Effects of bone marrow sparing radiotherapy on acute hematologic toxicity for patients with locoregionally advanced cervical cancer: a prospective phase II randomized controlled study.

OBJECTIVE: To evaluate effects of bone marrow sparing (BMS) radiotherapy on decreasing the incidence of acute hematologic toxicity (HT) for locoregionally advanced cervical cancer (LACC) patients treated by pelvic irradiation. MATERIALS AND METHODS: LACC patients were recruited prospectively from May 2021 to May 2022 at a single center and were evenly randomized into the BMS group and the control group. All patients received pelvic irradiation with concurrent cisplatin (40 mg/m2 weekly), followed by brachytherapy and BM V40 < 25% in the BMS group was additionally prescribed. Acute HT was assessed weekly. Binary logistic regression model and receiver operating characteristic (ROC) curve were used for predictive value analysis. The trial was registered with Chinese clinical trial registry (ChiCTR2200066485). RESULTS: A total of 242 patients were included in the analysis. Baseline demographic, disease and treatment characteristics were balanced between the two groups. In the intention-to-treat population, BMS was associated with a lower incidence of grade ≥ 2 and grade ≥ 3 acute HT, leukopenia and neutropenia s(72.70% v 90.90%, P < 0.001*; 16.50% vs. 65.30%, P < 0.001*; 66.10% vs. 85.10%, P = 0.001*; 13.20% vs. 54.50%, P < 0.001*; 37.20% vs. 66.10%, P < 0.001*; 10.70% vs. 43.80%, P < 0.001*). BMS also resulted in decreased dose delivered to the organs at risk (OARs) including rectum, bladder and left and right femoral head. Univariate and multivariate analyses showed that BM V40 was an independent risk factor for grade ≥ 3 acute HT (odds ratio [OR] = 2.734, 95% confidence interval [CI] = 1.959-3.815, P < 0.001*). Cutoff value was 25.036% and area under the curve (AUC) was 0.786. The nomogram was constructed, which was rigorously evaluated and internally cross-validated, showing good predictive performance. CONCLUSIONS: Receiving BMS pelvic irradiation could reduce the incidence of acute HT in LACC patients, and BM V40 < 25% may be a significant factor in reducing the risks of acute HT.

Hypermethylated ITGA8 Facilitate Bladder Cancer Cell Proliferation and Metastasis.

DNA methylation plays a vital role during the development of tumorigenesis. The purpose of this study is to identify candidate DNA methylation drivers during progression of bladder cancer (BLCA). The methylation spectrum in bladder cancer tissues was detected by CHARM analysis, and methylated ITGA8 was selected for further study due to its low expression. Methylation levels in BLCA tissues and cells were detected with methylated-specific PCR (MSP), while mRNA expression and methylation of ITGA8 were detected by qRT-PCR and MSP. After treatment with 5-Aza-dC (DNA methylation inhibitor), the proliferation, migration, and invasion abilities of BLCA cells were determined by MTT, wound healing, and transwell assays, respectively. Flow cytometric analysis was performed to evaluate any variance in the cell cycle. In addition, the effect of demethylated ITGA8 on BLCA tumor growth was verified with an in vivo xenograft tumor model. Based on the methylation profiling of BLCA, ITGA8 was identified to be hypermethylated. ITGA8 methylation levels in BLCA tissues and cells were upregulated, and 5-Aza-dC significantly suppressed ITGA8 methylation levels and increased ITGA8 mRNA expression. Furthermore, after treatment with 5-Aza-dC, the propagation, migration, and invasiveness of the cancer cells were inhibited, and more cancer cells were arrested at the G0/G1 phase. In vivo assays further demonstrated that 5-Aza-dC could impede BLCA tumor growth by repressing methylation levels of ITGA8 and increasing ITGA8 mRNA expression. Hypermethylated ITGA8 facilitated BLCA progression, and 5-Aza-dC treatment inhibited BLCA cell propagation and metastasis by decreasing methylation levels of ITGA8 and inducing cell cycle arrest.

LncRNA RHPN1-AS1 inhibition induces autophagy and apoptosis in prostate cancer cells via the miR-7-5p/EGFR/PI3K/AKT/mTOR signaling pathway.

