Personalized follow-up of circulating DNA in resected stage III/IV melanoma: PERCIMEL multicentric prospective study protocol.

BACKGROUND: With more than 15,000 new cases /year in France and 2,000 deaths, cutaneous melanoma represents approximately 4% of incidental cancers and 1.2% of cancer related deaths. In locally advanced (stage III) or resectable metastatic (stage IV) melanomas, medical adjuvant treatment is proposed and recent advances had shown the benefit of anti-PD1/PDL1 and anti-CTLA4 immunotherapy as well as anti-BRAF and anti-MEK targeted therapy in BRAF V600 mutated tumors. However, the recurence rate at one year is approximately 30% and justify extensive research of predictive biomarkers. If in metastatic disease, the follow-up of circulating tumor DNA (ctDNA) has been demonstrated, its interest in adjuvant setting remains to be precised, especially because of a lower detection rate. Further, the definition of a molecular response could prove useful to personalized treatment. METHODS: PERCIMEL is an open prospective multicentric study executed through collaboration of the Institut de Cancérologie de Lorraine (non-profit comprehensive cancer center) and 6 French university and community hospitals. A total of 165 patients with resected stage III and IV melanoma, eligible to adjuvant imunotherapy or anti-BRAF/MEK kinase inhibitors will be included. The primary endpoint is the presence of ctDNA, 2 to 3 weeks after surgery, defined as mutated ctDNA copy number calculated as the allelic fraction of a clonal mutation relative to total ctDNA. Secondary endpoints are recurrence-free survival, distant metastasis-free survival and specific survival. We will follow ctDNA along treatment, quantitatively through ctDNA mutated copy number variation, qualitatively through the presence of cfDNA and its clonal evolution. Relative and absolute variations of ctDNA during follow-up will be also analyzed. PERCIMEL study aims at provide scientific evidence that ctDNA quantitative and qualitative variations can be used to predict the recurrence of patients with melanoma treated with adjuvant immunotherapy or kinase inhibitors, thus defining the notion of molecular recurrence.

Cell-Free DNA in the Management of Colorectal Cancer.

Colorectal cancer is one of the leading cause of death by cancer worldwide in both men and women. Liquid biopsy belongs nowadays to the landscape of cancer management biological tools. In this chapter, we will describe and discuss the actual, potential and future applications of cfDNA analysis in plasma of patients with colorectal cancer in early or metastatic stage. During the last decade, the development of molecular biology assays like digital PCR or next-generation sequencing made the analysis of cfDNA in plasma possible with an excellent sensitivity and applications like early detection, diagnosis, prognosis, response to treatment, monitoring of an emerging resistance, mapping of the disease molecular landscape or evaluation of the residual disease are now feasible. cfDNA detection has several promising applications in the management of patients with colorectal cancer, but prospective randomised clinical trials are still lacking to make liquid biopsy ready for prime-time.

Identification of Specific Tumor Markers in Vulvar Carcinoma Through Extensive Human Papillomavirus DNA Characterization Using Next Generation Sequencing Method.

OBJECTIVES: A subset of vulvar carcinomas (VC) are associated with human papillomavirus (HPV) DNA. This trait can be used to identify tumor markers for patient's follow-up. A large diversity of HPV prevalence in VC has been reported, but no data are available concerning the insertional HPV status in this tumor type. Therefore, we have used an innovative next generation sequencing (NGS)-based CaptHPV method able to provide an extensive characterization of HPV DNA in tumors. MATERIAL AND METHODS: Tumor tissue specimens from 55 patients with VC were analyzed using p16 immunohistochemistry, in situ hybridization, polymerase chain reaction, and CaptHPV-NGS assays. RESULTS: Our analyses showed that 8 (14.5%) of 55 cases were associated with HPV 16 DNA. No other HPV genotypes were identified. The HPV genome was in a free episomal state only in one case and both episomal and integrated into the tumor cell genome in 7. There was a single insertion in 5 cases and multiple sites, scattered at different chromosomal loci in two. ISH data suggest that some of these might reflect tumor heterogeneity. Viral integration targeted cellular genes among which were TP63, CCDC148, LOC100133091, PKP1, and POLA2. Viral integration at the PKP1 locus was associated with partial gene deletion, and no PKP1 protein was detected in tumor tissue. CONCLUSIONS: Using the NGS-based innovative capture-HPV approach, we established a cartography of HPV 16 DNA in 8 VC cases and identified novel genes targeted by integration that may be used as specific tumor markers. In addition, we established a rationale strategy for optimal characterization of HPV status in VC.

