CA9 Expression Research Articles

Matching treatment strategies to clinical phenotype: Biomarker-driven selection for neoadjuvant therapy in pancreatic cancer.

Introduction: Up to 30% of patients who undergo pancreatic ductal adenocarcinoma (PDAC) resection face mortality within one year of surgery. We have previously demonstrated that a triple biomarker panel comprising S100A4, Ca-125 and mesothelin predicts median overall survival (mOS) after resection. We aimed to evaluate the association between this triple biomarker panel and transcriptomic PDAC subtypes, clinical PDAC phenotypes, and level of survival benefit received from a neoadjuvant treatment strategy. Method: Patients who underwent resection for resectable and borderline resectable PDAC were included for analysis. PDAC tumours for all patients were immunohistochemically evaluated for expression of S100A4, Ca-125 and mesothelin. Data regarding Bailey transcriptomic classification (progenitor, ADEX, immunogenic or squamous) was obtained from a subset of these patients whose tumour was included in the International Cancer Genomic Consortium (ICGC). Result: 226 patients from the ICGC cohort and 252 patients from a single institution were included for analysis. Triple negative biomarker status was associated with non-squamous PDAC subtypes (OR 5.804, p=0.020). Triple positive biomarker status was characterized by a higher risk of distant disease recurrence compared with triple negative disease (HR 2.136, p=0.002). A neoadjuvant treatment approach was associated with longer mOS compared with upfront surgery in triple biomarker positive patients with resectable and borderline resectable PDAC (29.5 vs 13.7 mths, p=0.021). On the other hand, a neoadjuvant treatment approach was associated with shorter mOS compared with upfront surgery in triple biomarker negative patients with resectable PDAC (22.2 vs. 33.3 mths, p=0.038). Conclusion: Patients with triple positive disease, with a higher risk of distant metastatic recurrence, achieve longer overall survival with a neoadjuvant approach compared with upfront surgery. On the other hand, patients with triple negative disease are likely to have non-squamous subtype PDAC, and achieve longer overall survival with an upfront surgical resection compared with a neoadjuvant treatment approach.

RMP promotes the proliferation and radioresistance of esophageal carcinoma.

RMP is a RNA polymerase II Subunit RPB-5 associated protein shown to act as an oncogene in several cancer. However, the mechanism of the involvement of RMP in esophageal cancer (EC) remains unclear. We analyzed RMP expression in EC cell lines and EC tissues. The connection between RMP and clinical pathological features of EC was also elucidated. To investigate the role of RMP in EC, We performed CCK-8 assay to evaluate cell proliferation, and Annexin V/PI double-staining to evaluate cell apoptosis. Effect of RMP on tumor progression in nude mouse models was assessed by measurement of volume and weight of tumors. Expression of RMP, CEA and CA199 in vivo were measured by Inmunohistochemical staining. First of all, our study showed that RMP was highly expressed in EC cell lines (compared with normal cells) and tumor tissues (compare with corresponding normal tissues). Then, we found that RMP was bound up with the status of nodal and T stage which indicating that RMP may be related to the growth and malignant degree of EC. Moreover upregulation of RMP could contribute to tumor growth in vitro and vivo. In addition, the results also showed that overexpression of RMP could significantly reduce the susceptibility to radiotherapy. Taken together, all these further suggested that RMP would play a chance-promoting in EC which may provide us a powerful goal for gene targeting treatment of esophageal cancer.

Clinical Significance of Serum Galectin-1 and Its Tissue Immunohistochemical Expression in Serous Ovarian Carcinoma Patients

Objectives: Serous ovarian carcinoma (SOC) is the commonest ovarian carcinoma type with poor prognosis due to early metastasis and first presentation with advanced stage. In this work, we investigated serum level of Galactin-1 (Gal-1) and its tissue immunohistochemical expression in SOC patients at different stages trying to find out its significance as a diagnostic and prognostic marker. Patients and methods: The study included 95 females I-Control group: Twenty five healthy females; II-Patients group: Seventy females diagnosed as SOC at different stages; Stage I: 8 cases, Stage II: 12 cases, Stage III: 32 cases and Stage VI:18 cases. Serum Galectin-1 and CA-125 were measured by ELIZA and tissue Galectin-1 was assessed by immunohistochemistry. All patients were followed for up to 3 years after surgery. Results: Serum Gal-1 and CA-125 levels were significantly higehr in SOC patients compared to controls (p 0.001). We found a direct positive statistically significant correlation between serum Gal-1 and CA125 levels (p 0.001). Serum Gal-1 at cut off value > 135 ng/ml was superior to CA-125 a cut off value > 49 u/ml with sensitivity, specificity of 100%, vs 88.57, 96% for CA-125. Serum Gal-1 was significantly associated with tumor stage (p 0.001). Immunohistochemistry showed that patients with strong Gal-1 expression had higher serum level (p = 0.002). Stromal and tumor Gal-1 expression were significantly correlated with tumor grade (p 0.001) and stage (p = 0.001). Serum Gal-1, CA-125 and IHC Gal-1 expression were associated with poor survival (p 0.001, p = 0.009 and p = 0.002) respectively. Conclusion: Serum Gal-1 and its tissue IHC expression are useful diagnostic and prognostic markers for SOC patients.