LncRNA RHPN1-AS1 (RHPN1-AS1) has been confirmed to promote tumor progression in multiple cancers and is upregulated in prostate cancer (PCa), but whether it has an effect on PCa progression remains unclear. In this study, we found that PCa patients with high RHPN1-AS1 expression had a shorter survival time, and RHPN1-AS1 was significantly upregulated in PCa tissues and cells. Based on informatics analysis we predicted that miR-7-5p binds to 3'UTR of RHPN1-AS1 and epidermal growth factor receptor (EGFR) and verified it by luciferase reporter gene assay. Subsequently, we transfected PCa cells with RHPN1-AS1 overexpression vector (RHPN1-AS1), knockdown plasmids (sh-RHPN1-AS1) and/or miR-7-5p mimics or inhibitor and/or overexpression vector (EGFR) or small interfering RNA of EGFR (si-EGFR) or its control, and found that overexpression of RHPN1-AS1 inhibited miR-7-5p expression and promoted EGFR expression, silencing RHPN1-AS1 inhibited proliferation and invasion, and induced G2/M arrest, apoptosis and autophagy in PCa cells. 3MA (an inhibitor of autophagy)-mediated autophagy inhibition attenuated RHPN1-AS1 inhibition-induced apoptosis. Overexpression miR-7-5p or silencing EGFR promoted LC3-I to LC3-II conversion, enhanced autophagy activity, induced cleaved-caspase-3 expression and apoptosis in PCa cells. Furthermore, overexpression of RHPN1-AS1 promoted phosphorylation of phosphatidylinositol 3-kinase (PI3K), AKT and mTOR, inhibited LC3-I to LC3-II conversion and reduced apoptosis in PCa cells, while GSK2126458 (an inhibitor of PI3K) reversed the effect of RHPN1-AS1 on PCa cells. In summary, RHPN1-AS1 acted as a ceRNA of miR-7-5p to upregulate EGFR expression, silencing RHPN1-AS1 suppressed PCa tumor progression by inducing autophagy and apoptosis in PCa cells through the miR-7-5p/EGFR/PI3K/AKT/mTOR pathway.

Matrine impedes colorectal cancer proliferation and migration by downregulating endoplasmic reticulum lipid raft associated protein 1 expression.

Matrine exhibits anti-tumor effect on the proliferation and invasion of colorectal cancer (CRC) cells by reducing the activity of the p38 signaling pathway. However, these studies were limited because the underlying mechanism by which matrine inhibited CRC progression remained unclear. In this study, we provided for the first time that endoplasmic reticulum lipid raft associated protein 1 (Erlin1) is a novel target of matrine. Erlin1 was significantly upregulated in tumors and its knockdown suppressed the proliferation and migration of CRC cells, while its overexpression promoted CRC cell growth and migration. Furthermore, Erlin1 overexpression promoted inhibited apoptosis. Importantly, matrine treatment could reverse the oncogenic function of Erlin1 on CRC cell proliferation and migration. When Erlin1 was knocked down, matrine exhibited a more obvious anti-tumor effect in CRC cells. Partly due to this, matrine functions as an important anti-tumor drug and the results discovered here may clarify the mechanisms of matrine application for CRC treatment. CRC patients with low expression of Erlin1 might be more suitable for the treatment of matrine. This study could promote the application of matrine to be a promising therapeutic strategy for CRC patients.

RSPO2 silence inhibits tumorigenesis of nasopharyngeal carcinoma by ZNRF3/Hedgehog-Gli1 signal pathway.

R-spondins 2 (RSPO2) protein is a member of RSPO family which plays an essential role in stem cell survival, development and tumorigenicity. There has several evidence suggested that RSPO2 involved in breast, gastric, liver and colorectal cancer. However, the specific function and mechanism of RSPO2 in nasopharyngeal carcinoma (NPC) remain unknown. In the present study, we first observed that RSPO2 expression was elevated in NPC cell lines SUNE-6-10B, SUNE-5-8F, and CNE-1 compared with the normal laryngeal epithelia cell line NP69. Knockdown of RSPO2 significantly inhibits SUNE-6-10B and CNE-1 cell survival and proliferation by using CCK-8 assay and Edu assay, respectively. Further studies verified that RSPO2 silence suppressed migration and invasion of SUNE-6-10B and CNE-1 cells. Further studies suggested that RSPO2 silence suppressed epithelial-to-mesenchymal transition (EMT) related protein E-cadherin expression and promoted Vimentin and N-cadherin expression both in SUNE-6-10B and CNE-1 cells. Molecular mechanism explorations showed that RSPO2 deletion increased ZNRF3 expression and inhibited Gli1 expression. Additionally, knockdown ZNRF3 expression or overexpression Gli1 both reversed the effects of RSPO2 silence on NPC growth and metastasis. Finally, RSPO2 depletion was impaired NPC tumor growth in vivo animal experiments. In conclusion, the present study confirmed that RSPO2 silence inhibits the tumorigenesis of NPC via ZNRF3/Hedgehog-Gli1 signal pathway.

