Antibody-dependent cellular cytotoxicity plays a pivotal role in antibody-based tumor therapies and is based on the recruitment of natural killer cells to antibody-bound tumor cells via binding of the Fc␥ receptor III (CD16). Here we describe the generation of chimeric DNA aptamers that simultaneously bind to CD16␣ and c-Met, a receptor that is overexpressed in many tumors. By application of the systematic evolution of ligands by exponential enrichment (SELEX) method, CD16␣ specific DNA aptamers were isolated that bound with high specificity and affinity (91 pM-195 nM) to their respective recombinant and cellularly expressed target proteins. Two optimized CD16␣ specific aptamers were coupled to each of two c-Met specific aptamers using different linkers. Bi-specific aptamers retained suitable binding properties and displayed simultaneous binding to both antigens. Moreover, they mediated cellular cytotoxicity dependent on aptamer and effector cell concentration. Displacement of a bi-specific aptamer from CD16␣ by competing antibody 3G8 reduced cytotoxicity and confirmed the proposed mode of action. These results represent the first gain of a tumoreffective function of two distinct oligonucleotides by linkage into a bi-specific aptamer mediating cellular cytotoxicity.Aptamers are structured single-stranded oligonucleotides that can bind to a large variety of targets with high affinity and specificity (1, 2). Aptamers can be isolated by an in vitro selection and an evolution process referred to as systematic evolution of ligands by exponential enrichment (SELEX) 2 (3, 4). Because aptamers have the capacity to inhibit protein-protein interactions with potencies similar to those observed with antibodies, aptamers can also trigger inhibition signals, e.g. by blocking receptor multimerization, and consequently act as therapeutic antagonists. Reversely, bi-and multivalent aptamers can activate co-stimulatory receptors, e.g. to enhance T cell reactivity (5, 6). Finally, aptamers can be applied in ligand-based targeted therapies to specifically deliver cytotoxic payloads (7,8) or siRNA (9) to tumor cells. Monoclonal antibodies serve as established and successful tumor therapeutics. However, naturally bivalent antibody formats comprise the risks of immunogenicity (10) and undesired activation by receptor dimerization (11). Development of monovalent therapeutic antibodies is elaborate and time-intensive (MetMAb (12)). Although antibodies exceed aptamers with proof as therapeutic molecules, high stability, and good pharmacokinetics, the potential advantages of aptamers are a rapid optimization, cost-effective and uniform synthesis, and a high probability of an absence of immunogenicity (5, 13). Approval of Macugen (pegaptanib sodium (14)) as the first therapeutic aptamer in 2007 as well as promising approaches in preclinical development and clinical trials (15, 16) only a few years after inception of the technology indicate aptamers as a promising new class of targeted therapeutics.Antibody-dependent cellular cytotoxicity (A...

Background: Cysteine biosynthesis is the exclusive entry point for reduced sulfur in cellular metabolism. Results: The mitochondrial cysteine synthase complex (mCSC) regulates serine acetyltransferase activity in response to cysteine availability. Conclusion:The mCSC is a sensor of sulfur availability and regulates cysteine synthesis. Significance: The integration of cysteine in the regulatory model of the CSC establishes a new sensory function for the mCSC.