Suppressive effects of iron chelation in clear cell renal cell carcinoma and their dependency on VHL inactivation

Increasing data implicate iron accumulation in tumorigenesis of the kidney, particularly the clear cell renal cell carcinoma (ccRCC) subtype. The von Hippel Lindau (VHL)/hypoxia inducible factor-α (HIF-α) axis is uniquely dysregulated in ccRCC and is a major regulator and regulatory target of iron metabolism, yet the role of iron in ccRCC tumorigenesis and its potential interplay with VHL inactivation remains unclear. We investigated whether ccRCC iron accumulation occurs due to increased cell dependency on iron for growth and survival as a result of VHL inactivation. Free iron levels were compared between four VHL-mutant ccRCC cell lines (786-0, A704, 769-P, RCC4) and two benign renal tubule epithelial cell lines (RPTEC, HRCEp) using the Phen Green SK fluorescent iron stain. Intracellular iron deprivation was achieved using two clinical iron chelator drugs, deferasirox (DFX) and deferoxamine (DFO), and chelator effects were measured on cell line growth, cell cycle phase, apoptosis, HIF-1α and HIF-2α protein levels and HIF-α transcriptional activity based on expression of target genes CA9, OCT4/POU5F1 and PDGFβ/PDGFB. Similar assays were performed in VHL-mutant ccRCC cells with and without ectopic wild-type VHL expression. Baseline free iron levels were significantly higher in ccRCC cell lines than benign renal cell lines. DFX depleted cellular free iron more rapidly than DFO and led to greater growth suppression of ccRCC cell lines (>90% at ~30–150 µM) than benign renal cell lines (~10–50% at up to 250 µM). Similar growth responses were observed using DFO, with the exception that a prolonged treatment duration was necessary to deplete cellular iron adequately for differential growth suppression of the less susceptible A704 ccRCC cell line relative to benign renal cell lines. Apoptosis and G1-phase cell cycle arrest were identified as potential mechanisms of chelator growth suppression based on their induction in ccRCC cell lines but not benign renal cell lines. Iron chelation in ccRCC cells but not benign renal cells suppressed HIF-1α and HIF-2α protein levels and transcriptional activity, and the degree and timing of HIF-2α suppression correlated with the onset of apoptosis. Restoration of wild-type VHL function in ccRCC cells was sufficient to prevent chelator-induced apoptosis and G1 cell cycle arrest, indicating that ccRCC susceptibility to iron deprivation is VHL inactivation-dependent. In conclusion, ccRCC cells are characterized by high free iron levels and a cancer-specific dependency on iron for HIF-α overexpression, cell cycle progression and apoptotic escape. This iron dependency is introduced by VHL inactivation, revealing a novel interplay between VHL/HIF-α dysregulation and ccRCC iron metabolism. Future study is warranted to determine if iron deprivation using chelator drugs provides an effective therapeutic strategy for targeting HIF-2α and suppressing tumor progression in ccRCC patients.

PADI4 stimulates esophageal squamous cell carcinoma tumor growth and up-regulates CA9 expression.

An increasing amount of evidence indicates that peptidylarginine deiminase isoform 4 (PADI4) plays an important role in tumorigenesis. However, the effects of PADI4 on tumor‐bearing mice are unknown, and no studies have investigated this tumorigenic pathway in an animal model. In the present study, ECA109 cells originating from esophageal squamous cell carcinoma (ESCC) were transfected with PADI4‐expressing lentivirus and were injected into BALB/c nude mice. Tumor size and weight were significantly increased in the mouse tumors established with PADI4‐overexpressing ECA109 cells. PCR array analysis revealed increased CA9 expression in ECA109 cells transfected with a PADI4‐expressing plasmid, while decreased CA9 expression levels were detected in cells transfected with anti‐PADI4 siRNA. Furthermore, up‐regulation of CA9 expression was detected in mouse tumors established with PADI4‐overexpressing cells. Immunohistochemistry detected the increased expression and co‐localization of PADI4 and CA9 in ESCC tissues compared with adjacent non‐tumor tissues and normal tissue controls. These results were verified using Western blotting. Cell proliferation significantly increased or decreased in ECA109 and EC9706 (another ESCC‐originating cell line) cells transfected with a PADI4‐expressing plasmid or anti‐PADI4 siRNA, respectively. The above findings suggest that increased PADI4 expression in ESCC stimulates tumor growth and up‐regulates CA9 expression, which is known to promote metastatic properties in tumor cells.

Estrogen receptor \u03b2 is associated with expression of cancer associated genes and survival in ovarian cancer

BackgroundIn ovarian cancer, the role of estrogen receptors (ERs), particularly of ERβ, being suggested as tumor suppressor in breast and prostate cancer, remains unclear. We examined the expression of nuclear and cytoplasmic ERβ in ovarian cancer and correlated it with expression of ovarian cancer markers CA125, CEA and CA72–4, steroid hormone receptors ERα and PR, cancer-associated genes EGFR, p53, HER2 and proliferation marker Ki-67. Additionally we examined to what extent expression of ERβ and the other proteins affects survival of ovarian cancer patients.MethodsWe established a tissue microarray from 171 ovarian cancer patients and performed immunohistochemical analyses of the mentioned proteins.ResultsNuclear ERβ was detected in 47.31% of the ovarian cancer tissues and cytoplasmic expression of this receptor was observed in 23.08%. Nuclear expression of ERβ was significantly decreased in the G3 subgroup compared to better differentiated cancers (p < 0.01) and correlated with ovarian cancer markers CEA (95% CI 0.1598–0.4465; p < 0.0001) and CA72–4 (95% CI 0.05953–0.3616; p < 0.01). Cytoplasmic ERβ expression correlated with EGFR levels (95% CI 0.1059–0.4049; p < 0.001). ERα expression was associated with expression of CA125 and PR. Overall survival of patients with tumors expressing cytoplasmic ERβ was significant longer compared to those with ERβ-negative ovarian cancer (chi-square statistic of the log-rank, p < 0.05). Progression-free survival was dependent on expression of PR (chi-square statistic of the log-rank, p < 0.05) and Ki-67 (p = 0.05).ConclusionsOur data suggest an important, but distinct role of nuclear and cytoplasmic ERβ expression in ovarian cancer and encourage further studies on its role in this cancer entity.

