Multiomics sequencing and immune microenvironment characteristics define three subtypes of small cell neuroendocrine carcinoma of the cervix.

Small cell cervical carcinoma (SCCC) is the most common neuroendocrine tumor in the female genital tract, with an unfavorable prognosis and lacking an evidence-based therapeutic approach. Until now, the distinct subtypes and immune characteristics of SCCC combined with genome and transcriptome have not been described. We performed genomic (n = 18), HPV integration (n = 18), and transcriptomic sequencing (n = 19) of SCCC samples. We assessed differences in immune characteristics between SCCC and conventional cervical cancer, and other small cell neuroendocrine carcinomas, through bioinformatics analysis and immunohistochemical assays. We stratified SCCC patients through non-negative matrix factorization and described the characteristics of these distinct types. We further validated it using multiplex immunofluorescence (n = 77) and investigated its clinical prognostic effect. We confirmed a high frequency of PIK3CA and TP53 alterations and HPV18 integrations in SCCC. SCCC and other small cell carcinoma had similar expression signatures and immune cell infiltration patterns. Comparing patients with SCCC to those with conventional cervical cancer, the former presented immune excluded or 'desert' infiltration. The number of CD8+ cells in the invasion margin of SCCC patients predicted favorable clinical outcomes. We identified three transcriptome subtypes: an inflamed phenotype with high-level expression of genes related to the MHC-II complex (CD74) and IFN-α/ß (SCCC-I), and two neuroendocrine subtypes with high-level expression of ASCL1 or NEUROD1, respectively. Combined with multiple technologies, we found that the neuroendocrine groups had more TP53 mutations and SCCC-I had more PIK3CA mutations. Multiplex immunofluorescence validated these subtypes and SCCC-I was an independent prognostic factor of overall survival. These results provide insights into SCCC tumor heterogeneity and potential therapies. © 2024 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Methyltransferase-like 3 promotes cervical cancer metastasis by enhancing cathepsin L mRNA stability in an N6-methyladenosine-dependent manner.

N6-methyladenosine (m6A) is a highly abundant RNA modification in eukaryotic cells. Methyltransferase-like 3 (METTL3), a major protein in the m6A methyltransferase complex, plays important roles in many malignancies, but its role in cervical cancer metastasis remains uncertain. Here, we found that METTL3 was significantly upregulated in cervical cancer tissue, and its upregulation was associated with a poor prognosis in cervical cancer patients. Knockdown of METTL3 significantly reduced cervical cancer cell migration and invasion. Conversely, METTL3 overexpression markedly promoted cervical cancer cell metastasis in vitro and in vivo. Furthermore, METTL3 mediated the m6A modification of cathepsin L (CTSL) mRNA at the 5'-UTR, and the m6A reader protein insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2) bound to the m6A sites and enhanced CTSL mRNA stability. Our results indicated that METTL3 enhanced CTSL mRNA stability through an m6A-IGF2BP2-dependent mechanism, thereby promoting cervical cancer cell metastasis. These findings provide insights into a novel m6A modification pattern involved in cervical cancer development.

The MYBL2-CCL2 axis promotes tumor progression and resistance to anti-PD-1 therapy in ovarian cancer by inducing immunosuppressive macrophages.

BACKGROUND: An immunosuppressive tumor microenvironment in ovarian cancer facilitates tumor progression and resistance to immunotherapy. The function of MYB Proto-Oncogene Like 2 (MYBL2) in the tumor microenvironment remains largely unexplored. METHODS: A syngeneic intraovarian mouse model, flow cytometry analysis, and immunohistochemistry were used to explore the biological function of MYBL2 in tumor progression and immune escape. Molecular and biochemical strategies-namely RNA-sequencing, western blotting, quantitative reverse transcription-polymerase chain reaction (qRT-PCR), enzyme-linked immunosorbent assay, multiplex immunofluorescence, chromatic immunoprecipitation assay (CHIP) and luciferase assay-were used to reveal the mechanisms of MYBL2 in the OVC microenvironment. RESULTS: We found tumor derived MYBL2 indicated poor prognosis and selectively correlated with tumor associated macrophages (TAMs) in ovarian cancer. Mechanically, C-C motif chemokine ligand 2 (CCL2) transcriptionally activated by MYBL2 induced TAMs recruitment and M2-like polarization in vitro. Using a syngeneic intraovarian mouse model, we identified MYBL2 promoted tumor malignancyand increased tumor-infiltrating immunosuppressive macrophages. Cyclin-dependent kinase 2 (CDK2) was a known upstream kinase to phosphorylate MYBL2 and promote its transcriptional function. The upstream inhibitor of CDK2, CVT-313, reprogrammed the tumor microenvironment and reduced anti-PD-1 resistance. CONCLUSIONS: The MYBL2/CCL2 axis contributing to TAMs recruitment and M2-like polarization is crucial to immune evasion and anti-PD-1 resistance in ovarian cancer, which is a potential target to enhance the efficacy of immunotherapy.

