Uterine Cervical Cancer Cells Research Articles

CLEFMA induces intrinsic and extrinsic apoptotic pathways through ERK1/2 and p38 signalling in uterine cervical cancer cells.

Although concurrent chemoradiotherapy is the cornerstone of treatment for locally advanced or recurrent uterine cervical cancer, treatment fails at a high rate. Therefore, the development of novel targeting agents is critical. This study investigated the action of CLEFMA, a potent, synthetic curcumin derivative, on cervical cancer cells and its mechanism of action. We found that CLEFMA negatively regulated the viability of cervical cancer cells, involving induction of cell apoptosis. Cleaved caspase-3, cleaved poly(adenosine diphosphate-ribose) polymerase, cleaved caspase-8, and cleaved caspase-9 expression were increased by treatment with CLEFMA. After U0126 (ERK1/2 inhibitor) and SB203580 (p38 inhibitor) were applied as cotreatment with CLEFMA, the expression of cleaved caspase-8, -9, and -3 was reduced significantly. In conclusion, CLEFMA activates both extrinsic and intrinsic apoptotic pathways through ERK1/2 and p38 signal transduction in cervical cancer cells.

Combination treatment with cyclosporin A and arsenic trioxide induce synergistic cell death via non-apoptotic pathway in uterine cervical cancer cells

Cyclosporin A is an immunosuppressive drug with anti-cancer effect. Arsenic trioxide (As2O3), a well-known cancer-inhibiting drug, induced cytotoxicity via apoptosis and autophagy. The aim of this study is to evaluate the effect of combinational treatment with cyclosporin A and arsenic trioxide on cell viability inhibition in cervical cancer cells. Using MTT assay and combination index, combinational treatment with cyclosporin A and arsenic trioxide induced a synergistic cytotoxic effect in Caski and SiHa cells. Cyclosporin A and arsenic trioxide triggered cell death via non-apoptotic pathway by using annexin V/propidium iodide (PI) assay. Cyclosporin A and arsenic trioxide combined treatment decreased mitochondrial membrane potential and increase reactive oxygen species (ROS) generation. This co-treatment increased LC3B-II expression and autophagosome formation in cervical cancer cells. This study first demonstrated that combinational treatment with cyclosporin A and As2O3 trigger synergistic cytotoxic effect via autophagy in cervical cancer cells.

Fyn expression is associated with the response of patients with locally advanced uterine cervical squamous cell carcinoma to neoadjuvant chemotherapy.

Neoadjuvant chemotherapy (NAC) followed by surgery is the current standard of treatment for locally advanced uterine cervical cancer; however, its value and outcomes remain contested. Identifying biomarkers that allow for the prediction of the effect of NAC efficacy before initiation of treatment is essential, in order to assist in choosing the optimum therapeutic regimen to maximize the beneficial outcomes of treatment. In the present retrospective study, 44 patients with locally advanced uterine cervical squamous cell carcinoma who underwent NAC were divided into two groups: A NAC successful group and a NAC failure group, depending on the efficacy of NAC. Subsequently, the association between Fyn expression, a non-receptor tyrosine kinase that is a member of the Src family kinases, and NAC efficacy was determined; Fyn expression was detected by immunohistochemistry and assessed using a weighted scoring method. Additionally, the effect of Fyn knockdown on the sensitivity of a uterine cervical cancer cell line to cisplatin was determined. Notably, there were no significant differences between the two groups of patients regarding their characteristics. Regarding overall survival, the NAC successful group had a significantly longer survival time than the NAC failure group (P=0.01). Furthermore, the expression levels of Fyn in tumor tissues were significantly lower in the NAC successful group compared with those in the NAC failure group (P=0.003). The patients were subsequently divided into two groups (high expression group and low expression group) according to a cutoff value of 3, which was determined by producing a receiver operating characteristic curve from the weighted scores. The low expression group was significantly more sensitive to NAC than the high expression group (P<0.001). In vitro experiments revealed that Fyn knockdown significantly enhanced the sensitivity of uterine cancer cells to cisplatin (P<0.05). In conclusion, Fyn expression may be a potentially useful biomarker for predicting the response to NAC in patients with locally advanced uterine cervical squamous cell carcinoma, and may also be a promising molecular target for the management of uterine cancer.