Biomarker panel predicts survival after resection in pancreatic ductal adenocarcinoma: A multi-institutional cohort study

BackgroundUp to 60% of patients who undergo curative-intent pancreatic ductal adenocarcinoma (PDAC) resection experience disease recurrence within six months. We recently published a systematic review of prognostic immunohistochemical biomarkers in PDAC and shortlisted a panel of those reported with the highest level of evidence, including p53, p16, Ca-125, S100A4, FOXC1, EGFR, mesothelin, CD24 and UPAR. This study aims to discover and validate the prognostic significance of a combinatorial panel of tumor biomarkers in patients with resected PDAC. MethodsPatients who underwent PDAC resection were included from a single institution discovery cohort and a multi-institutional validation cohort. Tumors in the discovery cohort were stained immunohistochemically for all nine shortlisted biomarkers. Biomarkers significantly associated with overall survival (OS) were reevaluated as a combinatorial panel in both discovery and validation cohorts for its prognostic significance. Results224 and 191 patients were included in the discovery and validation cohorts, respectively. In both cohorts, S100A4, Ca-125 and mesothelin expression were associated with shorter OS. In both cohorts, the number of these biomarkers expressed was significantly associated with OS (discovery cohort 36.8 vs. 26.4 vs 16.3 vs 12.8 months, P < 0.001; validation cohort 25.2 vs 18.3 vs 13.6 vs 11.9 months, P = 0.008 for expression of zero, one, two and three biomarkers, respectively). On multivariable analysis, expression of at least one of three biomarkers was independently associated with shorter OS. ConclusionCombinations of S100A4, Ca-125 and mesothelin expression stratify survival after resection of localized PDAC. Co-expression of all three biomarkers is associated with the poorest prognostic outcome.

RNA binding protein HuR regulates extracellular matrix gene expression and pH homeostasis independent of controlling HIF-1α signaling in nucleus pulposus cells

Nucleus pulposus (NP) cells reside in the hypoxic niche of the intervertebral disc. Studies have demonstrated that RNA-binding protein HuR modulates hypoxic signaling in several cancers, however, its function in the disc is unknown. HuR did not show cytoplasmic translocation in hypoxia and its silencing did not alter levels of Hif-1α or HIF-targets in NP cells. RNA-Sequencing data revealed that important extracellular matrix-related genes including several collagens, MMPs, aggrecan, Tgf-β3 and Sdc4 were regulated by HuR. Further analysis of HuR-silenced NP cells confirmed that HuR maintained expression of these matrix genes. We confirmed decreased levels of secreted collagen I and Sdc4 and increased pro-MMP13 in HuR-knockdown cells. In addition, messenger ribonucleoprotein immunoprecipitation demonstrated HuR binding to Tgf-β3 and Sdc4 mRNAs. Interestingly, while HuR bound to Hif-1α and Vegf mRNAs, it was clear that compensatory mechanisms sustained their expression when HuR was silenced. Noteworthy, despite the presence of multiple HuR-binding sites and reported interaction in other cell types, HuR showed no binding to Pgk1, Eno1, Pdk1 and Pfkfb3 in NP cells. Metabolic studies showed a significant decrease in the extracellular acidification rate (ECAR) and mitochondrial oxygen consumption rate (OCR) and acidic pH in HuR-silenced NP cells, without appreciable change in total OCR. These changes were likely due to decreased Ca12 expression in HuR silenced cells. Taken together, our study demonstrates for the first time that HuR regulates extracellular matrix (ECM) and pH homeostasis of NP cells and has important implications in the maintenance of intervertebral disc health.

Ovarian fibroma/fibrothecoma with elevated serum CA125 level: A cohort of 66 cases.

Ovarian fibroma/fibrothecoma with elevated serum OC125 antigen (CA125) is rarely encountered in clinical practice, and also easily misdiagnosed as epithelial ovarian carcinoma (EOC). The aim of this study was to investigate the clinicopathological features of ovarian fibroma/fibrothecoma with elevated serum CA125.In total, 580 patients who underwent primary surgery and pathologically diagnosed as ovarian fibroma/fibrothecoma were retrospectively analyzed. The clinicopathological parameters were collected and compared between the patients with elevated serum CA125 (>35 U/mL) and without. The immunoreactivity for CA125 in ovarian fibroma/fibrothecoma and epithelial cancer tissues was detected and compared by immunohistochemistry. Univariate and multivariate analyses were performed to identify factors associated with elevated serum CA125 level. The correlation between the immunoreactivity of CA125 in tissue and serum CA125 level was examined by Pearson correlation analysis.Elevated serum CA125 level (range 36.7–1848 u/m) was found in 66 of 580 (11.3%) ovarian fibroma/fibrothecoma patients. Univariate analysis showed that the elevated serum CA125 level was significantly correlated with tumor diameter ≥10 cm (P < .001), ascites (P < .001), and hydrothorax (P < .001). Multivariate analysis revealed that tumor diameter ≥10 cm and ascites were independently associated factors (P < .001 and < .001 respectively). Immunohistochemical staining showed that the expression of CA125 was negative in all fibroma/fibrothecoma tissues, but positive in all EOC tissues, and the immunoreactivity for CA125 was positively correlated with serum CA125 level in the EOC patients (P = .005).The elevated serum CA125 level in ovarian fibroma/fibrothecoma is nontumor originated and occurs more frequently in those with larger size tumor or Meigs syndrome.

