Dexamethasone promotes renal fibrosis by upregulating ILT4 expression in myeloid-derived suppressor cells.

Studies have shown that adoptive transfer of myeloid-derived suppressor cells (MDSCs) can alleviate various inflammatory diseases, including glomerulonephritis, but the long-term effects of the transferred MDSCs are still unclear. In addition, although glucocorticoids exert immunosuppressive effects on inflammatory diseases by inducing the expansion of MDSCs, the impact of glucocorticoids on the immunosuppressive function of MDSCs and their molecular mechanisms are unclear. In this study, we found that adoptive transfer of MDSCs to doxorubicin-induced focal segmental glomerulosclerosis (FSGS) mice for eight consecutive weeks led to an increase in serum creatinine and proteinuria and aggravation of renal interstitial fibrosis. Similarly, 8 weeks of high-dose dexamethasone administration exacerbated renal interstitial injury and interstitial fibrosis in doxorubicin-induced mice, manifested as an increase in serum creatinine and proteinuria, collagen deposition and α-SMA expression. On this basis, we found that dexamethasone could enhance MDSC expression and secretion of the fibrosis-related cytokines TGF-ß and IL-10. Mechanistically, we revealed that dexamethasone promotes the expression of immunoglobulin-like transcription factor 4 (ILT4), which enhances the T-cell inhibitory function of MDSCs and promotes the activation of STAT6, thereby strengthening the expression and secretion of TGF-ß and IL-10. Knocking down ILT4 alleviated renal fibrosis caused by adoptive transfer of MDSCs. Therefore, our findings demonstrate that the role and mechanism of dexamethasone mediate the expression and secretion of TGF-ß and IL-10 in MDSCs by promoting the expression of ILT4, thereby leading to renal fibrosis.

Novel, pathogenic insertion variant of GSDME associates with autosomal dominant hearing loss in a large Chinese pedigree.

Nonsyndromic hearing loss (NSHL) is a genetically diverse, highly heterogeneous condition characterised by deafness, and Gasdermin E (GSDME) variants have been identified as directly inducing autosomal dominant NSHL. While many NSHL cases associated with GSDME involve the skipping of exon 8, there is another, less understood pathogenic insertion variant specifically found in Chinese pedigrees that causes deafness, known as autosomal dominant 5 (DFNA5) hearing loss. In this study, we recruited a large Chinese pedigree, conducted whole-exome and Sanger sequencing to serve as a comprehensive clinical examination, and extracted genomic DNA samples for co-segregation analysis of the members. Conservation and expression analyses for GSDME were also conducted. Our clinical examinations revealed an autosomal dominant phenotype of hearing loss in the family. Genetic analysis identified a novel insertion variant in GSDME exon 8 (GSDME: NM_004403.3: c.1113_1114insGGGGTGCAGCTTACAGGGTGGGTGT: p. P372fs*36). This variant is segregated with the deafness phenotype of this pedigree. The GSDME gene was highly conserved in the different species we analysed, and its mRNA expression was ubiquitously low in different human tissues. In conclusion, we have successfully identified a novel pathogenic insertion variant of GSDME in a Chinese pedigree that causes deafness, shedding light on the genetic basis of hearing loss within this specific family. Our findings expand the spectrum of known variants associated with GSDME-related deafness and may further support both the underlying gain-of-function mechanism and functional associations of GSDME hearing loss variants.

Comprehensive analysis, diagnosis, prognosis, and cordycepin (CD) regulations for GSDME expressions in pan-cancers.