Silencing of Fused Toes Homolog (FTS) Increases Radiosensitivity to Carbon-Ion Through Downregulation of Notch Signaling in Cervical Cancer Cells.

The effects of Carbon ion radiation (C-ion) alone or in combination with fused toes homolog (FTS) silencing on Notch signaling were investigated in uterine cervical cancer cell lines (ME180 and CaSki). In both cell lines, upon irradiation with C-ion, the expression of Notch signaling molecules (Notch1, 2, 3 and cleaved Notch1), γ-secretase complex molecules and FTS was upregulated dose-dependently (1, 2 and 4 Gy) except Notch1 in ME180 cells where the change in expression was not significant. However, overexpression of these molecules was attenuated upon silencing of FTS. The spheroid formation, expression of stem cell markers (OCT4A, Sox2 and Nanog) and clonogenic cell survival were reduced by the combination as compared to FTS silencing or C-ion irradiation alone. Additionally, immunoprecipitation and immunofluorescence assay revealed interaction and co-localization of FTS with Notch signaling molecules. In conclusion, FTS silencing enhances the radio-sensitivity of the cervical cancer cells to C-ion by downregulating Notch signaling molecules and decreasing the survival of cancer stem cells.

PTEN downregulation induces apoptosis and cell cycle arrest in uterine cervical cancer cells.

The tumor suppressors PTEN and p53 are often downregulated in various human cancer types, which has been associated with a poor prognosis. Recent evidence implies that PTEN downregulation may induce growth arrest of kidney cells and cancer cells. In the present study, the role of PTEN in the proliferation and survival of cervical cancer cells was investigated. It was found that PTEN silencing promoted apoptosis and cell-cycle arrest, accompanied by a significant decrease in the proportion of cells in the S1 phase of the cell cycle. Moreover, PTEN silencing in cervical cancer cells increased levels of p53, p27, p21, phospho-ERK and cleaved caspase-3, and decreased levels of cyclin A2 and cyclin D1. Furthermore, PTEN knockdown significantly impacted the viability of cervical cancer cells. P53 silencing did not affect the ability of PTEN knockdown to induce apoptosis in cervical cancer cells. Taken together, the present study results imply that PTEN silencing induces apoptosis and decreases proliferation in cervical cancer cells; hence, PTEN inhibition may represent a promising strategy for the treatment of cervical cancer.

MicroRNA-101 Promotes the Proliferation, Migration, and Invasion of Uterine Cervix Cancer Cells by Targeting Cyclin-Dependent Kinase 8

Uterine cervical cancer (UCC), or carcinoma of the uterine cervix, is a familiar malignancy in gynecology. The expression of multiple microRNAs is abnormal in UCC. Research has demonstrated that the level of microRNA-101 (miR-101) was decreased in UCC, but the mechanism by which miR-101 regulates UCC is still unclear. The TargetScan software predicted that one of the target genes of miR-101 was CDK8. This study aims to explore whether miR-101 affects the migration, invasion, and proliferation of UCC cells through CDK8. First, nanoparticle-assisted PCR was used to determine the levels of miR-101 and CDK8 in UCC tissues, normal adjacent tissues, and two UCC cell lines, C-33A and Siha. Compared to the normal tissues, the level of miR-101 was decreased in UCC tissues, while the level of CDK8 was increased. The dual-luciferase reporter experiments confirmed that miR-101 directly interacted with the binding site in CDK8 3’UTR to regulate luciferase activity. The UCC cells were transfected with the MIR101 mimic construct to overexpress miR-101. It was demonstrated that cell migration, invasion, and proliferation was reduced in C-33A cells overexpressing miR-101. In the cells co-transfected with CDK8 and MIR101, the overexpression of CDK8 reversed the effect of MIR101 overexpression on cell migration, invasion, and proliferation. Therefore, miR-101 can regulate the migration, invasion, and proliferation of UCC cells by targeting CDK8. Therefore, miR-101 has potential applications in the treatment of UCC.