Abstract B50: YAP induces development of mesenchymal subtype of high-grade serous ovarian cancer from granulosa cells

Abstract The Hippo pathway plays critical roles in both organ development and tumorigenesis. Our previous studies demonstrated that Yes-Associated Protein 1 (YAP), a transcription coactivator and the major effector of the Hippo pathway, plays a critical role in ovarian cancer development. The present study investigates the role of the Hippo/YAP signaling pathway in the tumorigenesis of ovarian granulosa cells. Fluorescent and chromogenic immunohistochemistry were used to examine YAP expression in human and mouse ovaries. Western blot and qRT-PCR were used to determine protein and mRNA expression. Retrovirus-based YAP expressing constructs and YAP siRNAs were utilized to manipulate YAP levels in primary or immortalized human granulosa cells. A new three-dimensional hanging-drop culture system was used to mimic the growth of granulosa cells in vivo. The soft agar assay and xenograft mouse models were employed to investigate the role of YAP in transformation and tumorigenesis of granulosa cells. Immunohistochemical analyses showed that the active form of YAP (nuclear YAP) was expressed in proliferative granulosa cells in human and mouse ovaries, whereas the inactive form of YAP (cytoplasmic YAP) was detected predominantly in terminally differentiated luteal cells, indicating that YAP activity may be involved in the regulation of granulosa cell proliferation and differentiation. Overexpression of wild-type or constitutively active YAP compromised the expression of genes required for steroidogenesis and induced de-differentiation of differentiated granulosa cells. Ectopic expression of the constitutively active form of YAP in less-differentiated human granulosa cells derived from growing follicles induced tumors in a xenograft mouse model. Intriguingly, we found that these tumors resembled human high-grade serous ovarian cancer (HGSOC) morphologically, histologically, and genetically. The tumors were accompanied by accumulation of large amount of ascites in the peritoneal cavity. Highly proliferative and aggressive tumor cells invaded into omentum, visceral fat tissue, diaphragm, pancreas, liver, and gastrointestinal system. Deletion of the BRCA1 gene in these tumor cells promoted tumor progression and significantly reduced survival rate. Interestingly, these tumors have high expression of N-cadherin but very low expression of CA125 and E-cadherin, which are molecular characteristics of a recently identified mesenchymal subtype of HGSOC. Studies from The Cancer Genome Atlas and the Australian Ovarian Cancer Study Group indicate that the mesenchymal subtype of HGSOC has the poorest prognosis among all subtypes of ovarian cancers. In conclusion, results from this study indicate that YAP plays a critical role in regulating granulosa cell proliferation and differentiation. Overactivation of YAP can result in de-differentiation and reprogramming of ovarian granulosa cells, leading to development of mesenchymal subtype of HGSOC. Our results identify early-stage granulosa cells as a cell of origin of newly identified mesenchymal type of HGSOC. Citation Format: Xiangmin Lv, Chunbo He, Cong Huang, Guohua Hua, Bowen Ma, Xingcheng Chen, Jin Zhou, Zhengfeng Wang, Jixin Dong, Bo R. Rueda, John S. Davis, Cheng Wang. YAP induces development of mesenchymal subtype of high-grade serous ovarian cancer from granulosa cells. [abstract]. In: Proceedings of the AACR Conference: Addressing Critical Questions in Ovarian Cancer Research and Treatment; Oct 1-4, 2017; Pittsburgh, PA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(15_Suppl):Abstract nr B50.

Circulating hypoxia marker carbonic anhydrase IX (CA9) in patients with hepatocellular carcinoma and patients with cirrhosis.

Background and aimsExpression of carbonic anhydrase IX (CA9), an enzyme expressed in response to hypoxia, acidosis and oncogenic alterations, is reported to be a prognostic factor in HCC patients. Here we evaluated serum CA9 levels in HCC and cirrhosis patients.MethodsHCC and cirrhosis patients were prospectively recruited and CA9 levels were determined. CA9 levels were compared to stages of cirrhosis and HCC stages. The association of the CA9 levels and overall survival (OS) was assessed. Furthermore, immunohistochemical CA9 expression in HCC and cirrhosis was evaluated.Results215 patients with HCC were included. The median serum CA9 concentration in patients with HCC was 370 pg/ml and significantly higher than in a healthy cohort. Patients with advanced cancer stages (BCLC and ALBI score) had hid significant higher levels of CA9 in the serum. HCC patients with high serum CA9 concentrations (>400 pg/ml) had an increased mortality risk (hazard ratio (HR) 1.690, 95% confidence interval (CI) 1.017–2.809, P = 0.043). Serum CA9 concentration in cirrhotic patients did not differ significantly from HCC patients. Higher CA9 levels in cirrhotic patients correlated with portal hypertension and esophageal varices. Patients with ethanol induced cirrhosis had the highest CA9 levels in both cohorts. Levels of CA9 did not correlate with immunohistochemical expression.ConclusionsWe conclude that a high CA9 level is a possible prognostic indicator for a poor outcome in HCC patients. The high CA9 levels are probably mainly associated with portal hypertension. Ductular reactions might be a possible source of serum CA9.

