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. 2017 Oct 26;7(2):e1386828.
doi: 10.1080/2162402X.2017.1386828. eCollection 2018.

Increased Infiltration of M2-macrophages, T-cells and PD-L1 Expression in High Grade Leiomyosarcomas Supports Immunotherapeutic Strategies

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Free PMC article

Increased Infiltration of M2-macrophages, T-cells and PD-L1 Expression in High Grade Leiomyosarcomas Supports Immunotherapeutic Strategies

Marie Kostine et al. Oncoimmunology. .
Free PMC article

Abstract

Background: Immunotherapy may be a rational strategy in leiomyosarcoma (LMS), a tumor known for its genomic complexity. As a prerequisite for therapeutic applications, we characterized the immune microenvironment in LMS, as well as its prognostic value. Methods: CD163+ macrophages, CD3+ T-cells, PD-L1/PD-L2 and HLA class I expression (HCA2, HC10 and β2m) were evaluated using immunohistochemistry in primary tumors (n = 75), local relapses (n = 6) and metastases (n = 19) of 87 LMS patients, as well as in benign leiomyomas (n = 7). Correlation with clinicopathological parameters and survival analyses were assessed. Effect of LMS cells on macrophage differentiation was investigated using coculture of CD14+ monocytes with LMS cell lines or their conditioned media (CM). Results: 58% and 52% of the tumors were highly infiltrated with CD163+ macrophages and T-cells, respectively, with HLA class I expression observed in almost all tumors and PD-L1 expression in 30%. PD-L2 expression was also detected in some PD-L1+ tumors. All these immune markers correlated with high tumor grade but only CD163 associated with overall survival (p = 0.003) and disease-specific survival (p = 0.041). In vitro, CD163 was upregulated in the presence of LMS cells producing M-CSF, suggesting that this tumor drives macrophages towards the M2 phenotype. Conclusion: The clinical significance of M2 macrophages, possibly induced by LMS cell-secreted factors, suggests that 2/3 of high-grade LMS patients might benefit from macrophage-targeting agents. Furthermore, PD-L1 expression together with high T-cell infiltrate and HLA class I expression in around 30% of high grade LMS reflects an active immune microenvironment potentially responsive to immune checkpoint inhibitors.

Keywords: HLA; PD-L1, immunotherapy; leiomyosarcoma; tumor-associated macrophages; tumor-infiltrating lymphocytes.

Figures

Figure 1.
Figure 1.
CD163 infiltrate in leiomyosarcoma. Representative images of primary leiomyosarcoma with low CD163 infiltrate (≤ 20%) (A) and high CD163 infiltrate (>20%) (B) using immunohistochemistry (scale bars 50 μm). The same pattern of CD163 infiltration was observed in the primary tumor and in the associated relapse/metastasis (C). Overall, 60% of leiomyosarcomas were highly infiltrated with CD163-positive cells, which strongly correlated with tumor grade (D). CD14-positive cells were differentiated for 6 days with GM-CSF or M-CSF as controls for M1 and M2 phenotype, respectively, and with leiomyosarcoma cells (LMS04, LMS05) using transwell or their conditioned media (CM). Expression of the surface marker CD163 (M2) on differentiated CD14-positive cells was analyzed by flow cytometry (E). Bars indicate relative geometric mean fluorescence intensity (MFI) ± standard error of mean (SEM) of three independent healthy donors, normalized to the M-CSF condition. Nonparametric Mann-Whitney test or the Kruskal-Wallis test followed by Dunn's post-test were used to compare differences between conditions. *p < 0.05. M-CSF measurement using a Luminex assay on LMS04 and LMS05 conditioned media (F). Results are expressed in pg/ml (n = 2).
Figure 2.
Figure 2.
Prognostic significance of CD163, CD3 and PD-L1 in leiomyosarcoma. Kaplan-Meier survival curves for overall survival (A, D, G), disease-specific survival (B, E, H) and disease-free survival (C, F, I) according to CD163 infiltration (low n = 27; high n = 48), CD3 infiltration (low n = 30; high n = 43) and PD-L1 expression (negative n = 46; positive n = 28) in primary leiomyosarcomas. A high CD163 infiltrate (> 20%) is associated with poor overall and disease specific survival. P-value obtained by log-rank test.
Figure 3.
Figure 3.
HLA class I expression in leiomyosarcoma. Representative staining patterns of β2-microglobulin, HLA-A (HCA2 antibody) and HLA-B/C (HC10 antibody) expression in smooth muscle cells from blood vessel, leiomyoma and leiomyosarcoma using immunohistochemistry. In general, basal low HLA class I expression was found on normal smooth muscle cells while frequently heterogeneous or strongly positive expression was seen in leiomyosarcomas, suggesting that HLA class I is upregulated in the majority of these tumors. Scale bars, 50 μm.
Figure 4.
Figure 4.
Tumor-infiltrating lymphocytes and PD-L1 expression in leiomyosarcoma categorized with the TIME classification. Representative images for PD-L1 and CD3 immunostaining in leiomyosarcoma patients. Scale bars, 50 μm. According to the TIME classification, 39 tumors exhibited the T1 subtype (TILs, PD-L1), 23 the T2 subtype (PD-L1+, TIL+), 25 the T3 subtype (PD-L1, TIL+) and 6 the T4 subtype (PD-L1+, TIL). Distribution of tumor grades within each subtype is represented by a bar chart.

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References

    1. Lazar A, Evans H, Shipley J. Leiomyosarcoma In: Fletcher CDM, Bridge JA, Hogendoorn PC, Mertens F., editors. WHO Classification of Tumours of Soft Tissue and Bone. 4th ed Lyon, France: IARC Press, 2013; p. 111–13.
    1. ESMO/European Sarcoma Network Working Group Soft tissue and visceral sarcomas: ESMO Clinical Practice Guidelines for diagnosis, treatment and follow-up. Ann Oncol. 2014;25 Suppl 3:iii102–12. doi:10.1093/annonc/mdu254. PMID:25210080 - DOI - PubMed
    1. Duffaud F, Ray-Coquard I, Salas S, Pautier P. Recent advances in understanding and managing leiomyosarcomas. F1000prime Rep. 2015;7:55. doi:10.12703/P7-55. PMID:26097728 - DOI - PMC - PubMed
    1. Penel N, Italiano A, Isambert N, Bompas E, Bousquet G, Duffaud F, French Sarcoma Group. (Groupe Sarcome Français/Groupe d'Etude des Tumeurs Osseuses) Factors affecting the outcome of patients with metastatic leiomyosarcoma treated with doxorubicin-containing chemotherapy. Ann Oncol. 2010;21(6):1361–5. doi:10.1093/annonc/mdp485. PMID:19880438 - DOI - PubMed
    1. Nabeshima A, Matsumoto Y, Fukushi J, Iura K, Matsunobu T, Endo M, Fujiwara T, Iida K, Fujiwara Y, Hatano M, et al. Tumour-associated macrophages correlate with poor prognosis in myxoid liposarcoma and promote cell motility and invasion via the HB-EGF-EGFR-PI3 K/Akt pathways. Br J Cancer. 2015;112(3):547–55. doi:10.1038/bjc.2014.637. PMID:25562433 - DOI - PMC - PubMed

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