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Pancreatic cancer (PC) has a high mortality rate due to its poor prognosis and the possibility of surgical resection in patients with the disease. Importantly, adjuvant chemotherapy is necessary to improve PC prognosis. Chrysin, a natural product with anti-inflammatory, antioxidant, and anticancer properties, has been studied for several years. Our previous study demonstrated that chrysin induced G protein-coupled estrogen receptor (GPER) expression and regulated its activity in breast cancer. Herein, we investigated whether chrysin-induced GPER activation suppresses PC progression in MIA PaCa-2 cells and a xenograft model. To determine its mechanism of action, cytotoxicity and clonogenic assays, a FACS analysis, and Western blotting were performed. Furthermore, the delay in tumor growth was evaluated in the MIA PaCa-2-derived xenograft model. Tumor tissues were investigated by Western blotting, immunohistochemistry, and a proteomic analysis. Chrysin caused cell cycle arrest and significantly decreased cell viability. Following co-treatment with chrysin and 17β-estradiol, the inhibitory effect of chrysin on cell proliferation was enhanced. In the xenograft model, chrysin and G1 (a GPER agonist) significantly delayed tumor growth and reduced both Ki-67 (a proliferation marker) and c-Myc expressions in tumor tissues. The proteomic analysis of tumor tissues identified that rho-associated coiled-coil containing protein kinase 1 (ROCK1), transgelin 2 (TAGLN2), and FCH and Mu domain containing endocytic adaptor 2 (FCHO2) levels were significantly reduced in chrysin-treated tumor tissues. High ROCK1, TAGLN2, and FCHO2 expressions were indicative of low overall PC survival as found using the Kaplan–Meier plotter. In conclusion, our results suggest that chrysin suppresses PC progression through the activation of GPER and reductions in ROCK1, TAGLN2, and FCHO2 expressions.

Liver cancer, one of the leading death causes, has different incidence and mortality rates in men and women. The influencing factor is considered to estrogen. However, the role of estrogen in liver cancer remains controversial. In this study, we investigated the effects of estrogen on tumor progression. Total RNA sequencing was analyzed in SK-Hep1-derived tumor tissues, and 15 genes were expressed only in female mice. Among the differentially expressed genes, matrix metalloprotease 7 (MMP7), germ cell associated 1 (GSG1), and chromosome 6 open reading frame 15 (C6orf15) were associated with significantly different overall survival rates based on their expression level in liver cancer patients. Interestingly, exogenous estrogen aggravated SK-Hep1-derived tumor growth in ovariectomized (OVX) mice. When OVX mice were treated with exogenous estrogen, SK-Hep1-derived tumor tissues exhibited high MMP7 expression levels and low GSG1 and C6orf15 expression levels. These expression patterns were consistent with those of liver cancer patients with low overall survival rates. These results suggest that these genes are expected to be prognostic biomarkers of liver cancer. In conclusion, our results suggest that continuous estrogen exposure may promote tumor growth in OVX mice.

Gastric cancer (GC) is the most common cancer worldwide and the third leading cause of cancer death, with the fifth highest incidence. The development of effective chemotherapeutic agents is needed to decrease GC mortality. Policosanol (PC) extracted from Cuban sugar cane wax is a healthy functional food ingredient that helps improve blood cholesterol levels and blood pressure. Its various physiological activities, such as antioxidant, anti-inflammatory, and anticancer activities, have been reported recently. Nevertheless, the therapeutic efficacy of PC in gastric xenograft models is unclear. We aimed to investigate the anticancer effect of PC on human GC SNU-16 cells and a xenograft mouse model. PC significantly inhibited GC cell viability and delayed tumor growth without toxicity in the SNU-16-derived xenograft model. Therefore, we investigated protein expression levels in tumor tissues; the expression levels of Ki-67, a proliferation marker, and cdc2 were decreased. In addition, we performed proteomic analysis and found thirteen differentially expressed proteins. Our results suggested that PC inhibited GC progression via cdc2 suppression and extracellular matrix protein regulation. Notably, our findings might contribute to the development of novel and effective therapeutic strategies for GC.

Activation of sterol regulatory element-binding protein 1 (SREBP-1), a master lipogenic transcription factor, is associated with cancer metabolism and metabolic disorders. Neddylation, the process of adding NEDD8 to its substrate, contributes to diverse biological processes. Here, we identified SREBP-1 as a substrate for neddylation by UBC12 and explored its impact on tumor aggressiveness. In cell-based assays, SREBP-1 neddylation prolonged SREBP-1 stability with a decrease in ubiquitination. Consequently, NEDD8 overexpression facilitated proliferation, migration, and invasion of SK-Hep1 liver tumor cells. MLN4924 (an inhibitor of the NEDD8-activating enzyme-E1) treatment or UBC12 knockdown prevented SREBP-1 neddylation and tumor cell phenotype change. This effect was corroborated in an in vivo xenograft model. In human specimens, SREBP-1, UBC12, and NEDD8 were all upregulated in hepatocellular carcinoma (HCC) compared to nontumorous regions. Moreover, SREBP-1 levels positively correlated with UBC12. In GEO database analyses, SREBP-1 levels were greater in metastatic HCC samples accompanying UBC12 upregulation. In HCC analysis, tumoral SREBP-1 and UBC12 levels discriminated overall patient survival rates. Additionally, MLN4924 treatment destabilized SREBP-1 in MDA-MB-231 breast cancer cells and in the tumor cell xenograft. SREBP-1 and UBC12 were also highly expressed in human breast cancer tissues. Moreover, most breast cancers with lymph node metastasis displayed predominant SREBP-1 and UBC12 expressions, which compromised overall patient survival rates. In summary, SREBP-1 is neddylated by UBC12, which may contribute to HCC and breast cancer aggressiveness through SREBP-1 stabilization, and these events can be intervented by MLN4924 therapy. Our findings may also provide potential reliable prognostic markers for tumor metastasis.