Abstract 2941: APE1/Ref-1 redox signaling regulates HIF1a-mediated CA9 expression in hypoxic pancreatic cancer cells: Combination treatment in patient-derived pancreatic tumor models

Abstract Pancreatic ductal adenocarcinoma (PDAC) is a deadly disease characterized by aggressive metastasis and therapeutic resistance. Reactive stroma in PDAC tumors leads to fibrosis, inflammation, and hypoxia. Hypoxia signaling creates a more aggressive phenotype with increased potential for metastasis and decreased therapeutic efficacy. Carbonic anhydrase IX (CA9) functions as part of the cellular response to hypoxia by regulating intracellular pH to promote cell survival. Apurinic/Apyrimidinic Endonuclease-1-Reduction/oxidation Effector Factor 1 (APE1/Ref-1) is a multifunctional protein with endonuclease activity in DNA base excision repair and redox signaling activity. This redox activity is responsible for reducing oxidized cysteines on specific transcription factors, including hypoxia inducible factor 1 alpha (HIF1α), enabling them to bind target sequences in DNA. We evaluated the mechanisms underlying PDAC cell responses to hypoxia and APE1/Ref-1 redox signaling control of HIF1α, a critical factor in hypoxia-induced CA9 transcription. We hypothesized that obstructing the HIF-CA9 axis at two points via APE1/Ref-1 inhibition (which results in a decrease in CA9 expression) and direct CA9 inhibition results in enhanced PDAC cell killing under hypoxic conditions. In our studies, HIF1α-mediated induction of CA9 is significantly attenuated following APE1/Ref-1 knock down or redox signaling inhibition in patient-derived PDAC cells and pancreatic cancer-associated fibroblast cells using the APE1/Ref-1 redox signaling inhibitor APX3330 (currently in clinical trials). Additionally, dual-targeting of APE1/Ref-1 redox signaling activity and CA9 activity results in additive-to-synergistic enhancement of acidification and cytotoxicity of PDAC cells under hypoxic conditions as well as decreased tumor growth in an ex vivo 3-dimensional tumor co-culture model. These studies are clinically relevant as we used the CA9 inhibitor SLC-0111 (phase I clinical trial completed), as well as APX3330 (Apexian Pharmaceuticals: IND 125360), for which a phase I clinical trial has opened. Further experiments characterized novel analogs of APX3330: APX2009 and APX2014, which demonstrated up to 50-fold improved potency as measured by pH reduction, cytotoxicity, and inhibition of hypoxia-induced CA9 expression. An SLC-0111 analog, FC12-531A, demonstrated up to 75-fold improved potency as measured by cytotoxicity. In combination, these analogs resulted in synergistic inhibition of 3D tumor spheroid growth at nanomolar-to-low-micromolar concentrations. These results underscore the concept that proper combination therapy has significant clinical utility of blocking APE1/Ref-1 and CA9 function for novel PDAC therapeutic treatment. Citation Format: Derek Logsdon, Fenil Shah, Fabrizio Carta, Claudiu Supuran, Melissa Fishel, Mark R. Kelley. APE1/Ref-1 redox signaling regulates HIF1a-mediated CA9 expression in hypoxic pancreatic cancer cells: Combination treatment in patient-derived pancreatic tumor models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2941.

Abstract 639: Salt-inducible kinase 3 expression identifies long-term survivors of serous ovarian cancer

Abstract Background: Epithelial ovarian cancer (EOC) has a high tumor-associated mortality rate among the gynecological cancers because EOC is usually diagnosed during the advanced stage. Cancer antigen (CA) 125 is the most well-studied biomarker for ovarian cancer screening. However, CA125 is also elevated in numerous conditions, resulting in decreased specificity. Materials and Method: We collected ovarian cancer specimens from 204 patients and examined the SIK3 and CA125 expression levels. The expression percentage of SIK3 or CA125 in the tumor cells, pre-surgery serum CA125 was divided into two groups and compared with patient overall survival, progression-free survival, cancer stage, cancer type and chemotherapeutic resistance. Results: A total of 30.9% of the patients had stage I disease, 7.8% had stage II disease, 52.4% had stage III disease, and 8.9% had stage IV disease. The median OS and PFS were 49.5 months (range 0.25-205 months) and 28.4 months (0.25-182.4), respectively. The mean age of the patients was 52.8 years (range 25-82 years). The histological classification was based on the four main ovarian cancer types: serous (51.5%), endometrioid (14.2%), clear cell (20.6%), and mucinous (9.3%). In all stages of the disease, high expression of SIK3 (SIK3-H) was associated with a markedly better OS, compared with low expression of SIK3 (SIK3-L) (127.0 months and 46.0 months, respectively; hazard ratio [HR] for death, 0.59; 95% confidence interval [CI], 0.41 to 0.86; P=0.005). SIK3-H had a markedly better PFS rate than SIK3-L (66.0 months and 26.0 months, respectively; HR for disease progression or recurrence, 0.68; 95% CI, 0.47 to 0.99; P=0.04). The same results were evident for serous ovarian cancer, where SIK3-H had a better OS rate than SIK3-L (75.0 months and 34.0 months, respectively; HR for death, 0.57; 95% CI, 0.36 to 0.91; P=0.02) and a better PFS rate than SIK3-L (44 and 13 months, respectively; HR, 0.52; 95% CI, 0.32 to 0.83; P=0.006). Notably, for advanced stage serous ovarian cancer, Only SIK3-H had a better OS rate than SIK3-L (48.0 months and 28.0 months, respectively; HR for death, 0.56; 95% CI, 0.33 to 0.96; P=0.03) and PFS rate (32 and 13 months, respectively; HR, 0.57; 95% CI, 0.34 to 0.95; P=0.03). Tissue CA125 expression and the pre-surgery serum CA125 level did not show significant differences in OS and PFS. Our results demonstrated that high SIK3 expression correlated with better OS and PFS rates in ovarian cancer patients, especially in patients with advanced serous ovarian cancer. Conclusion: High expression of SIK3 indicates a better prognosis in primary ovarian cancer and serous disease, especially in advanced serous disease. Citation Format: Keng-Fu Hsu, Yu-Ling Liang, Chin-Han Wu, Neng-Yao Shih. Salt-inducible kinase 3 expression identifies long-term survivors of serous ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 639.

