Beyond bacteria: How the intratumor mycobiome modulates lung adenocarcinoma progression

Abstract

Increasing evidence suggests that tumors harbor diverse microbiomes, adding complexity to the tumor microenvironment. In this issue of Cancer Cell, Liu et al. highlight the role of the intratumor mycobiome, specifically Aspergillus sydowii, in promoting lung adenocarcinoma progression. A. sydowii enhances the recruitment and activation of myeloid-derived suppressor cells via IL-1β signaling driven by the β-glucan-mediated Dectin-1/CARD9 pathway.

Among the myriad of factors contributing to cancer pathogenesis and prognosis, the microbiome has emerged as a key player in modulating tumorigenesis and response to therapy. While many studies have underscored the role of bacteria and, to some extent, viruses in tumor progression, the influence of fungi or the mycobiome within this intricate landscape remains a largely uncharted territory.¹ Despite its lower biomass compared to the bacteriome, the mycobiome has an undeniable influence on health and disease. Not only are fungi significant commensals within the human body but they can also manifest as opportunistic pathogens particularly in individuals with immunocompromised immune systems, such as patients with cancer.² Remarkably, the mycobiome has been identified across a wide range of human cancers.³ Yet, despite these insights, in-depth studies interrogating the link between the fungal microbiome and cancer remain very few.^4-6

In the study by Liu et al., the researchers identified an enrichment of the fungus Aspergillus sydowii within tumors of patients with lung adenocarcinoma (LUAD).⁷ They also revealed that this fungus plays a pivotal role in lung tumor progression. Specifically, A. sydowii appears to promote tumor progression via IL-1β-mediated expansion and activation of myeloid-derived suppressor cells (MDSCs), which in turn suppress cytotoxic T lymphocyte activity and lead to an accumulation of T regulatory cells (Tregs) and PD-1+ CD8⁺ T cells.

First, the study identified a higher abundance of fungi in LUAD tissues compared to matched non-tumor tissues. Using shotgun metagenomic deep sequencing, the authors found a significant enrichment of A. sydowii within tumor tissues. Beyond mere statistical presence, Liu and colleagues verified viability of A. sydowii within tumors, emphasizing its potentially active role within the tumor ecosystem.

Next, delving deeper into functional studies, the authors utilized murine models to elucidate the role of A. sydowii in LUAD progression. Intratumoral injection of A. sydowii into the Lewis Lung Carcinoma (LLC) model accelerated LUAD progression by fostering an immunosuppressive tumor microenvironment (TME). This is underscored by the upregulation of genes related to immunosuppressive pathways and a notable increase in the proportion of MDSCs and Tregs. To further validate their findings in a setting that better emulates the TME of the lungs, the authors employed an orthotopic LUAD murine model using Kras^LSL–G12D/+/P53^fl/fl (KP) cells as well as a spontaneous KP LUAD mouse model. In line with findings from patient samples, tumors exposed to A. sydowii bore an elevated fungal load compared to matching normal tissues, emphasizing the need to identify the factors that promote such colonization within tumor environments. Notably, this fungal exposure led to increased IL-1β secretion, catalyzing the recruitment of MDSCs from bone marrow and their expansion. Furthermore, A. sydowii seemed to drive the functional maturation of these MDSCs as evidenced by augmented arginase activity and increased NO and ROS production. This robust MDSC activity was directly linked to the suppressed ability of T cells to kill LUAD cells in vitro, as well as driving the polarization of primary CD4⁺ T cells into Tregs. Additionally, A. sydowii-activated MDSCs, when co-cultured with CD8⁺ T cells, increased PD-1 expression, possibly underlying the observed PD1+ CD8⁺ T cells expansion in the TME (Figure 1). Indeed, oral and inhaled amphotericin B, an anti-fungal, proved to be effective in halting tumor growth. Furthermore, in vivo MDSC neutralization reduced tumor progression in the A. sydowii-treated orthotopic LUAD model, concurrently decreasing MDSCs, Tregs, and PD-1₊ CD8⁺ T cells. The authors showed that exposure to A. sydowii’s cell wall component, β-glucan, transforms bone marrow-derived macrophages (BMDMs) into MDSCs through the Dectin-1/CARD9 signaling pathway (Figure 1). Elevated IL-1β secretion by BMDMs upon exposure to A. sydowii or β-glucan was central to this transformation and, thus, neutralizing IL-1β was found to reduce MDSC differentiation and tumor growth. The group also showed that absence of CARD9 reduced tumor growth by decreasing MDSCs while increasing cytotoxic T lymphocytes in the TME. Collectively, these findings highlight that A. sydowii drives MDSC differentiation by stimulating IL-1β production via the β-glucan-mediated Dectin-1/CARD9 signaling pathway, leading to enhanced lung tumor growth and an immunosuppressive environment (Figure 1).