Metastasis decreases the survival rate of patients with liver cancer. Therefore, novel anti-metastatic strategies are needed. Korean Red Ginseng (KRG) is often ingested as a functional food with an immune-boosting effect. We investigated a combination of KRG and natural killer (NK) cells as a novel immunotherapy approach. SK-Hep1 cells were injected into the tail vein of NRGA mice to establish an experimental metastasis model. KRG, NK cells, or a combination of KRG and NK cells were administered. Tumor growth was observed using an in vivo imaging system, and metastatic lesions were evaluated by histological analysis and immunohistochemistry. Bioluminescence intensity was lower in the KRG and NK cell combination group than in the other groups, indicating that the combination treatment suppressed the progression of metastasis. CD56 expression was used as a NK cell marker and hematological analysis was performed. The combination treatment also decreased the expression of matrix metalloproteinases and the area of metastatic lesions in liver and bone tissues, as well as increased the eosinophil count. Expression of cytokines-related eosinophils and NK cells was determined by Western blotting analysis. The expression of interleukin 33 (IL33) was induced by the combination of KRG and NK cells. High IL33 expression was associated with prolonged overall survival in the Kaplan–Meier plotter. Our results suggest that KRG enhances the immune activity of NK cells by IL-33 through eosinophils and suppresses metastatic liver cancer progression.

Cancer is the leading cause of mortality worldwide. In cancer progression, sex hormones and their receptors are thought to be major factors. Many studies have reported the effects of estrogen and estrogen receptors (ERs) in cancer development and progression. Among them, G protein-coupled estrogen receptor (GPER), a G proteincoupled receptor, has been identified as an estrogen membrane receptor unrelated to nuclear ER. The mechanism of GPER, including its biological action, function, and role, has been studied in various cancer types. In this review, we discuss the relation between GPER and estrogen or estrogen agonists/antagonists and cancer progression.

RH1 (2, 5-diaziridinyl-3-(hydroxymethy)-6-methyl-1, 4-benzoquinone) is a bioreductive anticancer drug. The mechanism underlying its therapeutic properties has not yet been elucidated. In this study, we aimed to determine whether RH1 exerts its anticancer effect via p53-mediated apoptosis and senescence in vitro and in vivo. RH1 displayed dose-dependent biphasic effects in vitro, i.e., it induced apoptosis at higher dose and senescence at lower dose accompanied by marked activation of p53. Thus, RH1 primarily induced cell death by apoptosis. The cytotoxicity of RH1 was inhibited in A549 cells treated with the p53-inhibitor pifithrin-α or transfected p53 siRNA and in human colon cancer HCT116 isogenic (p53-/-) cells. At sub-lethal doses of RH1, the cells survived and underwent senescence. The senescent cells showed flattened and enlarged morphology, and exhibited blue color in senescence-associated β-galactosidase staining. These changes were found to be related to p53. RH1-induced senescence decreased in A549-E6 cells (suppressed p53 level) and HCT 116 p53-/- cells. The growth of A549 xenograft tumors in nude mice was significantly delayed by intraperitoneal injection of RH1, and senescent cells were observed in these xenograft tumors. These results suggest that the in vivo anticancer therapeutic effect of RH1 is mediated by senescence via p53 activation.

Gastric cancer has the fifth-highest incidence rate and is the third leading cause of cancer-related deaths worldwide. The incidence of gastric cancer is higher in men than in women, but for the diffuse types of gastric cancer, the trend is opposite. Estrogen is considered the prime culprit behind these differences. Nevertheless, the action of estrogen in gastric cancers remains unclear. In this study, we investigated the effect of estrogen on diffuse-type gastric cancer. Human female diffuse gastric cancer SNU-16 cells were transplanted into male and female mice to analyze the effect of endogenous estrogen on tumor growth. Furthermore, the effect of exogenous estrogen was evaluated in ovariectomized mice. Expressed genes were compared between female and male xenograft models using RNA sequencing analysis. Furthermore, human gene expression omnibus databases were utilized to examine the effect of our target genes on overall survival. SNU-16-derived tumor growth was faster in female mice than in male mice. In total RNA sequencing, interferon gamma receptor 2 (IFNGR2), IQ motif containing E (IQCE), transient receptor potential cation channel subfamily M member 4 (TRPM4), and structure-specific endonuclease subunit SLX4 (SLX4) were found. These genes could be associated with the tumor growth in female diffuse-type gastric cancer which was affected by endogenous estrogen. In an ovariectomized gastric cancer xenograft model, exogenous estrogen promoted tumor growth. Especially, our results indicated that estrogen induced G protein-coupled estrogen receptor expression in these mice. These results suggest that estrogen aggravates tumor progression in female diffuse gastric cancer.

