Mycobacterium bovis Bacillus Calmette-Guérin Alters Melanoma Microenvironment Favoring Antitumor T Cell Responses and Improving M2 Macrophage Function.

Ricardo D. Lardone,Peter A. Sieling,Leland J. Foshag,Agnes F. Lee,Mark B. Faries,Alfred A. Chan,Delphine J. Lee

doi:10.3389/fimmu.2017.00965

Abstract

Intralesional Mycobacterium bovis bacillus Calmette–Guérin (BCG) has long been a relatively inexpensive therapy for inoperable cutaneous metastatic melanoma (CMM), although intralesional BCG skin mechanisms remain understudied. We analyzed intralesional BCG-treated CMM lesions combined with in vitro studies to further investigate BCG-altered pathways. Since macrophages play a pivotal role against both cancer and mycobacterial infections, we hypothesized BCG regulates macrophages to promote antitumor immunity. Tumor-associated macrophages (M2) infiltrate melanomas and impair antitumor immunity. BCG-treated, in vitro-polarized M2 (M2-BCG) showed transcriptional changes involving inflammation, immune cell recruitment, cross talk, and activation pathways. Mechanistic network analysis indicated M2-BCG potential to improve interferon gamma (IFN-γ) responses. Accordingly, frequency of IFN-γ-producing CD4+ T cells responding to M2-BCG vs. mock-treated M2 increased (p < 0.05). Moreover, conditioned media from M2-BCG vs. M2 elevated the frequency of granzyme B-producing CD8+ tumor-infiltrating lymphocytes (TILs) facing autologous melanoma cell lines (p < 0.01). Furthermore, transcriptome analysis of intralesional BCG-injected CMM relative to uninjected lesions showed immune function prevalence, with the most enriched pathways representing T cell activation mechanisms. In vitro-infected MM-derived cell lines stimulated higher frequency of IFN-γ-producing TIL from the same melanoma (p < 0.05). Our data suggest BCG favors antitumor responses in CMM through direct/indirect effects on tumor microenvironment cell types including macrophages, T cells, and tumor itself.

Highlights

Cutaneous melanoma is predicted to cause approximately 9,700 deaths from metastatic disease in the United States in 2017 [1]
Our findings indicate that intralesional bacillus Calmette–Guerin (BCG) may stimulate the melanoma tumor microenvironment to promote antitumor immune responses by altering macrophage and T cell activities at the site of disease
To determine whether alternatively activated Mφs (M2)-BCG could increase interferon gamma (IFN-γ) production from T cells, we evaluated T cell responses using an IFN-γ-based Enzyme-Linked ImmunoSpot (ELISPOT) assay in which autologous T-cells were cocultured with different MΦs populations (Figure 4A)

Summary

Introduction

Cutaneous melanoma is predicted to cause approximately 9,700 deaths from metastatic disease in the United States in 2017 [1]. In-transit melanoma, metastasis is a pattern of lymphatic disease spread occurring in approximately 7% of patients [2]. Most patients with in-transit melanoma can suffer significant locoregional toxicity due to painful, bleeding, or necrotic lesions, which may become superinfected. They can develop stage IV systemic recurrence within 12–18 months [3]. Mycobacterium bovis bacillus Calmette–Guerin (BCG) is the best known agent for intralesional therapy of cutaneous melanoma metastases [5] and is a recommended therapeutic option in version 1.2017 NCCN Guidelines for inoperable stage III in-transit melanoma [6]. The use of BCG in cancer is not limited to melanoma; instillation of BCG is used as an intravesical therapy for superficial bladder cancer [12] and as an adjuvant in some cancer vaccines [13]

Methods

Results

Conclusion