The Gasdermin E gene (GSDME) plays roles in deafness and cancers. However, the roles and mechanisms in cancers are complex, and the same gene exhibits different mechanisms and actions in different types of cancers. Online databases, such as GEPIA2, cBioPortal, and DNMIVD, were used to comprehensively analyze GSDME profiles, DNA methylations, mutations, diagnosis, and prognosis in patients with tumor tissues and matched healthy tissues. Western blotting and RT-PCR were used to monitor the regulation of GSDME by Cordycepin (CD) in cancer cell lines. We revealed that GSDME expression is significantly upregulated in eight cancers (ACC, DLBC, GBM, HNSC, LGG, PAAD, SKCM, and THYM) and significantly downregulated in seven cancers (COAD, KICH, LAML, OV, READ, UCES, and UCS). The overall survival was longer only in ACC, but shorter in four cancers, including COAD, KIRC, LIHC, and STAD, when GSDME was highly expressed in cancers compared with the corresponding normal tissues. Moreover, the high expression of GSDME was negatively correlated with the poor prognosis of ACC, while the low expression of GSDME was negatively correlated with the poor prognosis of COAD, suggesting that GSDME might serve as a good prognostic factor in these two cancer types. Accordingly, results indicated that the DNA methylations of those 7 CpG sites constitute a potentially effective signature to distinguish different tumors from adjacent healthy tissues. Gene mutations for GSDME were frequently observed in a variety of tumors, with UCES having the highest frequency. Moreover, CD treatment inhibited GSDME expression in different cancer cell lines, while overexpression of GSDME promoted cell migration and invasion. Thus, we have systematically and successfully clarified the GSDME expression profiles, diagnostic values, and prognostic values in pan-cancers. Targeting GSDME with CD implies therapeutic significance and a mechanism for antitumor roles in some types of cancers via increasing the sensitivity of chemotherapy. Altogether, our study may provide a strategy and biomarker for clinical diagnosis, prognostics, and treatment of cancers by targeting GSDME.

Primary schwannoma of the thyroid gland: analysis of case characteristics and review of the literature.

BACKGROUND: To improve the characteristics of primary thyroid schwannomas (PTS) and to provide reference basis for clinical diagnosis and treatment. METHODS: PubMed was searched for case reports of PTS up to December 2022 using the search terms "Thyroid nerve sheath tumor" or "Thyroid schwannoma" or "Thyroid Neurilemmoma", respectively. 34 cases were screened. RESULTS: PTS can occur at any age, nodules averaged 3.9 cm. The most common symptoms were voice change and dysphagia. Fine needle aspiration cytology showing spindle-shaped cells should be considered for schwannoma. Most cases underwent thyroid lobectomy or nodule removal with a good prognosis. Tissue types with both Antoni A and Antoni B features are common. Positive immunohistochemical staining for S-100 protein, CD34 and waveform proteins helped confirm the diagnosis. CONCLUSIONS: Positive immunohistochemistry for S-100 and wave proteins helps confirm the diagnosis. Preoperative diagnosis is challenging, but pathology and immunohistochemical staining are the gold standard for diagnosis. The first choice of treatment is surgical resection of the nodules, the prognosis is good.

Impact of TMPRSS2 Expression, Mutation Prognostics, and Small Molecule (CD, AD, TQ, and TQFL12) Inhibition on Pan-Cancer Tumors and Susceptibility to SARS-CoV-2.

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.

Expressions and significances of the angiotensin-converting enzyme 2 gene, the receptor of SARS-CoV-2 for COVID-19.

The ACE2 gene is a receptor of SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) for COVID-19 (coronavirus disease 2019). To analyze the expression profiles and clinical significances for this gene in humans, RNA-seq data representing 27 different tissues were analyzed using NCBI; total RNA was extracted from different tissues of mouse and semi-quantitative reverse transcriptional-polymerase chain reaction (Q-RT-PCR) was carried out. Immunohistochemistry expression profiles in normal tissues and cancer tissues and TCGA survival analysis in renal and liver cancer were conducted. ACE2 was highly conserved in different species. In normal tissues, ACE2 expression distributions were organ-specific, mainly in the kidney, male testis and female breast, and cardiovascular and gastrointestinal systems. High level of expression in testis, cardiovascular and gastrointestinal system indicated that SARS-CoV-2 might not only attack the lungs, but also affect other organs, particularly the testes, thus it may severely damage male sexual development for younger male and lead to infertility in an adult male, if he contracted COVID-19. On the other side, high expression of ACE2 was correlated with increased survival rate in renal and liver cancer, indicating that ACE2 is a prognostic marker in both renal cancer and liver cancers. Thus, the ACE2 is a functional receptor for SARS-CoV-2 and has a potential anti-tumor role in cancer. Taken together, this study may not only provide potential clues for further medical pathogenesis of COVID-19 and male fertility, but also indicate the clinical significance of the role of the ACE2 gene in cancer.

Combined application of pharamcokinetic DCE-MRI and IVIM-DWI could improve detection efficiency in early diagnosis of ductal carcinoma in situ.