Long Non-Coding RNA GAS5 Suppresses Tumor Progression and Enhances the Radiosensitivity of Prostate Cancer Through the miR-320a/RAB21 Axis.

BACKGROUND: Long non-coding RNAs (lncRNAs) function as a class of significant mediators in prostate cancer (PCa), and this study mainly discussed the molecular mechanism of lncRNA growth arrest-specific 5 (GAS5) in PCa progression and radiosensitivity. MATERIALS AND METHODS: GAS5 and microRNA-320a (miR-320a) levels were determined by quantitative real-time polymerase chain reaction (qRT-PCR). Cell viability and migration were severally examined through 3-(4, 5-dimethylthiazol-2-y1)-2, 5-diphenyl tetrazolium bromide (MTT) and transwell assays. PCa cells were treated with X-ray irradiation. Cell survival and apoptosis rate were assayed using colony formation assay and flow cytometry, respectively. The apoptosis-related protein and Rab GTPase 21 (RAB21) protein levels were measured by Western blot. The relation between miR-320a and GAS5 or RAB21 was assessed via the dual-luciferase reporter assay. The effect of GAS5 on radiosensitivity of PCa in vivo was evaluated by xenotransplantation assay. RESULTS: GAS5 was down-regulated in PCa tissues and cells. GAS5 overexpression suppressed cell viability and migration while facilitated radiosensitivity of PCa cells. GAS5 was a molecular sponge of miR-320a. The effects of GAS5 up-regulation on PCa cells were accomplished by sponging miR-320a. MiR-320a targeted RAB21 and GAS5 up-regulated RAB21 expression via targeting miR-320a. RAB21 knockdown reversed the effects of miR-320a inhibition on PCa cells. GAS5 promoted the radiosensitivity of PCa by the miR-320a/RAB21 axis in vivo. CONCLUSION: Collectively, GAS5 restrained tumor development and expedited the radiosensitivity in PCa by the miR-320a/RAB21 axis, which provided a molecular regulatory mechanism of GAS5/miR-320a/RAB21 in PCa development and radioresistance.

Downregulation of the long non-coding RNA TUSC7 promotes NSCLC cell proliferation and correlates with poor prognosis.

BACKGROUND: Emerging evidence indicated that dysregulated long non-coding RNAs (lncRNAs) was implicated in the tumorigenesis and progression. LncRNA TUSC7 was involved in various malignancies. However, the role of TUSC7 in human non-small-cell lung cancer (NSCLC) remains unclear. METHODS: Expression of TUSC7 was analyzed in 112 cases of NSCLC tissues and six lung cancer cell lines by quantitative real-time PCR (qRT-PCR). Then the correlation of TUSC7 expression with clinicopathological features and prognosis was aslo studied. Furthermore, overexpression of TUSC7 was performed and its role in tumor progression was explored. RESULTS: The expression level of TUSC7 was lower in NSCLC tissues and lung cancer cells compared with their normal counterparts. Lower expression of TUSC7 in NSCLC tissues was associated with larger tumor size and higher TNM stage. Patients with lower TUSC7 expression had worse overall survival compared with the high expression cases. Univariate and multivariate analyses suggested that low expression of TUSC7 was an independent poor prognostic indicator for NSCLC patients. Moreover, upregulation of TUSC7 expression could inhibit proliferation of lung cancer cell in vitro. CONCLUSIONS: Our results suggested that TUSC7 was a potential biomarker for NSCLC prognosis, and the dysregulation of TUSC7 may play an important role in the NSCLC progression.

Matrine inhibits the invasive properties of human glioma cells by regulating epithelial­to­mesenchymal transition.