Abstract 4337: Identification of novel fusion gene, PUM1-TRAF3, as potent biomarker in bile duct cancer

Abstract Background: Gene fusion occurs when a part of one gene fuses with a part of another gene by genome rearrangement and may possess oncogenic properties. Bile duct cancer (BDC) are one of the most lethal malignancy, but lack of proper diagnostic or prognostic markers. Our aim was to investigate BDC specific fusion genes found in patient's specimens and its clinical significance. Method : We extracted total RNA from five BDC tissues and normal tissues from the same patients. RNA sequencing was performed and the result data was analyzed using ChimeraScan, Jaffa or FusionCatcher softwares to detect gene fusion. The expression of the detected novel fusion gene was confirmed by ddPCR using tissues from the operated BDC patients and its clinical significance was analyzed with medical information of the patients. Result : We found 52 BDC specific fusion genes by RNA sequencing and we identified novel BDC specific fusion gene, PUM1-TRAF3 by bioinformatics analysis. Total 55 patients with BDC who underwent curative surgery were evaluated to investigate clinical significance of PUM1-TRAF3. There were 28 patients (50.1%) who expressed PUM1-TRAF3 by ddPCR. Patients with PUM1-TRAF3 presented higher rate of female gender (57.1 vs. 25.9, p=0.019), advanced stage (25% vs. 3.7%, p=0.025) and recurrence rate (64.7 vs. 48.1%, p=0.080) compared to patients without PUM1-TRAF3. Disease free survival (DFS) was 12.1 vs. 29.8 (p=0.078) and overall survival was 26.1 vs. 36.6 (p=0.239) between the patients with PUM1-TRAF3 and without PUM1-TRAF3. In multivariated analysis by cox-regression, expression of PUM1-TRAF3 (HR 2.86, p=0.023), advanced stage (HR 2.69, p=0.041), higher CA19-9 (HR 1.00, p=0.011) and positive resection margin (HR 4.57, p=0.002) were significantly associated with short DFS.Conclusion : In this study, we identified novel BDC-specific fusion gene, PUM1-TRAF3. Expression of PUM1-TRAF3 was confirmed in patient's tissues and associated with short DFS in operated BDC patients. PUM1-TRAF3 can be a potent biomarker in BDC. Citation Format: Dawoon E. Jung, Kahee Kim, Chanyang Kim, Soo Been Park, Jung Hyun Jo, Hee Seung Lee, Si Young Song. Identification of novel fusion gene, PUM1-TRAF3, as potent biomarker in bile duct cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4337.

Co-culturing nucleus pulposus mesenchymal stem cells with notochordal cell-rich nucleus pulposus explants attenuates tumor necrosis factor-\u03b1-induced senescence

BackgroundCell therapy for the treatment of intervertebral disc degeneration (IDD) faces serious barriers since tissue-specific adult cells such as nucleus pulposus cells (NPCs) have limited proliferative ability and poor regenerative potential; in addition, it is difficult for exogenous adult stem cells to survive the harsh environment of the degenerated intervertebral disc. Endogenous repair by nucleus pulposus mesenchymal stem cells (NPMSCs) has recently shown promising regenerative potential for the treatment of IDD. Notochordal cells (NCs) and NC-conditioned medium (NCCM) have been proven to possess regenerative ability for the treatment of IDD, but this approach is limited by the isolation and passaging of NCs. Our previous study demonstrated that modified notochordal cell-rich nucleus pulposus (NC-rich NP) has potential for the repair of IDD. However, whether this can protect NPMSCs during IDD has not been evaluated.MethodsIn the current study, tumor necrosis factor (TNF)-α was used to mimic the inflammatory environment of IDD. Human NPMSCs were cocultured with NC-rich NP explants from healthy rabbit lumbar spine with or without TNF-α. Cell proliferation and senescence were analyzed to investigate the effect of NC-rich NP explants on TNF-α-treated NPMSCs. The expression of mRNA encoding proteins related to matrix macromolecules (such as aggrecan, Sox-9, collagen Iα, and collagen IIα), markers related to the nucleus pulposus cell phenotype (including CA12, FOXF1, PAX1, and HIF-1α), and senescence markers (such as p16, p21, and p53), senescence-associated proinflammatory cytokines (IL-6), and extracellular proteases (MMP-13, ADAMTS-5) was assessed. The protein expression of CA12 and collagen II was also evaluated.ResultsAfter a 7-day treatment, the NC-rich NP explant was found to enhance cell proliferation, decrease cellular senescence, promote glycosaminoglycan (GAG), collagen II, and CA12 production, upregulate the expression of extracellular matrix (ECM)-related genes (collagen I, collagen II, SOX9, and ACAN), and enhance the expression of nucleus pulposus cell (NPC) markers (HIF-1α, FOXF1, PAX1, and CA12).ConclusionModified NC-rich NP explants can attenuate TNF-α-induced degeneration and senescence of NPMSCs in vitro. Our findings provide new insights into the therapeutic potential of NC-rich NP for the treatment of IDD.

