Copy number gain of granulin-epithelin precursor (GEP) at chromosome 17q21 associates with overexpression in human liver cancer

BACKGROUND: Granulin-epithelin precursor (GEP), a secretory growth factor, demonstrated overexpression in various human cancers, however, mechanism remain elusive. Primary liver cancer, hepatocellular carcinoma (HCC), ranks the second in cancer-related death globally. GEP controlled growth, invasion, metastasis and chemo-resistance in liver cancer. Noted that GEP gene locates at 17q21 and the region has been frequently reported to be amplified in subset of HCC. The study aims to investigate if copy number gain would associate with GEP overexpression. METHODS: Quantitative Microsatellite Analysis (QuMA) was used to quantify the GEP DNA copy number, and fluorescent in situ hybridization (FISH) was performed to consolidate the amplification status. GEP gene copy number, mRNA expression level and clinico-pathological features were analyzed. RESULTS: GEP DNA copy number determined by QuMA corroborated well with the FISH data, and the gene copy number correlated with the expression levels (n = 60, r = 0.331, P = 0.010). Gain of GEP copy number was observed in 20% (12/60) HCC and associated with hepatitis B virus infection status (P = 0.015). In HCC with increased GEP copy number, tight association between GEP DNA and mRNA levels were observed (n = 12, r = 0.664, P = 0.019). CONCLUSIONS: Gain of the GEP gene copy number was observed in 20% HCC and the frequency comparable to literatures reported on the chromosome region 17q. Increased gene copy number contributed to GEP overexpression in subset of HCC. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12885-015-1294-x) contains supplementary material, which is available to authorized users

Abstract 882: Genomics approach to identify drug transporter ABCF1 associated with liver cancer recurrence and chemo-resistance

Abstract Background: Liver cancer is the third leading cause of cancer death globally. Surgery is the mainstay for early stage patients with the best survival outcome, but cancer recurrence is still common after curative surgery. Chemotherapy has been widely used to treat advanced liver cancer but with marginal efficacy. There is an urgent need to elucidate the key genes that regulate recurrence and chemo-resistance in the clinical situation. Recently, we have demonstrated that the novel growth factor granulin-epithelin precursor (GEP) is associated with chemo-resistance and recurrence-free survival. GEP has been identified as a potential therapeutic target for liver cancer in our earlier genomic studies. We demonstrated that GEP overexpressed in the majority of liver cancers, regulated proliferation, invasion and tumorigenicity. Importantly, the anti-GEP monoclonal antibody treatment inhibited the growth of in vivo human liver cancer. GEP modulated the expression of the drug transporter ABCB5, and blockage of ABCB5 sensitized the liver cancer cells to chemotherapeutic agents and attenuated the expression of hepatic cancer stem cell markers CD133 and EpCAM. Notably, GEP expression is detectable in all liver cancer tissues while only half show detectable ABCB5 transcript. Importantly, the majority of the patients were chemo-resistant. Therefore, there should be additional crucial drug transporters. Objective: To use genomics approach to systematically examine the drug transporters in association with GEP, recurrence-free survival outcome, and functional role on chemo-resistance. Methods: The liver cancer gene expression profiles reported previously were re-examined, and all the drug transporter family members were ranked in association with GEP expression patterns. The genes that have shown high correlation with GEP expressions in the microarray hybridization datasets were further validated in an independent cohort of clinical specimens using independent research platform real-time quantitative RT-PCR. Results: Here, we reported that the expression levels of drug transporter ABCF1 and GEP were significantly correlated in human liver tissues (P &amp;lt; 0.001). ABCF1 was detectable in all tumor tissues, upregulated in the tumor as compared with the adjacent non-tumor liver (P &amp;lt; 0.001), and that the increased ABCF1 level was associated with poor recurrence-free survival (log-rank test, P = 0.001). Functionally, GEP regulated ABCF1 expresson levels, and suppression of ABCF1 rendered the liver cancer cells sensitive to chemotherapeutic agents. Conclusion: In summary, chemo-resistance and cancer recurrence are dictated by a subset of cells expressing GEP and ABC transporters. Targeting the novel growth factor GEP and drug transporters, in combination with chemotherapy, could provide novel treatment modalities to eradicate the aggressive liver cancer cells. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 882. doi:1538-7445.AM2012-882</jats:p

