Chromodomain-helicase-DNA-binding protein 5 (CHD5) is a newly identified tumor suppressor that is frequently downregulated in a variety of human cancers. Our previous work revealed that the low ...expression of CHD5 in colorectal cancer is correlated with CHD5 promoter CpG island hypermethylation. In this study, we investigated the effect of microRNA-211 (miR-211)-regulated CHD5 expression on colorectal tumorigenesis.
miR-211 was predicted to target CHD5 by TargetScan software analysis. A stably expressing exogenous miR-211 colorectal cancer cell line (HCT-116(miR-211)) was generated using lentiviral transduction and used as a model for in vitro and in vivo studies. The expression level of miR-211 in HCT-116(miR-211) cells was upregulated by 16-fold compared to vector control cells (HCT-116(vector)). Exogenous miR-211 directly binds to the 3'-untranslated region (3'-UTR) of CHD5 mRNA, resulting in a 50% decrease in CHD5 protein level in HCT-116(miR-211) cells. The levels of cell proliferation, tumor growth, and cell migration of HCT-116(miR-211) cells were significantly higher than HCT-116(vector) cells under both in vitro and in vivo conditions, as determined using the methods of MTT, colony formation, flow cytometry, scratch assay, and tumor xenografts, respectively. In addition, we found that enforced expression of miR-211 in HCT-116 cells was able to alter p53 pathway-associated regulatory proteins, such as MDM2, Bcl-2, Bcl-xL, and Bax.
Our results demonstrate that CHD5 is a direct target of miR-211 regulation. Enforced expression of miR-211 promotes tumor cell growth at least in part by downregulating the expression level of the CHD5 tumor suppressor. Our results provide a better understanding of the association of between miR-211-regulated CHD5 expression and CHD5 function in colorectal tumorigenesis.
Hepatocellular carcinoma (HCC) is a lethal malignancy with high mortality. The inhibition of cyclin-dependent kinase 7 (CDK7) activity has shown therapeutic efficacy in HCC. However, the underlying ...molecular mechanisms remain elusive. Here, we show that three HCC lines, HepG2, Hep3B, and SK-Hep-1, were highly susceptible to the CDK7 inhibitor THZ1. In mouse models, THZ1 effectively reduced HepG2 tumor growth and tumor weight. THZ1 arrested cell cycle and triggered MYC-related apoptosis in HepG2. To evaluate how MYC protein levels affected THZ1-induced apoptotic cell death, we overexpressed MYC in HepG2 and found that exogenously overexpressed MYC promoted cell cycle progression and increased cells in the S phase. THZ1 drastically engendered the apoptosis of MYC-overexpressing HepG2 cells in the S and G2/M phases. Importantly, transcription-inhibition-induced apoptosis is associated with DNA damage, and exogenous MYC expression further enhanced the THZ1-induced DNA damage response in MYC-overexpressing HepG2 cells. Consistently, in the HepG2 xenografts, THZ1 treatment was associated with DNA-damage-induced cell death. Together, our data indicate that the converged effect of MYC-promoted cell cycle progression and CDK7 inhibition by THZ1 confers the hypersensitivity of HCC to DNA-damage-induced cell death. Our findings may suggest a new therapeutic strategy of THZ1 against HCC.
Abstract
The innate immune stimulator of interferon genes (STING) pathway is known to activate type I interferons (IFN-I) and participate in generating antitumor immunity. We previously produced ...hDT806, a recombinant diphtheria immunotoxin, and demonstrated its efficacy against head and neck squamous cell carcinoma (HNSCC). However, it’s unknown whether the tumor-intrinsic STING plays a role in the anti-HNSCC effects of hDT806. In this study, we investigated the innate immune modulation of hDT806 on HNSCC. hDT806 significantly upregulated the level of STING and the ratio of p-TBK1/TBK1 in the HNSCC cells. Moreover, intratumoral hDT806 treatment increased the expression of STING-IFN-I signaling proteins including IFNA1, IFNB, CXCL10 and MX1, a marker of IFN-I receptor activity, in the HNSCC xenografts. Overexpression of STING mimicked the hDT806-induced upregulation of the STING-IFN-I signaling and induced apoptosis in the HNSCC cells. In the mouse xenograft models of HNSCC with STING overexpression, we observed a significant suppression of tumor growth and reduced tumor weight with increased apoptosis compared to their control xenograft counterparts without STING overexpression. Collectively, our data revealed that hDT806 may act as a stimulator of tumor-intrinsic STING-IFN-I signaling to inhibit tumor growth in HNSCC.