Various strategies, such as optimization of surface chemistry, size, shape, and charge, have been undertaken to develop nanoparticles (NPs) as DDS (drug delivery system) nanocarriers for evading the reticuloendothelial system (RES) in vivo. We previously developed a hollow NP composed of hepatitis B virus (HBV) surface antigen L proteins and lipid bilayers, hereinafter referred to as bio-nanocapsule (BNC), as a nonviral DDS nanocarrier. Such a BNC harbors the HBV-derived human hepatic cell-specific infection mechanism, and intravenously injected BNCs by themselves were shown to avoid clearance by RES-rich organs and accumulate in target tissues. In this study, since the surface modification with albumins is known to prolong the circulation time of nanomedicines, we examined whether the polymerized albumin receptor (PAR) of BNCs contributes to RES evasion in mouse liver. Our results show that NPs conjugated with peptides possessing sufficient PAR activity were captured by Kupffer cells less efficiently in vitro and were able to circulate for a longer period of time in vivo. Comparing with polyethylene glycol, PAR peptides were shown to reduce the recognition by RES to equal content. Taken together, our results strongly suggest that the PAR domain of BNCs, as well as HBV, harbors an innate RES evasion mechanism. Therefore, the surface modification with PAR peptides could be an alternative strategy for improving the pharmacodynamics and pharmacokinetics of forthcoming nanomedicines.

Preclinical evaluation models have been developed for precision medicine, with patient-derived xenograft models (PDXs) and patient-derived organoids (PDOs) attracting increasing attention. However, each of these models has application limitations. In this study, an advanced xenograft model was established and used for drug screening. PDO and endothelial colony-forming cells (ECFCs) were cotransplanted in NRGA mice (PDOXwE) to prepare the model, which could also be subcultured in Balb/c nude mice. Our DNA sequencing analysis and immunohistochemistry results indicated that PDOXwE maintained patient genetic information and tumor heterogeneity. Moreover, the model enhanced tumor growth more than the PDO-bearing xenograft model (PDOX). The PDO, PDOXwE, and clinical data were also compared in the liver metastasis of a colorectal cancer patient, demonstrating that the chemosensitivity of PDO and PDOXwE coincided with the clinical data. These results suggest that PDOXwE is an improvement of PDOX and is suitable as an evaluation model for precision medicine.

Cancer-associated fibroblasts (CAFs) in the tumor microenvironment have been associated with tumor progression in breast cancer. Although crosstalk between breast cancer cells and CAFs has been studied, the effect of CAFs on non-neoplastic breast epithelial cells is not fully understood to date. Here, we investigated the effect of CAFs on aggressive phenotypes in non-neoplastic MCF10A breast epithelial cells. CAFs induced epithelial-to-mesenchymal transition (EMT) and invasive phenotype in MCF10A cells. S100A8, a potential prognostic marker in several cancers, was markedly increased in MCF10A cells by CAFs. S100A8 was crucial for CAFs-induced invasive phenotype of MCF10A cells. Among cytokines increased by CAFs, interleukin (IL)-8 induced S100A8 through transcription factors p65 NF-κB and C/EBPβ. In a xenograft mouse model with MCF10A cells and CAFs, tumor was not developed, suggesting that coinjection with CAFs may not be sufficient for in vivo tumorigenicity of MCF10A cells. Xenograft mouse tumor models with MDA-MB-231 breast carcinoma cells provided an in vivo evidence for the effect of CAFs on breast cancer progression as well as a crucial role of IL-8 in tumor growth and S100A8 expression in vivo. Using a tissue microarray of human breast cancer, we showed that S100A8 expression was correlated with poor outcomes. S100A8 expression was more frequently detected in cancer-adjacent normal human breast tissues than in normal breast tissues. Together, this study elucidated a novel mechanism for the acquisition of invasive phenotype of non-neoplastic breast cells induced by CAFs, suggesting that targeting IL-8 and S100A8 may be an effective strategy against breast cancer.

The incidence and mortality rates of gastric cancer rank among the highest five of all cancer types worldwide. The chemotherapeutic agent 5-fluorouracil (5-FU) is the gold standard for treating gastric cancer, but its efficacy is limited due to high rates of resistance. To improve the therapeutic efficacy of 5-FU and overcome its resistance, the synergistic effect of chrysin with 5-FU was investigated and its mechanism was elucidated. Chrysin was co-administered with 5-FU in AGS cells and 5-FU-resistant AGS cells (AGS/FR). Cytotoxicity was investigated using MTT assay, followed by calculating the combination index (CI). Several biomarkers were detected using western blotting analysis. Apoptosis and cell cycle distribution were measured by flow cytometry. The combination of chrysin and 5-FU significantly increased cytotoxicity more than chrysin or 5-FU alone. 5-FU induced apoptosis through p53-p21 activity, while chrysin arrested the cell cycle in the G2/M phase. The combination of chrysin and 5-FU showed an anticancer effect via S phase arrest. The results indicated that chrysin and 5-FU exhibited anticancer properties via different pathways. Furthermore, the present study found that chrysin enhanced the chemotherapeutic effect of 5-FU in AGS/FR cells. In the resistant cells, the combination of chrysin and 5-FU improved the anticancer effect via G2/M phase arrest. These findings indicated that chrysin potentiated the chemotherapeutic effect of 5-FU in gastric cancer AGS and AGS/FR cells via cell cycle arrest. Therefore, chrysin may be used to treat gastric cancers that have become resistant to 5-FU.