Functional non-invasive detection of glycolytic pancreatic ductal adenocarcinoma

Abstract Background: Pancreatic Ductal Adenocarcinoma (PDAC) lacks effective treatment options beyond chemotherapy. Although molecular subtypes such as classical and QM (quasi-mesenchymal)/basal-like with transcriptome-based distinct signatures have been identified, deduced therapeutic strategies and targets remain elusive. Gene expression data show enrichment of glycolytic genes in the more aggressive and therapy resistant QM subtype. However, whether the glycolytic transcripts are translated into functional glycolysis that could further be explored for metabolic targeting in QM subtype is still not known. Methods: We used different patient-derived PDAC model systems (conventional and primary patient derived cells, patient derived xenografts (PDX) and patient FFPE samples) and performed transcriptional and functional metabolic analysis. These included: RNAseq and Illumina HT12 bead array, in vitro Seahorse metabolic flux assays and metabolic drug targeting, and in vivo hyperpolarized [1-13C]pyruvate and [1-13C]lactate magnetic resonance spectroscopy (HP-MRS) in PDAC xenografts. Results: We found that glycolytic metabolic dependencies are not unambiguously functionally exposed in all QM PDACs. Metabolic analysis demonstrated functional metabolic heterogeneity in patient-derived primary cells and less so in conventional cell lines independent of molecular subtype. Importantly, we observed that the glycolytic product lactate is actively imported in the PDAC cells and used in mitochondrial oxidation in both classical and QM PDAC cells, although more actively in the QM cell lines. By using hyperpolarized 13C-magnetic resonance spectroscopy (HP-MRS), we were able to non-invasively identify highly glycolytic PDAC xenografts by detecting the last glycolytic enzymatic step and prominent intra-tumoral [1‑13C]pyruvate and [1-13C]lactate interconversion in vivo.Conclusion: Our study adds functional metabolic phenotyping to transcriptome-based analysis and proposes a functional approach to identify highly glycolytic PDACs as candidates for anti-metabolic therapeutic avenues.</jats:p

Functional metabolic phenotyping of human pancreatic ductal adenocarcinoma

AbstractPancreatic Ductal Adenocarcinoma (PDAC) lacks targeted treatment options. Although subtypes with transcriptome-based distinct lineage and differentiation features have been identified, deduced clinically actionable targets remain elusive. We here investigate functional metabolic features of the classical and QM (quasi-mesenchymal)/basal-like PDAC subtypes potentially exploitable for non-invasive subtype differentiation and therapeutic intervention.A collection of human PDAC cell lines, primary patient derived cells (PDC), patient derived xenografts (PDX) and patient PDAC samples were transcriptionally stratified into the classical and QM subtype. Functional metabolic analyses including targeted and non-targeted metabolite profiling (matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI)), seahorse metabolic flux assays and metabolic drug targeting were performed. Hyperpolarized 13C-magnetic resonance spectroscopy (HP-MRS) of PDAC xenografts was used for in vivo detection of intra-tumoral [1-13C]pyruvate and [1-13C]lactate metabolism.We identified glycolysis and lipid metabolism/fatty acid oxidation as transcriptionally preserved metabolic pathways in QM and classical PDAC subtype respectively. However, these metabolic cues were not unambiguously functionally linked to one subtype. Striking functional metabolic heterogeneity was observed especially in primary patient derived cells with only individual samples representing high dependence on glycolysis or mitochondrial oxidation. Of note, QM cells actively use the glycolytic product lactate as oxidative mitochondrial fuel. Using HP-MRS, we were able to non-invasively differentiate glycolytic tumor xenografts with high intratumoral [1-13C]pyruvate to [1-13C]lactate conversion in vivo.Although PDAC transcriptomes indicate molecular subtype-associated distinct metabolic pathways, we found substantial functional metabolic heterogeneity independent of the molecular subtype. Non-invasive identification of highly glycolytic tumors by [1-13C]pyruvate/lactate HP-MRS support individualized metabolic targeting approaches.</jats:p