Reshaping preoperative treatment of pancreatic cancer in the era of precision medicine.

This review summarises the recent evidence on preoperative therapeutic strategies in pancreatic cancer and discusses the rationale for an imminent need for a personalised therapeutic approach in non-metastatic disease. The molecular diversity of pancreatic cancer and its influence on prognosis and treatment response, combined with the failure of 'all-comer' treatments to significantly impact on patient outcomes, requires a paradigm shift towards a genomic-driven approach. This is particularly important in the preoperative, potentially curable setting, where a personalised treatment allocation has the substantial potential to reduce pancreatic cancer mortality.

Investigating the feasibility of tumour molecular profiling in gastrointestinal malignancies in routine clinical practice.

Background: Targeted capture sequencing can potentially facilitate precision medicine, but the feasibility of this approach in gastrointestinal (GI) malignancies is unknown. Patients and methods: The FOrMAT (Feasibility of a Molecular Characterisation Approach to Treatment) study was a feasibility study enrolling patients with advanced GI malignancies from February 2014 to November 2015. Targeted capture sequencing (mainly using archival formalin-fixed paraffin-embedded diagnostic/resection samples) was carried out to detect mutations, copy number variations and translocations in up to 46 genes which had prognostic/predictive significance or were targets in current/upcoming clinical trials. Results: Of the 222 patients recruited, 215 patients (96.8%) had available tissue samples, 125 patients (56.3%) had ≥16 genes successfully sequenced and 136 patients (61.2%) had ≥1 genes successfully sequenced. Sample characteristics influenced the proportion of successfully sequenced samples, e.g. tumour type (colorectal 70.9%, biliary 52.6%, oesophagogastric 50.7%, pancreas 27.3%, P = 0.002), tumour cellularity (high versus low: 78.3% versus 13.3%, P ≤ 0.001), tumour content (high versus low: 78.6% versus 27.3%, P = 0.001) and type of sample (resection versus biopsy: 82.4% versus 47.6%, P ≤ 0.001). Currently, actionable alterations were detected in 90 (40.5%) of the 222 patients recruited (66% of the 136 patients sequenced) and 2 patients subsequently received a targeted therapy. The most frequently detected currently actionable alterations were mutations in KRAS, BRAF, TP53 and PIK3CA. For the 205 patients with archival samples, the median time to obtain sequencing results was 18.9 weeks, including a median of 4.9 weeks for sample retrieval and 5.1 weeks for sequencing. Conclusions: Targeted sequencing detected actionable alterations in formalin-fixed paraffin-embedded samples, but tissue characteristics are of critical importance in determining sequencing success. Routine molecular profiling of GI tumours outside of clinical trials is not an effective use of healthcare resources unless more targeted drugs become available. ClinicalTrials.gov identifier: NCT02112357.

Regulation of miR-483-3p by the O-linked N-acetylglucosamine transferase links chemosensitivity to glucose metabolism in liver cancer cells.

The miR-483-3p is upregulated in several tumors, including liver tumors, where it inhibits TP53-dependent apoptosis by targeting the pro-apoptotic gene BBC3/PUMA. The transcriptional regulation of the miR-483-3p could be driven by the ß-catenin/USF1 complex, independently from its host gene IGF2, and we previously demonstrated that in HepG2 hepatoblastoma cells carrying wild-type TP53 the upregulation of the miR-483-3p overcomes the antitumoral effects of the tumor-suppressor miR-145-5p by a mechanism involving cellular glucose availability. Here we demonstrate that in HepG2 cells, the molecular link between glucose concentration and miR-483-3p expression entails the O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT), which stabilizes the transcriptional complex at the miR-483 promoter. HepG2 cells showed reduced miR-483-3p expression and increased susceptibility to 5-fluorouracil (5-FU)-induced apoptosis in presence of the inhibitor of glycolysis 2-deoxy-d-glucose (2-DG). However, in vivo experiments showed that HepG2 cells with higher miR-483-3p expression were selected during tumor progression regardless of 5-FU treatment. Furthermore, treatment with 2-DG alone did not significantly reduce HepG2 xenograft load in immunodeficient mice. In conclusion, we show that in HepG2 cells glucose uptake increases the expression of the oncogenic miR-483-3p through the OGT pathway. This suggests that depletion of the miR-483-3p may be a valuable therapeutic approach in liver cancer patients, but the use of inhibitors of glycolysis to achieve this purpose could accelerate the selection of resistant neoplastic cell clones.

Carbon dating cancer: defining the chronology of metastatic progression in colorectal cancer.

