The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been shown to affect cancer patients at a higher rate than healthy people. The role played by the host enzyme TMPRSS2 in infection and cancer is the focus of a new study available on Research Square* and under consideration at Journal of Translational Medicine that suggests this gene encoding this molecule is a tumor suppressor gene.
Introduction
Earlier studies showed that the transmembrane serine protease TMPRSS2 is key to the cell entry of SARS-CoV-2. The virus accomplishes host cell attachment and invasion via its spike glycoprotein. The S1 subunit binds the angiotensin-converting enzyme 2 (ACE2) host cell receptor, undergoing a conformational change that exposes the S2 subunit. This achieves membrane fusion and viral endocytosis, following which the virus undergoes replication and transcription to form new viral particles.
TMPRSS2 is found at higher levels in many tumors, including lung adenocarcinoma (LUAD) and lung squamous cell carcinoma (LUSC). An earlier study postulated that TMPRSS2 acted as a tumor suppressor and is downregulated in LUAD tissue compared to normal tissue.
In the current study, the researchers examined the associations between the level of TMPRSS2 expression and the immune phenotype in five LUAD cell types – CD8+ T cells, natural killer (NK) cells, CD4+ T regulatory cells (Tregs), myeloid-derived suppressor cells (MDSCs), and PD-L1. They also looked at oncogenic pathway activity in association with TMPRSS2 expression, as well as its link with tumor phenotype, genomic characteristics, and the outcome.
What did the study show?
TMPRSS2 expression was negatively correlated with the presence of all five types of immune cells. CD8+ T cells, as well as NK cells, are involved in antitumor immunity via the adaptive and innate arms, and the levels of both were increased with low levels of TMPRSS2 expression.
Immune cytolytic activity was also inversely related to TMPRSS2 expression, and so was PD-L-1. Meanwhile, CD4+ Tregs and MDSC infiltration, which denote tumor immunosuppression, showed a negative correlation with TMPRSS2. Overall, low levels of this molecule indicate high immune infiltration in LUAD.
The ratio of CD8+ T cells/PD-L1, representing immune-stimulatory and immune inhibitory cells, respectively, were positively associated with TMPRSS2 expression, showing that the latter falls more steeply with immunosuppressive phenotypes. The same negative correlation was present with cell cycle, mismatch repair, and other related signaling pathways.
Tumor proliferation marker MKI67 was also increased with low TMPRSS2 expression, indicating stemness of the tumor cell and therefore increased chances of a poor outcome. With low levels of this marker, TMPRSS2, LUAD tumors were more advanced in stage and larger in size, and thus the survival odds were lower.
Earlier research has established that the presence of EGFR mutations is associated with a better prognosis compared to the wild-type EGFR in LUAD. The former was associated with higher TMPRSS2 expression than the latter. Another favorable prognostic marker is TRU (terminal respiratory unit), which was also inversely associated with TMPRSS2 levels.
Genomic instability is an indicator of a poorer prognosis in cancer since it is associated with increased initiation, progression, and invasion. Either mutation numbers, aneuploidy, or copy numbers of somatic chromosomes may be involved. TMPRSS2 downregulation is linked to higher levels of gene instability.
Homologous recombination deficiency (HRD) promotes such instability and has been found to be higher with low TMPRSS2 expression.
Harmful mutations were previously found in nine DDR pathways in TCGA cancers. The current study classified LUAD into those with and without mutations in each pathway. This showed that TMPRSS2 levels were markedly lower in the tumors with mutated pathways for seven of the pathways, such as damage sensing, excision repair of nucleotides and of bases, mismatch repair, and homologous recombination.
TMPRSS2 expression was also lower in cases when DDR molecules were expressed at higher levels, this being the marker of increased genomic instability.
Several other genes sets dealing with the cell cycle, p53 signaling, mismatch repair, and homologous recombination pathways were all upregulated with lower TMPRSS2 expression levels. Six gene modules were highly enriched in LUADs with high TMPRSS2 levels, conversely forming an ECM signature that is central to cancer progression.
These findings were validated using in vitro human cell lines and in vivo mouse models. In other words, TMPRSS2 knockdown led to significantly higher tumor cell proliferation and invasiveness in the cell model and larger progressive tumors in the mouse experiments.
Bioinformatics also showed that immune infiltration in LUAD occurred more frequently with lower TMPRSS2 expression. At the same time, MHC class I genes were expressed at higher levels, and NK cells showed higher proliferation rates. In vivo too, CD8+ T cells and NK cells showed an increase in infiltration as well as higher activation levels, with TMPRSS2 knockdown, as signified by the production of TNF-α and IFN-γ. Exhaustion markers in these tumors were increased simultaneously.
PD-L1 is a biomarker of the tumor response to immune checkpoint inhibitors in cancer treatment. With TMPRSS2 knockdown, tumor volume reduced more than that of wild-type tumors following treatment with one such inhibitor. This agrees with the finding that PD-L1 expression increases with the knockdown of TMPRSS2.
In the same treatment scenario, both CD8+ T cells and NK cells infiltrating the tumor showed higher activity levels in TMPRSS2 knockdown tumors than in wild-type tumors. The inhibitor seems to enhance immune-mediated tumor cell elimination by blocking exhaustion of CD8+ and NK tumor-infiltrating lymphocytes in the LUAD with TMPRSS2 knockdown.
What are the implications?
The TMPRSS2 molecule jumped into the limelight with SARS-CoV-2, being essential for its successful entry into the target host cell. However, the current study shows that in LUAD, the most common type of lung cancer, TMPRSS2 is a tumor suppressor. With low levels of this enzyme, tumor-inducing pathways show enhanced activity.
With the downregulation of TMPRSS2, tumor cell proliferation increases with stem cell characteristics, genomic instability, and tumor stage advancement. The odds of survival dip. Experiments in vitro and in vivo confirmed the key role played by TMPRSS2 in LUAD proliferation and invasiveness.
This molecule appears to be lowest when immune inhibition phenotypes are present vs. immunostimulatory signatures. This could explain why patients with LUAD showing TMPRSS2 deficiency have worse outcomes. In vivo experiments demonstrated the increased sensitivity of TMPRSS2-knockdown tumors to the PD-1/PD-L1 inhibitor, making the former a positive marker of immunotherapy in this disease.
Given the tumor suppressor activity of TMPRSS2, its inhibition may tend to promote increased tumor growth and invasiveness. Surprisingly, non-smokers with LUAD had higher rates of infection with SARS-CoV-2 than smokers with this tumor.
TMPRSS2 is a tumor suppressor in LUAD, as evidenced by its downregulation correlated with increased genomic instability, tumor progression, and unfavorable clinical outcomes in LUAD. However, TMPRSS2 downregulation is a positive biomarker of immunotherapy for LUAD. Our data provide implications in the connection between lung cancer and pneumonia caused by SARS-CoV-2 infection.”
