Dose-Response Relationship between Head and Neck Radiation and Damages to Gustatory Cells in Mice.

Objective: To investigate the dose-response relationship between radiation to the head and neck regions and damage observed in mice gustatory cells. Materials and Methods: A total number of 45 mice (C57BL/6) (aged 8-12 weeks) were enrolled in this study. The head and neck regions of the mice were irradiated at doses of 8 Gy (low-dose group, n = 15), 16 Gy (moderate-dose group, n = 15), and 24 Gy (high-dose group, n = 15). Each time, 3 mice from each group were sacrificed before radiation and then 2-day, 4-day, 7-day, and 14-day post the irradiation, respectively. The immune-histochemical staining method was employed to obtain gustatory papilla tissues and mark gustatory cells. Careful calculation of the numbers of proliferative cells, taste buds, and type II gustatory cells was conducted. Results: A decrease in the number of Ki-67-marked proliferative cells was noted at 2 days postirradiation (DPI), and the number of cells was recovered to the normal level at 4-DPI in each group. The number of Ki-67-marked proliferative cells was hypercompensation (significantly higher than normal) in the moderate-dose and high-dose groups at 7-DPI and insufficient compensation (significantly lower than normal) in the high-dose group at 14-DPI. There was a significant reduction of taste buds and type II gustatory cells at 2-DPI and is lowest at 4-DPI in the moderate-dose and high-dose groups, while little change was observed in the low-dose group. Conclusion: Damages to Gustatory Cells after head and neck radiation were dose-related and compensation occurred in 14-DPI and may be insufficient when overdosed.

Theaflavin promoted apoptosis in nasopharyngeal carcinoma unexpectedly via inducing autophagy in vitro.

Objectives: This study aimed to investigate the mechanism of the anticancer effect of theaflavin (TF) in nasopharyngeal carcinoma. Materials and Methods: CNE2 cells were used to study the anticancer effect of TF. This study used Cell Counting Kit-8 (CCK8) assay on proliferation and used flow cytometry to detect apoptosis. The protein expression of Bcl-2, Bax, caspase 3, and caspase 9 was detected by Western blot, and autophagy-related proteins were also detected. Results: TF inhibited proliferation of CNE2 cells, promoted apoptosis, and up-regulated the expression of caspase 3, caspase 9, and Bax, and decreased the level of Bcl-2. Unexpectedly, TF induced autophagy rather than inhibiting autophagy through up-regulating the levels of the autophagy marker light chain 3 (LC3) and Lysosomal-associated membrane protein 1 (LAMP1) and reducing levels of the autophagosome cargo protein p62, and the effect was via the mTOR pathway. Besides, autophagy inhibitor Chloroquine (CQ) suppressed the effect of TF on Bax, Bcl-2 and activation of caspase 3 and caspase 9. Conclusion: TF promoted apoptosis of nasopharyngeal carcinoma cells, the mechanism was unexpectedly involved in inducing autophagy.

MiR-122 radiosensitize hepatocellular carcinoma cells by suppressing cyclin G1.

PURPOSE: Emerging evidence has shown that radiotherapy is an effective treatment for hepatocellular carcinoma (HCC), Micro(mi)RNAs are involved in regulating radiosensitivity in many cancers. MiR-122 accounts for approximately 70% of all cloned miRNAs in the liver, but there are few reports about whether it is involved in regulating of radiosensitivity in HCC cells. MATERIALS AND METHODS: HCC cells (HepG2 and Huh7) overexpressing miR-122 were constructed by transfecting them with lentiviral-miR-122. Then, their proliferation ability was analyzed by the MTT, and colony formation assays and a xenograft tumor model was used to detect their radiosensitivity. The expression of cyclin G1 mRNA and protein was detected by the quantitative real-time polymerase chain reaction and western blotting, respectively. RESULTS: Overexpression of miR-122 inhibited the proliferation of, and radiosensitized HCC cells. Cyclin G1 mRNA and protein level were suppressed in HepG2 tumors overexpression miR-122. CONCLUSION: MiR-122 may be useful as a potential radiosensitizer for HCC, and its mechanism is related to the regulation of cyclin G1.