Triple-negative breast cancer (TNBC) is associated with a high mortality rate among women globally. TNBC shows a high rate of recurrence and distant metastasis. Particularly, the chemotherapy is limited because hormone therapy of breast cancer is ineffective. Thus, an effective chemotherapeutic agent is needed for tumor suppression. Chrysin-nanoparticles (chrysin-NPs) were investigated for their inhibitory effect on a MDA-MB-231-derived xenograft model. To gain insight into the underlying mechanisms, we conducted human matrix metalloproteinase (MMP) array, western blot, and immunohistochemistry analysis. Furthermore, in vivo imaging was used to monitor the chemotherapeutic efficacy of chrysin-NPs in a metastasis mouse model. Chrysin-NPs significantly inhibited the proliferation of MDA-MB-231 cells via the PI3K/JNK pathway and induced cell death through the p53-apoptosis pathway, leading to delayed MDA-MB-231-derived tumor growth. Interestingly, chrysin-NPs significantly induced G protein-coupled estrogen receptor (GPER) expression, which suppresses MMPs and NF-κB expression. Chrysin-NPs acted as effective metastasis inhibitors. Our results suggest that chrysin-NPs may be used as an effective adjuvant formulation to inhibit TNBC progression.

Triple-negative breast cancer (TNBC) is an aggressive type of breast cancer with a poor prognosis for which no effective therapeutic measures are currently available. The present study aimed to investigate whether interactions with endothelial colony-forming cells (ECFCs) promote aggressive progression of TNBC cells. Herein, using an indirect co-culture system, we showed that co-culture increased the invasive and migratory phenotypes of both MDA-MB-231 TNBC cells and ECFCs. Through a cytokine antibody array and RT-PCR analysis, we revealed that co-culture markedly induced secretion of the chemokine C-C motif ligand (CCL)8 from ECFCs and that of interleukin (IL)-8 from MDA-MB-231 cells. CCL8 was crucial for ECFC-induced IL-8 secretion and invasion of MDA-MB-231 cells as well as for MDA-MB-231-enhanced MMP-2 secretion and angiogenesis of ECFCs. We suggest c-Jun as a transcription factor for CCL8-induced IL-8 expression in MDA-MB-231 cells. IL-8 was important for co-culture-induced CCL8 and MMP-2 upregulation and invasion of ECFCs. Notably, our findings reveal a positive feedback loop between CCL8 and IL-8, which contributes to the aggressive phenotypes of both ECFC and TNBC cells. Using an MDA-MB-231 cell-based xenograft model, we show that tumor growth and metastasis are increased by co-injected ECFCs in vivo. Increased expression of IL-8 was observed in tissues with bone metastases in mice injected with conditioned media from co-cultured cells. High IL-8 levels are correlated with poor recurrence-free survival in TNBC patients. Together, these results suggest that CCL8 and IL-8 mediate the crosstalk between ECFCs and TNBC, leading to aggravation of tumorigenicity in TNBC.

Bio-nanocapsules (BNCs) consisting of hepatitis B virus surface antigen (HBsAg) L proteins and phospholipids are used as efficient non-viral carriers for liver-specific delivery of genes and drugs. Considering the administration to HB vaccinees and HB patients, endogenous anti-HBsAg immunoglobulins (HBIGs) may reduce the delivery efficacy and prevent repetitive administration. Therefore, low immunogenic BNCs were generated by inserting two point mutations in the HBsAg L protein, which were found in HBV escape mutants. Escape mutant-type BNC (emBNC) showed 50% lower HBIG binding capacity than that of parental BNC (wtBNC). It induced HBIG production to a lesser extent than that associated with wtBNC in BALB/c mice. The emBNC could accumulate into human hepatocyte-derived tumor in mice pre-treated with HBIGs. The complex of emBNC and cationic liposomes could deliver plasmid DNA to HepG2 cells efficiently in the presence of HBIGs. Thus, emBNC could evade HBIG-neutralizing antibodies, expanding the clinical utility of BNC-based nanomedicine.

Human hepatocellular carcinoma (HCC) is the most common and even worse at prognosis. The patients with HCC which accompanied by other diseases, such as cirrhosis, can be limited in various treatments, such as chemotherapy, not HCC patients without other diseases. NLRP3 inflammasome plays an important role in the innate immune response, but emerging evidence has indicated that the NLRP3 inflammasome is implicated in all stages of cancer development. Various cells express NLRP3 protein through the autocrine or paracrine signaling in their environment, but NK cells do not. The expanding evidence shows that patients who suffer from liver cancers have a low frequency of natural killer (NK) cells, and the function of these cells is also impaired. Thus, we examined how the expression of NLRP3 in HCC cells affects cancer surveillance by NK cells in a state of a co-culture of both cells. When the expression of NLRP3 in HCC cells was ablated, MICA/B on the surface of HCC cells was upregulated through the lowered expression of matrix metalloproteinase. The expression of MICA on the surface of HCC cells interacted with the NKG2D receptor on NK-92 cells, which led to NK cytotoxicity. Furthermore, in a xenograft mice model, NLRP3 KO HCC cells delayed tumor development and metastasis as well as increased the sensitivity to NK cell cytotoxicity. Taken together, NLRP3 KO in HCC could enhance NK immunosurveillance through an interaction of NKG2D-MICA.