APOBEC3B up-regulation independently predicts ovarian cancer prognosis: a cohort study

BackgroundOvarian cancer is a heterogeneous disease with a high degree of genomic instability, pro-/antitumor immunity and inflammation, and remains the most lethal gynecologic cancer worldwide. APOBEC3B, a member of the AID/APOBEC family, is part of the innate immune system which plays a key role in combating exogenous infection especially viral infection. Studies have shown that APOBEC3B expression is elevated in a variety of cancer tissues and cell lines, and plays a prominent role in the genesis and evolution of various cancers. However, the clinical relevance of APOBEC3B in ovarian cancer needs to be further investigated. The current study aimed to evaluate the predictive value of APOBEC3B in ovarian cancer clinical outcome, and to explore possible molecular mechanisms contributing to ovarian cancer progression.MethodsThe expression of APOBEC3B in biopsy tissue specimens from 88 ovarian cancer patients was examined using immunohistochemistry. In addition, ovarian cancer cell lines were transfected with APOBEC3B siRNA or pLenti-APOBEC3B construct. Western blotting and SRB assay were performed to explore the role of APOBEC3B in ovarian cancer.ResultsPatients were followed for a median of 74.77 months following the time of surgery. Forty-two patients had died, 5 had relapsed but were still alive at the end of study, and 41 patients remained alive and had no recurrence. Over-expression of APOBEC3B was associated with advanced FIGO stage and elevated CA125 (both p< 0.05). Univariate analysis result showed that histological subtype, FIGO stage, intravascular tumor thrombus, CA125 and APOBEC3B expression were associated with overall survival and disease-free survival of ovarian cancer patients. Multivariate analysis result showed that higher APOBEC3B expression were an independent prognostic factor to predict both worse overall survival (hazard ratio: 5.18, 95% confidence interval: 1.40–11.95, p= 0.003) and disease-free survival (hazard ratio: 4.23, 95% confidence interval: 1.60–11.17, p= 0.004) of ovarian cancer patients. Furthermore, knockdown of APOBEC3B expression in ovarian cancer cells caused an decrease in cell line viability.ConclusionsAPOBEC3B expression is an independent prognostic factor in ovarian cancer patients. Knockdown of APOBEC3B expression affects ovarian cancer viability.

Abstract A27: Exploring the role of glycosylation in pancreatic disease

Abstract Pancreatic ductal adenocarcinoma (PDA) is almost uniformly lethal and surgical intervention is the only cure. Unfortunately, most patients are ineligible for resection because of the advanced stage of disease by the time of diagnosis. This is due in part to the lack of diagnostic tools, especially for families with elevated risk. The PDA biomarker, CA19-9, is measured in the blood to follow tumor burden longitudinally, but is neither sensitive nor specific enough to be used for diagnosis. The use of CA19-9 in PDA diagnosis is problematic given the elevation of CA19-9 in benign pancreatic disease, such as pancreatitis. While CA19-9 has been traditionally used as a diagnostic, retrospective studies reported that PDA patients who maintain a CA19-9 negative/low status have a significantly longer survival relative to those with higher CA19-9 levels in multivariate analyses. The functional significance of CA19-9 to PDA initiation, maintenance, and progression remains unclear due in part to the absence of this carbohydrate modification in mice. We found that expression of CA19-9 in the mouse pancreas is sufficient to induce pancreatitis, a benign proliferative condition that often confounds the diagnosis of PDA. Specifically, CA19-9 elevation resulted in rapid elevation of pancreatic enzymes in the blood, pancreatic infiltration of immune cells, acinar-to-ductal metaplasia and atrophy, as well as increased proliferation. Furthermore, we explored the utility of CA19-9 as a therapeutic target for both acute and chronic pancreatitis. This avenue of treatment strategy exhibits potential given that a pilot study demonstrated that turning off CA19-9 expression results in the normalization of pancreatic enzyme levels within four days following an acute episode of pancreatitis. Future work will focus on how elevation of this glycosylation modification mediates the development of pancreatitis by identifying the signaling pathways that are altered upon CA19-9 expression. In addition, we will explore the efficacy of therapeutically targeting CA19-9 in both pancreatitis and PDA with a larger goal of delineating the role of CA19-9 in pancreatic disease. Citation Format: Dannielle Engle, Hervé Tiriac, Arnaud Pommier, Christina Schoepfer, Brandon Da Silva, Melissa Yao, Youngkyu Park, Michael A. Hollingsworth, David Tuveson. Exploring the role of glycosylation in pancreatic disease [abstract]. In: Proceedings of the AACR Special Conference: Advances in Modeling Cancer in Mice: Technology, Biology, and Beyond; 2017 Sep 24-27; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(10 Suppl):Abstract nr A27.