Matrine is reported to be effective in tumor therapies; however, the anti­metastatic effect and molecular mechanism(s) of matrine on glioma remain poorly understood. Therefore, the purpose of this study was to assess the effects of matrine on glioma and the associated mechanism(s). In the study, we demonstrated that matrine inhibited the proliferation of glioma cells. We also observed that matrine inhibited the migration and invasion of glioma cells at non­toxic concentrations. Matrine also decreased the expression of E­cadherin and increased the expression of N­cadherin. These results suggest that the anti­metastatic effect of matrine may be correlated with epithelial­to­mesenchymal transition (EMT). Moreover, matrine could reduce the phosphorylation levels of p38 and AKT proteins. In conclusion, these results suggest matrine may be a potential alternative against invasive glioma cells via the p38 MAPK and AKT signaling­dependent inhibition of EMT.

Saikosaponin-d increases the radiosensitivity of smmc-7721 hepatocellular carcinoma cells by adjusting the g0/g1 and g2/m checkpoints of the cell cycle.

BACKGROUND: Saikosaponin-d (SSd), a monomer terpenoid purified from the Chinese herbal drug Radix bupleuri, has multiple effects, including anticancer properties. However, the effect of SSd on tumors exposed to radiation is largely unknown. To investigate the radiosensitizing effect of SSd and its possible mechanism, we combined SSd with radiation therapy to treat SMMC-7721 hepatocellular carcinoma cells under oxia and hypoxia. METHODS: Cell growth, apoptosis, and cell cycle distribution were examined after treatment with SSd alone, radiation alone, and their combinations under oxia and hypoxia. The protein and mRNA levels of p53, Bcl2, and BAX were measured using western blot analysis and RT-PCR, respectively. RESULTS: Treatment with SSd alone and radiation alone inhibited cell growth and increased apoptosis rate at the concentration used. These effects were enhanced when SSd was combined with radiation. Moreover, SSd potentiated the effects of radiation to induce G0/G1 arrest in SMMC-7721 cells, and reduced the G2/M-phase population under hypoxia. However, under oxia, SSd only potentiated the effects of radiation to induce G0/G1 arrest, but not G2/M-phase arrest. These effects of SSd alone, radiation alone, and their combination, were accompanied by upregulated expression of p53 and BAX and downregulation of Bcl2 expression under oxia and hypoxia. CONCLUSION: SSd potentiates the effects of radiation on SMMC-7721 cells; thus, it is a promising radiosensitizer. The radiosensitizing effect of SSd may contribute to its effect on the G0/G1 and G2/M checkpoints of the cell cycle.

The effect of RHIZOMA COPTIDIS and COPTIS CHINENSIS aqueous extract on radiation-induced skin injury in a rat model.

BACKGROUND: Radiation-induced skin injury is a common complication of radiotherapy. The RHIZOMA COPTIDIS and COPTIS CHINENSIS aqueous extract (RCE) can ameliorate radiation-induced skin injury in our clinical observation. But, the protective mechanism of RHIZOMA COPTIDIS and COPTIS CHINENSIS in radiation-induced skin injury remains unclear. METHODS: In this experiment, we developed a radiation-induced skin injury rat model to study the mechanism. The animals were randomly divided into control group, treatment group, radiation group, and treatment and radiation group. 5 rats in each group were separately executed on 2 d and 49 d post-radiation. The semi-quantitative skin injury score was used to measure skin reactions by unblinded observers, and hematoxylin and eosin staining was used to evaluate the damage areas by irradiation. The MDA content, SOD activity of skin and serum were measured to detect the oxidative stress. RESULTS: Acute skin reactions were caused by a single dose of 45 Gy of ß-ray irradiation, and the skin injury could be found in all rats receiving irradiation based on the observation of HE staining of skin at different time-points, while RCE could significantly ameliorate those changes. The MDA content in serum and skin of control rats was 4.13±0.12 mmol/ml and 4.95±0.35 mmol/mgprot on 2 d post-radiation. The rats receiving radiation showed an increased content of MDA (5.54±0.21 mmol/ml and 7.10±0.32 mmol/mgprot), while it was 4.57±0.21 mmol/ml and 5.95±0.24 mmol/mgprot after treated with RCE (p<0.05). Similar changes of the MDA content could be seen on 49 d post-radiation. However, the SOD activity of rats receiving radiation decreased compared with control group on both time-points, which was inhibited by RCE (p<0.05). Meanwhile, no valuable changes could be found between control group and treatment group on 2 d and 49 d. CONCLUSIONS: Our study provides evidences for the radioprotective role of RCE against radiation-induced skin damage in rats by modulating oxidative stress in skin, which may be a useful therapy for radiation-induced skin injury.