Differences in Linear Energy Transfer Affect Cell-killing and Radiosensitizing Effects of Spread-out Carbon-ion Beams.

The cell-killing and radiosensitizing effects of carbon-ion (C-ion) beams with low linear energy transfer (LET) are underexplored. We aimed to demonstrate the cell-killing effects of 60Co gamma rays and C-ion beams at various LET values and the radiosensitizing effect of C-ion beams at various LET and cisplatin levels. Human uterine cervical cancer cells were irradiated with 60Co gamma rays and C-ion beams at different levels of LET, with and without cisplatin treatment. Low-LET C-ion beams had a superior cell-killing effect compared to 60Co gamma rays. Survival curves under low-LET C-ion beams were more similar to that of 60Co gamma rays than that of high-LET C-ion beams. Cisplatin significantly reduced cell survival after 1, 2, and 3 Gy C-ion beam irradiations at LET values of 13/30/70 keV/μm, 13/30 keV/μm, and 13 keV/μm, respectively. Low-LET C-ion beams combined with cisplatin have higher radiosensitizing effects than high-LET C-ion beams.

Regio-specific biotransformation of alizarin to alizarin methoxide with enhanced cytotoxicity against proliferative cells.

Alizarin has been reported to have an antigenotoxic activity along with an inhibitory effect on the tumor cell growth of human colon carcinoma cells. Alizarin was biotransformed into an O-methoxide derivative using O-methyltransferase from Streptomyces avermitilis MA4680 (SaOMT2) to enhance its bioefficacy. The biotransformed product was extracted, purified, and characterized using various chromatographic and spectroscopic analyses, and confirmed to be an alizarin 2-O-methoxide. The antiproliferative activity of the compound against gastric cancer cells (AGS), uterine cervical cancer (Hela), liver cancer (HepG2), and normal cell lines was investigated. Alizarin 2-O-methoxide showed an inhibitory effect on all three cancer-cell lines at very low concentrations, from 0.078µM, with no cytotoxicity against 267B1 (human prostate epithelial) and MRC-5 (normal human fetal lung fibroblast).

Cell-type-specific Modulation of Non-homologous End Joining of Gamma Ray-induced DNA Double-strand Breaks by cAMP Signaling in Human Cancer Cells.

BackgroundCyclic AMP (cAMP) signaling is activated by various hormones and neurotransmitters and regulates numerous physiological phenomena, including energy metabolism, gene expression, and proliferation. cAMP signaling plays a role in the repair of DNA damage, but its specific function is inconsistent in the literature. The present study aimed to investigate the mechanism of the different roles of cAMP signaling in DNA repair by analyzing the cell-type differences in the modulation of DNA repair by cAMP signaling following γ-ray irradiation.MethodscAMP signaling was activated in human malignant melanoma cells (SK-MEL-2 and SK-MEL-28), human uterine cervical cancer cells (HeLa and SiHa) and human non-small cell lung cancer cells (H1299 and A549) by expressing a constitutively active mutant of the long-form stimulatory α subunit of GTP-binding protein or by treating with isoproterenol and prostaglandin E2 before γ-ray irradiation. DNA damage was quantitated by western blot analysis of γ-H2AX, and non-homologous end joining (NHEJ) was assessed by fluorescent reporter plasmid repair assay and immunofluorescence of microscopic foci of XRCC4 and DNA-ligase IV.ResultscAMP signaling modulated DNA damage, apoptosis and the NHEJ repair following γ-ray irradiation differently depending upon the cell type. cAMP signaling regulated the phosphorylation of DNA-dependent protein kinase catalytic subunit (DNA-PKcs) at Ser2056 and Thr2609 in cell-type-specific manners following γ-ray irradiation, an activity that was mediated by protein kinase A.ConclusioncAMP signaling modulates the NHEJ repair of γ-ray-induced DNA damage in melanoma cells, uterine cervical cancer cells and lung cancer cells in a cell-type-specific manner, and the modulation is likely mediated by protein kinase A-dependent phosphorylation of DNA-PKcs. This study suggests that cell- and tissue-specific modulation of DNA damage repair by cAMP signaling may contribute to improve the therapeutic efficiency of radiation therapy.