Postresection CA19-9 and margin status as predictors of recurrence after adjuvant treatment for pancreatic carcinoma: Analysis of NRG oncology RTOG trial 9704

PurposeNRG Oncology RTOG 9704 was the first adjuvant trial to validate the prognostic value of postresection CA19-9 levels for survival in patients with pancreatic carcinoma. The data resulting from this study also provide information about predictors of recurrence that may be used to tailor individualized management in this disease setting. This secondary analysis assessed the prognostic value of postresection CA19-9 and surgical margin status (SMS) in predicting patterns of disease recurrence. Methods and materialsThis multicenter cooperative trial included participants who were enrolled as patients at oncology treatment sites in the United States and Canada. The study included 451 patients analyzable for SMS, of whom 385 were eligible for postresection CA19-9 analysis. Postresection CA19-9 was analyzed at cut points of 90, 180, and continuously. Patterns of disease recurrence included local/regional recurrence (LRR) and distant failure (DF). Multivariable analyses included treatment, tumor size, and nodal status. To adjust for multiple comparisons, a P value of ≤ .01 was considered statistically significant and > .01 to ≤ .05 to be a trend. ResultsFor CA19-9, 132 (34%) patients were Lewis antigen–negative (no CA19-9 expression), 200 (52%) had levels <90, and 220 (57%) had levels <180. A total of 188 patients (42%) had negative margins, 152 (34%) positive, and 111 (25%) unknown. On univariate analysis, CA19-9 cut at 90 was associated with increases in LRR (trend) and DF. Results were similar at the 180 cut point. SMS was not associated with an increase in LRR on univariate or multivariate analyses. On multivariable analysis, CA19-9 ≥ 90 was associated with increased LRR and DF. Results were similar at the 180 cut point. ConclusionsIn this prospective evaluation, postresection CA19-9 was a significant predictor of both LRR and DF, whereas SMS was not. These findings support consideration of adjuvant radiation therapy dose intensification in patients with elevated postresection CA19-9.

The clinical significance of the combined detection of serum Smac, HE4 and CA125 in endometriosis-associated ovarian cancer.

This study aims to investigate the clinical significance of serum Smac, HE4 and CA125 alone or combined for detecting endometriosis-associated ovarian cancer (EAOC). The level of serum Smac, HE4 and CA125 in 40 healthy controls, 40 cases of benign endometriosis ovarian tumor, and 60 cases of EAOC were detected by ELISA and electrochemical immune method. Serum Smac expression level was significantly lower in the EAOC group than in the control group and benign ovarian tumor group (P< 0.05), while HE4 and CA125 expression levels were significantly higher in the EAOC group than the other two groups. The sensitivity of Smac single detection was up to 91.67%, and the specificity of HE4 was up to 98.75%. Furthermore, the sensitivity of Smac + HE4 + CA125 combined was the highest, which reached up to 98.33%; but the specificity was low, which reached up to 75%. The serum expression level differences before and after surgery were statistically significant. As the number of chemotherapies increases, the Smac level increased, and HE4 and CA125 levels gradually decreased. Furthermore, Smac increased to normal at the end of the 2nd period of chemotherapy, while HE4 and CA125 decreased to normal in 2nd and 3rd period of chemotherapy, respectively. Serum Smac, HE4 and CA125 may play an important role in predicting EAOC and in monitoring the prognosis of postoperative EAOC.

Prognostic significance of mucinous component in gastric adenocarcinoma after radical D2 gastrectomy.

BackgroundThe mucinous component is a special histologic factor in gastric adenocarcinoma. The aim of this study was to assess the prognostic significance of mucinous component in gastric adenocarcinoma according to proportion.Patients and methodsCandidate patients with gastric adenocarcinoma were given radical D2 gastrectomies from September 2008 to May 2015 in our division. Clinicopathologic data and prognosis were monitored and analyzed among gastric adenocarcinoma patients with various proportions of mucinous component.ResultsA total of 690 gastric adenocarcinomas with various proportions of mucinous component from 6,025 gastric adenocarcinoma patients were included. Higher numbers of patients with mucinous component came from: young patients, females, those with drinking history, at lower locations, Borrmann type III and IV, T4 stage, and positive for dissected lymph nodes. Tumors and pathological molecular markers showed more positivity in CEA, CA19-9, S100, and CD34. As the various proportions increased, more mucinous component seemed to be accompanied by more Borrmann type III and IV, T4 stage, and more positive expression of CEA and CA19-9. However, no significant difference in 5-year overall survival rate was observed among various proportions or existence of mucinous component. Also, proportion or existence of mucinous component was not an independent prognostic factor in multivariate analysis.ConclusionMucinous component was not a prognostic factor for gastric adenocarcinoma after radical D2 gastrectomy, no matter what proportion the component comprised. However, gastric adenocarcinoma with mucinous component showed specific clinicopathological characteristics, such as more advanced tumor stage, different age and sex, and more positive rate of molecular markers, which might provide a new strategy for optimal individual diagnosis and therapies.