The incidence of hepatocellular carcinoma (HCC) has continued to increase worldwide, and advanced HCC is difficult to treat using the currently available therapeutics. Chemoradiotherapy with cisplatin (cis-diamminedichloroplatinum, CDDP) is expected to confer a curative benefit on HCC patients; however, its application is limited due to side-effects such as acute nephrotoxicity as well as the conventionally limited application of chemoradiotherapy for HCC. For the practical application of this drug in the clinical setting, we formulated a novel drug carrier-comprising bio-nanocapsule (BNC) and liposomal CDDP (BNC-LP-CDDP) that recognizes the human liver and releases CDDP. BNC-LP-CDDP showed selectively high cytotoxicity for HCC cells, and markedly reduced the survival fractions of HCC when combined with ionizing radiation (IR) treatment in in vitro assays. In particular, the treatment of mice bearing human HCC with BNC-LP-CDDP and 3 Gy IR showed 95.68% growth inhibition, whereas IR treatment alone showed 65.6% growth inhibition. Moreover, BNC-LP-CDDP led to the withdrawal of CDDP-induced nephrotoxicity. These results indicate that BNC-LP-CDDP in combination with IR markedly enhanced the chemo-radiotherapeutic efficacy and eliminated CDDP induced nephrotoxicity, thus, suggesting the potential for its clinical application as human HCC therapy.

Reactive oxygen species (ROS)-mediated DNA adducts as well as DNA strand breaks are highly mutagenic leading to genomic instability and tumorigenesis. DNA damage repair pathways and oxidative stress response signaling have been proposed to be highly associated, but the underlying interaction remains unknown. In this study, we employed mutant strains lacking Rad51, the homolog of E. coli RecA recombinase, and Yap1 or Skn7, two major transcription factors responsive to ROS, to examine genetic interactions between double-strand break (DSB) repair proteins and cellular redox regulators in budding yeast Saccharomyces cerevisiae. Abnormal expression of YAP1 or SKN7 aggravated the mutation rate of rad51 mutants and their sensitivity to DSB- or ROS-generating reagents. Rad51 deficiency exacerbated genome instability in the presence of increased levels of ROS, and the accumulation of DSB lesions resulted in elevated intracellular ROS levels. Our findings suggest that evident crosstalk between DSB repair pathways and ROS signaling proteins contributes to cell survival and maintenance of genome integrity in response to genotoxic stress.

There is a wide disparity in the incidence, malignancy and mortality of different types of cancer between each sex. The sex-specificity of cancer seems to be dependent on the type of cancer. Cancer incidence and mortality have been demonstrated as sex-specific in a number of different types of cancer, such as liver cancer, whereas sex-specificity is not noticeable in certain other types of cancer, including colon and lung cancer. The present study aimed to elucidate the molecular basis for sex-biased gene expression in cancer. The mRNA expression of the epithelial-to-mesenchymal transition-associated genes was investigated, including E-cadherin (also termed CDH1), vimentin (VIM), discoidin domain receptor 1 (DDR1) and zinc finger E-box binding homeobox 1 (ZEB1) in female- and male-derived cancer cell lines by reverse transcription (RT)-PCR and the Broad-Novartis Cancer Cell Line Encyclopedia (CCLE) database analysis. A negative correlation was observed between DDR1 and ZEB1 only in the female-derived cancer cell lines via RT-PCR analysis. A negative correlation between DDR1 index (defined by the logarithmic value of DDR1 divided by ZEB1, based on the mRNA data from the RT-PCR analysis) and an invasive phenotype was observed in cancer cell lines in a sex-specific manner. Analysis of the CCLE database demonstrated that DDR1 and ZEB1, which are already known to be sex-biased, were negatively correlated in female-derived liver cancer cell lines, but not in male-derived liver cancer cell lines. In contrast, cell lines of colon and lung cancer did not reveal any sex-dependent difference in the correlation between DDR1 and ZEB1. Kaplan-Meier survival curves using the transcriptomic datasets such as Gene Expression Omnibus, European Genome-phenome Archiva and The Cancer Genome Atlas databases suggested a sex-biased difference in the correlation between DDR1 expression pattern and overall survival in patients with liver cancer. The results of the present study indicate that sex factors may affect the regulation of gene expression, contributing to the sex-biased progression of the different types of cancer, particularly liver cancer. Overall, these findings suggest that analyses of the correlation between DDR1 and ZEB1 may prove useful when investigating sex-biased cancers.

Interleukin (IL)-6 plays a crucial role in the progression, invasion, and metastasis of breast cancer. Triple-negative breast cancer (TNBC) cell line MDA-MB-231 is known for its aggressive metastasis. Epithelial to mesenchymal transition (EMT) is a critical process in cancer metastasis. The positive correlation between IL-6 and EMT in tumor microenvironment is reported. We found significantly upregulated IL-6 expression in MDA-MB-231 cells. A blockade of IL-6 expression decreased levels of phosphorylated signal transducer and activator of transcription 3 (pSTAT3), phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K), phosphorylated protein kinase B (pAkt), and cell cycle-related molecules, including cyclin-dependent kinases (CDKs) and cyclins in MDA-MB-231 cells. A short-hairpin RNA (shRNA)-mediated blockade of IL-6 expression inhibited migration and N-cadherin expression and induced E-cadherin expression in MDA-MB-231 cells. Growth rate was slower for the tumors derived from IL-6 shRNA-treated MDA-MB-231 cells than for those derived from control shRNA-treated MDA-MB-231 cells. The expression of pSTAT3, phosphorylated extracellular signal-regulated kinase (pERK), PI3K, pAkt, snail, vimentin, and N-cadherin was significantly lower in tumors from IL-6 shRNA-treated MDA-MB cells. In addition, apigenin treatment significantly inhibited the growth of MDA-MB-231-derived xenograft tumors along with the protein expressions of pSTAT3, pERK, IL-6, PI3K, pAkt, and N-cadherin. Our results demonstrate that the anti-invasive effect of apigenin in MDA-MB-231-derived xenograft tumors is mediated by the inhibition of IL-6-linked downstream signaling pathway.