The Application of Arsenic Trioxide in Ameliorating ABT-737 Target Therapy on Uterine Cervical Cancer Cells through Unique Pathways in Cell Death.

ABT-737, a B cell lymphoma-2 (Bcl-2) family inhibitor, activates apoptosis in cancer cells. Arsenic trioxide is an apoptosis activator that impairs cancer cell survival. The aim of this study was to evaluate the effect of a combination treatment with ABT-737 and arsenic trioxide on uterine cervical cancer cells. MTT (3-(4,5-dimethylthiazol-2-yl)-25-diphenyltetrazolium bromide) assay revealed that ABT-737 and arsenic trioxide induced a synergistic effect on uterine cervical cancer cells. Arsenic trioxide enhanced ABT-737-induced apoptosis and caspase-7 activation and the ABT-737-mediated reduction of anti-apoptotic protein Mcl-1 in Caski cells. Western blot assay revealed that arsenic trioxide promoted the ABT-737-mediated reduction of CDK6 and thymidylate synthetase in Caski cells. Arsenic trioxide promoted ABT-737-inhibited mitochondrial membrane potential and ABT-737-inhibited ANT expression in Caski cells. However, ABT-737-elicited reactive oxygen species were not enhanced by arsenic trioxide. The combined treatment induced an anti-apoptosis autophagy in SiHa cells. This study is the first to demonstrate that a combination treatment with ABT-737 and arsenic trioxide induces a synergistic effect on uterine cervical cancer cells through apoptosis. Our findings provide new insights into uterine cervical cancer treatment.

CXCL3 overexpression promotes the tumorigenic potential of uterine cervical cancer cells via the MAPK/ERK pathway.

CXCL3 belongs to the CXC-type chemokine family and is known to play a multifaceted role in various human malignancies. While its clinical significance and mechanisms of action in uterine cervical cancer (UCC) remain unclear. This investigation demonstrated that the UCC cell line HeLa expressed CXCL3, and strong expression of CXCL3 was detected in UCC tissues relative to nontumor tissues. In addition, CXCL3 expression was strongly correlated with CXCL5 expression in UCC tissues. In vitro, HeLa cells overexpressing CXCL3, HeLa cells treated with exogenous CXCL3 or treated with conditioned medium from WPMY cells overexpressing CXCL3, exhibited enhanced proliferation and migration activities. In agreement with these findings, CXCL3 overexpression was also associated with the generation of HeLa cell tumor xenografts in athymic nude mice. Subsequent mechanistic studies demonstrated that CXCL3 overexpressing influenced the expression of extracellular signal-regulated kinase (ERK) signaling pathway associated genes, including ERK1/2, Bcl-2, and Bax, whereas the CXCL3-induced proliferation and migration effects were attenuated by exogenous administration of the ERK1/2 blocker PD98059. The data of the current investigation support that CXCL3 appears to hold promise as a potential tumor marker and interference target for UCC.

Effects of ABT-737 combined with irradiation treatment on uterine cervical cancer cells.