SMAD4 and the TGFß Pathway in Patients with Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of cancer death worldwide. PDAC is an aggressive disease with an 11-month median overall survival and a five-year survival of less than 5%. Incidence of PDAC is constantly increasing and is predicted to become the second leading cause of cancer in Western countries within a decade. Despite research and therapeutic development, current knowledge about PDAC molecular mechanisms still needs improvements and it seems crucial to identify novel therapeutic targets. Genomic analyses of PDAC revealed that transforming growth factor ß (TGFß) signaling pathways are modified and the SMAD4 gene is altered in 47% and 60% of cases, respectively, highlighting their major roles in PDAC development. TGFß can play a dual role in malignancy depending on the context, sometimes as an inhibitor and sometimes as an inducer of tumor progression. TGFß signaling was identified as a potent inducer of epithelial-to-mesenchymal transition (EMT), a process that confers migratory and invasive properties to epithelial cells during cancer. Therefore, aberrant TGFß signaling and EMT are linked to promoting PDAC aggressiveness. TGFß and SMAD pathways were extensively studied but the mechanisms leading to cancer promotion and development still remain unclear. This review aims to describe the complex role of SMAD4 in the TGFß pathway in patients with PDAC.

HPV insertional pattern as a personalized tumor marker for the optimized tumor diagnosis and follow-up of patients with HPV-associated carcinomas: a case report.

BACKGROUND: In clinical oncology, only a few applications have been developed using HPV as a personalized tumor marker, a lack most probably related to the limited information obtained by the classical Polymerase Chain Reaction (PCR) approach. To overcome this limitation, we have recently developed the capture-based Next-Generation Sequencing (NGS) "CaptHPV" assay, designed to provide an extensive and comprehensive molecular characterization of HPV DNA sequences associated with neoplasias, ie the sequence of the viral genome (245 genotypes), its physical state, viral load, integration site and genomic alterations at integration locus. These data correspond to highly specific tumor markers that can be used to improve diagnosis and patient's follow-up. CASE PRESENTATION: We report here a case that is a straightforward and practical illustration of the power of the CaptHPV method. A patient developed successively a carcinoma of the anal canal and of the tongue. The two tumors were squamous cell carcinoma, found associated with HPV16 using PCR. In order to document a possible metastasis to the tongue from the anal cancer, we performed CaptHPV analysis on the two tumors. The analysis of the anal carcinoma found 55 viral/human hybrid reads allowing the identification of the HPV16 DNA integration in the 4q25 chromosomal band locus with a 178,808 bp deletion in the cell genome. Molecular analysis of the tongue tumor disclosed 6110 reads of HPV16, with a viral pattern strictly identical to that of the anal tumor. A total of 131 hybrid reads between HPV16 and the cell genome were found, corresponding exactly to the same locus of integration of viral DNA at the 4q25 site. The 178,808 bp genomic deletion was also found in the lingual tumor. The exact identity of HPV insertional signatures in the two tumors, demonstrates unambiguously that the tongue tumor derived from the anal cancer whereas neither histological immunophenotyping nor classical viral analysis using PCR could allow a definitive diagnosis. CONCLUSION: Our observation indicates that the establishment of a detailed cartography of HPV DNA sequences in a tumor specimen provides crucial information for the design of specific biomarkers that can be used for diagnostic, prognostic or predictive purposes.

Emerging Roles of DDB2 in Cancer.

