Henry, C. J. | bioRxiv Science

bioRxiv2020

In situ humoral selection in human lupus tubulointerstitial inflammation

ABSTRACTIn human lupus nephritis, tubulointerstitial inflammation (TII) is associated with in situ expansion of B cells expressing anti-vimentin antibodies (AVAs). The mechanism by which AVAs are selected is unclear. Herein, we demonstrate that AVA somatic hypermutation and selection increase affinity for vimentin. However, enzyme-linked immunosorbent assays (ELISAs) suggested that affinity maturation might be a non-specific consequence of increasing polyreactivity. Subsequent multi-color confocal microscopy studies indicated that while TII AVAs often appeared polyreactive by ELISA, they bound selectively to vimentin fibrils in whole cells or inflamed renal tissue. Using a novel machine learning pipeline (CytoSkaler) to quantify the cellular distribution of antibody staining, we demonstrated that TII AVAs were selected for both enhanced binding and specificity in situ. These data suggest a new approach to assess and define antibody polyreactivity based on quantifying the distribution of binding to native and contextually relevant antigens.Competing Interest StatementThe authors have declared no competing interest.View Full Text

immunology↗

bioRxiv2020

Rapid Assembly and Screening of Multivalent Immune Cell-Redirecting Therapies for Leukemia

Therapies that bind with immune cells and redirect their cytotoxic activity towards diseased cells represent a promising and versatile approach to immunotherapy with applications in cancer, lupus, and other diseases; traditional methods for discovering these therapies, however, are often time-intensive and lack the throughput of related target-based discovery approaches. Inspired by the observation that the cytokine, IL-12, can enhance antileukemic activity of the clinically approved T cell redirecting therapy, blinatumomab, here we describe the structure and assembly of a chimeric immune cell-redirecting agent which redirects the lytic activity of primary human T cells towards leukemic B cells and simultaneously co-targets the delivery of T cell-stimulating IL-12. We further describe a novel method for the parallel assembly of compositionally diverse libraries of these bi-specific T cell engaging cytokines (BiTEokines) and their high-throughput phenotypic screening, requiring just days for hit identification and the analysis of structure-function relationships. Using this approach, we identified CD19 x CD3 x IL12 compounds that exhibit ex vivo lytic activity comparable to current FDA-approved therapies for leukemia and correlated drug treatment with specific cell-cell contact, cytokine delivery, and leukemia cell lysis. Given the modular nature of these multivalent compounds and their rapid assembly/screening, we anticipate facile extension of this therapeutic approach to a wide range of immune cells, diseased cells, and soluble protein combinations in the future.

bioengineering↗