The aim of the present study was to examine the role of ABT-737, an inhibitor of B-cell lymphoma 2 (Bcl-2), in enhancing the effect of irradiation on uterine cervical cancer. Based on The Cancer Genomic Atlas (TCGA), Bcl-2 mRNA expression was associated with the Tumor-Node-Metastasis stage of cervical cancer. Therefore, it was hypothesized that Bcl-2 inhibition may decrease the progression of cervical cancer. ABT-737 was added to irradiation treatment to evaluate its effectiveness in inhibiting cancer cell progression. SiHa and CaSki cervical cancer cells were selected for in vitro assays. Patients with advanced stage III uterine cancer had slightly increased mRNA expression levels of Bcl-2 compared with patients with stage I cancer, although the difference was not significant. ABT-737 and radiation administration induced a synergistic cytotoxic effect based on the MTT assay and flow cytometry results, where an increase in apoptosis was observed. The apoptotic percentages were significantly increased in the cells treated with a combination of ABT-737 and irradiation. Loss of mitochondrial membrane potential and gain of reactive oxygen species (ROS) were detected by flow cytometry in CaSki and SiHa cells treated with ABT-737 and radiation. Additionally, the protein expression levels of the cleaved forms of poly ADP ribose polymerase and caspase-7 were increased following the combined treatment. In conclusion, ABT-737 and irradiation may induce apoptosis via loss of mitochondrial membrane potential and a ROS-dependent apoptotic pathway in CaSki and SiHa cells. The present study indicates that ABT-737 may be a potential irradiation adjuvant when treating cervical cancer.

DNA binding interaction studies of flavonoid complexes of Cu(II) and Fe(II) and determination of their chemotherapeutic potential

The present study is based on molecular level investigation of DNA binding of three flavonoids (Fls), morin (mor), quercetin (quer) and primuletin (prim) and their metal (Cu (II) and Fe (III)) complexes through UV–Vis spectroscopy, fluorescence spectroscopy, thermal analysis and cyclic voltammetry (CV). The spectroscopic experiments were performed by titrating compounds with DNA and the results were in good agreement with those obtained from titration of DNA by the compounds. The Fe-mor, Cu-quer, Fe-quer and prim which exhibited hypsochromic/hyperchromic shifts in UV–Vis spectra, showed negligible effects over florescent spectra of NR-DNA (intercalating Neil red dye–DNA complex), intact Tm (thermal melting temperature) of DNA and the shift of peak potential (Ep) towards less positive value were designated to exhibit electrostatic mode of binding with dsDNA having binding constant, Kb ≈ 102 M−1. The mor, Cu-mor, Cu-prim and Fe-prim showed intercalating mode of interaction with dsDNA which was recognized from hypochromic/bathochromic shifts in the spectra, Ep shifts towards higher potential in CV, significant decay in fluorescence intensity, distinct shift in Tm of DNA and binding constant (Kb) of the order of 104 M−1. The mor exhibited mixed binding mode (intercalation and groove binding) at different concentrations. The quer displayed increase in peak current and hyperchromism thus depicted groove binding into dsDNA (Kb ≈ 103 M−1). The binding modes were found to be correlated with the apoptotic activity of the Fls and M-Fls (flavonoids and metal-flavonoids) assayed against human breast cancer cell lines (MCF-7) and human uterine cervical cancer cell lines (HeLa). The correlation is interpreted in terms of, the intercalation mode as cytostatic activity and the groove binding with no apoptosis. However, electrostatic binding of (M-)Fls to dsDNA did not produce significant anti-proliferation activity against Hela cell lines with exception of Fe-mor (cytostatic activity) and Cu-quer (cytotoxicity) against MCF-7 cell line.

Prostaglandin E2 produced by myeloid-derived suppressive cells induces cancer stem cells in uterine cervical cancer.

Myeloid-derived suppressor cells (MDSCs) enhance tumor progression by suppressing tumor-specific T cell responses, stimulating tumor angiogenesis, or promoting tumor cell metastasis. However, the biology of MDSCs have not been fully investigated. In the current study, we investigated the role of MDSCs in inducing cancer stem-like cells and explored a clinically feasible approach for targeting MDSCs-mediated cancer stem-like cells induction. In vitro and in vivo experiments revealed that MDSCs induced by tumor-derived G-CSF enhanced the stemness of cervical cancer cells by producing Prostaglandin E2 (PGE2). We also demonstrated that anti-Gr-1 neutralizing antibody or celecoxib inhibited the induction of cancer stem-like cells and enhanced the efficacy of cisplatin in cervical cancer. In clinical samples, MDSCs, PGE2, and CSCs had positive correlations. In conclusion, G-CSF-induced MDSCs enhance the stemness of uterine cervical cancer cells by producing PGE2. Targeting MDSCs or PGE2 might be a reasonable strategy for enhancing the efficacies of treatments.