Transcriptional response of wolfberry to infestation with the endophytic Fusarium nematophilum strain NQ8GII4.

Fusarium nematophilum NQ8GII4 is an endophytic fungus isolated from the root of healthy wolfberry (Lycium barbarum). Previous studies have reported that NQ8GII4 could dwell in wolfberry roots and enhance the defense responses in wolfberry against root rot, which is caused by F. oxysporum. To further elucidate the molecular mechanism of wolfberry disease resistance induced by NQ8GII4, in the present study, we adopted RNA sequencing analysis to profile the transcriptome of wolfberry response to NQ8GII4 infestation over a time course of 3 and 7 days post-inoculation (dpi). Gene ontology (GO) enrichment analysis revealed that DEGs were enriched related to biological regulation, response to stimulus, signaling, detoxification, immune system process, transporter activity, electron carrier activity, transcription factor activity, nucleic acid binding transcription factor, and antioxidant activity. Through Kyoto encyclopedia of genes and genomes (KEGG) analysis, it was found that many of these DEGs were enriched in pathways related to plant-pathogen interactions, hormone signal transduction, and phenylpropanoid biosynthesis pathway in wolfberry. This suggests that innate immunity, phytohormone signaling, and numerous phenylpropanoid compounds, which comprise a complex defense network in wolfberry. Chloroplast 50S ribosomal proteins (50S RP) were consistently located at the core position of the response in wolfberry following infestation with NQ8GII4 analyzed by protein-protein interaction (PPI) network. This study elucidated the molecular mechanism underlying the interaction between NQ8GII4 and wolfberry, clarified the wolfberry immune response network to endophytic fungi infestation, identified candidate resistance genes in wolfberry, and provided a fundamental date for subsequent work.

Community ecological succession of endophytic fungi associates with medicinal compound accumulation in Sophora alopecuroides.

Endophytic fungi of medicinal plants are symbiotic with the host and play an important role in determining metabolites. To understand the relationship between the accumulation of Sophora alopecuroides' medicinal bioactive compounds and the ecological succession of endophytic fungi, here we collected samples from S. alopecuroides at four developmental stages (adult, flowering, podding, and mature) and different organs (roots, stems, leaves, and seeds) at the mature stage. We then used high-performance liquid chromatography-mass spectrometry and high-throughput sequencing on the internal transcribed spacer region to identify the medicinal compounds and endophytic fungal communities in each sample. The endophytic fungal community characteristics and accumulation of medicinally bioactive compounds of S. alopecuroides varied with the host's developmental stages and organs, with the highest total alkaloids content of 111.9 mg/g at the mature stage. Membership analysis and network connection analysis showed a total of 15 core endophytic fungi in different developmental stages and 16 core endophytic fungi in different organs at the mature stage. The unclassified Ascomycota, Aspergillus, and Alternaria were significantly and positively correlated with the medicinal compounds of S. alopecuroides at the mature stage (r > 0.6 or r < -0.6; P < 0.05). In this study, we identified key endophytic fungal resources that affect the content of medicinally bioactive compounds in S. alopecuroides. This discovery could lay the foundation for enhancing the yield of medicinally bioactive compounds in S. alopecuroides and the development and application of functional endophytic fungi.IMPORTANCESophora alopecuroides is a traditional Chinese herbal medicine. The major medicinal chemicals are considered to be quinolizidine alkaloids. Quinolizidine alkaloids have been widely used for the treatment of tumors, dysentery, and enteritis. Previous studies have found that endophytic fungi in S. alopecuroides can promote the accumulation of host quinolizidine alkaloids. However, the relationship between the accumulation of S. alopecuroides' medicinal bioactive compounds and the ecological succession of endophytic fungi remains unclear. In this study, we screened the key endophytic fungal resources affecting the content of medicinally bioactive compounds and laid the foundation for subsequent research on the mechanism by which endophytic fungi promote the accumulation of medicinally bioactive compounds in S. alopecuroides.

