Philippe Pourquier

Protein arginine (N)-methyl transferase 7 (PRMT7) as a potential target for the sensitization of tumor cells to camptothecins

PRMT7 belongs to the protein arginine methyl‐transferases family. We show that downregulation of PRMT7α and β isoforms in DC‐3F hamster cells was associated with increased sensitivity to the Top1 inhibitor camptothecin (CPT). This effect was not due to a change in Top1 contents or catalytic activity, or to a difference in the reversal of DNA breaks. Overexpression of PRMT7α and β in DC‐3F cells had no effect on CPT sensitivity, whereas it conferred a resistance to DC‐3F/9‐OH‐E cells for which both isoforms are reduced by two‐ to three‐fold as compared to DC‐3F parental cells. Finally, downregulation of the human PRMT7 could also sensitize HeLa cells to CPT, suggesting that it could be used as a target to potentiate CPT derivatives.

Abstract 785: ETS-mediated expression of miR-24 regulates Top1 levels and resistance of prostate cancer cell lines to camptothecin

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Human nuclear topoisomerase I (Top1) is a crucial enzyme involved in the removal of DNA supercoiling during replication, transcription and chromosome segregation. These roles rely on its capability to introduce reversible single-strand breaks in duplex DNA. Top1 is the unique target of camptothecin (CPT) derivatives such as Irinotecan and Topotecan that are approved for the treatment of colon, ovarian and lung cancers. These inhibitors stabilize covalent Top1-DNA complexes resulting in cytotoxic double-strand breaks by collision of stabilized complexes with advancing replication forks. The mechanisms of resistance to CPT derivatives are complex and not fully understood yet, though it is known that the amount of Top1 can directly affect tumor cell response to these agents. Advanced forms of prostate cancers (PCa) that usually become resistant to castration are treated with chemotherapy using taxane-based regimen with disappointing response rates. Unfortunately, they also display an intrinsic resistance to many other cytotoxic agents including CPT derivatives. In this study, we identified one resistance mechanism linked to the regulation of Top1 levels via the overexpression of transcription factors from the ETS family (mainly ERG and ETV1) that is detected in 50-80% of PCa. We first showed that siRNA-mediated transient repression of ERG or ETV1 in ERG-overexpressing VCaP cells and in ETV1-overexpressing LNCaP cells, respectively, resulted in increased cell sensitivity to CPT, confirming the role of these two ETS transcription factors in PCa cell response to Top1 inhibitors. Using the CHiP-Seq database from UCSC, we could identify potential ETS binding domains in the promoter region of a cluster of miRNAs containing miR-24, which expression level across the NCI-60 cell line panel was found to be negatively correlated with the sensitivity to CPT derivatives. We then evaluated the modulation of miR-24 expression on cell sensitivity to CPT. We found that overexpression of miR-24 reduced sensitivity of LNCaP cells to CPT, whereas overexpression of an anti-miR-24 had the opposite effect, confirming previously published bioinformatics predictions. Interestingly, miR-24 overexpression in LNCaP cells was accompanied with a reduction in Top1 protein levels whereas transfection with anti-miR-24 resulted in increased Top1 levels. Together, our preliminary results provide the first functional evidences that Top1 levels and subsequent resistance of PCa cell lines to CPT are, at least in part, regulated via ETS-mediated expression of miR-24. They further represent a rational basis for the clinical development of miR-24 inhibitors for the sensitization of CPT-resistant tumors such as advanced PCa. Citation Format: Samer Kayali, Emmanuel Roche, Daniele Montaudon, Philippe Pourquier, Nadine Houede. ETS-mediated expression of miR-24 regulates Top1 levels and resistance of prostate cancer cell lines to camptothecin. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 785. doi:10.1158/1538-7445.AM2014-785

Redox mechanism of levobupivacaine cytostatic effect on human prostate cancer cells