Identification of lncRNA-miRNA-mRNA Networks Linked to Non-small Lung Cancer Resistance to Inhibitors of Epidermal Growth Factor Receptor.

Background: Tyrosine kinase inhibitors that act against epidermal growth factor receptor (EGFR) show strong efficacy against non-small cell lung cancer (NSCLC) involving mutated EGFRs. However, most such patients eventually develop resistance to EGFR-TKIs. Numerous researches have reported that messenger RNAs (mRNAs) and non-coding RNAs (ncRNAs) may be involved in EGFR-TKI resistance, but the comprehensive expression profile and competitive endogenous RNA (ceRNA) regulatory network between mRNAs and ncRNAs in EGFR-TKI resistance of NSCLC are incompletely known. We aimed to define a ceRNA regulatory network linking mRNAs and non-coding RNAs that may mediate this resistance. Methods: Using datasets GSE83666, GSE75309 and GSE103352 from the Gene Expression Omnibus, we identified long non-coding RNAs (lncRNAs), microRNAs (miRNAs) and mRNAs differentially expressed between NSCLC cells that were sensitive or resistant to EGFR-TKIs. The potential biological functions of the corresponding differentially expressed genes were analyzed based KEGG pathways. We combined interactions among lncRNAs, miRNAs and mRNAs in the RNAInter database with KEGG pathways to generate transcriptional regulatory ceRNA networks associated with NSCLC resistance to EGFR-TKIs. Kaplan-Meier analysis was used to assess the ability of core ceRNA regulatory sub-networks to predict the progression-free interval and overall survival of NSCLC. The expression of two core ceRNA regulatory sub-networks in NSCLC was validated by quantitative real-time PCR. Results: We identified 8,989 lncRNAs, 1,083 miRNAs and 3,191 mRNAs that were differentially expressed between patients who were sensitive or resistant to the inhibitors. These DEGs were linked to 968 biological processes and 31 KEGG pathways. Pearson analysis of correlations among the DEGs of lncRNAs, miRNAs and mRNAs identified 12 core ceRNA regulatory sub-networks associated with resistance to EGFR-TKIs. The two lncRNAs ABTB1 and NPTN with the hsa-miR-150-5p and mRNA SERPINE1 were significantly associated with resistance to EGFR-TKIs and survival in NSCLC. These lncRNAs and the miRNA were found to be down-regulated, and the mRNA up-regulated, in a resistant NSCLC cell line relative to the corresponding sensitive cells. Conclusion: In this study, we provide new insights into the pathogenesis of NSCLC and the emergence of resistance to EGFR-TKIs, based on a lncRNA-miRNA-mRNA network.

Efficacy of Concurrent Chemoradiotherapy With S-1 vs Radiotherapy Alone for Older Patients With Esophageal Cancer: A Multicenter Randomized Phase 3 Clinical Trial.

IMPORTANCE: Most older patients with esophageal cancer cannot complete the standard concurrent chemoradiotherapy (CCRT). An effective and tolerable chemoradiotherapy regimen for older patients is needed. OBJECTIVE: To evaluate the efficacy and toxic effects of CCRT with S-1 vs radiotherapy (RT) alone in older patients with esophageal cancer. DESIGN, SETTING, AND PARTICIPANTS: A randomized, open-label, phase 3 clinical trial was conducted at 23 Chinese centers between June 1, 2016, and August 31, 2018. The study enrolled 298 patients aged 70 to 85 years. Eligible participants had histologically confirmed esophageal cancer, stage IB to IVB disease based on the 6th edition of the American Joint Committee on Cancer (stage IVB: only metastasis to the supraclavicular/celiac lymph nodes) and an Eastern Cooperative Oncology Group performance status of 0 to 1. Data analysis was performed from August 1, 2020, to March 10, 2021. INTERVENTIONS: Patients were stratified according to age (<80 vs ≥80 years) and tumor length (<5 vs ≥5 cm) and randomly assigned (1:1) to receive either CCRT with S-1 or RT alone. MAIN OUTCOMES AND MEASURES: The primary end point was the 2-year overall survival rate using intention-to-treat analysis. RESULTS: Of the 298 patients enrolled, 180 (60.4%) were men. The median age was 77 (interquartile range, 74-79) years in the CCRT group and 77 (interquartile range, 74-80) years in the RT alone group. A total of 151 patients (50.7%) had stage III or IV disease. The CCRT group had a significantly higher complete response rate than the RT group (41.6% vs 26.8%; P = .007). Surviving patients had a median follow-up of 33.9 months (interquartile range: 28.5-38.2 months), and the CCRT group had a significantly higher 2-year overall survival rate (53.2% vs 35.8%; hazard ratio, 0.63; 95% CI, 0.47-0.85; P = .002). There were no significant differences in the incidence of grade 3 or higher toxic effects between the CCRT and RT groups except that grade 3 or higher leukopenia occurred in more patients in the CCRT group (9.5% vs 2.7%; P = .01). Treatment-related deaths were observed in 3 patients (2.0%) in the CCRT group and 4 patients (2.7%) in the RT group. CONCLUSIONS AND RELEVANCE: In this phase 3 randomized clinical trial, CCRT with S-1 was tolerable and provided significant benefits over RT alone in older patients with esophageal cancer. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02813967.