Targeting the Cdc2-like kinase 2 for overcoming platinum resistance in ovarian cancer.

Platinum resistance represents a major barrier to the survival of patients with ovarian cancer (OC). Cdc2-like kinase 2 (CLK2) is a major protein kinase associated with oncogenic phenotype and development in some solid tumors. However, the exact role and underlying mechanism of CLK2 in the progression of OC is currently unknown. Using microarray gene expression profiling and immunostaining on OC tissues, we found that CLK2 was upregulated in OC tissues and was associated with a short platinum-free interval in patients. Functional assays showed that CLK2 protected OC cells from platinum-induced apoptosis and allowed tumor xenografts to be more resistant to platinum. Mechanistically, CLK2 phosphorylated breast cancer gene 1 (BRCA1) at serine 1423 (Ser1423) to enhance DNA damage repair, resulting in platinum resistance in OC cells. Meanwhile, in OC cells treated with platinum, p38 stabilized CLK2 protein through phosphorylating at threonine 343 of CLK2. Consequently, the combination of CLK2 and poly ADP-ribose polymerase inhibitors achieved synergistic lethal effect to overcome platinum resistance in patient-derived xenografts, especially those with wild-type BRCA1. These findings provide evidence for a potential strategy to overcome platinum resistance in OC patients by targeting CLK2.

DNAJC8: a prognostic marker and potential therapeutic target for hepatocellular carcinoma.

Background: Hepatocellular carcinoma (HCC) is the most common type of liver cancer, accounting for ~90% of the total cases. DnaJ heat shock protein family member C8 (DNAJC8), belonging to the heat shock protein 40 (HSP40) family, is known to regulate cancer biology function. However, the role of DNAJC8 on HCC development remains unknown. Methods: The Cancer Genome Atlas, GTEx, cBioPortal, and Human Protein Atlas were used to analyze the expression and clinical significance of DNAJC8 in HCC. Two HCC cell lines, MHCC-97H and Huh-7, were utilized to determine the biological function of DNAJC8. Results: DNAJC8 expression was upregulated in HCC tissues and correlated with poor clinical prognosis. It was closely related to spliceosome, nucleocytoplasmic transport, and cell cycle and might be involved in the formation of tumor immunosuppressive microenvironment. Knockdown of DNAJC8 severely inhibited HCC cell proliferation and induced apoptosis. Conclusion: Our study demonstrate that DNAJC8 functions as an oncogene in HCC and hence may be used as a potential therapeutic target and prognostic marker for HCC.

A Nomogram for Preoperative Prediction of the Risk of Lymph Node Metastasis in Patients with Epithelial Ovarian Cancer.

OBJECTIVE: To develop a nomogram for predicting lymph node metastasis (LNM) in patients with epithelial ovarian cancer (EOC). METHODS: Between December 2012 and August 2022, patients with EOC who received computed tomography (CT) and serological examinations and were treated with upfront staging or debulking surgery were included. Systematic pelvic and para-aortic lymphadenectomy was performed in all patients. Univariate and multivariate analysis was used to identify significant risk factors associated with LNM. A nomogram was then constructed to assess the risk of LNM, which was evaluated with respect to its area under the receiver operating characteristic curve (AUC), calibration, and clinical usefulness. RESULTS: Of 212 patients enrolled in this study, 78 (36.8%) had positive LNs. The nomogram integrating CT-reported LN status, child-bearing status, tumour laterality, and stage showed good calibration and discrimination with an AUC of 0.775, significantly improving performance over the CT results (0.699, p = 0.0002) with a net reclassification improvement of 0.593 (p < 0.001) and integrated discrimination improvement of 0.054 (p < 0.001). The decision curve analysis showed the nomogram was of clinical use. CONCLUSIONS: A nomogram was constructed and internally validated, which may act as a decision aid in patients with EOC being considered for systemic lymphadenectomy.

Tislelizumab for cervical cancer: A retrospective study and analysis of correlative blood biomarkers.