Anti-cancer effects of local anesthetics have been reported but the mode of action remains elusive. Here, we examined the bioenergetic and REDOX impact of levobupivacaine on human prostate cancer cells (DU145) and corresponding non-cancer primary human prostate cells (BHP). Levobupivacaine induced a combined inhibition of glycolysis and oxidative phosphorylation in cancer cells, resulting in a reduced cellular ATP production and consecutive bioenergetic crisis, along with reactive oxygen species generation. The dose-dependent inhibition of respiratory chain complex I activity by levobupivacaine explained the alteration of mitochondrial energy fluxes. Furthermore, the potency of levobupivacaine varied with glucose and oxygen availability as well as the cellular energy demand, in accordance with a bioenergetic anti-cancer mechanism. The levobupivacaine-induced bioenergetic crisis triggered cytostasis in prostate cancer cells as evidenced by a S-phase cell cycle arrest, without apoptosis induction. In DU145 cells, levobupivacaine also triggered the induction of autophagy and blockade of this process potentialized the anti-cancer effect of the local anesthetic. Therefore, our findings provide a better characterization of the REDOX mechanisms underpinning the anti-effect of levobupivacaine against human prostate cancer cells.

Abstract 4794: Oligoamide-based mimics of double-stranded B-DNA as a new class of DNA topoisomerase I catalytic inhibitors

Since the discovery of Topoisomerase I as a specific target for the treatment of cancers more than 30 years ago, only two inhibitors derived from the natural compound camptothecin (CPT) have been approved in the clinic for the treatment of colon, lung and ovarian cancers: topotecan and irinotecan. The cytotoxicity of these Top1 poisons relies on their capability to stabilize covalent Top1-DNA complexes, leading to replication-mediated DNA double-strand breaks. However tumor cells develop multiple resistance mechanisms that are limiting their efficacy. Though new CPT derivatives and other Top1 poisons with various chemical structures have been developed, they share the same resistance mechanisms. Aside from conventional approaches focus on compounds with higher potency to stabilize Top1-DNA complexes, we investigated the possibility to inhibit Top1 binding to DNA and/or DNA cleavage by using DNA mimics that would act as decoys, a strategy that was never explored before. To this aim, single chain oligoamides composed by iteration of mQQ units (Q: 8-amino-2-quinoline carboxylic acid; mQ: 8-aminomethyl-2-quinoline carboxylic acid) were synthesized. These molecules fold into helices that are stabilized by electrostatic repulsions and hydrogen bonds between the amide functions and endocyclic nitrogen atoms at adjacent residues. When Q and mQ precursors are functionalized by negatively charged residues, the positions of these residues in the folded structure match the position of phosphate residues in duplex B-DNA. Because distances between residues in mQQ and QmQ units are slightly different, it is possible to define a major groove and a minor groove, as in B-DNA, with the possibility to impact on groove width by changing the positions of the substituents (Q4 or Q5 for substitution in position 4 or 5 of the quinoline monomer, respectively), clearly establishing (mQQ5)n and (mQQ4)n as potential DNA-mimics of an unprecedented kind. We found that both (mQQ5)n and (mQQ4)n oligomers inhibited the activity of Top1-mediated relaxation of supercoiled DNA in vitro. Inhibition increased with the length of the oligoamide to reach an IC50 value (concentration inhibiting 50% of Top1-mediated DNA relaxation) around the nM range for the longest oligomer that was tested i.e. (mQQ4)16, corresponding to a double-strand DNA of 16 base pairs. By comparison, CPT had an IC50 of ∼10 μM in the same conditions. Top1 inhibition was rather selective as a (mQQ4)8 oligomer moderately inhibited Top2-mediated activity and did not affect the activity of various DNA interacting enzymes such as restriction enzymes XhoI and NdeI or nucleases such as DNAse I, S1 nuclease, benzonase or Flap-endonuclease I. Because these oligomers are resistant to proteases and nucleases and can be synthesized rapidly by solid phase synthesis with the possibility to modify its substituents without altering their helicity, they represent good candidates for drug development. Citation Format: Philippe Pourquier, Krzysztof Ziach, Celine Chollet, Vincent Parissi, Mathieu Marchivie, Panchami Prabhakaran, Partha P. Bose, Katta Laxmi-Reddy, Frederic Godde, Stephane Chaignepain, Jean Marie Schmitter, Ivan Huc. Oligoamide-based mimics of double-stranded B-DNA as a new class of DNA topoisomerase I catalytic inhibitors. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4794.