By analyzing the expression profile of microRNAs in head and neck squamous cell carcinomas (HNSCC), we found that the expression level of miR-124 was 4.59-fold lower in tumors than in normal tissues. ...To understand its functions, we generated a miR-124-expressing subline (JHU-22miR124) and a mock vector-transfected subline (JHU-22vec) by transfecting the mimic of miR-124 into JHU-22 cancer cells. Restored expression of miR-124 in JHU-22miR124 cells led to reduced cell proliferation, delayed colony formation, and decreased tumor growth, indicating a tumor-suppressive effect of miR-124. Subsequent target search revealed that the 3'-UTR of SphK1 mRNA carries a complementary site for the seed region of miR-124. SphK1 was also detected to be overexpressed in HNSCC cell lines, but down-expressed in JHU-22miR124 cells and tumor xenografts. These results suggest that SphK1 is a target of miR-124. To confirm this finding, we constructed a 3'-UTR-Luc-SphK1 vector and a binding site-mutated luciferase reporter vector. Co-transfection of 3'-UTR-Luc-SphK1 with miR-124 expression vector exhibited a 9-fold decrease in luciferase activity compared with mutated vector, suggesting that miR-124 inhibits SphK1 activity directly. Further studies on downstream signaling demonstrated accumulation of ceramide, increased expression of the pro-apoptotic Bax, BAD and PARP, decreased expression of the anti-apoptotic Bcl-2 and Bcl-xL, and enhanced expression of cytochrome c and caspase proteins in JHU-22miR124 compared with JHU-22vec cells and tumor xenografts. We conclude that miR-124 acts as a tumor suppressor in HNSCC by directly inhibiting SphK1 activity and its downstream signals.
Incidence of head and neck squamous cell carcinoma (HNSCC) has continuously increased in past years while its survival rate has not been significantly improved. There is a critical need to better ...understand the genetic regulation of HNSCC tumorigenesis and progression. In this study, we comprehensively analyzed the function of miRNA-128 (miR-128) in the regulation of HNSCC growth and its putative targets in vitro and in vivo systems.
The function and targets of miR-128 were investigated in human HNSCC cell lines (JHU-13 and JHU-22), which were stably transfected with the miR-128 gene using a lentiviral delivery system. The expression levels of miR-128 and its targeted proteins were analyzed with qRT-PCR, Western blotting and flow cytometry. The binding capacity of miRNA-128 to its putative targets was determined using a luciferase report assay. MTT, colony formation, and a tumor xenograft model further evaluated the effects of miR-128 on HNSCC growth.
We generated two miR-128 stably transfected human HNSCC cell lines (JHU-13miR-128 and JHU-22miR-128). Enforced expression of miR-128 was detected in both cultured JHU-13miR-128 and JHU-22miR-128 cell lines, approximately seventeen to twenty folds higher than in vector control cell lines. miRNA-128 was able to bind with the 3'-untranslated regions of BMI-1, BAG-2, BAX, H3f3b, and Paip2 mRNAs, resulting in significant reduction of the targeted protein levels. We found that upregulated miR-128 expression significantly inhibited both JHU-13miR-128 and JHU-22miR-128 cell viability approximately 20 to 40%, and the JHU-22miR-128 tumor xenograft growth compared to the vector control groups.
miR-128 acted as a tumor suppressor inhibiting the HNSCC growth by directly mediating the expression of putative targets. Our results provide a better understanding of miRNA-128 function and its potential targets, which may be valuable for developing novel diagnostic markers and targeted therapy.
Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer with limited treatment options. It is urgent to develop new therapeutics against this disease. Salvinolic acid B (Sal-B) ...is a leading bioactive component of Salvia
Bunge, a well-known Chinese medicine for treating various diseases without appreciable adverse effects. To understand the antitumor properties of Sal-B against TNBC, we analyzed its effects on the cell viability, cell cycle and apoptosis of triple-negative MDA-MB-231 cells with the hormone receptor-positive MCF-7 cells as the control. The
analysis showed that Sal-B could significantly reduce the cell viability and suppress the proliferation of both MDA-MB-231 and MCF-7 cells with decreased cyclin B1 expression, but with no noticeable cell cycle phase change. In mouse models, Sal-B markedly inhibited the growth, decreased the PCNA expression, and increased the cell apoptosis of MDA-MB-231 tumor xenografts. To understand the antitumor mechanisms, we analyzed the expression levels of ceramides, and anti-apoptotic (Bcl-xL and survivin) and pro-apoptotic (caspase-3 and caspase-8) proteins. We found that Sal-B enhanced the ceramide accumulation and inhibited the anti-apoptotic protein expression. Interestingly, the ceramide accumulation was accompanied by decreased expression of glucosylceramide and GM3 synthases, two key enzymes regulating ceramide metabolism. These findings indicate that Sal-B exerts its antitumor effects at least partially by inducing the ceramide accumulation and ceramide-mediated apoptosis via inhibiting the expression of glucosylceramide and GM3 synthases, which was independent of estrogen receptor α. Sal-B appears to be a promising therapeutic agent against TNBC.