Damage-specific DNA-binding protein 2 (DDB2) was originally identified as a DNA damage recognition factor that facilitates global genomic nucleotide excision repair (GG-NER) in human cells. DDB2 also contributes to other essential biological processes such as chromatin remodeling, gene transcription, cell cycle regulation, and protein decay. Recently, the potential of DDB2 in the development and progression of various cancers has been described. DDB2 activity occurs at several stages of carcinogenesis including cancer cell proliferation, survival, epithelial to mesenchymal transition, migration and invasion, angiogenesis, and cancer stem cell formation. In this review, we focus on the current state of scientific knowledge regarding DDB2 biological effects in tumor development and the underlying molecular mechanisms. We also provide insights into the clinical consequences of DDB2 activity in cancers.

Dual Epigenetic Regulation of ERα36 Expression in Breast Cancer Cells.

Breast cancer remains the major cause of cancer-induced morbidity and mortality in women. Among the different molecular subtypes, luminal tumors yet considered of good prognosis often develop acquired resistance to endocrine therapy. Recently, misregulation of ERα36 was reported to play a crucial role in this process. High expression of this ERα isoform was associated to preneoplastic phenotype in mammary epithelial cells, disease progression, and enhanced resistance to therapeutic agents in breast tumors. In this study, we identified two mechanisms that could together contribute to ERα36 expression regulation. We first focused on hsa-miR-136-5p, an ERα36 3'UTR-targeting microRNA, the expression of which inversely correlated to the ERα36 one in breast cancer cells. Transfection of hsa-miR136-5p mimic in MCF-7 cells resulted in downregulation of ERα36. Moreover, the demethylating agent decitabine was able to stimulate hsa-miR-136-5p endogenous expression, thus indirectly decreasing ERα36 expression and counteracting tamoxifen-dependent stimulation. The methylation status of ERα36 promoter also directly modulated its expression level, as demonstrated after decitabine treatment of breast cancer cell and confirmed in a set of tumor samples. Taken together, these results open the way to a direct and an indirect ERα36 epigenetic modulation by decitabine as a promising clinical strategy to counteract acquired resistance to treatment and prevent relapse.

Nuclear factor I X is a recurrent target for HPV16 insertions in anal carcinomas.

Anal carcinomas (AC) are associated with human papillomavirus (HPV) DNA sequences, but little is known about the physical state of the viral genome in carcinoma cells. To define the integration status and gene(s) targeted by viral insertions in AC, tumor DNAs extracted from 35 tumor specimen samples in patients with HPV16-associated invasive carcinoma were analyzed using the detection of integrated papillomavirus sequences-PCR approach. The genomic status at integration sites was assessed using comparative genomic hybridization-array assay and gene expression using reverse transcription quantitative PCR (RT-qPCR). HPV16 DNA was found integrated in 25/35 (71%) cases and the integration locus could be determined at the molecular level in 19 cases (29 total integration loci). HPV DNA was inserted on different chromosomes, but 5 cases harbored viral sequences at 19p13.2, within the nuclear factor I X (NFIX) locus. Viral DNA mapped between the most distal and the two proximal alternatively expressed exons of this gene in three cases (CA21, CA04, and CA35) and upstream of this gene (663 kb and 2.3 Mb) in the others. CGH arrays showed genomic gains/amplifications at the NFIX region, associated with HPV within the gene and RT-qPCR, revealed NFIX mRNA overexpression. Other genes targeted by integration were IL20RB, RPS6KA2, MSRA1, PIP5K1B, SLX4IP, CECR1, BCAR3, ATF6, CSNK1G1, APBA2, AGK, ILF3, PVT1, TRMT1, RAD51B, FASN, CCDC57, DSG3, and ZNF563. We identified recurrent targeting of NFIX by HPV16 insertion in anal carcinomas, supporting a role for this gene in oncogenesis, as reported for non-HPV tumors.

Tumor-derived cell-free DNA and circulating tumor cells: partners or rivals in metastasis formation?