Background: Tislelizumab is an anti-programmed cell death 1 (PD-1) monoclonal antibody engineered to minimize binding to Fcγ receptors. It has been used to treat several solid tumors. However, its efficacy and toxicity, and the predictive and prognostic value of baseline hematological parameters in patients with recurrent or metastatic cervical cancer (R/M CC) receiving tislelizumab remain unclear. Methods: We reviewed 115 patients treated for R/M CC with tislelizumab from March 2020 to June 2022 in our institute. The antitumor activity of tislelizumab was assessed using RECIST v1.1. Associations between the baseline hematological parameters and efficacy of tislelizumab in these patients were analyzed. Results: With a median follow-up of 11.3 months (range, 2.2-28.7), the overall response rate was 39.1% (95% CI, 30.1-48.2) and the disease control rate was 77.4% (95% CI, 69.6-85.2). The median progression-free survival (PFS) was 19.6 months (95% CI, 10.7 to not reached). The median overall survival (OS) was not reached. Treatment-related adverse events (TRAEs) of any grade occurred in 81.7% of the patients and only 7.0% of the patients experienced grade 3 or 4 TRAEs. Univariate and multivariate regression analyses showed that the level of pretreatment serum C-reactive protein (CRP) was an independent risk factor for the response (complete or partial response) to tislelizumab and the PFS of R/M CC patients treated with tislelizumab (P = 0.0001 and P = 0.002, respectively). R/M CC patients with elevated baseline CRP levels had a short PFS (P = 0.0005). Additionally, the CRP-to-albumin ratio (CAR) was an independent risk factor for the PFS and OS of R/M CC patients treated with tislelizumab (P = 0.001 and P = 0.031, respectively). R/M CC patients with an elevated baseline CAR had short PFS and OS (P < 0.0001 and P = 0.0323, respectively). Conclusions: Tislelizumab showed promising antitumor activity and tolerable toxicity in patients with R/M CC. The baseline serum CRP levels and CAR showed potential for predicting the efficacy of tislelizumab and the prognosis of R/M CC patients receiving tislelizumab.

The Endophytic Fungi Diversity, Community Structure, and Ecological Function Prediction of Sophora alopecuroides in Ningxia, China.

Sophora alopecuroides L. has great medicinal and ecological value in northwestern China. The host and its microbiota are mutually symbiotic, collectively forming a holobiont, conferring beneficial effects to the plant. However, the analysis of diversity, mycobiota composition, and the ecological function of endophytic fungi in the holobiont of S. alopecuroides is relatively lacking. In this article, the fungal community profiling of roots, stems, leaves, and seeds of S. alopecuroides (at the fruit maturity stage) from Huamachi and Baofeng in Ningxia, China were investigated based on the ITS1 region, using high-throughput sequencing technology. As a result, a total of 751 operational taxonomic units (OTUs) were obtained and further classified into 9 phyla, 27 classes, 66 orders, 141 families, 245 genera, and 340 species. The roots had the highest fungal richness and diversity, while the stems had the highest evenness and pedigree diversity. There also was a significant difference in the richness of the endophytic fungal community between root and seed (p < 0.05). The organ was the main factor affecting the community structure of endophytic fungi in S. alopecuroides. The genera of unclassified Ascomycota, Tricholoma, Apiotrichum, Alternaria, and Aspergillus made up the vast majority of relative abundance, which were common in all four organs as well. The dominant and endemic genera and biomarkers of endophytic fungi in four organs of S. alopecuroides were different and exhibited organ specificity or tissue preference. The endophytic fungi of S. alopecuroides were mainly divided into 15 ecological function groups, among which saprotroph was absolutely dominant, followed by mixotrophic and pathotroph, and the symbiotroph was the least. With this study, we revealed the diversity and community structure and predicted the ecological function of the endophytic fungi of S. alopecuroides, which provided a theoretical reference for the further development and utilization of the endophytic fungi resources of S. alopecuroides.

Correlation in endophytic fungi community diversity and bioactive compounds of Sophora alopecuroides.

