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T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive form of leukemia that currently requires intensive chemotherapy. While childhood T-ALL is associated with high cure rates, adult T-ALL is not, and both are associated with significant short- and long-term morbidities. Thus, less toxic and effective strategies to treat T-ALL are needed. CD99 is overexpressed on T-ALL blasts at diagnosis and at relapse. Although targeting CD99 with cytotoxic antibodies has been proposed, the molecular features required for their activity are undefined. We identified human antibodies that selectively bound to the extracellular domain of human CD99, and the most potent clone, 10A1, shared an epitope with a previously described cytotoxic IgM antibody. We engineered clone 10A1 in bivalent, trivalent, tetravalent, and dodecavalent formats. Increasing the antibody valency beyond two had no effects on binding to T-ALL cells. In contrast, a valency of ≥3 was required for cytotoxicity, suggesting a mechanism of action in which an antibody clusters ≥3 CD99 molecules to induce cytotoxicity. We developed a human IgG-based tetravalent version of 10A1 that exhibited cytotoxic activity to T-ALL cells but not to healthy peripheral blood cells. The crystal structure of the 10A1 Fab in complex with a CD99 fragment revealed that the antibody primarily recognizes a proline-rich motif (PRM) of CD99 in a manner reminiscent of SH3-PRM interactions. This work further validates CD99 as a promising therapeutic target in T-ALL and defines a pathway toward the development of a selective therapy against T-ALL., Competing Interests: Conflict of interest LAR, TH, CYP and SK are listed as inventors of a patent application filed by New York University (US 63/174,131) related to this study. S.K. is a scientific advisory board member and holds equity in and receives consulting fees from Black Diamond Therapeutics and receives research funding from Puretech Health and Argenx BVBA., (Copyright © 2021 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Aims: Smokers have lower plasma concentrations of high-density lipoprotein (HDL) cholesterol and apolipoprotein A-I (apo A-I) compared with nonsmokers. To determine the molecular basis of this observation, the effect of activation of the aryl hydrocarbon receptor (AhR) on apo A-I gene expression was examined., Main Methods: HepG2 cells were treated with AhR receptor agonists benzo(a)pyrene (BaP) and CAY10465, and AhR receptor antagonist CAY10464 and apo A-I protein, mRNA levels and promoter activity were measured. The effect of nicotine on apo A-I protein secretion was also tested. Using a series or apo A-I gene promoter deletion constructs, a xenobiotic response element (XRE) was identified., Key Findings: Treatment of HepG2 cells with the AhR receptor agonists BaP and CAY10465, inhibited apo A-I protein synthesis while nicotine, which does not bind AhR had no effect. Benzo(a)pyrene treatment also suppressed apo A-I mRNA and gene promoter activity. Treatment of HepG2 cells with the AhR receptor antagonist CAY10464 reversed the suppressive effect of BaP on apo A-I gene expression. A putative xenobiotic response element (XRE) was identified between nucleotides -325 and -186 (relative to the transcriptional start site, +1)., Significance: These results suggest that the cigarette smoking related environmental contaminant BaP promotes hypoalphalipoproteinemia in part through activation of the hepatic AhR., (Copyright © 2012 Elsevier Inc. All rights reserved.)