The origin of metastases is a topic that has sparked controversy. Despite recent advancements, metastatic disease continues to pose challenges. The first admitted model of how metastases develop revolves around cells breaking away from the primary tumor, known as circulating tumor cells (CTCs). These cells survive while circulating through the bloodstream and subsequently establish themselves in secondary organs, a process often referred to as the "metastatic cascade". This intricate and dynamic process involves various steps, but all the mechanisms behind metastatic dissemination are not yet comprehensively elucidated. The "seed and soil" theory has shed light on the phenomenon of metastatic organotropism and the existence of pre-metastatic niches. It is now established that these niches can be primed by factors secreted by the primary tumor before the arrival of CTCs. In particular, exosomes have been identified as important contributors to this priming. Another concept then emerged, i.e. the "genometastasis" theory, which challenged all other postulates. It emphasizes the intriguing but promising role of cell-free DNA (cfDNA) in metastasis formation through oncogenic formation of recipient cells. However, it cannot be ruled out that all these theories are intertwined. This review outlines the primary theories regarding the metastases formation that involve CTCs, and depicts cfDNA, a potential second player in the metastasis formation. We discuss the potential interrelationships between CTCs and cfDNA, and propose both in vitro and in vivo experimental strategies to explore all plausible theories.

Prediction of Adjuvant Gemcitabine Sensitivity in Resectable Pancreatic Adenocarcinoma Using the GemPred RNA Signature: An Ancillary Study of the PRODIGE-24/CCTG PA6 Clinical Trial.

PURPOSE: GemPred, a transcriptomic signature predictive of the efficacy of adjuvant gemcitabine (GEM), was developed from cell lines and organoids and validated retrospectively. The phase III PRODIGE-24/CCTG PA6 trial has demonstrated the superiority of modified folinic acid, fluorouracil, irinotecan, and oxaliplatin (mFOLFIRINOX) over GEM as adjuvant therapy in patients with resected pancreatic ductal adenocarcinoma at the expense of higher toxicity. We evaluated the potential predictive value of GemPred in this population. PATIENTS AND METHODS: Routine formalin-fixed paraffin-embedded surgical specimens of 350 patients were retrieved for RNA sequencing and GemPred prediction (167 in the GEM arm and 183 in the mFOLFIRINOX [mFFX] arm). Survival analyses were stratified by resection margins, lymph node status, and cancer antigen 19-9 level. RESULTS: Eighty-nine patients' tumors (25.5%) were GemPred+ and were thus predicted to be gemcitabine-sensitive. In the GEM arm, GemPred+ patients (n = 50, 30%) had a significantly longer disease-free survival (DFS) than GemPred- patients (n = 117, 70%; median 27.3 v 10.2 months, hazard ratio [HR], 0.43 [95% CI, 0.29 to 0.65]; P < .001) and cancer-specific survival (CSS; median 68.4 v 28.6 months, HR, 0.42 [95% CI, 0.27 to 0.66]; P < .001). GemPred had no prognostic value in the mFFX arm. DFS and CSS were similar in GemPred+ patients who received adjuvant GEM and mFFX (median 27.3 v 24.0 months, and 68.4 v 51.4 months, respectively). The statistical interaction between GEM and GemPred+ status was significant for DFS (P = .008) and CSS (P = .004). GemPred+ patients had significantly more adverse events of grade ≥3 in the mFFX arm (76%) compared with those in the GEM arm (40%; P = .001). CONCLUSION: This ancillary study of a phase III randomized trial demonstrates that among the quarter of patients with a GemPred-positive transcriptomic signature, survival was comparable with that of mFOLFIRINOX, whereas those receiving adjuvant gemcitabine had fewer adverse events.

ESR1 Gene Mutations and Liquid Biopsy in ER-Positive Breast Cancers: A Small Step Forward, a Giant Leap for Personalization of Endocrine Therapy?