Sophora alopecuroides L. is a traditional Chinese medicine used for the treatment of several different disease states including bacillary dysentery and enteritis. But importantly, it also plays a role as an anti-tumor agent. That said, little is known about the role endophytes play regarding the clinically bioactive metabolites in S. alopecuroides. In order to explore the effects of endophytic fungi on the accumulation, quality, and correlation in the content of the medicinal compounds, the structural diversity of endophytic fungi in S. alopecuroides was analyzed. The relationship between endophytes and quinolizidine alkaloids (QAs), housed within the seeds of S. alopecuroides, which were interpreted based on established methods of high-throughput sequencing and high-performance liquid chromatography. A total of 1,034,418 effective sequence reads and 257 operational taxonomic units (OTUs) were obtained from 33 samples which were sourced from 11 different sampling sites and further classified into 9 phyla, 20 classes, 45 orders, 85 families, and 118 genera. Ascomycota was found to be the dominant phylum of endophytic fungi in S. alopecuroides, with a relative abundance ranging from 60.85 to 98.30%. Alternaria, Cladosporium, Filobasidium, and an unidentified Ascomycota were the core-shared endophytes, accounting for 49.96, 27.12, 14.83, and 7.88%, respectively. Correlation analysis showed that the Simpson's diversity index of endophytic fungal community in S. alopecuroides was significantly positively correlated with the Oxymatrine (OMA) content in different areas, while the Chao and Shannoneven indexes were significantly negatively correlated with OMA. The endophytic fungi of Alternaria were positively correlated with the content of OMA, Oxysophocarpine (OSC), and total QAs. This study has mastered the endophytic fungi resources of S. alopecuroides, explored potential functional endophytic fungi, and provided a scientific basis for using biological fertilization strategies to improve the quality of S. alopecuroides.

Inhibition of miR-574-5p suppresses cell growth and metastasis and enhances chemosensitivity by targeting RNA binding protein QKI in cervical cancer cells.

Cervical cancer is the fourth most common female malignancy worldwide and microRNA (miR)-574-5p is a candidate oncogene in multiple cancers. The present study aimed to investigate the role and mechanism of miR-574-5p in cervical cancer. miR-574-5p inhibitors or mimics were transfected into cervical cancer cells to study the function of miR-574-5p. The effects of miR-574-5p on cell growth, invasion, and chemosensitivity were evaluated using CCK8, flow cytometry, transwell, immunofluorescence, and Western blotting analysis. Further depletion or forced expression of QKI was performed to explore the regulatory mechanism of miR-574-5p in cervical cancer. Up-regulation of miR-547-5p and down-regulation of QKI were observed in 30 cervical cancer tissues versus 30 adjacent normal tissues. Silencing of miR-574-5p increased apoptosis, inhibited proliferation, cell cycle progression, and cell invasiveness, as well as enhanced chemosensitivity towards cisplatin and doxorubicin in cervical cancer cells. Overexpression of miR-574-5p exerted promoting effect on cancer progression and metastasis. Knockdown of miR-574-5p induced an up-regulation of E-cadherin and down-regulation of cyclinD1, N-cadherin, matrix metallopeptidase 9 (MMP-9), and ß-catenin in cervical cancer cells Moreover, QKI was verified as a target of miR-574-5p and involved in regulation of miR-574-5p-induced cervical cancer cell progression and metastasis. miR-574-5p functions to be oncogenic in cervical cancer, and its inhibition suppresses cervical cancer progression and metastasis as well as enhances chemosensitivity by targeting QKI. Therefore, miR-574-5p is suggested as a potential therapeutic target for cervical cancer treatment.

Long non-coding RNA DLEU1 exerts an oncogenic function in non-small cell lung cancer.

OBJECTIVE: To explore the role of long non-coding RNA DLEU1 in the tumorigenesis and progression of non-small cell lung cancer (NSCLC) and the underlying mechanisms. METHODS: The expression of DLEU1 in NSCLC tumor tissues and adjacent normal tissues was evaluated using bioinformatics analysis and qPCR. The effects of DLEU1 overexpression or deficiency on cell proliferation, apoptosis, migration and invasion were explored experimentally. Additionally, the impact of DLEU1 up-regulation on tumourigenesis was also assessed in vivo. RESULTS: The expression of DLEU1 was up-regulated in NSCLC tumor tissues. DLEU1 overexpression promoted the proliferation, migration, and invasion, but inhibited the apoptosis of NSCLC cells by upregulating CDK1 expression, binding with SRC and altering the expression of P70(S6K), MMP2 and E-cadherin. Besides, xenograft tumors in nude mice demonstrated that DLEU1 overexpression accelerated tumor growth. CONCLUSIONS: DLEU1 promoted tumorigenesis and progression of NSCLC, and might be a promising therapeutic target for NSCLC.