Functional analysis of protein disulfide isomeraseP5 in glioblastoma cells as a novel anticancer target.

P5, which is a member of the protein disulfide isomerase family, possesses isomerase and chaperone activity invitro; however, the physiological functions of this enzyme in cells remain unclear. To understand the important roles of P5 in cancer cells, the present study examined its expression on the surface of normal and cancer cell lines by flow cytometry using an affinity‑purified anti‑P5 antibody labeled with 6‑(fluorescein‑5‑carboxamido) hexanoic acid succinimidyl ester. P5 expression was increased on the surface of various cancer cell lines, including leukemia cells, and glioblastoma, breast, colon, ovarian and uterine cervical cancer cells, compared with normal cells. However, P5 was constantly expressed within both normal and cancer cell lysates, and its total expression levels were not significantly different between the cells. P5 knockdown in glioblastoma cells by small interfering RNA affected Bip promoter activation during cancer cell growth, and significantly inhibited cancer cell growth and migration. Immunoprecipitation using an anti‑P5 antibody in cancer and normal cells demonstrated that vimentin was bound to P5, predominantly in U251 glioblastoma cells. P5 knockdown in glioblastoma cells did not affect the protein expression levels of vimentin; however, it did affect the expression of numerous epithelial‑mesenchymal transition markers, including Snail and Slug. These results suggested that P5 may serve an important role in cancer cell growth, and may be considered an attractive and potent target for the treatment of glioblastoma.

Effects of 60kDa prolactin and estradiol on metabolism and cell survival in cervical cancer: Co‑expression of their hormonal receptors during cancer progression.

Estrogens and estrogen receptors (ERs), such as ERα and ERβ, prolactin (PRL) and prolactin receptor (PRLR) have been reported to be involved in the physiopathology of uterine cervical cancer (UCC). The 60kDa PRL is an isoform of PRL, which is produced by UCC‑derived cells. The present study aimed to evaluate the expression of hormonal receptors in different degrees of cervical lesions, and to determine whether 60kDa PRL and 17β‑estradiol (E2) modulated cell survival and metabolism in UCC cells, and in HaCaT cells transduced with human papillomavirus (HPV)16 and18E6/E7 oncogenes. ERα, ERβ, PRLR, Ki67 and B‑cell lymphoma2 expression levels were analyzed in biopsies of precursor lesions and UCC using immunohistochemistry. In addition, HeLa, SiHa and C33A cells, and transduced HaCaT cells, were stimulated with 60kDa PRL, E2 or a combination of both. Proliferation was evaluated using the xCELLigence platform, apoptosis was analyzed by flow cytometry and cell metabolism was determined using the MTT assay. The results revealed that ERα, ERβ, PRLR and Ki67 expression levels were increased during the progression of cancer. Invitro, 60kDa PRL alone significantly increased proliferation of SiHa cells. Furthermore, E2 alone or in combination with 60kDa PRL increased the sensitivity of SiHa cells to cisplatin and increased the percentage of apoptosis; in HaCaT cells, these treatment strategies had the opposite effect on cisplatin sensitivity. Treatment with E2 increased mitochondrial activity in HeLa and SiHa cells, and in HaCaT cells transduced with HPV16E6/E7 and HPV18E6 oncogenes. PRL had a similar effect on HeLa cells, and on HaCaT cells transduced with HPV18E6 and HPV16E7. The co‑expression of these receptors demonstrated the hormonal dependence of UCC. In addition, E2 and the 60kDa PRL significantly impacted the metabolism, but not the survival, of cells.

Annexin A5 overexpression might suppress proliferation and metastasis of human uterine cervical carcinoma cells.