The predominant forms of breast cancer (BC) are hormone receptor-positive (HR+) tumors characterized by the expression of estrogen receptors (ERs) and/or progesterone receptors (PRs). Patients with HR+ tumors can benefit from endocrine therapy (ET). Three types of ET are approved for the treatment of HR+ BCs and include selective ER modulators, aromatase inhibitors, and selective ER downregulators. ET is the mainstay of adjuvant treatment in the early setting and the backbone of the first-line treatment in an advanced setting; however, the emergence of acquired resistance can lead to cancer recurrence or progression. The mechanisms of ET resistance are often related to the occurrence of mutations in the ESR1 gene, which encodes the ER-alpha protein. As ESR1 mutations are hardly detectable at diagnosis but are present in 30% to 40% of advanced BC (ABC) after treatment, the timeline of testing is crucial. To manage this resistance, ESR1 testing has recently been recommended; in ER+ HER2- ABC and circulating cell-free DNA, so-called liquid biopsy appears to be the most convenient way to detect the emergence of ESR1 mutations. Technically, several options exist, including Next Generation Sequencing and ultra-sensitive PCR-based techniques. In this context, personalization of ET through the surveillance of ESR1 mutations in the plasma of HR+ BC patients throughout the disease course represents an innovative way to improve the standard of care.

Integrated Molecular Characterization of HER2-Low Breast Cancer Using Next Generation Sequencing (NGS).

Based on immunohistochemistry (IHC) and in situ hybridization (ISH), HER2-low breast cancers (BC) subtype-defined as IHC1+ or IHC2+/ISH- tumors-emerged and represent more than half of all BC. We evaluated the performance of NGS for integrated molecular characterization of HER2-low BC, including identification of actionable molecular targets, copy number variation (CNV), and microsatellite instability (MSI) analysis. Thirty-one BC specimens (11 HER2+, 10 HER2-, and 10 HER2-low) were routinely analyzed using IHC and ISH, and were selected and analyzed using NGS for gene mutations including ESR1, PIK3CA, AKT1, ERBB2, TP53, BRCA1, and BRCA2, CNV, and MSI. CNV values for the ERBB2 gene were significantly (p < 0.001) different between HER2+, and either HER2-low or HER2- tumors with mean values of 7.8 (SD = 6.8), 1.9 (SD = 0.3), and 2.0 (SD = 0.3), respectively. Using 3.25 as the cutoff value, 96.8% overall concordance of HER2 status was achieved between IHC and NGS compared to IHC and ISH. Using NGS, gene mutations and amplifications were detected in 68% (21/31) and 19% (6/31) of the cases, respectively. One case of MSI was detected in a HER2-negative and ISH unamplified case. Beside IHC, NGS allows the identification of HER2-low subtype simultaneously, with the detection of multiple actionable gene mutations being helpful for molecular board treatment selection.

CRISPR/Cas9-mediated knock-in of BRCA1/2 mutations restores response to olaparib in pancreatic cancer cell lines.

Pancreatic cancer is one of the most aggressive diseases with a very poor outcome. Olaparib, a PARP inhibitor, as maintenance therapy showed benefits in patients with metastatic pancreatic adenocarcinoma bearing germline BRCA1/2 mutations. However, germline BRCA mutation has been described in only 4-7% of patients with pancreatic adenocarcinoma. A CRISPR/Cas9-mediated system was used to knock-in the c.763G > T p.(Glu255*) and c.2133C > A p.(Cys711*) mutations in cell lines to obtain truncated BRCA1 and BRCA2 proteins, respectively. A CRISPR/Cas9 ribonucleoprotein complex was assembled for each mutation and transfected into two pancreatic cell lines (T3M4 and Capan-2) and into a breast cancer cell lines (MCF7) as control. BRCA protein levels were significantly decreased in all BRCA-depleted cells (P < 0.05), proving the transfection efficiency of our CRISPR/Cas9 systems. As expected, the calculated olaparib IC50 were significantly reduced for all cell lines harbored BRCA1 or BRCA2 mutations compared to wild-type BRCA1/2 cells (P < 0.01). Furthermore, we observed a higher induction of apoptosis after 72 h olaparib treatment in BRCA-depleted cells than in wild-type cells. This strategy might offer new insights into the management of patients with pancreatic cancer and open up new perspectives based on the in vivo use of CRISPR/Cas9 strategy.

Validation of the Idylla GeneFusion assay to detect fusions and MET exon-skipping in non-small cell lung cancers.