Liver cancer shows noticeable differences in the incidence rate and mortality between genders. To investigate the estrogen effect on tumor progression in liver cancer, we developed a xenograft model using estrogen pellets. SK-Hep1 cells (human male liver carcinoma) were inoculated into male or female nude mice. Subsequently, estrogen pellets were subcutaneously implanted into these xenograft models. Interestingly, the marked adverse effect of estrogen pellets (0.5 mg/21 days) were observed in the male-derived xenograft model, with increased ulcerative dermatitis in male mice than in female mice. Additionally, necrosis was observed in male mice with SK-Hep1-derived tumors. However, the estrogen pellet (0.5 mg/60 days) did not exhibit these adverse effects. Tumor growth in female mice was significantly suppressed by estrogen (0.5 mg/60 days). Tumor growth was also suppressed in male mice implanted with estrogen (0.5 mg/60 days), but the suppression was not significant. We found that estrogen-induced skin damage was more severe in male mice than female mice. The tumor suppression of estrogen was effective in female mice compared to male mice bearing liver cancer. The results suggest that the sex difference affects estrogen activity and thus should be considered in the preclinical assessment. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s43188-019-00020-6) contains supplementary material, which is available to authorized users.

// Si Yeol Song 1, 3 , Kyu-pyo Kim 2 , Seong-Yun Jeong 3, 4 , Jin Park 3, 4 , Jaesook Park 3, 4 , Joohee Jung 5 , Hye Kyung Chung 6 , Sa-Won Lee 7 , Min Hyo Seo 7 , Jung-shin Lee 2, 3 , Kyung Hae Jung 2 , Eun Kyung Choi 1, 3 1 Department of Radiation Oncology, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Korea 2 Department of Oncology, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Korea 3 Institute for Innovative Cancer Research, Asan Medical Center, Seoul, Korea 4 Asan Institute for Life Sciences, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Korea 5 College of Pharmacy, Duksung Women’s University, Seoul, Korea 6 Korea Institute of Radiological and Medical Sciences, National Project to Establish Platform to Develop The New Concept Therapy, Seoul, Korea 7 Department of Parenteral Delivery Program, Samyang Pharmaceuticals R&D, Daejeon, Korea Correspondence to: Eun Kyung Choi, email: ekchoi@amc.seoul.kr Kyung Hae Jung, email: khjung@amc.seoul.kr Keywords: PNP-DTX, phase I, MTD, efficacy, pancreas Received: March 27, 2016 Accepted: September 25, 2016 Published: October 14, 2016 ABSTRACT We assessed the efficacy of the polymeric nanoparticle containing docetaxel (PNP-DTX) in preclinical mouse models and determined the maximum tolerated dose (MTD) through clinical study. Subcutaneous and orthotopic mouse models were dedicated. Tumor growth delay in orthotopic model and quantification of in vivo imaging in orthotopic model were evaluated. Phase I clinical study was a single-center, prospective, open-label trial in advanced solid tumors. PNP-DTX was injected intravenously and the starting dose was 20 mg/m 2 escalated to 35 mg/m 2 , 45 mg/m 2 , 60 mg/m 2 and 75 mg/m 2 . Pharmacokinetics, tumor response, toxicities were evaluated. Preclinical result revealed the more potent cytotoxic effect of PNP-DTX than docetaxel (DTX). However, there was no difference between PNP-DTX and DTX in subcutaneous model. Tubulin polymerization assay showed that PNP-DTX preserved original mode of action of DTX. For phase I clinical trial, 18 patients were analyzed. The dose of 75 mg/m 2 was tentatively determined as the MTD and the most common toxicity was grade 4 neutropenia not lasting over 7days. The C max of 60 mg/m 2 PNP-DTX and AUC last of 45 mg/m 2 PNP-DTX were measured to be comparable to those of 75 mg/m 2 DTX. Partial remission (PR) was achieved in 4 (22%) patients. The potency of PNP-DTX was revealed especially in orthotopic mouse model. The MTD of PNP-DTX could not be confirmed, but 75 mg/m 2 was tentatively determined. The PNP-DTX of 45 mg/m 2 had the same pharmacokinetic profile with that of 75 mg/m 2 DTX.

Chrysin is a flavone found in several plants, mushroom, and honeycomb. This constituent is broadly used in herbal medicine in Asia. Since its biological activities were identified in various studies, the focus has shifted to the development of chrysin as a complementary medicine for health promotion. Chrysin is known to have chemopreventive and therapeutic effects in skin aging, atherosclerosis, inflammation, diabetes, AIDS, and cancer. However, its poor bioavailability is a bottleneck for pharmaceutical applications. To overcome the limitations and enhance the bioactive effects, methods like nanoencapsulation or conjugation have been attempted. In this review, current trends of chrysin use in the biomedical field are summarized.