Silencing the Expression of Cyclin G1 Enhances the Radiosensitivity of Hepatocellular Carcinoma In Vitro and In Vivo by Inducing Apoptosis.

Radiotherapy plays an important role in the treatment of hepatocellular carcinoma (HCC). Cyclin G1 is a novel member of the cyclin family, and it is abnormally expressed in HCC. In this study we investigated the role of cyclin G1 in the radiotherapy of HCC cells. The expression of cyclin G1 was silenced by transfection of cyclin G1-siRNA into HepG2 cells and Huh7 cells, and the expression of cyclin G1 mRNA and protein was measured by qRT-PCR and Western blot analysis. The proliferation was analyzed using MTT assay, and the radiosensitivity of HCC cells was detected using colony formation assay and a xenograft tumor model. The expression of apoptosis-related proteins (Bcl-2 and Bax) was detected by Western blot analysis, and caspase-3 was detected using fluorimetry. The expression of cyclin G1 mRNA and protein in HepG2/Huh7-cyclin G1-siRNA cells was found to be significantly decreased compared to that in HepG2/Huh7 cells. Silencing the expression of cyclin G1 inhibited the proliferation of HCC cells and enhanced radiosensitivity in HCC cells in vitro and in vivo. Knockdown of cyclin G1 expression significantly decreased Bcl-2 expression, and increased Bax expression and caspase-3 activity in HCC cells. Silencing of cyclin G1 expression enhances the radiosensitivity of HCC cells in vitro and in vivo. The mechanism for this may be related to the regulation of apoptosis-related proteins.

Suppression of CCT3 inhibits the proliferation and migration in breast cancer cells.

BACKGROUND: CCT3 is a subunit of chaperonin-containing TCP-1 (CCT), which folds many proteins involved in cancer development and plays an important role in many cancers. However, the role of CCT3 in breast cancer is still unclear. METHODS: CCT3 expression was knocked down by transfecting breast cancer cells with lentiviral shRNA. The proliferation of breast cancer cells (HCC1937 and MDA-MB-231) was detected by Celigo image cytometry and MTT assay, the migration of the cells was measured by Transwell analysis, cell cycle distribution and apoptosis was detected by flow cytometry, and changes in signal transduction proteins were detected by western blot analysis. RESULTS: The expression of CCT3 was significantly suppressed by transduction with lentiviral shRNA; CCT3 knockdown significantly reduced the proliferation and metastasis ability of breast cancer cells (HCC 1937 and MDA-MB-231), increased the proportion of cells in S phase, and decreased the proportion of cells in G1 phase compared to those in shControl cells. There was no significant change in the number of cells in the G2/M phase. Apoptosis analysis showed that knockdown of CCT3 induced apoptosis in breast cancer cells. Western blot analysis showed that the expression of many signal transduction proteins was changed after suppression of CCT3. A rescue experiment showed that overexpression of NFκB-p65 rescued the cell proliferation and migration affected by CCT3 in breast cancer cells. CONCLUSION: CCT3 is closely related to the proliferation and migration of breast cancer and may be a novel therapeutic target.