PURPOSE: Ductal carcinoma in situ (DCIS) is a precursor of invasive ductal breast carcinoma (IDC). This study aimed to use pharamcokinetic dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) and intravoxel incoherent motion diffusion-weighted imaging (IVIM-DWI) for the early diagnosis of DCIS. METHODS: Forty-seven patients, including 25 with DCIS (age: 28-70 yr, mean age: 48.7 yr) and 22 with benign disease (age: 25-67 yr, mean age: 43.1 yr) confirmed by pathology, underwent pharamcokinetic DCE-MRI and IVIM-DWI in this study. The quantitative parameters Ktrans , Kep , Ve , Vp , and D, f, D* were obtained by processing of DCE-MRI and IVIM-DWI images with Omni-Kinetics and MITK-Diffusion softwares, respectively. Parameters were analyzed statistically using GraphPad Prism and MedCalc softwares. RESULTS: All low-grade DCIS lesions demonstrated mass enhancement with clear boundaries, while most middle-grade and high-grade DCIS lesions showed non-mass-like enhancement (NMLE). DCIS lesions were significantly different from benign lesions in terms of Ktrans , Kep , and D (t = 5.959, P < 0.0001; t = 5.679, P < 0.0001; and t = 5.629, P < 0.0001, respectively). The AUC of Ktrans , Kep , D and the combined indicator of Ktrans , Kep, and D were 0.936, 0.902, 0.860, and 0.976, respectively. There was a significant difference in diagnostic efficacy only between D and the combined indicator (Z = 2.408, P = 0.016). CONCLUSION: DCE-MRI and IVIM-DWI could make for the early diagnosis of DCIS, and reduce the misdiagnosis of DCIS and over-treatment of benign lesions.

Establishment of stable cell line for inducing KAP1 protein expression.

Generation of the stable cell lines is a highly efficient tool in functional studies of certain genes or proteins, where the particular genes or proteins are inducibly expressed. The KRAB-associated protein-1 (KAP1) is an important transcription regulatory protein, which is investigated in several molecular biological studies. In this study, we have aimed to generate a stable cell line for inducing KAP1 expression. The recombinant plasmid pcDNA5/FRT/TO-KAP1 was constructed at first, which was then transfected into Flp-In™T-REx™-HEK293 cells to establish an inducible pcDNA5/FRT/TO-KAP1-HEK293 cell line. The Western blot analysis showed that the protein level of KAP1 is over-expressed in the established stable cell line by doxycycline induction, both dose and time dependently. Thus we have successfully established stable pcDNA5/FRT/TO-KAP1-HEK293 cell line, which can express KAP1 inducibly. This inducible cell line might be very useful for KAP1 functional studies.

Comprehensive analysis and immunohistochemistry localization of NRP1 expression in pancancer and normal individual tissues in relation to SARS­CoV­2 susceptibility.

Neuropilin 1 (NRP1/CD304) is a typical membrane-bound co-receptor for vascular endothelial growth factor, semaphorin family members and viral severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, NRP1 expression levels across cancer types and the potential role of SARS-CoV-2 infection in patients with cancer are not clear. Online databases, such as The Cancer Genome Atlas database of Human Protein Atlas, Gene Expression Profiling Interactive Analysis and cBioPortal were used for the expression analysis in this study. Immunohistochemical (IHC) staining for NRP1 was performed in the tissues of patients with non-small cell carcinoma. As a result, it was found that NRP1 mRNA and protein expression levels were highest in the female reproductive tissues and the respiratory system, specifically in the nasopharynx, bronchus and fallopian tube, as well as in adipocytes, hepatic stellate cells, Sertoli cells, endothelial cells and dendritic cells. IHC showed that the NRP1 protein was mainly localized to the cytoplasm and membrane in the tissues of patients with non-small cell carcinoma, demonstrating its role in lung infection by SARS-CoV-2, due to invasion of cell membranes by the virus. Levels of NRP1 mRNA were significantly increased in lymphoid neoplasm diffuse large B-cell lymphoma, esophageal carcinoma, glioblastoma multiforme, head and neck squamous cell carcinoma, kidney renal clear cell carcinoma (KIRC), pancreatic adenocarcinoma, stomach adenocarcinoma and thymoma, and significantly decreased in cervical squamous cell carcinoma and endocervical adenocarcinoma, kidney chromophobe, lung squamous cell carcinoma, ovarian serous cystadenocarcinoma, uterine corpus endometrial carcinoma and uterine carcinosarcoma, compared with corresponding healthy tissues in pancancer, indicating roles for viral invasion in most cancer types. Moreover, low NRP1 expression was significantly associated with long overall survival (OS) time in adrenocortical carcinoma, brain lower grade glioma, stomach adenocarcinoma and uveal melanoma, but with short OS time in KIRC only. The ENST00000374867.6 (NRP1-202) isoform is most highly expressed in most cancer types and thus could be involved in tumorigenesis and SARS-CoV-2 invasion in cancer patients. NRP1 may be involved in SARS-CoV-2 invasion in patients with cancer, including those with lung cancer.