Gene fusions and MET exon skipping drive oncogenesis in 8-9% and 3% of non-small cell lung cancers (NSCLC) respectively. Their detection are essential for the management of patients since they confer sensitivity to specific targeted therapies with significant clinical benefit over conventional chemotherapy. Immunohistochemistry (IHC) and fluorescent in situ hybridization (FISH) account for historical reference techniques however molecular-based technologies (RNA-based sequencing and RT-PCR) are emerging as alternative or complementary methods. Here, we evaluated the analytical performance of the fully-automated RT-PCR Idylla GeneFusion assay compared to reference methods using 35 fixed NSCLC samples. Idylla demonstrated overall agreement, sensitivity and specificity of 100% compared to RNASeq. Interestingly, it succeeded in retrieving 10 out of 11 samples with inconclusive results due to insufficient RNA quality for sequencing. Idylla showed an overall agreement, sensitivity and specificity of 90.32%, 91.67% and 89.47% compared to IHC/FISH respectively. Using commercial standards, the limit of detection of the Idylla system for the most frequent fusions and exon skipping ranges between 5 and 10 ng RNA input. These results support that the Idylla assay is a reliable and rapid option for the detection of these alterations, however a particular attention is needed for the interpretation of the expression imbalance.

GIInger predicts homologous recombination deficiency and patient response to PARPi treatment from shallow genomic profiles.

Homologous recombination deficiency (HRD) is a predictive biomarker for poly(ADP-ribose) polymerase 1 inhibitor (PARPi) sensitivity. Routine HRD testing relies on identifying BRCA mutations, but additional HRD-positive patients can be identified by measuring genomic instability (GI), a consequence of HRD. However, the cost and complexity of available solutions hamper GI testing. We introduce a deep learning framework, GIInger, that identifies GI from HRD-induced scarring observed in low-pass whole-genome sequencing data. GIInger seamlessly integrates into standard BRCA testing workflows and yields reproducible results concordant with a reference method in a multisite study of 327 ovarian cancer samples. Applied to a BRCA wild-type enriched subgroup of 195 PAOLA-1 clinical trial patients, GIInger identified HRD-positive patients who experienced significantly extended progression-free survival when treated with PARPi. GIInger is, therefore, a cost-effective and easy-to-implement method for accurately stratifying patients with ovarian cancer for first-line PARPi treatment.

Deciphering Tumour Heterogeneity: From Tissue to Liquid Biopsy.

Human solid malignancies harbour a heterogeneous set of cells with distinct genotypes and phenotypes. This heterogeneity is installed at multiple levels. A biological diversity is commonly observed between tumours from different patients (inter-tumour heterogeneity) and cannot be fully captured by the current consensus molecular classifications for specific cancers. To extend the complexity in cancer, there are substantial differences from cell to cell within an individual tumour (intra-tumour heterogeneity, ITH) and the features of cancer cells evolve in space and time. Currently, treatment-decision making usually relies on the molecular characteristics of a limited tumour tissue sample at the time of diagnosis or disease progression but does not take into account the complexity of the bulk tumours and their constant evolution over time. In this review, we explore the extent of tumour heterogeneity with an emphasis on ITH and report the mechanisms that promote and sustain this diversity in cancers. We summarise the clinical strikes of ITH in the management of patients with cancer. Finally, we discuss the current material and technological approaches that are relevant to adequately appreciate ITH.

Intraoperative prediction of non-sentinel lymph node metastases in breast cancer using cytokeratin 19 mRNA copy number: A retrospective analysis.

One-step nucleic acid amplification (OSNA) is a molecular procedure used intraoperatively for the detection of sentinel lymph node (SLN) metastases. The aim of the present study was to define a cut-off of cytokeratin (CK)19 mRNA copy number predictive of positive completion axillary lymph node dissection (ALND). The OSNA procedure was employed for SLN analysis in 812 patients with T1-T2 N0 breast cancer. A total of 197 patients with SLN metastases were retrospectively analyzed. A total of 40 patients (20%) had non-SLN metastases. Receiver operating characteristics curve analysis established a cut-off of 5,000 CK19 mRNA copy number with 75% sensitivity and 72% specificity. The positive and negative predictive values were 40.5 and 92%, respectively. Multivariate analysis showed that this cut-off and tumor localization in the outer or lower-outer quadrant of the breast were significantly associated with non-SNL involvement (P<0.001 and P=0.025, respectively). The findings of the present study support the conventional cut-off of 5,000 copies for intraoperative decision to perform ALND, whereas ALND can safely be avoided in patients with tumor located outside the outer or lower-outer quadrant of the breast if the CK19 mRNA copy number is <5,000.