Abstract 3813: Role of the ERG transcription factor in the resistance of prostate cancer cells to the topoisomerase I inhibitor camptothecin

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Prostate cancer (PCa) is a major public health issue as it is the second cause of death among cancers in industrialized countries, and its incidence is rising. Surgery is the main option for the curative treatment of localized forms of PCa and recurrences are managed with androgen deprivation strategies. In the case of advanced metastatic PCa and when tumors become resistant to castration (mCRPC), chemotherapy with taxanes is initiated. However, response rates remain low with a restricted number of alternatives due to the intrinsic resistance of CRPC to a wide range of cytotoxic agents. Advanced PCa are particularly resistant to topoisomerase I (Top1) inhibitors from the camptothecin (CPT) family, but the mechanistic bases of this resistance have not been investigated in details. Top1 is a nuclear enzyme that induces transient single strand breaks in duplex DNA to ensure the removal of torsional constraints associated with crucial DNA transactions such as replication, transcription, chromosome segregation or DNA recombination. We have previously shown that interaction of Top1 with DNA-PKcs, a kinase involved in non-homologous end-joining, could regulate the cellular response to CPT independently of DNA repair. This was further confirmed by the fact that NU7441, a specific inhibitor of the kinase activity of DNA-PKcs, had no effect on Hela cell sensitivity to CPT. Recently, it was shown that, in PCa cells, DNA-PKcs could interact with ERG, a transcription factor of the ETS family. As advanced PCa are often characterized by a TMPRSS2-ERG fusion leading to wild-type ERG overexpression, we investigated whether ERG could play a role in the resistance of PCa cells to CPT by regulating Top1/DNA-PKcs interaction. We showed that VCaP cells harboring the TMPRSS2-ERG fusion resulting in a 6-fold increase of ERG protein as compared to ERG fusion-negative (control) DU145 or PC3 cells were >100-fold more resistant to CPT. RWPE-1 stably overexpressing low levels of wild-type ERG (2-fold increase) did not show a major change in resistance to CPT as compared to control RWPE-1 cells stably expressing lacZ. Using two specific siRNA, we also showed that transient downregulation of ERG drastically reduced the sensitivity of VCaP cells to CPT, this effect being dependent on the efficacy of ERG repression. Interestingly, this effect was not due to a change in Top1 levels, suggesting that ERG downregulation may affect Top1-DNA-PKcs interaction. We detected this interaction in VCaP cells by immunoprecipitation in the presence of ethidium bromide, indicating that it was independent of DNA and it remains to be seen whether it is disrupted following ERG repression. Our preliminary findings provide evidences for a new mechanism by which ERG and its interaction with DNA-PKcs could explain, at least in part, the intrinsic resistance of PCa to Top1 inhibitors by the regulation of Top1-DNA-PKcs interaction. Citation Format: Emmanuel Roche, Daniele Montaudon, Samer Kayali, Nadine Houede, Philippe Pourquier. Role of the ERG transcription factor in the resistance of prostate cancer cells to the topoisomerase I inhibitor camptothecin. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3813. doi:10.1158/1538-7445.AM2014-3813

Abstract 4478: SHMT2 modulates DNA methylation and differentially affects prostate cancer cell response to platinum derivatives.

Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC Prostate cancer (PCa) is one of the leading causes of death from cancer in men. Several prognostic factors allow differentiating low-grade from high-grade PCa that are often refractory to chemical castration but are still treated with hormone therapy to which docetaxel or cabazitaxel are added when they become resistant to the anti-androgen. Despite many clinical trials with other chemotherapeutic agents, response rates remain low, pointing towards the need for new alternative therapies to treat these aggressive tumors. Using a rational in silico approach based on the NCI60-cell line panel, we recently identified a signature of 6 genes, the expression of which could predict at the functional level, sensitivity to oxaliplatin but not to cisplatin in DU145, LNCaP and C42B prostate cancer cell lines (Puyo et al. Mol. Pharmacol, 2012). Among them, we focused on SHMT2, the mitochondrial isoform of serine hydroxymethyl transferase involved in the biosynthesis of purines. Downregulation of SHMT2 or absence of SHMT2 catalytic activity was associated with a resistance to oxaliplatin, whereas it had no effect on cisplatin sensitivity, a selectivity that was attributed to the DACH moiety of platinum derivatives. Here, we investigated the role of DNA methylation in this selective response to platinum compounds since SHMTs can indirectly regulate the methylation status of DNA. Variations in global level of methylation as measured by the methylation status of LINE-1 retrotransposon, was associated with differences in sensitivity of PCa cells to platinum compounds. Using our in silico approach, we found significant correlations between SHMT2 expression and cell sensitivity to both demethylating agents azacytidine and decitabine. We also found that treatment of DU145 cells, with high level of global DNA methylation, sensitized cells to platinum compounds. In order to evaluate whether methylation could impair the formation of Pt-adducts in vitro, we used purified oligonucleotides containing a unique site of platination. We show that methylation at specific CpG in the vicinity of the platination site could reduce the kinetics of DNA adducts formation. This inhibition was more pronounced for DACH platin than for cisplatin. We also assessed the effect of transient repression of SHMT2 in LNCaP cells on the methylation status of ∼450,000 CpG sites using the Infinium Meth450K beadchip from Illumina. Preliminary results show that significant changes in methylation status was only observed in a reduced number of CpGs which were not located in genes that are known to be involved in cell response to Pt adducts formation. Together our results demonstrate that SHMT2-mediated specific response to oxaliplatin can be due to both methylation in the vicinity of the platination site and the modulation of the expression of genes that are not directly linked to the processing of DNA-adducts. Citation Format: Stephane Puyo, Nadine Houede, Marina Hamant, Pierre Richaud, Jacques Robert, Philippe Pourquier. SHMT2 modulates DNA methylation and differentially affects prostate cancer cell response to platinum derivatives. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4478. doi:10.1158/1538-7445.AM2013-4478

Abstract 5064: A gene expression signature which could predict high grade prostate cancer response to oxaliplatin

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL Prostate cancer is one of the leading causes of death from cancer in men. The prognosis of patients is defined according to staging, PSA levels and Gleason score, which differentiates low grade (Gleason < 4) and high grade (Gleason ≥ 4) cancers. High-grade tumors are associated with high metastatic potential and poor prognosis. They are treated with hormone therapy, to which microtubule poisons are added when they become resistant to the anti-androgen. Despite many clinical trials with other chemotherapeutic agents, response rates remain low. Moreover, none of these trials took into account the tumor grade. We therefore envisaged a new rational in silico approach to screen for drug candidates that could be used as an alternative to docetaxel, based on an expression signature of 86 genes that could distinguish low-grade and high-grade tumors, with a reliability of 81% (PNAS 2006 103:10991-6). We explored the NCI databases, which allow access to both gene expression profiles of 60 human tumor cell lines and their in vitro sensitivity to thousands of anticancer drugs and extracted the expression profiles of the 86 genes’ signature. We calculated for each gene the Pearson coefficients of correlation (r) between their expression level in the 60 cell lines and cell sensitivity to 152 core anticancer compounds. We found that the expression of 11 genes was associated with sensitivity to oxaliplatin. They include DPM1, PCCB, ATP5G3, and SHMT2 genes involved in metabolism; RHOT2, CD59 and JUN genes involved in signal transduction, the CDKN2C cyclin-dependent kinase inhibitor gene, RPL13 and EIF4A1 genes involved in translation, and the unknown [FLJ35093][1] gene. This signature seems specific to DACH platinum derivatives since no correlation was found with the sensitivity to cisplatin. Functional validation of this signature was performed in vitro using the prostate cancer cell lines DU145 and LNCaP and the benign prostate hyperplasia (BPH) cells as “normal” cells. We measured the effect of siRNA-mediated downregulation of each of the 11 genes on the sensitivity to oxaliplatin or cisplatin by clonogenic or MTT assays. Preliminary data confirmed our in silico results for 7 out of the 11 genes, for which donwregulation induced a significant change in IC50 for oxaliplatin (but not cisplatin) in DU145 and/or LNCaP cells. For EIF4A1 we found a correlation opposite to that obtained from in silico results. Also, inhibition of CDKN2C could affect the sensitivity to cisplatin in LNCaP cells. Conversely, no effect of each siRNA tested could be observed in BPH cells, suggesting that functional validation should be tested in a tumoral background. Our new rational approach allowed the identification of oxaliplatin as an alternative therapy for high grade prostate cancers. It also identified a potential gene expression signature that could be used to predict tumor response to oxaliplatin (and potentially DACH platinum derivatives) in the clinic. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5064. doi:10.1158/1538-7445.AM2011-5064 [1]: /lookup/external-ref?link_type=GENPEPT&access_num=FLJ35093&atom=%2Fcanres%2F71%2F8_Supplement%2F5064.atom

