ABBV-011, A Novel, Calicheamicin-Based Antibody-Drug Conjugate, Targets SEZ6 to Eradicate Small Cell Lung Cancer Tumors.

In the past year, four antibody-drug conjugates (ADC) were approved, nearly doubling the marketed ADCs in oncology. Among other attributes, successful ADCs optimize targeting antibody, conjugation chemistry, and payload mechanism of action. Here, we describe the development of ABBV-011, a novel SEZ6-targeted, calicheamicin-based ADC for the treatment of small cell lung cancer (SCLC). We engineered a calicheamicin conjugate that lacks the acid-labile hydrazine linker that leads to systemic release of a toxic catabolite. We then screened a patient-derived xenograft library to identify SCLC as a tumor type with enhanced sensitivity to calicheamicin ADCs. Using RNA sequencing (RNA-seq) data from primary and xenograft SCLC samples, we identified seizure-related homolog 6 (SEZ6) as a surface-expressed SCLC target with broad expression in SCLC and minimal normal tissue expression by both RNA-seq and IHC. We developed an antibody targeting SEZ6 that is rapidly internalized upon receptor binding and, when conjugated to the calicheamicin linker drug, drives potent tumor regression in vitro and in vivo. These preclinical data suggest that ABBV-011 may provide a novel treatment for patients with SCLC and a rationale for ongoing phase I studies (NCT03639194).

In vitro Fab display: a cell-free system for IgG discovery.

Selection technologies such as ribosome display enable the rapid discovery of novel antibody fragments entirely in vitro. It has been assumed that the open nature of the cell-free reactions used in these technologies limits selections to single-chain protein fragments. We present a simple approach for the selection of multi-chain proteins, such as antibody Fab fragments, using ribosome display. Specifically, we show that a two-chain trastuzumab (Herceptin) Fab domain can be displayed in a format which tethers either the heavy or light chain to the ribosome while retaining functional antigen binding. Then, we constructed synthetic Fab HC and LC libraries and performed test selections against carcinoembryonic antigen (CEA) and vascular endothelial growth factor (VEGF). The Fab selection output was reformatted into full-length immunoglobulin Gs (IgGs) and directly expressed at high levels in an optimized cell-free system for immediate screening, purification and characterization. Several novel IgGs were identified using this cell-free platform that bind to purified CEA, CEA positive cells and VEGF.

Development of an enzyme-linked immunosorbent assay (ELISA) to measure the level of tyrosine hydroxylase protein in brain tissue from Parkinson's disease models.

Tyrosine hydroxylase (TH) catalyses the rate-limiting step in the biosynthesis of catecholamines. TH expression is regulated in a tissue-specific manner during neuronal development and differentiation. Because of its key regulatory role in central and peripheral catecholamine synthesis, TH is associated with the pathogenesis of several neurological and psychiatric diseases, including Parkinson's disease, dystonia, schizophrenia, affective disorders, and cardiovascular diseases. Therefore, developing a quantitative method to monitor the changes in TH expression in disease models could facilitate the identification and characterisation of neuromodulatory and neuroprotective therapeutic agents. The present report describes the generation and characterisation of a new set of monoclonal TH antibodies and the development of a novel sandwich ELISA for the quantitative detection of the TH protein in rodent brain tissue. This ELISA exhibits excellent reproducibility and good linearity in the analysis of complex brain tissue lysates. The cross-validation of the TH ELISA using semi-quantitative TH Western blot methods and HPLC measurement of dopamine levels suggests that the new TH ELISA is sufficiently sensitive to detect small-to-moderate region-specific differences, developmental changes, and Parkinson's disease-related changes in TH expression in rodent brains. This new TH ELISA also offers greater flexibility than conventional HPLC-based dopamine assays because the optimal tissue lysis buffer used for the detection of TH in brain tissue is also compatible with the analysis of other proteins associated with Parkinson's disease, such as α-synuclein, suggesting that this TH ELISA could be used in a multiplexed format.

Laminin-411 is a vascular ligand for MCAM and facilitates TH17 cell entry into the CNS.

TH17 cells enter tissues to facilitate pathogenic autoimmune responses, including multiple sclerosis (MS). However, the adhesion molecules involved in the unique migratory capacity of TH17 cells, into both inflamed and uninflamed tissues remain unclear. Herein, we characterize MCAM (CD146) as an adhesion molecule that defines human TH17 cells in the circulation; following in vitro restimulation of human memory T cells, nearly all of the capacity to secrete IL-17 is contained within the population of cells expressing MCAM. Furthermore, we identify the MCAM ligand as laminin 411, an isoform of laminin expressed within the vascular endothelial basement membranes under inflammatory as well as homeotstatic conditions. Purified MCAM-Fc binds to laminin 411 with an affinity of 27 nM, and recognizes vascular basement membranes in mouse and human tissue. MCAM-Fc binding was undetectable in tissue from mice with targeted deletion of laminin 411, indicating that laminin 411 is a major tissue ligand for MCAM. An anti-MCAM monoclonal antibody, selected for inhibition of laminin binding, as well as soluble MCAM-Fc, inhibited T cell adhesion to laminin 411 in vitro. When administered in vivo, the antibody reduced TH17 cell infiltration into the CNS and ameliorated disease in an animal model of MS. Our data suggest that MCAM and laminin 411 interact to facilitate TH17 cell entry into tissues and promote inflammation.