Tumor-associated macrophages (TAMs) are major components of tumor microenvironment that promote invasion and metastasis of cancer cells. In this study, we investigated the effect of TAMs on phenotypic conversion of non-neoplastic MCF10A human breast epithelial cells using an indirect co-culture system. Co-culture with TAMs induced epithelial-to-mesenchymal transition, invasive phenotype, and MMP-9 upregulation in MCF10A cells. Comparative proteomic analysis revealed that endoplasmic reticulum oxidoreductase (ERO)1-α was increased in MCF10A cells co-cultured with TAMs compared to that in mono-cultured cells. ERO1-α was crucial for TAMs-induced invasive phenotype and MMP-9 upregulation involving transcription factors c-fos and c-Jun. Cytokine array analysis showed that levels of interleukin (IL)-6, C-X-C motif ligand (CXCL)1, C-C motif ligand (CCL)2, growth-regulated protein (GRO), IL-8, and granulocyte-macrophage colony-stimulating factor (GM-CSF) were increased in conditioned media of co-cultured cells. Among these cytokines increased in conditioned media of co-cultured cells, CCL2 was secreted from TAMs, leading to induction of ERO1-α, MMP-9 upregulation, and invasiveness in MCF10A cells. Our findings elucidated a molecular mechanism underlying the aggressive phenotypic change of non-neoplastic breast cells by co-culture with TAMs, providing useful information for prevention or treatment of recurrent breast cancer.

C-reactive protein (CRP) is an acute phase protein synthesized upon the inflammatory responses, associated with breast cancer. The process of tumor cell invasion and metastasis involves the adherence of cells to the extracellular matrix via integrin as a receptor for matrix molecules. The present study investigated the role of CRP in the adhesive phenotype of breast cells and the underlying mechanisms. Here, we first showed that CRP induces adhesion of MCF10A human breast epithelial cells through the activation of integrin α2 signaling. Expression of integrin α2 was induced by CRP in which transcription factors c-fos and SP1 may be involved. Binding of CRP with integrin α2 leads to the activation of focal adhesion kinase (FAK), paxillin and ERKs. CRP also binds to an Fcγ receptor Fcγ receptor I (FcγRI), and induces activation of paxillin, FAK and ERKs. Integrin α2 and FAK have crucial roles in the adhesive and invasive phenotypes as well as MMP-9 upregulation induced by CRP in MCF10A cells. Treatment with an inflammatory lipid sphingosine-1-phosphate induced CRP, which may be secreted and exert an autocrine effect by binding to FcγRI and integrin α2. Involvement of CRP in adhesion, invasion, anchorage-independent growth and upregulation of integrin α2, paxillin and FAK was observed in MDA-MB-231 triple-negative human breast cancer (TNBC) cells. Using an in vivo invasion model and an orthotopic mouse tumor model with MDA-MB-231 cells, we showed that CRP has an important role in intravasation and tumor growth in vivo, demonstrating the in vivo relevance of our in vitro results. The present study elucidates a critical molecular basis between CRP, integrin α2 and FcγRI pathways in MCF10A breast cells and MDA-MB-231 TNBC cells, thereby providing useful information on CRP-induced aggressiveness of breast cells in the inflammatory microenvironment.

Docetaxel (DTX) is an effective commercial anticancer agent for chemotherapy in non–small cell lung cancer (NSCLC), breast cancer, gastric cancer, and prostate cancer, but its adverse effects including edema, neurotoxicity, and hair loss limit its application. To improve the chemotherapeutic efficacy of DTX and reduce adverse effects, combination therapy is one of the alternative methods. So chrysin, which has various biological activities including anticancer effects, was considered. In vitro, the combination of chrysin and DTX was investigated in A549 cells. Increased cytotoxicity, suppressed cellular proliferation, and induced apoptosis were observed with posttreatment of chrysin following DTX treatment. In vivo, chrysin enhanced the tumor growth delay of DTX and increased DTX-induced apoptosis in the A549-derived xenograft model. Furthermore, chrysin prevented DTX-induced edema in ICR mouse. These results indicated that chrysin strengthened the therapeutic efficacy of DTX and diminished the adverse effect of DTX, suggesting chrysin could be exploited as an adjuvant therapy for NSCLC.

The goal of this study was to identify immunological markers for use in antigen-specific assays that predict long-term survival after renal allograft and distinguish stable-functioning (SP) patients from poorly functioning (PP) patients. For this prospective study, 20 patients were enrolled. Eight SP and six PP patients were enrolled in this study. Serum cytokine/chemokine levels were analyzed by the Luminex multiplex assay. To detect indirect alloreactive T cells, we performed indirect mixed lymphocyte reaction using donor-antigen-pulsed autologous dendritic cells as stimulators. Serum induced protein-10 levels were significantly higher in the serum of PP patients, whereas sCD40L levels were higher in SP patients. The PP patients had significantly higher numbers of donor-specific CD4+CD43highCD45RO+ T cells after indirect allostimulation, whereas this cell population was unchanged in SP patients. The donor-specific CD4+CD43highCD45RO+ T cells had the effector memory T cell phenotype. Prospectively, we studied whether these cells influence graft outcome and found that their strong proliferation in pre-transplant patients is related to a poorly functioning graft. Indirectly allostimulated CD4+CD43highCD45RO+ T cells may not only contribute to chronic allograft nephropathy development but may also have a role in the progression of acute rejection. Thus, these cells may have potential use as immune-monitoring markers in a noninvasive in vitro assay that predicts graft outcome.