Abstract 2332: The new protein FAM110B regulates the expression of E-cadherin and is involved in cell migration and invasion

FAM110B was first identified in a functional screen using genetic suppressor elements to identify new genes involved in the sensitivity to topoisomerase II poisons (Gros et al. Cancer Res, 2003 63: 164-71). FAM110B is extremely conserved in vertebrates. It shares strong sequence homologies with the two other members of this new family (FAM110A & C), but has no known functional domains. We generated a polyclonal antibody and showed that endogenous FAM110B had a MW of ≈40 kDa and was expressed in various cancer cell lines with a predominant cytoplasmic localization. To identify its biological function, we evaluate the different genes/pathways that it could modulate, using specific lentiviral shRNA to target FAM110B in HeLa cells. We found that transient downregulation of FAM110B induced a net decrease in cell proliferation and cell survival. It also induced a change in cell morphology which was accompanied by an increase in E-cadherin and a decrease in N-cadherin expression which are two markers of a reverse epithelial to mesenchymal (EMT) transition phenotype. We also measured the levels of SNAIL and SLUG protein levels, two known repressors of E-cadherin expression and found that only SLUG was decreased in shFAM110B-transfected cells, suggesting that regulation of E-cadherin expression by FAM110B involves, at least in part, its transcriptional repression by SLUG. Increased expression of E-cadherin was confirmed at the mRNA level by microarray analyses comparing control cells to shFAM110B-transfected cells. This analysis also revealed the presence of various genes involved in EMT among the 1000 most differentially expressed genes (adjusted t-value > 7 or Our data were confirmed at the functional level by analyzing the effects of FAM110B repression on the motility/migratory behaviour of HeLa cells using the wound model. While cells expressing control shRNA were able to invade the wound within 7-10h, cells with reduced levels of FAM110B had barely started to move towards the wound. Similarly, shFAM110B-transfected cells displayed a 2-fold decrease in their capacity to invade and migrate through collagen gels as compared to control cells. Using the Cignal Finder Cancer 10-Pathway Reporter Assay, we assess whether other signalling pathways could be deregulated following FAM110B downregulation. We found that Wnt, TGF-β and the Notch pathways, which are all involved in EMT, were significantly downregulated in shFAM110B-transfected cells, which is in accordance with our previous observations. Together, our results provide new evidences on the potential role of FAM110B in the regulation of E-cadherin expression and more generally in crucial cellular processes such as cell motility, migration, invasion and metastasis which makes it a new potential target for cancer therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2332. doi:10.1158/1538-7445.AM2011-2332

Abstract 1639: Targeting the p38 MAPK pathway inhibits irinotecan resistance in colon adenocarcinoma