3D pseudocontinuous arterial spin-labeling perfusion imaging detected crossed cerebellar diaschisis in acute, subacute and chronic intracerebral hemorrhage.

OBJECTIVE: We aimed to evaluate the value of 3D pseudocontinuous arterial spin-labeling (pCASL) perfusion imaging detected crossed cerebellar diaschisis (CCD) at different stages of intracerebral hemorrhage (ICH). MATERIALS AND METHODS: We assessed bilateral cerebral blood flow (CBF) values of different brain regions and the relationships between the CCD and clinical status of 16 ICH patients. RESULTS: The ICH patients had significantly lower CBF values in the contralateral cerebellum in acute, subacute and chronic stages. The subacute CCD had a significant correlation with clinical status. CONCLUSIONS: 3D pCASL may be an ideal tool to study the phenomenon and clinical consequences of ICH with CCD.

L-asparaginase-based regimen as a first-line treatment for newly diagnosed nasal type extranodal natural killer cell/T-cell lymphoma.

The aim of the present study was to compare the efficacy of an L-asparaginase-based regimen and a CHOP regimen followed by radiotherapy as first-line treatments for newly diagnosed nasal type extranodal natural killer cell/T-cell lymphoma (ENKTL). A total of 69 patients received the CHOP regimen as the first-line treatment and 112 patients received the L-asparaginase-based regimen. All patients received radical radiotherapy following two cycles of chemotherapy. The overall response rates of the L-asparaginase-based and CHOP treatment groups were 90.18 and 72.46%, respectively (P=0.002). The one, two, and five-year overall survival (OS) rates and progression-free survival (PFS) rates of the L-asparaginase group were 96.0, 88.3, 65.1, 94.2, 79.8 and 50.0%, respectively. The one, two, and five-year OS and PFS rates of the CHOP group were 82.6, 61.9, 25.8, 63.8, 44.0 and 21.0%, respectively (P<0.001). Compared with CHOP treatment, L-asparaginase-based chemotherapy combined with radiotherapy was a safe and highly effective treatment for newly diagnosed ENKTL.

Caspase-3 expression in metastatic lymph nodes of esophageal squamous cell carcinoma is prognostic of survival.

AIM: To assess whether differential expression of caspase-3 in paired metastatic lymph nodes (LNs) is prognostic of survival in patients with resectable esophageal squamous cell carcinoma (ESCC). METHODS: Capases-3 expression was evaluated immunohistochemically in 122 pairs of primary ESCCs and regional metastatic LNs assembled on tissue microarrays. The impact of caspase-3 expression on survival outcomes was analyzed by the Kaplan-Meier method and Cox proportional hazards regression model. RESULTS: The level of caspase-3 expression was significantly higher in LN metastases than in primary tumors (P < 0.001). Caspase-3 expression in the primary tumors was associated with longer median survival (23 mo vs 21 mo, P = 0.033), whereas higher expression in paired metastatic LNs was associated with shorter median survival (20 mo vs 22 mo, P = 0.043). Multivariate analysis showed that both were independent prognostic factors. CONCLUSION: Caspase-3 expression in metastatic LNs may be a potential independent predictor of poorer overall survival in patients with resected ESCC and LN metastasis. Protein expression in metastatic tumors may be a biomarker prognostic of survival.

Promising outcomes of definitive chemoradiation and cetuximab for patients with esophageal squamous cell carcinoma.

This study investigated cetuximab added to definitive concurrent chemoradiation for esophageal squamous cell carcinoma (ESCC). Previously untreated patients with stage II-IVa ESCC received cetuximab (400 mg/m(2) per week in week 1, then 250 mg/m(2) per week during weeks 2-8), paclitaxel (45 mg/m(2) per week) and cisplatin (20 mg/m(2) per week) in weeks 2-8 with 59.4 Gy radiotherapy. Epidermal growth factor receptor (EGFR) status in tumor specimens was assessed. Thirty-one patients were enrolled and evaluated for toxicity. Of the 29 patients assessable for a response, 20 (69.0%) had a clinical complete response (CR). Over a median follow up of 23.6 months, disease progression was observed in seven patients. The 1- and 2-year progression-free survival (PFS) rates were 85.5% and 75.1%, respectively. The PFS was shorter for patients with lymphatic metastatic disease than for those with locally confined tumor; the 1-year PFS rates were 78.7% and 92.3%, respectively (P = 0.038). Sixteen (55.2%) patients were immunohistochemically positive for EGFR. The patients with EGFR-expressing tumor had a CR rate of 75.0% compared with 61.5% in those with negative EGFR expression (P = 0.024). The PFS for patients with EGFR-expressing tumor was longer compared with the PFS of patients with negative EGFR (P = 0.133). The patients with prominent cetuximab-induced rash (≥grade 2) had a better CR rate and PFS than those with no or grade 1 rash (P < 0.05). The rates of grades 3/4 esophagitis, hematological and dermatological toxicities were 9.7%, 29.0% and 16.1%, respectively. The regimen of definitive chemoradiation plus cetuximab achieved good clinical response and has an acceptable safety profile in Chinese ESCC patients.