Cordyceps extract has been reported to have various pharmacological activities including an anti-cancer effect. We investigated the inhibitory effect of Cordyceps militaris on hepatitis C virus-infected human hepatocarcinoma 7.5 cells (J6/JFH1-huh 7.5 cells). The huh7.5 cells with or without HCV infection were treated with various concentrations of ethanol extract of Cordyceps militaris (CME) for 48 h and the cytotoxicity was measured by CCK-8 assay. Both J6/JFH1-huh7.5 cells and huh7.5 cells were highly susceptible to CME. To examine the molecular mechanisms of the inhibitory effect on huh7.5 cells, the effect of CME on cell apoptosis was measured using flow cytometry and the expressions of p53, Bim, Bax, PARP, (cleaved) caspase-9, and (cleaved) caspase- 3 in huh 7.5 cells were detected by western blot assays. CME significantly increased early apoptosis and up-regulated the expression of Bim, Bax, cleaved PARP, cleaved caspase 9 and cleaved caspase-3. We also found the decrease of HCV Core or NS3 protein by CME in HCV-infected huh 7.5 cells.

A new series of 3-alkylseleno-6-alkylaminopyridazines, 3-alkylseleno-6-alkyliminopyridazines, and 3-alkylseleno-6-dialkylaminopyridazines was synthesized from 3,6-dichloropyridazine for development of new anticancer agents. The process involves the selenylation, Se-alkylation, and aralkylamination (or imination). The alkylamines such as ethylamine and the dialkylamines such as dipropylamine were introduced into the 3-position of the pyridazine ring. These new compounds showed antiproliferative activities against breast cancer (MCF-7) and hepatocellular carcinoma (Hep3B) cells in CCK-8 assays and could be promising candidates for chemotherapy of carcinomas.

Radiotherapy (RT) is one of the major modalities for non‑small cell lung cancer (NSCLC), but its efficacy is often compromised by cellular resistance caused by various mechanisms including the overexpression of epidermal growth factor receptor (EGFR). Although cis‑diamminedichloroplatinum(Ⅱ) (cisplatin, CDDP) has been well characterized as an effective radiosensitizer, its clinical application is limited by its severe nephrotoxic effects. In our current study, we developed a CDDP‑incorporated liposome (LP) conjugated with EGFR antibodies (EGFR:LP‑CDDP) and evaluated its potential to radiosensitize EGFR‑overexpressing cells without exerting nephrotoxic effects. EGFR:LP‑CDDP showed higher cytotoxicity than non‑targeting liposomal CDDP (LP‑CDDP) in the cells expressing EGFR in vitro. In an A549 cell‑derived xenograft tumor mouse model, increased delays in tumor growth were observed in the mice treated with a combination of EGFR:LP‑CDDP and radiation. Notably, the EGFR:LP‑CDDP‑treated animals showed no differences in body weight loss, survival rates of nephrotoxicity compared with untreated control mice. In contrast, the use of CDDP caused lower body weights and poorer survival outcomes accompanied by a significant level of nephrotoxicity [e.g., decreased kidney weight, increased blood urea nitrogen (BUN) and creatinine, and pathological change]. These findings suggest the feasibility of using EGFR:LP‑CDDP to radiosensitize cells in a targeted manner without inducing nephrotoxic effects. This compound may therefore have clinical potential as part of a tailored chemoradiotherapy strategy.

Kyoung Mee Kim,1,2 Hyun Kyung Lim,1,2 Sang Hee Shim,1,2 Joohee Jung1,2 1College of Pharmacy, 2Innovative Drug Center, Duksung Women’s University, Seoul, Republic of Korea Abstract: Chrysin is a flavone that is found in several plants and in honeycomb and possesses various biological activities. However, its low solubility means it has poor bioavailability, which must be resolved to enable its pharmaceutical applications. In the present study, chrysin was incorporated into methoxy poly(ethylene glycol)-β-polycaprolactone nanoparticles (chrysin-NPs) using the oil-in-water technique in order to overcome problems associated with chrysin. The properties of chrysin-NPs were analyzed, and their anticancer effects were investigated in vitro and in vivo. Chrysin-NPs were 77nm sized (as determined by dynamic laser light scattering) and showed a monodisperse distribution. The zeta potential of chrysin-NPs was –2.22mV, and they were spherically shaped by cryo-transmission electron microscopy (cryo-TEM). The loading efficiency of chrysin-NPs was 46.96%. Chrysin-NPs retained the cytotoxicity of chrysin in A549 cells. The therapeutic efficacies of chrysin-NPs were compared with those of chrysin in an A549-derived xenograft mouse model. Chrysin-NPs were intravenously injected at a 10 times lower dosage than chrysin 3 times per week (q2d×3/week). However, free chrysin was orally administrated 5 times per week (q1d×5/week). Chrysin-NP-treated group showed significant tumor growth delay, which was similar to that of chrysin-treated group, despite the considerably lower total dosage. These results suggest that the injectable chrysin-NPs enhance therapeutic efficacy in vivo and offer a beneficial formulation for chemotherapy. Keywords: chrysin, nanoparticle, chemotherapeutic efficacy, non-small-cell lung cancer, in vivo model