Annexin A5 (ANXA5) is a kind of Ca2+-dependent phospholipid binding protein which is involved in cell membrane dynamics and organization. Recent data showed that ANXA5 might involve in tumorigenesis. To explore what role ANXA5 play in human uterine cervical carcinoma. In this study, a recombined ANXA5 plasmid was constructed and uterine cervical carcinoma cell lines HeLa and SiHa were transfected with it. After ANXA5 overexpression was determined by Western Blot, cell proliferation test was detected by MTT assay and colony formation assay respectively. FACS assay and Hochest33258 staining methods were employed to detect cell apoptosis. To further investigate whether ANXA5 influence cell migration and invasion, wound healing assay and transwell assay were applied. At the same time, the relative mechanism was investigated. When ANXA5 expression increased, cell proliferation was inhibited by regulating the expression of bcl-2 and bax while cell metastasis was suppressed by regulating E-cadherin and MMP-9 expression. ANXA5 overexpression in the uterine cervical carcinoma might play important roles in cell proliferation and metastasis of uterine cervical cancer cells and act as an anti-cancer gene in uterine cervical cancer.

Sensitive molecular detection of small nodal metastasis in uterine cervical cancer using HPV16-E6/CK19/MUC1 cancer biomarkers.

Metastatic nodal involvement is a critical prognostic factor in uterine cervical cancer (UCC). To improve current methods of detecting UCC metastases in lymph nodes (LNs), we used quantitative PCR (qPCR) to assess mRNA expression of potential metastatic biomarkers. We found that expression of HPV16-E6, cytokeratin19 (CK19), and mucin1 (MUC1) is consistently upregulated in tumors and metastatic tissues, supporting a role for these genes in UCC progression. These putative biomarkers were able to predict the presence of histologically positive metastatic LNs with respective sensitivities and specificities of 82% and 99% (CK19), 76% and 95% (HPV16-E6), and 76% and 78% (MUC1). While the biomarkers failed to detect 1.7% to 2.2% of the histologically positive LNs when used individually, combining CK19 and HPV16-E6 enhanced sensitivity and specificity to 100% and 94%, respectively. To explore the sensitivity of qPCR-based detection of varying proportions of invading HPV16-positive UCC cells, we designed a LN metastasis model that achieved a fresh cell detection limit of 0.008% (1:12500 HPV16-positive to HPV16-negative cells), and a paraffin-embedded, formalin-fixed (PEFF) detection limit of 0.02% (1:5000 HPV16-positive to HPV16-negative cells), both of which are within the theoretical detection limit for micrometastasis. Thus, HPV E6/E7 oncogenes may be useful targets for the ultrasensitive detection of nodal involvements like micrometastases in fresh or archived tissue samples. Moreover, our results suggest that the biomarker combination of CK19/HPV-E6 could support a real-time intraoperative strategy for the detection of small, but potentially lethal, metastatic nodal involvements in fresh UCC tissues.

XPA expression is a predictive marker of the effectiveness of neoadjuvant chemotherapy for locally advanced uterine cervical cancer.

The standard treatment for locally advanced uterine cervical cancer is concurrent chemoradiotherapy. Successful neoadjuvant chemotherapy (NAC) may reduce tumor size and facilitate a hysterectomy, thereby improving the prognosis for patients with locally advanced cervical cancer. In contrast, unsuccessful NAC may worsen the prognosis because if a hysterectomy is not possible, the change in treatment plan may delay the initiation of core treatment. Therefore, there is a need to identify biomarkers that predict the efficacy of NAC in patients with uterine cervical cancer. The xeroderma pigmentosum complementation group A (XPA) protein serves a major role in nucleotide excision repair, which is a key DNA damage response pathway involved in cisplatin resistance. In the present study, the association between XPA expression in tumor tissue and the efficacy of NAC for locally advanced uterine cervical cancer was investigated. Data from 56 patients aged <70 years with locally advanced uterine cervical cancer (FIGO stages IIIA or IIIB) who were classified into two groups based on effective (n=31) and ineffective (n=25) responses to NAC treatment was evaluated. Tumor tissue samples were obtained by punch biopsy prior to NAC and XPA expression was examined immunohistochemically and scored using a weighted scoring system. In addition, the effects of RNA interference-mediated downregulation of XPA on the cisplatin sensitivity of uterine cervical cancer cells was investigated in vitro. It was revealed that the NAC effective group had significantly lower weighted XPA scores than the NAC ineffective group (P=0.001). Similarly, low tumor expression of XPA was significantly associated with higher sensitivity to NAC (P=0.001). Additionally, the downregulation of XPA expression in cervical cancer cells significantly increased their sensitivity to cisplatin in vitro. The results of the present study suggest that low XPA expression may be a predictive biomarker of NAC efficacy for patients with locally advanced uterine cervical cancer, which may be helpful for improving their prognosis.