Development of a Clinical-Biological Model to Assess Tumor Progression in Metastatic Pancreatic Cancer: Post Hoc Analysis of the PRODIGE4/ACCORD11 Trial.

Background: The follow-up of pancreatic cancer (PC) is based on computed tomography (CT) assessment; however, there is no consensus on the use of clinical and biological criteria in tumor progression. We aimed to establish a clinical−biological model to highlight the progression of metastatic PC during first-line treatment. Methods: The patients treated with first-line chemotherapy in the phase 2/3 PRODIGE4/ACCORD11 clinical trial were evaluated retrospectively. Clinical and biological markers were evaluated at the time of CT scans and during treatment to determine tumor progression. Results: In total, 196 patients were analyzed, with 355 available tumor assessments. The clinical and biological factors associated with tumor progression in multivariate analysis included gemcitabine, global health status ≤ 33 (OR = 3.38, 95%CI [1.15; 9.91], p = 0.028), quality of life score between 34 and 66 (OR = 2.65, 95%CI [1.06; 6.59], p = 0.037), carcinoembryonic antigen (CEA) ≥ 3 times the standard value without any increase in the CEA level from inclusion (OR = 2.22, 95%CI [1.01; 4.89], p = 0.048) and with an increase in the CEA level from inclusion (OR = 6.56, 95%CI [2.73; 15.78], p < 0.001), and an increase in the carbohydrate antigen 19-9 level from inclusion (OR = 2.59, 95%CI [1.25; 5.36], p = 0.016). Conclusions: The self-assessment of patients' general health status alongside tumor markers is an interesting approach to the diagnosis of the tumor progression of metastatic pancreatic cancer patients during first-line treatment.

DDB2 represses epithelial-to-mesenchymal transition and sensitizes pancreatic ductal adenocarcinoma cells to chemotherapy.

Introduction: Damage specific DNA binding protein 2 (DDB2) is an UV-indiced DNA damage recognition factor and regulator of cancer development and progression. DDB2 has dual roles in several cancers, either as an oncogene or as a tumor suppressor gene, depending on cancer localization. Here, we investigated the unresolved role of DDB2 in pancreatic ductal adenocarcinoma (PDAC). Methods: The expression level of DDB2 in pancreatic cancer tissues and its correlation with patient survival were evaluated using publicly available data. Two PDAC cell models with CRISPR-modified DDB2 expression were developed: DDB2 was repressed in DDB2-high T3M4 cells (T3M4 DDB2-low) while DDB2 was overexpressed in DDB2-low Capan-2 cells (Capan-2 DDB2-high). Immunofluorescence and qPCR assays were used to investigate epithelial-to-mesenchymal transition (EMT) in these models. Migration and invasion properties of the cells were also determined using wound healing and transwell assays. Sensitivity to 5-fluorouracil (5-FU), oxaliplatin, irinotecan and gemcitabine were finally investigated by crystal violet assays. Results: DDB2 expression level was reduced in PDAC tissues compared to normal ones and DDB2-low levels were correlated to shorter disease-free survival in PDAC patients. DDB2 overexpression increased expression of E-cadherin epithelial marker, and decreased levels of N-cadherin mesenchymal marker. Conversely, we observed opposite effects in DDB2 repression and enhanced transcription of SNAIL, ZEB1, and TWIST EMT transcription factors (EMT-TFs). Study of migration and invasion revealed that these properties were negatively correlated with DDB2 expression in both cell models. DDB2 overexpression sensitized cells to 5-fluorouracil, oxaliplatin and gemcitabine. Conclusion: Our study highlights the potential tumor suppressive effects of DDB2 on PDAC progression. DDB2 could thus represent a promising therapeutic target or biomarker for defining prognosis and predicting chemotherapy response in patients with PDAC.