Over 90% of head and neck squamous cell carcinoma (HNSCC) overexpresses the epidermal growth factor receptor (EGFR). However, the EGFR-targeted monotherapy response rate only achieves 10-30% in ...HNSCC. Recombinant immunotoxin (RIT) often consists of an antibody targeting a tumor antigen and a toxin (e.g., diphtheria toxin DT) that kills cancer cells. We produced a humanized RIT, designated as hDT806, targeting overexpressed EGFR and investigated its effects in HNSCC. Distinct from the EGFR-targeted tyrosine kinase inhibitor erlotinib or antibody cetuximab, hDT806 effectively suppressed cell proliferation in the four HNSCC lines tested (JHU-011, -013, -022, and -029). In JHU-029 mouse xenograft models, hDT806 substantially reduced tumor growth. hDT806 decreased EGFR protein levels and disrupted the EGFR signaling downstream effectors, including MAPK/ERK1/2 and AKT, while increased proapoptotic proteins, such as p53, caspase-9, caspase-3, and the cleaved PAPR. The hDT806-induced apoptosis of HNSCC cells was corroborated by flow cytometric analysis. Furthermore, hDT806 resulted in a drastic inhibition in RNA polymerase II carboxy-terminal domain phosphorylation critical for transcription and a significant increase in the γH2A.X level, a DNA damage marker. Thus, the direct disruption of EGFR signaling, transcription inhibition, DNA damage, as well as apoptosis induced by hDT806 may contribute to its antitumor efficacy in HNSCC.
The photophysical properties of a chlorin, isobacteriochlorin and bacteriochlorin built on a core tetrapentafluorophenylporphyrin (TPPF20) and the nonhydrolyzable para thioglycosylated conjugates of ...these chromophores are presented. The photophysical characterization of these compounds was done in three different solvents to correlate with different environments in cells and tissues. Compared with TPPF20 other dyes have greater absorption in the red region of the visible spectrum and greater fluorescence quantum yields. The excited state lifetimes are from 3 to 11 ns. The radiative and nonradiative rate constants for deactivation of the excited state were estimated from the fluorescence quantum yield and excited state lifetime. The data indicate that the bacteriochlorin has strong absorption bands near 730 nm and efficiently enters the triplet manifold. The isobacteriochlorin has a 40–70% fluorescence quantum yield depending on solvent, so it may be a good fluorescent tag. The isobacteriochlorins also display enhanced two‐photon absorption, thereby allowing the use of 860 nm light to excite the compound. While the two‐photon cross section of 25 GM units is not large, excitation of low chromophore concentrations can induce apoptosis. The glycosylated compounds accumulate in cancer cells and a head and neck squamous carcinoma xenograft tumor model in mice. These compounds are robust to photobleaching.
Click‐type chemistry allows tetraglycosylation of the perfluorophenyl derivatives of a chlorin, bacteriochlorin, and an isobacteriochlorin in high yields. The four non‐hydrolysable sugars target the chromophores to mouse models of head and neck squamous cell carcinoma. Here the glycosylated chlorin allows fluorescence imaging of these tumors.
Public understanding of human papillomavirus (HPV)-associated oropharyngeal cancer (OPC) is minimally understood. Therefore uncovering communication gaps between the public and healthcare ...professionals regarding this disease is vital. Social media provide an unobtrusive way to understand public perception about health issues.
Computer-assisted quantitative content analysis.
Tweets about HPV-associated OPC (N = 3,112) were collected for 40 weeks using the standard real-time streaming Application Programming Interface (API). The collection of tweets was not limited to one specific geographic location but worldwide. All tweets were entered into nVivo 12.0 to conduct computer-assisted quantitative content analysis. We used an inductive method to develop a coding scheme and examined the frequency of specific keywords, terms, and phrases in texts.
Findings show that (a) the majority of discourse on Twitter focused on risk factors and prevention with little information on diagnosis, treatment, and prognoses; (b) many tweets promoted HPV vaccination among boys and emphasized the risk of HPV-associated OPC among males; (c) the role of dental care professionals in the prevention and detection of OPC minimally appeared; (d) the public referred to OPC as oral cancer, head and neck cancer, or throat cancer; and (e) health organizations in New Zealand, Australia, and the United Kingdom led the discussion on HPV-associated OPC on Twitter.
The current study unravels the utility of social media data and data mining techniques in understanding public perception and understanding of HPC-associated OPC. The outcomes from the current study provide baseline knowledge of where communication gaps exist in terms of HPV-associated OPC, without which the planning of potential interventions and much-needed social media-based campaigns cannot be effectively undertaken.
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