Figure 1. Aspergillus sydowii-mediated immune modulation in lung adenocarcinoma progression.

Liu et al. describe how the intratumor mycobiome, especially Aspergillus sydowii, influences lung adenocarcinoma progression. A. sydowii triggers the expansion and activation of myeloid-derived suppressor cells through IL-1β signaling initiated by the β-glucan-triggered Dectin-1/CARD9 pathway. This then reduces cytotoxic T lymphocyte activity and results in more T regulatory cells (Tregs) and PD-1₊ CD8⁺ T cells, ultimately leading to tumor progression.

The translational significance of these findings was reflected by examining tumor samples from 92 patients with LUAD. The presence of A. sydowii directly correlated with increased immune suppression within tumors (MDSCs, Tregs, and PD-1₊ CD8⁺ T cells), aligning with findings from murine studies. Clinically, increased levels of A. sydowii were associated with aggressive tumor traits, lymph node invasion, more advanced stage, and poor prognosis. Together, the study not only underscores the significance of A. sydowii as a target to halt LUAD progression but also suggests its potential as a valuable clinical biomarker for patient prognosis.

The study mechanistically dissects the relationship between A. sydowii and host immune interactions. The emphasis on functional mechanistic studies is important, given the inherent challenges of contamination in microbiome research. The mechanism unveiled in this study both complements and builds upon preexisting knowledge derived from pancreatic ductal adenocarcinoma (PDAC). Certain PDAC models expressing the oncogenic Kras^G12D show a surge in IL-33 levels, a change influenced by the intratumoral fungal mycobiome. This elevation leads to the activation of T helper 2 (Th2) T and ILC2 cells, contributing to PDAC progression.⁴ A separate study has underscored possible fungal translocation from the gut to the pancreas. Consequently, PDACs display a unique mycobiotic composition, predominantly enriched with Malassezia species. Notably, the ligation of mannose-binding lectin (MBL) within the complement cascade has been identified as a crucial step in tumor progression.⁵ With multiple mechanisms now identified, it becomes imperative to discern the determinants guiding these specific mycobiome-immune system interactions. Are they modulated by distinct microbial taxa, or are there host genetic or epigenetic factors that impinge on the fungal-immune interface? A comprehensive understanding of these myriad interactions will be instrumental in harnessing the therapeutic potential of the mycobiome.

While the study provides invaluable insights into the role of the intratumor mycobiome, there are still gaps to be addressed. The study underscores the pivotal role of MDSCs in LUAD progression. Given the heterogeneity of this population, further studies are warranted to determine which specific subset is responsible for A. sydowii-mediated cancer progression.⁸ Crucial interactions between A. sydowii and other microbes are also inadequately characterized. Although genomic features and roles of A. sydowii in LUAD progression were explored, a deeper understanding of its virulence, commensal fitness, and interactions within the tumor microenvironment remains elusive. Notably, little is known of the temporal order between A. sydowii colonization and tumorigenesis; does the colonization by this fungus act as a precursor to tumor formation, or do tumors create a milieu conducive to A. sydowii establishment? One hypothesis is that inflammation or injury may promote fungal dysbiosis in the lung, which could, in turn, catalyze tumor progression. In this context, mechanisms by which A. sydowii reaches human lung tumors are yet to be delineated. It is also pertinent to explore how carcinogens like cigarette smoke could functionally influence early protumor roles of Aspergillus in the lung. Importantly, while the current correlations were drawn from a limited clinical cohort, they call for validation in larger and diverse patient groups to affirm their significance. Lastly, translating these findings into actionable therapeutics will need to be realized.