Among different cancer immunotherapy approaches, bispecific antibodies (BsAbs) are of great interest due to their ability to recruit immune cells to kill tumor cells directly. Various BsAbs against Her2 tumor cells have been proposed with potent cytotoxic activities. However, most of these formats require extensive processing to obtain heterodimeric bispecific antibodies. In this study, we describe a bispecific antibody, BiHC (bispecific Her2-CD3 antibody), constructed with a single-domain anti-Her2 and a single-chain Fv (variable fragment) of anti-CD3 in an IgG-like format. In contrast to most IgG-like BsAbs, the two arms in BiHC have different molecular weights, making it easier to separate hetero- or homodimers. BiHC can be expressed in Escherichia coli and purified via Protein A affinity chromatography. The purified BiHC can recruit T cells and induce specific cytotoxicity of Her2-expressing tumor cells in vitro. The BiHC can also efficiently inhibit the tumor growth in vivo. Thus, BiHC is a promising candidate for the treatment of Her2-positive cancers. View Publication

In several clinical contexts, the measurement of ATP concentration in T lymphocytes has been proposed as a biomarker of immune status, predictive of secondary infections. However, the use of such biomarker in lymphopenic patients requires some adaptations in the ATP dosage protocol. We used blood from healthy volunteers to determine the optimal experimental settings. We investigated technical aspects such as the type of anticoagulant for blood sampling, the effect of freeze and thaw cycles, the reagent and sample mixing sequence, and the optimal dilution buffer. We also shortened the incubation time to 8h, and even showed that a 30min incubation may be sufficient. To evaluate the ATP rise upon lymphocyte activation, the optimal dose of stimulant was defined to be 4μg/mL of phytohaemagglutinin. Lastly, we determined that the number of T cells needed for this measurement was as low as 50,000, which is compatible with the existing lymphopenia in clinical settings. This optimized protocol appears ready to be assessed in lymphopenic patients to further investigate the interconnection between T lymphocyte metabolism and impaired phenotype and functions. View Publication