Abstract 529: The significance of activated PI3K/AKT pathway in FGFR3-TACC3 fusion positive cervical cancer

Abstract Uterine cervical cancer is one of the most common cancer in women worldwide, and the prognosis in advanced or recurrent cases remains poor because of no effective molecular target therapies for this disease. The aim of our study is to identify and validate therapeutically targetable gene fusions in uterine cervical cancer, leading to the development of new therapeutic strategies. We have analyzed RNA sequencing data of 253 TCGA cervical cancer samples to search for gene fusions by using PRADA algorithm, and validated recurrent fusions in our Japanese dataset (n = 100) by RT-PCR and sanger sequencing. In TCGA samples, we detected 358 fusion transcripts and extracted 3 kinds of recurrent in-frame fusion transcripts (FGFR3-TACC3, ARL8B-ITPR1 and NXN-ABR). We focused on FGFR3-TACC3 fusion as a candidate of therapeutic kinase fusion and identified FGFR3-TACC3 fusion in 2 of 253 (0.8%) TCGA samples and 2 of 100 (2%) Japanese samples. All of fusion positive samples, which were histologically diagnosed as squamous cell carcinoma, showed high expression of both FGFR3 and TACC3 genes. To evaluate the possibility as therapeutically targetable fusions in uterine cervical cancer, we transfected FGFR3-TACC3, wild type FGFR3 and TACC3 transcripts to normal immortalized cervical keratinocytes derived from ectocervix (Ect1/E6E7) and four uterine cervical cancer cell lines (SiHa, ME180, HeLa, CaSki). Continuous expression of FGFR3-TACC3 fusion transcript led to anchorage-independent growth in Ect1/E6E7 and the dramatically alteration of cell proliferation in all cancer cell lines. On the other hand, no obvious phenotype change was observed in FGFR3 or TACC3 transfected cells. Western blotting analysis demonstrated that MAPK pathway was activated in all the fusion transfected cell lines but PI3K/AKT pathway was activated only in ME180 and CaSki harboring PIK3CA mutation. Fusion transfected cell lines exhibited high sensitivity to fusion specific siRNA and FGFR inhibitor compared to control. Although PI3K/AKT activated cell lines (ME180 and CaSki) showed less effective against FGFR inhibitor compared to PI3K/AKT neutral cell lines (SiHa and HeLa), combined inhibition of FGFR and AKT had a synergic effect in PI3K/AKT activated cell lines. Our findings suggest FGFR3-TACC3 fusion gene is an oncogenic driver event and therapeutic target in a fraction of cervical cancer. When FGFR3-TACC3 fusion positive cervical cancer is treated by FGFR inhibitor, genomic background such as PI3K/AKT status should be considered. Citation Format: Ryo Tamura, Kosuke Yoshihara, Tetsuya Saito, Ryosuke Ishimura, Emmanuel Martínez-Ledesma, Yutaro Mori, Kaoru Yamawaki, Kazuaki Suda, Tatsuya Ishiguro, Yoichi Aoki, Seiya Sato, Hiroaki Itamochi, Masaaki Komatsu, Roeland Verhaak, Takayuki Enomoto. The significance of activated PI3K/AKT pathway in FGFR3-TACC3 fusion positive cervical cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 529. doi:10.1158/1538-7445.AM2017-529