Evidence is presented showing that a neuronal isoform of nitric oxide synthase (NOS) is expressed in rat pancreatic islets and INS-1 cells. Sequencing of the coding region indicated a 99.8% homology with rat neuronal NOS (nNOS) with four mutations, three of them resulting in modifications of the amino acid sequence. Doubleimmunofluorescence studies demonstrated the presence of nNOS in insulin-secreting ␤-cells. Electron microscopy studies showed that nNOS was mainly localized in insulin secretory granules and to a lesser extent in the mitochondria and the nucleus. We also studied the mechanism involved in the dysfunction of the ␤-cell response to arginine and glucose after nNOS blockade with N Gnitro-L-arginine methyl ester. Our data show that miconazole, an inhibitor of nNOS cytochrome c reductase activity, either alone for the experiments with arginine or combined with sodium nitroprusside for glucose, is able to restore normal secretory patterns in response to the two secretagogues. Furthermore, these results were corroborated by the demonstration of a direct enzymesubstrate interaction between nNOS and cytochrome c, which is strongly reinforced in the presence of the NOS inhibitor. Thus, we provide immunochemical and pharmacological evidence that ␤-cell nNOS exerts, like brain nNOS, two catalytic activities: a nitric oxide production and an NOS nonoxidating reductase activity, both of which are essential for normal ␤-cell function. In conclusion, we suggest that an imbalance between these activities might be implicated in ␤-cell dysregulation involved in certain pathological hyperinsulinic states. T he short-lived free radical gas nitric oxide (NO) is synthesized from L-arginine by a family of enzymes known as NO synthases (NOSs). Three NOS isoenzymes, encoded by three separate genes, have been described, including the Ca 2ϩ /calmodulin-dependent and constitutively expressed neuronal NOS (nNOS) and endothelial NOS (eNOS) enzymes and a calmodulin-independent cytokine-inducible NOS (iNOS) enzyme found in various cell types (rev. in 1). The small amounts of NO, produced by the constitutive forms in response to increases in intracellular calcium, play a crucial role in a number of physiological functions, including neurotransmission (2), vascular tone (3) and platelet aggregation (4), whereas the large amounts, produced by iNOS in a calcium-independent manner over prolonged periods of time, are implicated in pathological functions, such as cytotoxicity of activated macrophages (5).Even though the inducible isoform has been cloned in insulin-producing cells after induction by cytokines (6), it is unclear whether pancreatic ␤-cells express a constitutive NOS. Both NADPH-diaphorase (NADPH-d) histochemical staining previously shown to be specific for NOS (7) and immunohistochemical studies using various nNOS antisera yielded apparently conflicting data. Positive NADPH-d and immunoreactive nNOS stainings have been found to be colocalized in most pancreatic endocrine cells (8), a finding not confirmed in other studies...

AXL receptor tyrosine kinase (RTK) is implicated in proliferation and invasion of many cancers, particularly in pancreatic ductal adenocarcinoma (PDAC), for which new therapeutic options are urgently required. We investigated whether inhibition of AXL activity by specific monoclonal antibodies (mAbs) is efficient in limiting proliferation and migration of pancreatic cancer cells. Expression of AXL was evaluated by immunohistochemistry in 42 PDAC. The AXL role in oncogenesis was studied using the short hairpin RNA approach in a pancreatic carcinoma cell line. We further generated antihuman AXL mAbs and evaluated their inhibitory effects and the AXL downstream signaling pathways first in vitro, in a panel of pancreatic cancer cell lines and then in vivo, using subcutaneous or orthotopic pancreatic tumor xenografts. AXL receptor was found expressed in 76% (32/42) of PDAC and was predominantly present in invasive cells. The AXL-knockdown Panc-1 cells decreased in vitro cell migration, survival and proliferation, and reduced in vivo tumor growth. Two selected anti-AXL mAbs (D9 and E8), which inhibited phosphorylation of AXL and of its downstream target AKT without affecting growth arrest-specific factor 6 (GAS6) binding, induced downexpression of AXL by internalization, leading to an inhibition of proliferation and migration in the four pancreatic cancer cell lines studied. In vivo, treatment by anti-AXL mAbs significantly reduced growth of both subcutaneous and orthotopic pancreatic tumor xenografts independently of their KRAS mutation status. Our in vitro and preclinical in vivo data demonstrate that anti-human AXL mAbs could represent a new approach to the pancreatic cancer immunotherapy.

Each year, breast cancer accounts for more than 400,000 new cancer cases and more than 130,000 cancer deaths in Europe. Prognosis of nonmetastatic breast cancer patients is directly related to the extent of the disease, mainly nodal spreading and tumor size, and to the molecular profile, particularly HER2 over-expression. In patients with HER2-over-expressing tumors, different studies have shown cellular and/or humoral immune responses against HER2 associated with a lower tumor development at early stages of the disease. These findings have led to the hypothesis that the generation of an anti-HER2 immune response should protect patients from HER2-over-expressing tumor growth. Taken together with the clinical efficiency of trastuzumab-based anti-HER2 passive immunotherapy, these observations allowed to envisage various vaccine strategies against HER2. The induction of a stable and strong immunity by cancer vaccines is expected to lead to establishment of immune memory, thereby preventing tumor recurrence. However, an immunological tolerance against HER2 antigen exists representing a barrier to effective vaccination against this oncoprotein. As a consequence, the current challenge for vaccines is to find the best conditions to break this immunological tolerance. In this review, we will discuss the different anti-HER2 vaccine strategies currently developed; considering the strategies having reached the clinical phases as well as those still in preclinical development. The used antigen can be either composed of tumoral allogenic cells or autologous cells, or specific to HER2. It can be delivered by dendritic cells or in a DNA, peptidic or proteic form. Another area of research concerns the use of antiidiotypic antibodies mimicking HER2.