TGF-ß orchestrates the phenotype and function of monocytic myeloid-derived suppressor cells in colorectal cancer.

BACKGROUND: Monocytic myeloid-derived suppressor cells (M-MDSCs) are significantly expanded in the blood of colorectal cancer (CRC) patients. However, their presence and underlying mechanisms in the tumour microenvironment of CRC have not been examined in detail. METHODS: Tumour tissues and peripheral blood from CRC patients were analysed for the presence of M-MDSCs. The mechanisms of suppression were analysed by blocking pathways by which MDSCs abrogate T cell proliferation. Co-culture of CRC cells with monocytes were performed with and without cytokine blocking antibodies to determine the mechanism by which CRC cells polarise monocytes. Multi-spectral IHC was used to demonstrate the intra-tumoral location of M-MDSCs. RESULTS: Tumour tissues and blood of CRC patients contain M-MDSCs which inhibit T cell proliferation. Whilst inhibition of arginase and nitric oxide synthase 2 fail to rescue T cell proliferation, blockade of IL-10 released by these HLA-DR- cells abrogates the suppresivity of M-MDSCs. Tumour conditioned media (TCM) significantly reduces HLA-DR expression, increases IL-10 release from monocytes and causes them to become suppressive. TGF-ß is highly expressed in the TCM and accumulates in the plasma. TGF-ß reduces HLA-DR expression and drives monocyte immunosuppressivity. The invasive margin of CRC is enriched in CD14+ HLA-DR- cells in close proximity to T cells. CONCLUSIONS: Our study demonstrates the cross-talk between CRC cells, M-MDSCs and T cells. Characterisation of CRC M-MDSCs point to therapeutic avenues to target these cells in addition to TGF-ß blockade.

MDSC targeting with Gemtuzumab ozogamicin restores T cell immunity and immunotherapy against cancers.

BACKGROUND: Targeting of MDSCs is a major clinical challenge in the era of immunotherapy. Antibodies which deplete MDSCs in murine models can reactivate T cell responses. In humans such approaches have not developed due to difficulties in identifying targets amenable to clinical translation. METHODS: RNA-sequencing of M-MDSCs and G-MDSCs from cancer patients was undertaken. Flow cytometry and immunohistochemistry of blood and tumours determined MDSC CD33 expression. MDSCs were treated with Gemtuzumab ozogamicin and internalisation kinetics, and cell death mechanisms determined by flow cytometry, confocal microscopy and electron microscopy. Effects on T cell proliferation and CAR-T cell anti-tumour cytotoxicity were identified in the presence of Gemtuzumab ozogamicin. FINDINGS: RNA-sequencing of human M-MDSCs and G-MDSCs identified transcriptomic differences, but that CD33 is a common surface marker. Flow cytometry indicated CD33 expression is higher on M-MDSCs, and CD33+ MDSCs are found in the blood and tumours regardless of cancer subtype. Treatment of human MDSCs leads to Gemtuzumab ozogamicin internalisation, increased p-ATM, and cell death; restoring T cell proliferation. Anti-GD2-/mesothelin-/EGFRvIII-CAR-T cell activity is enhanced in combination with the anti-MDSC effects of Gemtuzumab ozogamicin. INTERPRETATION: The study identifies that M-MDSCs and G-MDSCs are transcriptomically different but CD33 is a therapeutic target on peripheral and infiltrating MDSCs across cancer subtypes. The immunotoxin Gemtuzumab ozogamicin can deplete MDSCs providing a translational approach to reactivate T cell and CAR-T cell responses against multiple cancers. In the rare conditions of HLH/MAS gemtuzumab ozogamicin provides a novel anti-myeloid strategy. FUND: This work was supported by Cancer Research UK, CCLG, Treating Children with Cancer, and the alumni and donors to the University of Birmingham.