Tripartite motif-containing 28 (TRIM28) expression and cordycepin inhibition in progression, prognosis, and therapeutics of patients with breast invasive carcinoma.

Breast cancer (BC) is the most common tumor in women worldwide. TRIM28 (RNF96) plays pleiotropic biological functions, such as silencing target genes, facilitating DNA repair, stimulating cellular proliferation and differentiation, and contributing to cancer progression. TRIM28 plays an increasingly crucial role in cancer, but its impact on BC, including breast invasive carcinoma, remains poorly understood. In the current study, analyses of online databases, quantitative real-time quantitative PCR, immunohistochemistry, and western blotting were performed on patients with breast invasive carcinoma (BRCA). Cordycepin (CD) was used to monitor BC progression and TRIM28 expression in vivo. As a result, we observed that TRIM28 is highly expressed in breast invasive carcinoma tissues compared with the corresponding normal tissues and is correlated with metastatic / invasive progression. High expression of TRIM28 might serve as a prognostic marker for long-term survival in triple-negative BC, advanced BC, or breast invasive carcinoma. Although TRIM28 methylation in tumor tissues of breast invasive carcinoma is not significantly changed compared to the matched normal tissues, the expressions and methylation of TRIM28 are significantly reversely correlated. TRIM28 expression was inhibited by CD in the mouse model, indicating its role in preventing BC progression. Thus, TRIM28 might be a potentially valuable molecular target for forecasting the progression / prognosis of patients with breast invasive carcinoma. CD, which represses BC growth/metastasis, may be involved partially through suppressing TRIM28 expression.

Effect of DPP4/CD26 expression on SARS­CoV­2 susceptibility, immune response, adenosine (derivatives m62A and CD) regulations on patients with cancer and healthy individuals.

The worldwide COVID­19 pandemic was brought on by a new coronavirus (SARS Cov­2). A marker/receptor called Dipeptidyl peptidase 4/CD26(DPP4/CD26) may be crucial in determining susceptibility to tumors and coronaviruses. However, the regulation of DPP4 in COVID­invaded cancer patients and its role on small molecule compounds remain unclear. The present study used the Human Protein Atlas, Monaco, and Schmiedel databases to analyze the expression of DPP4 in human tissues and immune cells. The association between DPP4 expression and survival in various tumor tissues was compared using GEPIA 2. The DNMIVD database was used to analyze the correlation between DPP4 expression and promoter methylation in various tumors. On the cBioPortal network, the frequency of DPP4 DNA mutations in various cancers was analyzed. The correlation between DPP4 expression and immunomodulators was analyzed by TISIDB database. The inhibitory effects of cordycepin (CD), N6, N6­dimethyladenosine (m62A) and adenosine (AD) on DPP4 in cancer cells were evaluated. DPP4 was mainly expressed in endocrine tissue, followed by gastrointestinal tract, female tissue (mainly in placenta), male tissue (mainly in prostate and seminal vesicle), proximal digestive tract, kidney, bladder, liver, gallbladder and respiratory system. In immune cells, DPP4 mRNA was mainly expressed in T cells, and its expression was upregulated in esophageal carcinoma, kidney renal papillary cell carcinoma (KIRP), liver hepatocellular carcinoma (LIHC), lung adenocarcinoma, pancreatic adenocarcinoma, prostate adenocarcinoma, stomach adenocarcinoma, thyroid carcinoma and thymoma. However, it was downregulated in breast invasive carcinoma, kidney chromophobe, lung squamous cell carcinoma and skin cutaneous melanoma. Thus, DPP4 is involved in viral invasion in most types of cancer. The expression of DPP4 could be inhibited by CD, m62A and AD in different tumor cells. Moreover, CD significantly inhibited the formation of GFP­positive syncytial cells. In vivo experiments with AD injection further showed that AD significantly inhibited lymphocyte activating factor 3 expression. These drugs may have potential to treat COVID­19 by targeting DPP4. Thus, DPP4 may be medically significant for SARS­CoV­2­infected cancer patients, providing prospective novel targets and concepts for the creation of drugs against COVID­19.