Separation and characterization of clindamycin phosphate and related impurities in injection by liquid chromatography/electrospray ionization mass spectrometry.

A simple high-performance liquid chromatography/electrospray ionization tandem mass spectrometric (HPLC/ESI-MS/MS) method has been developed for the rapid identification of clindamycin phosphate and its degradation products or related impurities in clindamycin phosphate injection. Detection was performed by quadrupole time-of-flight mass spectrometry (Q-TOFMS) via an ESI source in positive mode. Clindamycin phosphate and its related substances lincomycin, 7-epilincomycin-2-phosphate, lincomycin-2-phosphate, clindamycin B, clindamycin B-2-phosphate, and clindamycin were identified simultaneously by HPLC/ESI-MS/MS results. Based on the MS/MS spectra of their quasi-molecular ions, the fragmentation pathways of clindamycin phosphate and its related substances were compared and proposed, which are specific and useful for the identification of the lincosamide antibiotics and related impurities. The method was rapid, sensitive and specific and can be used to identify clindamycin phosphate and its related impurities in clindamycin phosphate injection without control compounds.

Separation and identification of purine nucleosides in the urine of patients with malignant cancer by reverse phase liquid chromatography/electrospray tandem mass spectrometry.

Urinary-modified nucleosides have a potential role as cancer biomarkers for a number of malignant diseases. High performance liquid chromatography (HPLC) was combined with full-scan mass spectrometry, MS/MS analysis and accurate mass measurements in order to identify purine nucleosides purified from urine. Potential purine nucleosides were assessed by their evident UV absorbance in the HPLC chromatogram and then further examined by the mass spectrometric techniques. In this manner, numerous modified purine nucleosides were identified in the urine samples from cancer patients including xanthine, adenosine, N1-methyladenosine, 5'-deoxy-5'-methylthioadenosine, 2-methyladenosine, N6-threonylcarbamoyladenosine, inosine, N1-methylinosine, guanosine, N1-methylguanosine, N7-methylguanine, N2-methylguanosine, N2,N2-dimethyguanosine, N2,N2,N7-trimethylguanosine. Furthermore, a number of novel purine nucleosides were tentatively identified via critical interpretation of the combined mass spectrometric data including N3-methyladenosine, N7-methyladenine, 5'-dehydro-2'-deoxyinosine, N3-methylguanine, O6-methylguanosine, N1,N2,N7-trimethylguanosine, N1-methyl-N2-ethylguanosine and N7-methyl-N1-ethylguanosine.

Analysis of modified nucleosides in the urine of patients with malignant cancer by liquid chromatography/electrospray ionization mass spectrometry.

As modified nucleosides reflect altered tRNA turnover which seems to be impaired in the body of cancer patients, they have been evaluated as potential tumor markers. High-performance liquid chromatography/electrosprary ionization quadrupole time-of-flight mass spectrometry (HPLC/ESI-Q-TOFMS) was used to identify nucleosides purified from urine in positive ionization mode. Potential nucleosides were assessed by their evident UV absorbance in HPLC and then further examined by mass spectrometric techniques. In this manner, 21 nucleosides were detected in the urine of a patient with lymphoid cancer including three modified nucleosides 5'-dehydro-2-deoxyinosine, N1,N2,N7-trimethylguanosine and N1-methyl-N2-ethylguanosine, which had never been reported previously.