As a cellular protease, transmembrane serine protease 2 (TMPRSS2) plays roles in various physiological and pathological processes, including cancer and viral entry, such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Herein, we conducted expression, mutation, and prognostic analyses for the TMPRSS2 gene in pan-cancers as well as in COVID-19-infected lung tissues. The results indicate that TMPRSS2 expression was highest in prostate cancer. A high expression of TMPRSS2 was significantly associated with a short overall survival in breast invasive carcinoma (BRCA), sarcoma (SARC), and uveal melanoma (UVM), while a low expression of TMPRSS2 was significantly associated with a short overall survival in lung adenocarcinoma (LUAD), demonstrating TMPRSS2 roles in cancer patient susceptibility and severity. Additionally, TMPRSS2 expression in COVID-19-infected lung tissues was significantly reduced compared to healthy lung tissues, indicating that a low TMPRSS2 expression may result in COVID-19 severity and death. Importantly, TMPRSS2 mutation frequency was significantly higher in prostate adenocarcinoma (PRAD), and the mutant TMPRSS2 pan-cancer group was significantly associated with long overall, progression-free, disease-specific, and disease-free survival rates compared to the wild-type (WT) TMPRSS2 pan-cancer group, demonstrating loss of functional roles due to mutation. Cancer cell lines were treated with small molecules, including cordycepin (CD), adenosine (AD), thymoquinone (TQ), and TQFL12, to mediate TMPRSS2 expression. Notably, CD, AD, TQ, and TQFL12 inhibited TMPRSS2 expression in cancer cell lines, including the PC3 prostate cancer cell line, implying a therapeutic role for preventing COVID-19 in cancer patients. Together, these findings are the first to demonstrate that small molecules, such as CD, AD, TQ, and TQFL12, inhibit TMPRSS2 expression, providing novel therapeutic strategies for preventing COVID-19 and cancers.

COVID-19 is an acute respiratory disease caused by SARS-CoV-2 that has spawned a worldwide pandemic. ADAM17 is a sheddase associated with the modulation of the receptor ACE2 of SARS-CoV-2. Studies have revealed that malignant phenotypes of several cancer types are closely relevant to highly expressed ADAM17. However, ADAM17 regulation in SARS-CoV-2 invasion and its role on small molecules are unclear. Here, we evaluated the ADAM17 inhibitory effects of cordycepin (CD), thymoquinone (TQ), and N6, N6-dimethyladenosine (m⁶₂A), on cancer cells and predicted the anti-COVID-19 potential of the three compounds and their underlying signaling pathways by network pharmacology. It was found that CD, TQ, and m⁶₂A repressed the ADAM17 expression upon different cancer cells remarkably. Moreover, CD inhibited GFP-positive syncytia formation significantly, suggesting its potential against SARS-CoV-2. Pharmacological analysis by constructing CD-, TQ-, and m⁶₂A-based drug-target COVID-19 networks further indicated that ADAM17 is a potential target for anti-COVID-19 therapy with these compounds, and the mechanism might be relevant to viral infection and transmembrane receptors-mediated signal transduction. These findings imply that ADAM17 is of potentially medical significance for cancer patients infected with SARS-CoV-2, which provides potential new targets and insights for developing innovative drugs against COVID-19.

NRP1/CD304 is a typical membrane-bound co-receptor for the vascular endothelial cell growth factor (VEGF), semaphorin family members, and viral SARS-CoV-2. Cordycepin (CD) is a natural product or active gradient from traditional Chinese medicine (TCM) from Cordyceps militaris Link and Ophiocordyceps sinensis (Berk.). However, NRP1 expression regulation via CD in cancers and the potential roles and mechanisms of SARS-CoV-2 infection are not clear. In this study, online databases were analyzed, Western blotting and quantitative RT-PCR were used for NRP1 expression change via CD, molecular docking was used for NRP/CD interaction, and a syncytial formation assay was used for CD inhibition using a pseudovirus SARS-CoV-2 entry. As a result, we revealed that CD inhibits NRP1 expressed in cancer cells and prevents viral syncytial formation in 293T-hACE2 cells, implying the therapeutic potential for both anti-cancer and anti-viruses, including anti-SARS-CoV-2. We further found significant associations between NRP1 expressions and the tumor–immune response in immune lymphocytes, chemokines, receptors, immunostimulators, immune inhibitors, and major histocompatibility complexes in most cancer types, implying NRP1’s roles in both anti-cancer and anti-SARS-CoV-2 entry likely via immunotherapy. Importantly, CD also downregulated the expression of NRP1 from lymphocytes in mice and downregulated the expression of A2AR from the lung cancer cell line H1975 when treated with CD, implying the NRP1 mechanism probably through immuno-response pathways. Thus, CD may be a therapeutic component for anti-cancer and anti-viral diseases, including COVID-19, by targeting NRP1 at least.

TQFL12 is a novel derivative designed and synthesized on the basis of Thymoquinone (TQ) which is extracted from Nigella sativa seeds. We have demonstrated that TQFL12 was more effective in the treatment of TNBC than TQ. In order to directly reflect the acute toxicity of TQFL12 in vivo, in this study, we designed, synthesized, and compared it with TQ. The mice were administered drugs with different concentration gradients intraperitoneally, and death was observed within one week. The 24 h median lethal dose (LD₅₀) of TQ was calculated to be 33.758 mg/kg, while that of TQFL12 on the 7th day was 81.405 mg/kg, and the toxicity was significantly lower than that of TQ. The liver and kidney tissues of the dead mice were observed by H&E staining. The kidneys of the TQ group had more severe renal damage, while the degree of the changes in the TQFL12 group was obviously less than that in the TQ group. Western blotting results showed that the expressions of phosphorylated levels of adenylate-activated protein kinase AMPKα were significantly up-regulated in the kidneys of the TQFL12 group. Therefore, it can be concluded that the acute toxicity of TQFL12 in vivo is significantly lower than that of TQ, and its anti-toxicity mechanism may be carried out through the AMPK signaling pathway, which has a good prospect for drug development.

Although thymoquinone (TQ) has been reported as an anti-tumor small molecule well investigated in numerous tumors. In this study, we designed and synthesized a novel TQ derivative, TQFL28, with a molecular formula of C₂₀H₂₃NO₂. TQFL28 showed stronger cytotoxicity or anti-proliferative activities against triple-negative breast cancer (TNBC) cell lines (BT549, MDA-MB-231, or 4T1) than TQ but is lower in the normal mammary epithelial cells, MCF10A. TQFL28 exhibited lower IC₅₀ values toward BT549 (38.78 ± 1.589) and MDA-MB-231 (39.63 ± 1.598) cells compared to TQ, indicating its efficacy for TNBC cytotoxicity. TQFL28 inhibited the growth, migration, and invasiveness of TNBC cells of 4T1 and BT549 in vitro and tumor progression and metastasis in a 4T1 allograft animal model in vivo. Moreover, TQFL28 presents lower toxicity than TQ in mice, showing a 7-day half-lethal dose (LD₅₀) of 59.43 mg/kg (41.6–83.6, 95% confidence interval). Altogether, our study obtained. In addition, TQFL28 induced a significant reduction in tumor volumes in the mouse model in comparison to the vehicle group. TQFL28, a novel small molecule, has a superior inhibitory effect and lower toxicity on TNBC both in vitro and in vivo. Thus, TQFL28 might have potential as a therapeutic small molecule for breast cancer, especially in TNBC.