic extracts of different parts (bulbs, bulblets, flower bulblets, flowers, and leaves) of three Allium spontaneous species which are endemic for Italian flora: Allium neapolitanum Cyr., Allium subhirsutum L., Allium roseum L. and to compare it with the in vitro antioxidant activity of aged 15% hydroethanolic extracts of bulbs and leaves of garlic. The antioxidant potential of aged extracts of all species has been evaluated using two different spectrophotometric assays: 2,2-diphenylpicrylhydrazyl (DPPH) test and the ferric reducing/antioxidant power (FRAP) assay. Furthermore the polyphenol content was determined. The aged extracts obtained from the leaves showed the best antioxidant activity, followed by flowers and then by bulbs in both used tests, while flower bulblets and bulblets exhibited lower results or no activity. The polyphenol content was generally directly correlated with antioxidant/antiradical activity. This study confirms the data obtained in previous researches, the wild-type species of Allium and in particular organs other than bulbs are more active and efficacious than garlic bulb. Surely leaves of these Allium spp. deserve special attention.

Cancer vaccines directed against tumor associate antigen (TAA) have produced encouraging results in preclinical models but not in cancer patients. A major limitation of this strategy is the relative degree of tolerance to these antigens and the low and heterogeneous tumor cell expression of TAA and major histocompatibility complex (MHC). Previous studies have shown that 5‐fluorouracil (5‐FU) can upregulate the expression of membrane‐associated carcino‐embryonic antigen (CEA), and MHC molecules in colon and breast carcinoma cell lines. We have investigated whether this drug can also enhance their sensitivity to the lytic effects of CEA‐peptide specific Cytotoxic T cell lymphocytes (CTL). The CEA peptide‐specific CTLs generated in our laboratory from normal HLA‐A(*)02.01⁺ donor PBMCs, were able to kill HLA‐A(*)02.01⁺/CEA⁺ breast (MCF‐7‐T103) and colon (HLA‐A(*)02.01 gene‐transfected HT‐29 and C22.20) carcinoma cells in HLA‐A(*)02.01 restricted manner. The treatment of target cells with 5‐FU, enhanced their CEA expression and susceptibility to CTL‐mediated lysis. Cold competition assays confirmed these results, thus supporting the hypothesis that immune target cell lysis and 5‐FU mediated enhancement were dependent on CEA peptide presentation by cancer cells. 5‐FU treatment of functionally “mature” CTL after in vitro expansion, did not reduce their cytolytic activity against MT‐2 target cells but, when the anti‐metabolite was added during the immune‐sensitization phase, CTL generation was significantly inhibited. These results provide a rationale for investigating a possible new role of 5‐FU as an immuno targeting amplifier agent in breast and colorectal cancer patients immunized with CEA‐directed cancer vaccines.

Context: Preeclampsia (PE) is a disorder that occurs only during pregnancy. The placenta has a controlling role in this condition. Recent literature suggests that the oxidative stress is a component of PE and plays a main role in the link between decreased placental perfusion and the impaired function of maternal endothelium. Objective: Because the human placenta expresses endothelin-1 (ET-1) and its circulating levels are high in pregnancies complicated with PE, the present study investigated the role of ET-1 on placental oxidative stress pathways. Design: Human placental explants, JEG-3, and primary cytotrophoblast cells were cultured with increasing ET-1 concentrations for 6 and 24 h. Setting: The study was conducted at tertiary clinical care centers in Siena and Padova, Italy Interventions: Human placental explants, JEG-3, and primary cytotrophoblast cells were used to test ET-1 effect. Main Outcome Measure(s): The main outcome measure was ET-1 mRNA and its receptor mRNAs, type A and B, detection by RT-PCR. The common markers of oxidative stress [malondialdehyde (MDA), glutathione (GSH), glutathione disulfide (GSSG), ascorbic acid (AA)] as well as cell proliferation and vitality were measured after stimulation periods. Results: ET-1 inhibits cell proliferation and vitality and triggers oxidative stress in the human placenta by altering the balance between oxidant (increased MDA levels) and antioxidant (decreased GSH, GSSG, and AA) forces in favor of oxidation. Conclusions: Because MDA damages endothelial cells, whereas GSH, GSSG, and AA protect them, we postulate that ET-1 may be one of the key links between primary placental disorders and the systemic endothelial dysfunction of PE.