Background: Patients often ask oncologists how long a cancer has been present before causing symptoms or spreading to other organs. The evolutionary trajectory of cancers can be defined using phylogenetic approaches but lack of chronological references makes dating the exact onset of tumours very challenging. Patients and methods: Here, we describe the case of a colorectal cancer (CRC) patient presenting with synchronous lung metastasis and metachronous thyroid, chest wall and urinary tract metastases over the course of 5 years. The chest wall metastasis was caused by needle tract seeding, implying a known time of onset. Using whole genome sequencing data from primary and metastatic sites we inferred the complete chronology of the cancer by exploiting the time of needle tract seeding as an in vivo 'stopwatch'. This approach allowed us to follow the progression of the disease back in time, dating each ancestral node of the phylogenetic tree in the past history of the tumour. We used a Bayesian phylogenomic approach, which accounts for possible dynamic changes in mutational rate, to reconstruct the phylogenetic tree and effectively 'carbon date' the malignant progression. Results: The primary colon cancer emerged between 5 and 8 years before the clinical diagnosis. The primary tumour metastasized to the lung and the thyroid within a year from its onset. The thyroid lesion presented as a tumour-to-tumour deposit within a benign Hurthle adenoma. Despite rapid metastatic progression from the primary tumour, the patient showed an indolent disease course. Primary cancer and metastases were microsatellite stable and displayed low chromosomal instability. Neo-antigen analysis suggested minimal immunogenicity. Conclusion: Our data provide the first in vivo experimental evidence documenting the timing of metastatic progression in CRC and suggest that genomic instability might be more important than the metastatic potential of the primary cancer in dictating CRC fate.

Sorafenib for the Treatment of Advanced Hepatocellular Cancer - a UK Audit.

AIMS: Sorafenib is the current standard treatment for advanced hepatocellular carcinoma. We carried out a national audit of UK patients treated with sorafenib as standard-of-care and those treated with systemic therapy in first-line trials. MATERIALS AND METHODS: Sorafenib-treated and trial-treated patients were identified via the Cancer Drugs Fund and local databases. Data were collected retrospectively from medical records according to a standard case report form. The primary outcome measure was overall survival, estimated by the Kaplan-Meier method. RESULTS: Data were obtained for 448 sorafenib-treated patients from 15 hospitals. The median age was 68 years (range 17-89) and 75% had performance status ≤ 1. At baseline, 77% were Child-Pugh A and 16.1% Child-Pugh B; 38% were albumin-bilirubin grade 1 (ALBI-1) and 48% ALBI-2; 23% were Barcelona Clinic Liver Classification B (BCLC-B) and 72% BCLC-C. The median time on sorafenib was 3.6 months, with a mean daily dose of 590 mg. The median overall survival for 448 evaluable sorafenib-treated patients was 8.5 months. There were significant differences in overall survival comparing Child-Pugh A versus Child-Pugh B (9.5 versus 4.6 months), ALBI-1 versus ALBI-2 (12.9 versus 5.9 months) and BCLC-B versus BCLC-C (13.0 versus 8.3 months). For trial-treated patients (n=109), the median overall survival was 8.1 months and this was not significantly different from the sorafenib-treated patients. CONCLUSION: For Child-Pugh A patients with good performance status, survival outcomes were similar to those reported in global randomised controlled trials. Patients with ALBI grade > 1, Child-Pugh B or poor performance status seem to derive limited benefit from sorafenib treatment.

Prognostic role of the LCS6 KRAS variant in locally advanced rectal cancer: results of the EXPERT-C trial.

BACKGROUND: Lethal-7 (let-7) is a tumour suppressor miRNA which acts by down-regulating several oncogenes including KRAS. A single-nucleotide polymorphism (rs61764370, T > G base substitution) in the let-7 complementary site 6 (LCS-6) of KRAS mRNA has been shown to predict prognosis in early-stage colorectal cancer (CRC) and benefit from anti-epidermal growth factor receptor monoclonal antibodies in metastatic CRC. PATIENTS AND METHODS: We analysed rs61764370 in EXPERT-C, a randomised phase II trial of neoadjuvant CAPOX followed by chemoradiotherapy, surgery and adjuvant CAPOX plus or minus cetuximab in locally advanced rectal cancer. DNA was isolated from formalin-fixed paraffin-embedded tumour tissue and genotyped using a PCR-based commercially available assay. Kaplan-Meier method and Cox regression analysis were used to calculate survival estimates and compare treatment arms. RESULTS: A total of 155/164 (94.5%) patients were successfully analysed, of whom 123 (79.4%) and 32 (20.6%) had the LCS-6 TT and LCS-6 TG genotype, respectively. Carriers of the G allele were found to have a statistically significantly higher rate of complete response (CR) after neoadjuvant therapy (28.1% versus 10.6%; P = 0.020) and a trend for better 5-year progression-free survival (PFS) [77.4% versus 64.5%: hazard ratio (HR) 0.56; P = 0.152] and overall survival (OS) rates (80.3% versus 71.9%: HR 0.59; P = 0.234). Both CR and survival outcomes were independent of the use of cetuximab. The negative prognostic effect associated with KRAS mutation appeared to be stronger in patients with the LCS-6 TT genotype (HR PFS 1.70, P = 0.078; HR OS 1.79, P = 0.082) compared with those with the LCS-6 TG genotype (HR PFS 1.33, P = 0.713; HR OS 1.01, P = 0.995). CONCLUSION: This analysis suggests that rs61764370 may be a biomarker of response to neoadjuvant treatment and an indicator of favourable outcome in locally advanced rectal cancer possibly by mitigating the poor prognosis of KRAS mutation. In this setting, however, this polymorphism does not appear to predict cetuximab benefit.