CTSL, a prognostic marker of breast cancer, that promotes proliferation, migration, and invasion in cells in triple-negative breast cancer.

Introduction: In the world, the incidence of breast cancer has surpassed that of lung cancer, and it has become the first malignant tumor among women. Triple-negative breast cancer (TNBC) shows an extremely heterogeneous malignancy toward high recurrence, metastasis, and mortality, but there is a lack of effective targeted therapy. It is urgent to develop novel molecular targets in the occurrence and therapeutics for TNBC, and novel therapeutic strategies to block the recurrence and metastasis of TNBC. Methods: In this study, CTSL (cathepsin L) expression in tissues and adjacent tissues of TNBC patients was monitored by immunohistochemistry and western blots. The correlations between CTSL expressions and clinicopathological characteristics in the patient tissues for TNBC were analyzed. Cell proliferation, migration, and invasion assay were also performed when over-expressed or knocked-down CTSL. Results: We found that the level of CTSL in TNBC is significantly higher than that in the matched adjacent tissues, and associated with differentiated degree, TNM Stage, tumor size, and lymph node metastatic status in TNBC patients. The high level of CTSL was correlated with a short RFS (p<0.001), OS (p<0.001), DMFS (p<0.001), PPS (p= 0.0025) in breast cancer from online databases; while in breast cancer with lymph node-positive, high level of CTSL was correlated with a short DMFS (p<0.001) and RFS (p<0.001). Moreover, in vitro experiments showed that CTSL overexpression promotes the abilities for proliferation, migration, and invasion in MCF-7 and MDA-MB-231 cell lines, while knocking-down CTSL decreases its characteristics in MDA-MB-231 cell lines. Conclusion: CTSL might involve into the regulation of the proliferation, invasion, and metastasis of TNBC. Thus, CTSL would be a novel, potential therapeutic, and prognostic target of TNBC.

The regulation of ISG20 expression on SARS-CoV-2 infection in cancer patients and healthy individuals.

ISG20 inhibits viruses such as SARS-CoV-2 invasion; however, details of its expression and regulation with viral susceptibility remain to be elucidated. The present study analyzed ISG20 expression, isoform information, survival rate, methylation patterns, immune cell infiltration, and COVID-19 outcomes in healthy and cancerous individuals. Cordycepin (CD) and N6, N6-dimethyladenosine (m6 2A) were used to treat cancer cells for ISG20 expression. We revealed that ISG20 mRNA expression was primarily located in the bone marrow and lymphoid tissues. Interestingly, its expression was significantly increased in 11 different types of cancer, indicating that cancer patients may be less vulnerable to SARS-CoV-2 infection. Among them, higher expression of ISG20 was associated with a long OS in CESC and SKCM, suggesting that ISG20 may be a good marker for both viral prevention and cancer progress. ISG20 promoter methylation was significantly lower in BLCA, READ, and THCA tumor tissues than in the matched normal tissues, while higher in BRCA, LUSC, KIRC, and PAAD. Hypermethylation of ISG20 in KIRC and PAAD tumor tissues was correlated with higher expression of ISG20, suggesting that methylation of ISG20 may not underlie its overexpression. Furthermore, ISG20 expression was significantly correlated with immune infiltration levels, including immune lymphocytes, chemokine, receptors, immunoinhibitors, immunostimulators, and MHC molecules in pan-cancer. STAD exhibited the highest degree of ISG20 mutations; the median progression-free survival time in months for the unaltered group was 61.84, while it was 81.01 in the mutant group. Isoforms ISG20-001 and ISG20-009 showed the same RNase_T domain structure, demonstrating the functional roles in tumorigenesis and SARS-CoV-2 invasion inhibition in cancer patients. Moreover, CD and m6 2A increase ISG20 expression in various cancer cell lines, implying the antiviral/anti-SARS-CoV-2 therapeutic potential. Altogether, this study highlighted the value of combating cancer by targeting ISG20 during the COVID-19 pandemic, and small molecules extracted from traditional Chinese medicines, such as CD, may have potential as anti-SARS-CoV-2 and anticancer agents by promoting ISG20 expression.

