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Summary The cell fate decision between interferon-producing plasmacytoid DC (pDC) and antigen-presenting classical DC (cDC) is controlled by the E protein transcription factor TCF4 (E2-2). We report that TCF4 comprises two transcriptional isoforms, both of which are required for optimal pDC development in vitro. The long Tcf4 isoform is expressed specifically in pDCs, and its deletion in mice impaired pDCs development and led to the expansion of non-canonical CD8+ cDCs. The expression of Tcf4 commenced in progenitors and was further upregulated in pDCs, correlating with stage-specific activity of multiple enhancer elements. A conserved enhancer downstream of Tcf4 was required for its upregulation during pDC differentiation, revealing a positive feedback loop. The expression of Tcf4 and the resulting pDC differentiation were selectively sensitive to the inhibition of enhancer-binding BET protein activity. Thus, lineage-specifying function of E proteins is facilitated by lineage-specific isoform expression and by BET-dependent feedback regulation through distal regulatory elements.

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Substantial evidence suggests a role for immunotherapy in treating Alzheimer's disease (AD). Several monoclonal antibodies targeting aggregated forms of beta amyloid, have been shown to reduce amyloid plaques and in some cases, mitigate cognitive decline in early-stage AD patients. We sought to determine if genetically engineered macrophages could improve the targeting and degradation of amyloid plaques. Chimeric antigen receptor macrophages (CAR-Ms), which show promise as a cancer treatment, are an appealing strategy to enhance target recognition and phagocytosis of amyloid plaques in AD. We genetically engineered macrophages to express a CAR containing the anti-amyloid antibody aducanumab as the external domain and the Fc receptor signaling domain internally. CAR-Ms recognize and degrade beta amyloid; in vitro and on APP/PS1 brain slices ex vivo; however, when injected intrahippocampally, these first-generation CAR-Ms have limited persistence and fail to reduce plaque load. We overcame this limitation by creating CAR-Ms that secrete M-CSF and self-maintain without exogenous cytokines. These CAR-Ms have greater survival in the brain niche, and significantly reduce plaque load locally in vivo. These proof-of-principle studies demonstrate that CAR-Ms, previously only applied to cancer, may be utilized to target and degrade unwanted materials, such as amyloid plaques in the brains of AD mice.

Diagnostic tests that detect antibodies (AB) against SARS-CoV-2 for evaluation of seroprevalence and guidance of health care measures are important tools for managing the COVID-19 pandemic. Current tests have certain limitations with regard to turnaround time, costs and availability, particularly in point-of-care (POC) settings. We established a hemagglutination-based AB test (HAT) that is based on bi-specific proteins which contain a dromedary-derived antibody (nanobody) binding red blood cells (RBD) and a SARS-CoV-2-derived antigen, such as the receptor-binding domain of the Spike protein (Spike-RBD). While the nanobody mediates swift binding to RBC, the antigen moiety directs instantaneous, visually apparent hemagglutination in the presence of SARS-CoV-2-specific AB generated in COVID-19 patients or vaccinated individuals. Method comparison studies with assays cleared by emergency use authorization (EUA) demonstrate high specificity and sensitivity. To further increase objectivity of test interpretation, we developed an image analysis tool based on digital image acquisition (via a cell phone) and a machine learning algorithm based on defined sample-training and –validation datasets. Preliminary data, including a small clinical study, provides proof of principle for test performance in a POC setting. Together, the data support the interpretation that this AB test format, which we refer to as ‘NanoSpot.ai’, is suitable for POC testing, can be manufactured at very low costs and, based on its generic mode of action, can likely be adapted to a variety of other pathogens.