microRNA-29 can regulate expression of the long non-coding RNA gene MEG3 in hepatocellular cancer.

The human genome is replete with long non-coding RNAs (lncRNA), many of which are transcribed and likely to have a functional role. Microarray analysis of >23,000 lncRNAs revealed downregulation of 712 (~3%) lncRNA in malignant hepatocytes, among which maternally expressed gene 3 (MEG3) was downregulated by 210-fold relative to expression in non-malignant hepatocytes. MEG3 expression was markedly reduced in four human hepatocellular cancer (HCC) cell lines compared with normal hepatocytes by real-time PCR. RNA in situ hybridization showed intense cytoplasmic expression of MEG3 in non-neoplastic liver with absent or very weak expression in HCC tissues. Enforced expression of MEG3 in HCC cells significantly decreased both anchorage-dependent and -independent cell growth, and induced apoptosis. MEG3 promoter hypermethylation was identified by methylation-specific PCR and MEG3 expression was increased with inhibition of methylation with either 5-Aza-2-Deoxycytidine, or siRNA to DNA Methyltransferase (DNMT) 1 and 3b in HCC cells. MiRNA-dependent regulation of MEG3 expression was studied by evaluating the involvement of miR-29, which can modulate DNMT 1 and 3. Overexpression of mir-29a increased expression of MEG3. GTL2, the murine homolog of MEG3, was reduced in liver tissues from hepatocyte-specific miR-29a/b1 knock-out mice compared with wild-type controls. These data show that methylation-dependent tissue-specific regulation of the lncRNA MEG3 by miR-29a may contribute to HCC growth and highlight the inter-relationship between two classes of non-coding RNA, miRNAs and lncRNAs, and epigenetic regulation of gene expression.

Molecular targets in Gastrointestinal Stromal Tumors (GIST) therapy.

Gastrointestinal Stromal Tumors (GISTs) are the most common mesenchimal tumors of the gastrointestinal tract. Such tumors usually have activating mutations in either KIT (75-80%) or Platelet Derived Growth Factor Receptor alpha (PDGFRa) (5-10%) which lead to ligand-independent signal transduction. Targeting these activated proteins with Imatinib mesylate, a small-molecule kinase inhibitor, has proven useful in the treatment of recurrent or metastatic GISTs. However, more than half of patients develop resistance to Imatinib after about 2 years. Therefore, other targets have been studying in order to implement the therapeutical armamentarium for this disease. Sunitinib malate is an oral multikinase inhibitor that targets several receptor tyrosine kinases and has proved to prolong survival in Imatinib-resistant patients. Other molecules, such as Nilotinib, Sorafenib and Dasatinib were shown to be useful in Imatinib resistant mutant cell lines and the results of their activity in humans are being awaited. Recent evidence suggests that GIST cells acquire the capability to escape from the control of KIT and PDGFRa through the activation of alternative pathways. Therefore, further effort should be invested in the discovery of new signaling pathways, such as AXL, MET, IGF-R, which might be involved in the evolution of the disease. After a description of KIT and PDGFRa as known targets of anti-GIST treatments, we review other mechanisms and mediators that might be potential targets of new therapies, providing a comprehensive revision of the new molecular strategies under investigation.

Insulin-like growth factor (IGF) 1 and 2 help to predict disease outcome in GIST patients.

