After identifying the binding epitope of MEDI-565, we evaluated the impact of polymorphisms and isoforms in the A2 domain of CEA on the activity of MEDI-565. Two nonsynonymous SNPs were identified in the binding epitope of MEDI-565 but occurred at a very low frequency in the general population and were considered to have little or no impact on MEDI-565 binding to CEA. In addition, a single splice variant was identified lacking a portion of the A2 domain critical for MEDI-565 binding. Efforts to understand the association of splice variant expression to disease found that its expression occurred in a substantial percentage of primary human colorectal and gastro-esophageal tumors, and to a lesser extent in lung and breast tumors in a pattern that was concordant with full length CEA expression. However, since MEDI-565 did not bind the CEA splice variant due to loss of the antibody-binding epitope, it was clear that the splice variant itself is not a target for MEDI-565 in primary human tumors that also express full length CEA. Because CEA could participate in homotypic interactions on adjacent cells, it remained possible that expression of the CEA splice variant may interfere with the binding of MEDI-565 and the subsequent tumor cell lysis through its interaction with full length CEA. By coexpressing the CEA splice variant with full length CEA, we formally demonstrated that this was not the case, as MEDI-565 binding to full length CEA and potency of CEA-directed lysis were not significantly affected by simultaneous expression of the splice variant and full length CEA. Therefore, the expression of the CEA splice variant by primary human tumor cells is not anticipated to interfere with MEDI-565 binding to full-length CEA, nor should it inhibit MEDI-565-mediated T-cell killing of tumor cells expressing full-length CEA. However, discrimination of full-length CEA from the CEA splice variant may be important while monitoring the status of CEA positive tumors via changes in serum CEA levels in clinical study patients that receive MEDI-565. Figure S1 Western blot of CEA protein expressed by CHO cell lines. The full-length CEA protein is indicated by a filled arrowhead, and the CEA splice variant by an open arrowhead; both proteins were detected using a CEACAM5-specific mAb. Molecular weights (kilodaltons; KDa) of the protein standard are indicated to the left of the image. Lanes of CHO FL and CHO FL+SV cell lysates contain a band at ,100 kDa that is presumed to be a non-glycosylated form of CEA. Equal amounts of protein loaded into each lane of the gel were controlled by detecting  High Throughput Screening for Small Molecule Enhancers of the Interferon Signaling Pathway to Drive Next-Generation Antiviral Drug Discovery Most of current strategies for antiviral therapeutics target the virus specifically and directly, but an alternative approach to drug discovery might be to enhance the immune response to a broad range of viruses. Based on clinical observation in humans and successful genetic strategies in experimental models, we reasoned that an improved interferon (IFN) signaling system might better protect against viral infection. Here we aimed to identify small molecular weight compounds that might mimic this beneficial effect and improve antiviral defense. Accordingly, we developed a cell-based high-throughput screening (HTS) assay to identify small molecules that enhance the IFN signaling pathway components. The assay is based on a phenotypic screen for increased IFN-stimulated response element (ISRE) activity in a fully automated and robust format (Z′>0.7). Application of this assay system to a library of 2240 compounds (including 2160 already approved or approvable drugs) led to the identification of 64 compounds with significant ISRE activity. From these, we chose the anthracycline antibiotic, idarubicin, for further validation and mechanism based on activity in the sub-µM range. We found that idarubicin action to increase ISRE activity was manifest by other members of this drug class and was independent of cytotoxic or topoisomerase inhibitory effects as well as endogenous IFN signaling or production. We also observed that this compound conferred a consequent increase in IFN-stimulated gene (ISG) expression and a significant antiviral effect using a similar dose-range in a cell-culture system inoculated with encephalomyocarditis virus (EMCV). The antiviral effect was also found at compound concentrations below the ones observed for cytotoxicity. Taken together, our results provide proof of concept for using activators of components of the IFN signaling pathway to improve IFN efficacy and antiviral immune defense as well as a validated HTS approach to identify small molecules that might achieve this therapeutic benefit. There has been significant progress in the development of vaccines and therapeutics against viruses, but there are still major gaps in medical therapy for some of the most common types of viral infections. For these types of infections, vaccines can still be ineffective due to new and emergent strains and can exhibit significant off-target effects [1, 2] . Similarly, the efficacy of antiviral therapeutics can often be limited by pathogen resistance as another sign of the difficulty in keeping up with rapidly evolving viral genomes [3] [4] [5] [6] [7] [8] [9] . An alternative to agents that specifically and directly target the virus itself is the possibility of improving natural host defense against a broad range of viruses. Although antiviral defense exhibits significant complexity and redundancy, one system that stands out as a useful target for improvement is the one based on the action of interferons (IFNs). And within this IFN system, which is similarly complex, the STAT1 transcription factor is remarkable as a central component that is critical for the functional activity of each type of IFN ( Figure 1 ). Consequently, genetic loss of STAT1 function causes a marked susceptibility to viral infection in mice and humans [10] [11] [12] . Moreover, modification of STAT1 to a form) that improves the efficiency of IFN signal transduction can result in improved control of viral infection [13] . These observations indicate that the IFN-signaling pathway is subject to a so-called ''rheo-STAT'' adjustment wherein downregulation causes increased susceptibility to viral infection whereas up-regulation might lead to increased efficiencies for IFNstimulated gene (ISG) expression and control of infection [14] .