No abstract

Gliomas are highly invasive, lethal brain tumors. Tumor-associated proteases play an important role in glioma progression. Annexin A2 is overexpressed in many cancers and correlates with increased plasmin activity on the tumor cell surface, which mediates degradation of extracellular matrix and promotes neoangiogenesis to facilitate tumor growth. In this study, we used two glioma cell lines, mouse GL261-EGFP and rat C6/lacZ, as well as stable clones transfected with an annexin A2 knockdown construct. We find that the annexin A2 knockdown decreased glioma cell migration in vitro and decreased membrane-bound plasmin activity. In vivo we injected the glioma cells into the rodent brain and followed glioma progression. Knockdown of annexin A2 in glioma cells decreased tumor size and slowed tumor progression, as evidenced by decreased invasion, angiogenesis and proliferation, as well as increased apoptosis in the tumor tissue of the annexin A2 knockdown group. Moreover, we report that the levels of expression of annexin A2 in human glioma samples correlate with their degree of malignancy. Taken together, our findings demonstrate that inhibition of annexin A2 expression in glioma cells could become a new target for glioma therapy.

Exposure of neurons to high concentrations of excitatory neurotransmitters causes them to undergo excitotoxic death via multiple synergistic injury mechanisms. One of these mechanisms involves actions undertaken locally by microglia, the CNS-resident macrophages. Mice deficient in the serine protease plasmin exhibit decreased microglial migration to the site of excitatory neurotransmitter release and are resistant to excitotoxic neurodegeneration. Microglial chemotaxis can be signaled by the chemokine monocyte chemoattractant protein-1 (MCP-1)/CCL2 (CC chemokine ligand 2). We show here that mice genetically deficient for MCP-1 phenocopy plasminogen deficiency by displaying decreased microglial recruitment and resisting excitotoxic neurodegeneration. Connecting these pathways, we demonstrate that MCP-1 undergoes a proteolytic processing step mediated by plasmin. The processing, which consists of removal of the C terminus of MCP-1, enhances the potency of MCP-1 in in vitro migration assays. Finally, we show that infusion of the cleaved form of MCP-1 into the CNS restores microglial recruitment and excitotoxicity in plasminogen-deficient mice. These findings identify MCP-1 as a key downstream effectorintheexcitotoxicpathwaytriggeredbyplasminandidentifyplasminasanextracellularchemokineactivator.Finally,ourresultsprovide a mechanism that explains the resistance of plasminogen-deficient mice to excitotoxicity.

Background-Ischemic stroke is a leading cause of morbidity and mortality worldwide and recombinant human tissue-type Plasminogen Activator (tPA) is the prominent among very few therapeutics used in its treatment. Due to complications attributed to the drug, most notably transformation of ischemia to hemorrhage, tPA is used only in a small number of ischemic stroke cases, albeit significantly more often in specialized stroke centers.

CitationSetting This is a pre-or post-print of an article published in Schneider, C.K., Vleminckx, C., Gravanis, I., Ehmann, F., Trouvin, J.-H., Weise, M., Thirstrup, S. Setting the stage for biosimilar monoclonal antibodies (2012) Nature Biotechnology, 30 (12), pp. 1179-1185. Advice more than once). It is interesting to note that the scope of questions in these confidential Scientific Advice procedures for mAbs has shifted over time (Box 1). Development of a biosimilar frameworkThe current concept of development of biosimilar mAbs follows the principle that any analysis that compares the biosimilar mAb with its reference mAb aims to detect differences using state of the art technology, thus excluding to the best possible extent that there are any relevant differences (see Table 1 for a summary of the basic concepts). Slight differences, within known and controlled ranges, are a given fact for biologicals-including innovative mAbs in their batch-to-batch microheterogeneity and variability-and thus do not preclude the conclusion that a given mAb is a 'true' biosimilar, as recently defined 8 .The highest burden for a biosimilar development program is on the physicochemical and biological characterization (in regulatory jargon the 'quality programme') with various state-ofthe-art methodologies. First, the biosimilar mAb has to fulfil all relevant requirements for a mAb of the concerned class (stand alone properties); second, it has to be comparable with the reference mAb. In other words, the analytical package for a biosimilar mAb submission is considerably larger than for a 'stand-alone' mAb (blue plus orange segment in quality portion shown in Fig. 2a). Since the objective of a biosimilar development is to establish biosimilarity, not to re-establish benefit, the non-clinical and clinical development program is more focused and 'tailored' toward this objective 8-10 ( Fig. 2a), making in part reference to the already licensed original mAb. We have provided a comparison of regulatory reviews of (non-mAb) biosimilars to those of stand-alone biotechnological medicinal products, as this may throw light on issues that will be important for biosimilar mAbs (Box 2).The current concepts for biosimilar mAbs have evolved over time; indeed, several previous guidelines on specific product classes of biosimilars 11 introduced important milestones on non-clinical and clinical aspects that paved the way for more complex biosimilars, including mAbs (see Table 2 for an overview of the relevant guidelines). One area that illustrates this evolution is EMA stance on whether evidence from a clinical trial in one particular clinical condition can be considered sufficient for a biosimilar to be equally effective and safe in another clinical condition.This 'extrapolation' of indications was first applied to a wider extent to biosimilar low-molecular provide the basis for use in the prevention of arterial thromboembolism. Several guidelines introduced concepts around pharmacodynamic markers that are usually more sensitive as clinical endp...