Epigenetic regulation of cell state by H2AFY governs immunogenicity in high-risk neuroblastoma
Divya Nagarajan, Rebeca T. Parracho, David Corujo, Minglu Xie, Ginte Kutkaite, Thale K. Olsen, Marta Rubies Bedos, Maede Salehi, Ninib Baryawno, Michael P. Menden, Xingqi Chen, Marcus Buschbeck, Yumeng Mao
Divya Nagarajan, Rebeca T. Parracho, David Corujo, Minglu Xie, Ginte Kutkaite, Thale K. Olsen, Marta Rubies Bedos, Maede Salehi, Ninib Baryawno, Michael P. Menden, Xingqi Chen, Marcus Buschbeck, Yumeng Mao
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Research Article Immunology Oncology

Epigenetic regulation of cell state by H2AFY governs immunogenicity in high-risk neuroblastoma

Abstract

Childhood neuroblastoma with MYCN amplification is classified as high risk and often relapses after intensive treatments. Immune checkpoint blockade therapy against the PD-1/L1 axis shows limited efficacy in patients with neuroblastoma, and the cancer intrinsic immune regulatory network is poorly understood. Here, we leverage genome-wide CRISPR/Cas9 screens and identify H2AFY as a resistance gene to the clinically approved PD-1 blocking antibody nivolumab. Analysis of single-cell RNA-Seq datasets reveals that H2AFY mRNA is enriched in adrenergic cancer cells and is associated with worse patient survival. Genetic deletion of H2afy in MYCN-driven neuroblastoma cells reverts in vivo resistance to PD-1 blockade by eliciting activation of the adaptive and innate immunity. Mapping of the epigenetic and translational landscape demonstrates that H2afy deletion promotes cell transition to a mesenchymal-like state. With a multiomics approach, we uncovered H2AFY-associated genes that are functionally relevant and prognostic in patients. Altogether, our study elucidates the role of H2AFY as an epigenetic gatekeeper for cell states and immunogenicity in high-risk neuroblastoma.

Authors

Divya Nagarajan, Rebeca T. Parracho, David Corujo, Minglu Xie, Ginte Kutkaite, Thale K. Olsen, Marta Rubies Bedos, Maede Salehi, Ninib Baryawno, Michael P. Menden, Xingqi Chen, Marcus Buschbeck, Yumeng Mao

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Figure 5

Genetic deletion of H2afy in NB cancer cells reverted resistance to PD-1 blockade.

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Genetic deletion of H2afy in NB cancer cells reverted resistance to PD-1...
(A) The H2afy gene was targeted by CRISPR/Cas9 in the MYCN-driven 9464D cancer cells. Expression of the H2AFY protein was detected using Western blotting at different cell passages. Representative blot of 2 biological repeats. (B) Proliferation of control (ctrl) and KO 9464D cells was compared using the Incucyte live-cell imaging system. A representative experiment of 3 biological repeats. (C) Treatment schedule of mice bearing ctrl or KO 9464D cells. (D–F) Comparison of average tumor volumes (mean±SEM), growth of individual tumors and survival between mice bearing s.c. ctrl 9464D tumors that were treated i.p. with a rat IgG2a isotype control (clone 2A3) or an αPD-1 antibody (clone RMP1-14) at 200 μg per mouse, 9 or 10 mice per group. (G–I) Comparison of average tumor volumes (mean±SEM), growth of individual tumors and survival between mice bearing s.c. H2afy-KO 9464D tumors that were treated with the rat IgG isotype or αPD-1 at 200 μg per mouse, 9 or 10 mice per group. 1 day before IgG or α-PD1 treatment, mice bearing H2afy-KO 9464D tumors were treated with depletion antibodies against (J) CD4+ T cells (clone GK1.5) or (K) CD8+ T cells (clone 2.43) at 100 μg per mouse (i.p.) every 5 days, 6–8 mice per group. Tumor growth was compared among groups using 2-way ANOVA. Survival of mice in different groups was depicted using Kaplan-Meier curves with a Log-rank (Mantel-Cox) test.