The Correlation Between Immune Invasion and SARS-COV-2 Entry Protein ADAM17 in Cancer Patients by Bioinformatic Analysis.

SARS-Cov-2 caused the COVID-19 pandemic worldwide. ADAM17 functions as a disintegrin and transmembrane metalloproteinase domain protein involved in the regulation of SARS-CoV-2 receptor ACE2. However, its impact on cancer patients infected with COVID-19 and its correlation with immune cell infiltration is unclear. This study compared ADAM17 expression between normal and tumor tissues based on GEPIA. The correlations between ADAM17 expression and immune cell infiltration and immunomodulators were investigated. Besides, treated drugs for targeting ADAM17 were searched in the TISDB database. We found that ADAM17 was highly conserved in many species and was mainly expressed in lung, brain, female tissues, bone marrow and lymphoid tissues. It was also highly expressed in respiratory epithelial cells of rhinitis and bronchus. ADAM17 expression in tumors was higher than that in several paired normal tissues and was negatively correlated with the prognosis of patients with malignant tumors. Interestingly, ADAM17 expression significantly correlated with immunomodulators and immune cell infiltration in normal and tumor tissues. Moreover, eight small molecules targeting ADAM17 only demonstrate therapeutic significance. These findings imply important implications for ADAM17 in cancer patients infected with COVID-19 and provide new clues for development strategy of anti-COVID-19.

COVID-19 receptor and malignant cancers: Association of CTSL expression with susceptibility to SARS-CoV-2.

CTSL is expressed by cancerous tissues and encodes a lysosomal cysteine proteinase that regulates cancer progression and SARS-CoV-2 entry. Therefore, it is critical to predict the susceptibility of cancer patients for SARS-CoV-2 and evaluate the correlation between disease outcomes and the expression of CTSL in malignant cancer tissues. In the current study, we analyzed CTSL expression, mutation rate, survival and COVID-19 disease outcomes in cancer and normal tissues, using online databases. We also performed immunohistochemistry (IHC) to test CTSL expression and western blot to monitor its regulation by cordycepin (CD), and N6, N6-dimethyladenosine (m62A), respectively. We found that CTSL is conserved across different species, and highly expressed in both normal and cancer tissues from human, as compared to ACE2 or other proteinases/proteases. Additionally, the expression of CTSL protein was the highest in the lung tissue. We show that the mRNA expression of CTSL is 66.4-fold higher in normal lungs and 54.8-fold higher in cancer tissues, as compared to ACE2 mRNA expression in the respective tissues. Compared to other proteases/proteinases/convertases such as TMPRSS2 and FURIN, the expression of CTSL was higher in both normal lungs and lung cancer samples. All these data indicate that CTSL might play an important role in COVID-19 pathogenesis in normal and cancer tissues of the lungs. Additionally, the CTSL-002 isoform containing both the inhibitor_I29 and Peptidase_C1 domains was highly prevalent in all cancers, suggesting its potential role in tumor progression and SARS-CoV-2 entry in multiple types of cancers. Further analysis of the expression of CTSL mutant showed a correlation with FURIN and TMPRSS2, suggesting a potential role of CTSL mutations in modulating SARS-CoV-2 entry in cancers. Moreover, high expression of CTSL significantly correlated with a short overall survival (OS) in lung cancer and glioma. Thus, CTSL might play a major role in the susceptibility of lung cancer and glioma patients to SARS-CoV-2 uptake and COVID-19 severity. Furthermore, CD or m62A inhibited CTSL expression in the cancer cell lines A549, MDA-MB-231, and/or PC3 in a dose dependent manner. In conclusion, we show that CTSL is highly expressed in normal tissues and increased in most cancers, and CD or m62A could inhibit its expression, suggesting the therapeutic potential of targeting CTSL for cancer and COVID-19 treatment.

COVID-19 disease and malignant cancers: The impact for the furin gene expression in susceptibility to SARS-CoV-2.