BACKGROUND: The expression of the insulin-like growth factor (IGF) system has never been studied in gastrointestinal stromal tumors (GISTs). PATIENTS AND METHODS: We studied the immunohistochemical expression of IGF1 receptor (IGFR-I), IGF1 and IGF2 in 94 samples of GISTs. IGF1 and IGF2 expression was scored in three classes: negative (N), moderate (M) and strong (S), according to staining intensity and extent. RESULTS: IGFR-I was overexpressed in all cases. IGF1 and IGF2 expression was absent in 25 and 48 cases, moderate in 29 and 16 cases and strong in 40 and 30 cases, respectively. Strong IGF1 expression significantly correlated with higher mitotic index (P = 0.0001), larger (P = 0.01), higher risk (P = 0.0002), metastatic (P = 0.0001) and relapsed (P = 0.04) GISTs. Strong IGF2 expression correlated with higher mitotic index (P = 0.05) and higher risk GISTs (P = 0.001). The Kaplan-Meier analysis (N versus M versus S) showed a significant worsening of the disease-free survival (DFS) with the increase of IGF1 (P = 0.02) and IGF2 (P = 0.02) expression. In the subgroup of patients with operated high-risk GISTs, there was a better trend in DFS for patients affected by GISTs with negative IGF1 and IGF2. CONCLUSIONS: The expression of IGF1 and IGF2 seems to predict relapse in GIST patients.

KIT and PDGFRalpha mutations in 104 patients with gastrointestinal stromal tumors (GISTs): a population-based study.

BACKGROUND: The prognostic significance of KIT or platelet-derived growth factor receptor alpha (PDGFRalpha) mutations in gastrointestinal stromal tumors (GISTs) is still controversial. PATIENTS AND METHODS: In all, 104 patients were diagnosed with GISTs by KIT immunoreactivity; tumor DNA was sequenced for the presence of mutations in KIT exons 9, 11, 13 and 17 and in PDGFRalpha exons 12 and 18. Disease-free survival (DFS) was analyzed in 85 radically resected patients. RESULTS: KIT mutations occurred in exon 11 (69), in exon 9 (11) and in exon 17 (1). PDGFRalpha mutations were detected in exon 18 (10) and in exon 12 (3). Ten GISTs were wild type. Exon 11 mutations were as follows: deletions in 42 cases and point mutations in 20 cases and insertions and duplications, respectively, in 2 and 5 cases. A better trend in DFS was evident for duplicated and point-mutated exon 11 KIT GISTs. There was a significant association between PDGFRalpha mutations, gastric location and lower mitotic index. Moreover, PDGFRalpha-mutated GISTs seemed to have a better outcome. CONCLUSIONS: Point mutations and duplications in KIT exon 11 are associated with a better clinical trend in DFS. PDGFRalpha-mutated GISTs are preferentially localized in the stomach and seem to have a favorable clinical behavior.

Anemia may influence the outcome of patients undergoing neo-adjuvant treatment of rectal cancer.

BACKGROUND: We hypothesized that anemia could represent one of the major factors influencing the outcome of patients undergoing neo-adjuvant treatment of rectal cancer. PATIENTS AND METHODS: This analysis included all the consecutive patients who underwent neo-adjuvant treatment (chemotherapy and/or radiotherapy) before surgery for rectal cancer in three oncology/radiotherapy departments from June 1996 to December 2003. RESULTS: Three hundred and seventeen patients were eligible for our analysis. Median age at diagnosis was 64 years (range 26-88 years); male/female ratio was 184/133. Two hundred and eighty-five patients (89.9%) were diagnosed with adenocarcinoma, while 32/317 (10.1%) with mucinous adenocarcinoma. Neo-adjuvant treatments carried out were as follows: radiotherapy alone in 75/317 patients (23.7%), radiotherapy plus chemotherapy in 242/317 patients (76.3%). At univariate and multivariate analysis, only the hemoglobin (Hb) level (group 1: < or=12 g/dl versus group 2: >12 g/dl) resulted in a significant factor for disease-free survival. The role of the Hb level seemed to be confirmed further by the clinical downstaging obtained in approximately 55% of patients in group 2, in comparison with 35% of the patients achieving a significant downstaging in group 1. CONCLUSION: Our results indicated that anemia could represent an important parameter able to influence the outcome in patients receiving neo-adjuvant treatment of rectal cancer.

[Paraneoplastic syndromes: a review]. / Le sindromi paraneoplastiche: revisione critica della letteratura.

Modern oncology often obtains good results against earlier neoplasms, whilst it's still in difficulties against the advanced ones. The knowledge of paraneoplastic syndromes is crucial both to cure patients and to do an earlier diagnosis. When we recognize a paraneoplastic syndrome that comes before the clinic beginning of a neoplasm, perhaps we save a life. This review discusses all the main paraneoplastic syndromes, focusing mainly on their clinical aspect and reminding the most commonly associated cancers.