Despite recent advances in the treatment of colon cancer, tumor resistance is a frequent cause of chemotherapy failure. To better elucidate the molecular mechanisms involved in resistance to irinotecan (and its active metabolite SN38), we established SN38-resistant clones derived from the colon adenocarcinoma HCT-116 and SW48 cell lines. These clones show various levels (6 to 60 fold) of resistance to SN-38 and display enhanced levels of activated MAPK p38 as compared to the corresponding parental cells. Because four different isoforms of p38 have been described, we then studied the effect of p38 overexpression or downregulation of each isoform on cell sensivity to SN38 and found that both alpha and beta isoforms are involved in the development of resistance to SN38. In this line, we show that cell treatment with SB202190, which specifically inhibits p38 alpha and beta, enhanced the cytotoxic activity of SN38, but not of 5-FU or oxaliplatin. Moreover, p38 inhibition sensitized tumor cells derived from both SN38-sensitive and -resistant HCT116 cells to irinotecan treatment in xenograft models. Finally, we detected less phosphorylated p38 in primary colon cancer of patients sensitive to irinotecan-based treatment, compared to non-responder patients. This indicates that enhanced level of expression of phosphorylated p38 could predict the absence of clinical response to irinotecan. Altogether, our results show for the first time that the p38 MAPK pathway is involved in irinotecan sensitivity and suggest that phosphorylated p38 expression level could be used as a marker of clinical resistance to irinotecan. They further suggest that targeting the p38 pathway may be a potential strategy to overcome resistance to irinotecan-based chemotherapies in colorectal cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1639. doi:10.1158/1538-7445.AM2011-1639

Abstract 2699: ERCC5 (XPG) status and clinical activity of trabectedin in patients with advanced soft-tissue sarcoma

Trabectedin is approved in Europe for the treatment of advanced soft tissue sarcoma (STS) and provides objective response and disease stabilisation rates ranging from 5%-10% and 30-40%, respectively. Although the precise mechanism of action of trabectedin is not fully elucidated, various in vitro studies have shown that its activity depends, at least in part, on the nucleotide excision repair (NER) status of the cells. Indeed, NER-deficient cell lines are less sensitive to trabectedin than their wild-type counterparts. Our aim was to determine whether the status of ERCC5 (XPG), a 3′-endonuclease which plays a crucial role in the excision of the damaged DNA, was associated with the clinical activity of trabectedin in advanced STS patients. We analyzed both, the single nucleotide polymorphism (Asp1104His, G>C, exon 15) and the expression status (real-time quantitative RT-PCR) of ERCC5 in tumors from a cohort of 119 patients with advanced STS (median age at the start of treatment: 49 years old). All patients included in this retrospective analysis were treated between 1999 and 2007 in phase I-II clinical trials or in the context of a compassionate-use program. Trabectedin was given at different doses (0.5-3 mg/m2) with either a 3-h infusion or a 24-h continuous infusion schedule. The two most frequent histological subtypes were leiomyosarcoma (non-uterine: 21, uterine: 11) and liposarcoma (myxoid round cell: 24, other: 15). Overall, tumour control (CR, PR, and SD ≥ 6 months) was achieved in 42 patients (35%, 95% CI 26-44). The median progression-free survival (PFS) and overall survival (OS) of the entire patient group were 3.3 months (95% CI 2.5-4.1) and 12 months (95% CI 7.8-16.1), respectively. Variant allele carriers for ERCC5 were found in 58 cases: G/C (40; 33%), C/C (17; 15%). ERCC5 mRNA was found to be overexpressed in 48% of analyzed cases. In patients with tumors homozygous for the wild-type allele (Asp), ERRC5 mRNA overexpression was associated with significantly higher median PFS: 17.1 months (95% CI 4.7-29.4) versus 1.8 months (95% CI 1.4-2.2), p=0.002. In the group of patients with tumors heterozygous or homozygous for the variant allele (His), median PFS was poor regardless to the expression status of ERRC5 mRNA: 2.8 months (95% CI 1.3-4.2) versus 3.4 months (95% CI 1.4-5.5), p=0.50. We also compared the effect of wild-type and variant ERCC5 expression in human 94RD27 XPG-deficient fibroblasts (deletion of the last 261 aa) on the sensitivity to trabectedin. Preliminary results show that cells stably expressing wild-type ERCC5 are more sensitive to trabectedin than cells expressing the variant allele of ERCC5. Altogether, our data suggest that overexpression of wild-type ERCC5 might be predictive of clinical benefit from trabectedin in advanced STS patients. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2699.