Furin is a proprotein convertase that activates different kinds of regulatory proteins, including SARS-CoV-2 spike protein which contains an additional furin-specific cleavage site. It is essential in predicting cancer patients' susceptibility to SARS-CoV-2 and the disease outcomes due to varying furin expressions in tumor tissues. In this study, we analyzed furin's expression, methylation, mutation rate, functional enrichment, survival rate and COVID-19 outcomes in normal and cancer tissues using online databases, and our IHC. As a result, furin presented with biased expression profiles in normal tissues, showing 12.25-fold higher than ACE2 in the lungs. The furin expression in tumors were significantly increased in ESCA and TGCT, and decreased in DLBC and THYM, indicating furin may play critical mechanistic functions in COVID-19 viral entry into cells in these cancer patients. Line with furin over/downexpression, furin promoter hypo-/hyper-methylation may be the regulatory cause of disease and lead to pathogenesis of ESCA and THYM. Furthermore, presence of FURIN-201 isoform with functional domains (P_proprotein, Peptidase_S8 and S8_pro-domain) is highest in all cancer types in comparison to other isoforms, demonstrating its use in tumorigenesis and SARS-Cov-2 entry into tumor tissues. Furin mutation frequency was highest in UCES, and its mutation might elevate ACE2 expression in LUAD and UCEC, reduce ACE2 expression in COAD, elevate HSPA5 expression in PAAD, and elevate TMPRSS2 expression in BRCA. These results showed that furin mutations mostly increased expression of ACE2, HSPA5, and TMPRSS2 in certain cancers, indicating furin mutations might facilitate COVID-19 cell entry in cancer patients. In addition, high expression of furin was significantly inversely correlated with long overall survival (OS) in LGG and correlated with long OS in COAD and KIRC, indicating that it could be used as a favorable prognostic marker for cancer patients' survival. GO and KEGG demonstrated that furin was mostly enriched in genes for metabolic and biosynthetic processes, retinal dehydrogenase activity, tRNA methyltransferase activity, and genes involving COVID-19, further supporting its role in COVID-19 and cancer metabolism. Moreover, Cordycepin (CD) inhibited furin expression in a dosage dependent manner. Altogether, furin's high expression might not only implies increased susceptibility to SARS-CoV-2 and higher severity of COVID-19 symptoms in cancer patients, but also it highlights the need for cancer treatment and therapy during the COVID-19 pandemic. CD might have a potential to develop an anti-SARS-CoV-2 drug through inhibiting furin expression.

Novel compound heterozygous missense variants (c.G955A and c.A1822C) of CACNA2D4 likely causing autosomal recessive retinitis pigmentosa in a Chinese patient.

Retinitis pigmentosa (RP) is a rare and heterogeneous group of inherited ocular diseases. However, the relationship between CACNA2D4 mutations and RP is not well understood. In this study, a Chinese autosomal recessive retinitis pigmentosa (arRP) pedigree was enrolled and targeted next-generation sequencing was employed for identifying the causative gene in the proband. These steps were followed by confirmatory Sanger sequencing and segregation analysis. RNA-sequencing (RNA-seq) data and semi-quantitative reverse transcription polymerase chain reaction analysis were then applied to examine the expressions in the human and mouse tissues. Novel compound heterozygous, deleterious missense variants of the CACNA2D4 gene, NM_172364.4: c.G955A (p.D319N) and c.A1822C (p.I608L), were identified in the arRP pedigree, co-segregating with the clinical phenotype in the patient. The CACNA2D4 protein is highly conserved among species. The CACNA2D4 mRNA expression showed the highest expression in the retina of humans and in the later four developmental stages/times of retinal tissues in mice, indicating its role in retina/eye functions and developments. This study is the first to identify novel compound heterozygous mutations c.G955A (p.D319N) and c.A1822C (p.I608L) in the CACNA2D4 gene. These might be disease-causing mutations, thereby extending the mutational spectra. The identification of pathogenic CACNA2D4 variants is expected to enhance our understanding of the genotype-phenotype correlations of arRP for disease diagnosis and genetic counseling. The relationship between the CACNA2D4 variants and diseases/phenotypes other than RP has also been reviewed and discussed in this paper.

Evaluation and characterization of HSPA5 (GRP78) expression profiles in normal individuals and cancer patients with COVID-19.

