Lysosomal processing of sulfatide analogs alters target NKT cell specificity and immune responses in cancer
Kumiko Nishio, … , Amy R. Howell, Jay A. Berzofsky
Kumiko Nishio, … , Amy R. Howell, Jay A. Berzofsky
Published December 21, 2023
Citation Information: J Clin Invest. 2024;134(4):e165281. https://doi.org/10.1172/JCI165281.
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Research Article Cell biology Immunology

Lysosomal processing of sulfatide analogs alters target NKT cell specificity and immune responses in cancer

Abstract

In a structure-function study of sulfatides that typically stimulate type II NKT cells, we made an unexpected discovery. We compared analogs with sphingosine or phytosphingosine chains and 24-carbon acyl chains with 0-1-2 double bonds (C or pC24:0, 24:1, or 24:2). C24:1 and C24:2 sulfatide presented by the CD1d monomer on plastic stimulated type II, not type I, NKT cell hybridomas, as expected. Unexpectedly, when presented by bone marrow–derived DCs (BMDCs), C24:2 reversed specificity to stimulate type I, not type II, NKT cell hybridomas, mimicking the corresponding β-galactosylceramide (βGalCer) without sulfate. C24:2 induced IFN-γ–dependent immunoprotection against CT26 colon cancer lung metastases, skewed the cytokine profile, and activated conventional DC subset 1 cells (cDC1s). This was abrogated by blocking lysosomal processing with bafilomycin A1, or by sulfite blocking of arylsulfatase or deletion of this enyzme that cleaves off sulfate. Thus, C24:2 was unexpectedly processed in BMDCs from a type II to a type I NKT cell–stimulating ligand, promoting tumor immunity. We believe this is the first discovery showing that antigen processing of glycosylceramides alters the specificity for the target cell, reversing the glycolipid’s function from stimulating type II NKT cells to stimulating type I NKT cells, thereby introducing protective functional activity in cancer. We also believe our study uncovers a new role for antigen processing that does not involve MHC loading but rather alteration of which type of cell is responding.

Authors

Kumiko Nishio, Lise Pasquet, Kaddy Camara, Julia DiSapio, Kevin S. Hsu, Shingo Kato, Anja Bloom, Stewart K. Richardson, Joshua A. Welsh, Tianbo Jiang, Jennifer C. Jones, Susanna Cardell, Hiroshi Watarai, Masaki Terabe, Purevdorj B. Olkhanud, Amy R. Howell, Jay A. Berzofsky

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Figure 6

Sulfatide analogs stimulate type I NKT cells but lose their ability to stimulate type II NKT cells when presented by BMDCs.

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Sulfatide analogs stimulate type I NKT cells but lose their ability to s...
The reactivity of type I or type II NKT cells stimulated with BMDCs prepulsed with sulfatide analogs was evaluated by ELISA to determine IL-2 levels in the culture media. (A) The type II NKT hybridoma clone XV19 (left) and the type I NKT hybridoma clone DN32 (right) were stimulated for 16 hours with BMDCs prepulsed with sulfatide analogs (10 μM) or KRN7000 (25 nM), or with 0.25 μg plate-bound anti-CD3 antibody in the presence of 10 μg/mL rat IgG or anti-CD1d antibody (20H2). Data indicate the mean ± SD of triplicate cultures. (B–D) The type II NKT hybridoma clone XV19 (B) and the type I NKT hybridoma clones DN32 (C) and 24.9E (D) were stimulated for 16 hours with BMDCs prepulsed with sulfatide analogs (10 μM) or KRN7000 (25 nM), or with 0.25 μg plate-bound anti-CD3 antibody in the presence of culture media, 0.03% DMSO, or 50 nM bafilomycin A1 (BAF). Data represent 2 experiments and the mean ± SD of triplicate cultures.