HSPA5 (BiP, GRP78) has been reported as a potential host-cell receptor for SARS-Cov-2, but its expression profiles on different tissues including tumors, its susceptibility to SARS-Cov-2 virus and severity of its adverse effects on malignant patients are unclear. In the current study, HSPA5 has been found to be expressed ubiquitously in normal tissues and significantly increased in 14 of 31 types of cancer tissues. In lung cancer, mRNA levels of HSPA5 were 253-fold increase than that of ACE2. Meanwhile, in both malignant tumors and matched normal samples across almost all cancer types, mRNA levels of HSPA5 were much higher than those of ACE2. Higher expression of HSPA5 significantly decreased patient overall survival (OS) in 7 types of cancers. Moreover, systematic analyses found that 7.15% of 5,068 COVID-19 cases have malignant cancer coincidental situations, and the rate of severe events of COVID-19 patients with cancers present a higher trend than that for all COVID-19 patients, showing a significant difference (33.33% vs 16.09%, p<0.01). Collectively, these data imply that the tissues with high HSPA5 expression, not low ACE2 expression, are susceptible to be invaded by SARS-CoV-2. Taken together, this study not only indicates the clinical significance of HSPA5 in COVID-19 disease and cancers, but also provides potential clues for further medical treatments and managements of COVID-19 patients.

Radiomic features of Pk-DCE MRI parameters based on the extensive Tofts model in application of breast cancer.

To explore radiomic features of pharmacokinetic dynamic contrast-enhanced (Pk-DCE) MRI on the extensive Tofts model to diagnose breast cancer and predict molecular phenotype. Breast lesions enrolled must undergo Pk-DCE MRI before treatment or puncture, and be identified as primary lesions by pathology. Ktrans, Kep, Ve and Vp were generated on the extensive Tofts model. Radiomic features (histogram, geometry and texture features) were extracted from parametric maps and selected by LASSO. The subjects were divided into training and validation cohort with a ratio of 4:1 to construct model in diagnosis of breast cancer. Feature analysis was made to predict the molecular phenotype. Area under curve (AUC), sensitivity, specificity and accuracy were used to evaluate radiomic features. DeLong's test was performed to compare AUC values. 228 breast lesions met the criteria were used to discrimination and 126 malignant lesions were used to study molecular phenotypes. The number of training cohort and validation cohort were 182 and 46, respectively. The AUC of Ktrans, Kep, Ve, and Vp was 0.95, 0.93, 0.89, and 0.96, and their accuracy was 85%, 89%, 89%, 94% respectively in diagnosis of breast lesions, while their AUC was 0.71 to 0.77, 0.61 to 0.68, and 0.67 to 0.74 to predict ER/PR, Her-2, and Ki-67. There was no significant difference among parameters (P > 0.05). Radiomic features based on Pk-DCE MRI have an advantage to diagnose breast cancer and less ability to predict molecular phenotypes, which are beneficial to guide clinical treatment of breast lesions in some extent.

Targeted Next-Generation Sequencing Identified Novel Compound Heterozygous Variants in the CDH23 Gene Causing Usher Syndrome Type ID in a Chinese Patient.

Usher syndrome includes a group of genetically and clinically heterogeneous autosomal recessive diseases, such as retinitis pigmentosa (RP) and sensorineural hearing loss. Usher syndrome type I (USHI) is characterized by profound hearing impairment beginning at birth, vestibular dysfunction, and unintelligible speech in addition to RP. The relationships between the Usher syndrome causing genes and the resultant phenotypes of Usher syndrome have not yet been fully elucidated. In the present study, we recruited a Chinese family with Usher syndrome and conducted paneled next-generation sequencing, Sanger sequencing, segregation analysis, and expression profile analysis. The functional effects of the identified cadherin-related 23 (CDH23) pathogenic variants were analyzed. The M101 pedigree consisted of a proband and seven family members, and the proband was a 39-year-old Chinese male who claimed that he first began to experience night blindness 11 years ago. We revealed novel, missense compound heterozygous variants c. 2572G > A (p.V858I) and c. 2891G > A (p.R964Q) in the CDH23 gene, which co-segregated with the disease phenotype causing Usher syndrome type ID (USH1D) in this Chinese pedigree. CDH23 mRNA was highly expressed in the retina, and this protein was highly conserved as revealed by the comparison of Homo sapiens CDH23 with those from nine other species. This is the first study to identify the novel, missense compound heterozygous variants c. 2572G > A (p.V858I) and c.2891G > A (p.R964Q) of CDH23, which might cause USH1D in the studied Chinese family, thereby extending CDH23 mutation spectra. Identifying CDH23 pathogenic variants should help in the detailed phenotypic characterization of USH1D.