An expression signature of the angiogenic response in gastrointestinal neuroendocrine tumours: correlation with tumour phenotype and survival outcomes.

BACKGROUND: Gastroenteropancreatic neuroendocrine tumours (GEP-NETs) are heterogeneous with respect to biological behaviour and prognosis. As angiogenesis is a renowned pathogenic hallmark as well as a therapeutic target, we aimed to investigate the prognostic and clinico-pathological role of tissue markers of hypoxia and angiogenesis in GEP-NETs. METHODS: Tissue microarray (TMA) blocks were constructed with 86 tumours diagnosed from 1988 to 2010. Tissue microarray sections were immunostained for hypoxia inducible factor 1α (Hif-1α), vascular endothelial growth factor-A (VEGF-A), carbonic anhydrase IX (Ca-IX) and somatostatin receptors (SSTR) 1–5, Ki-67 and CD31. Biomarker expression was correlated with clinico-pathological variables and tested for survival prediction using Kaplan–Meier and Cox regression methods. RESULTS: Eighty-six consecutive cases were included: 51% male, median age 51 (range 16–82), 68% presenting with a pancreatic primary, 95% well differentiated, 51% metastatic. Higher grading (P=0.03), advanced stage (P<0.001), high Hif-1α and low SSTR-2 expression (P=0.03) predicted for shorter overall survival (OS) on univariate analyses. Stage, SSTR-2 and Hif-1α expression were confirmed as multivariate predictors of OS. Median OS for patients with SSTR-2+/Hif-1α-tumours was not reached after median follow up of 8.8 years, whereas SSTR-2-/Hif-1α+ GEP-NETs had a median survival of only 4.2 years (P=0.006). CONCLUSION: We have identified a coherent expression signature by immunohistochemistry that can be used for patient stratification and to optimise treatment decisions in GEP-NETs independently from stage and grading. Tumours with preserved SSTR-2 and low Hif-1α expression have an indolent phenotype and may be offered less aggressive management and less stringent follow up

Qualification of tumour mutational burden by targeted next-generation sequencing as a biomarker in hepatocellular carcinoma

Background & Aims: Tumour mutational burden (TMB) predicts improved response and survival to immunotherapy. In this pilot study, we optimized targeted next-generation sequencing (tNGS) to estimate TMB in hepatocellular carcinoma (HCC). Methods: We sequenced 48 non-paired samples (21 fresh-frozen [FF] and 27 paraffin-embedded [FFPE]), among which 11 FFPE samples were pretreated with uracil-DNA glycosylase (UDG). Thirty samples satisfied post-sequencing quality control. High/low TMB was defined by median number of mutations/Mb (Mut/Mb), across different minimum allele frequency (MAF) thresholds ( 650.05, 650.1 and 650.2). Results: Eligible patients (n\ua0=\ua029) were cirrhotic (84%) with TNM stage I-II HCC (75%). FFPE samples had higher TMB (median 958.39 vs 2.51 Mut/Mb, P\ua0\ua0T transitions at CpG sites (median 60.3% vs 9.1%, P\ua0=.002) compared to FF. UDG-treated samples had lower TMB (median 4019.92 vs 353 Mut/Mb, P\ua0=.041) and deamination counts (median 6393.5 vs 328.5, P\ua0=.041) vs untreated FFPE. At 0.2 MAF threshold with UDG treatment, median TMB was 5.48 (range 1.68-16.07) and did not correlate with salient pathologic features of HCC, including survival. Conclusion: While tNGS on fresh HCC samples appears to be the optimal source of tumour DNA, the low median TMB values observed may limit the role of TMB as a predictor of response to immunotherapy in HCC

Improved neuron culture using scaffolds made of three-dimensional PDMS micro-lattices

Tissue engineering strives to create functional components of organs with different cell types in vitro. One of the challenges is to fabricate scaffolds for three-dimensional (3D) cell culture under physiological conditions. Of particular interesting is to investigate the morphology and function of the central nervous system (CNS) cultured using such scaffolds. Here, we used an elastomer, polydimethylsiloxane (PDMS), to produce lattice-type scaffolds from a photolithography defined template. The photomask with antidot arrays was spin-coated by a thick layer of resist and downward mounted on a rotating stage at angle of 45\ub0. After exposure for three or more times keeping the same exposure plan but rotated by the same angle, the photoresist was developed to produce a 3D porous template. Afterward, a pre-polymer mixture of PDMS was poured in and cured, followed by a resist etch, resulting in lattice-type PDMS features. Before cell culture, the PDMS lattices were surface functionalized. Culture test has been done using NIH-3T3 cells and primary hippocampal cells from rats, showing homogenously cell infiltration and 3D attachment. As expected, a much higher cell number was found in 3D PDMS lattices than in 2D culture. We also found a higher neuron to astrocyte ratio and a higher degree of cell ramification in 3D culture compared to 2D culture, due to the change of scaffold topography and the elastic properties of the PDMS micro-lattices. Our results demonstrate that the 3D PDMS micro-lattices improve the survival and growth of cells as well as the network formation of neurons. We believe that such an enabling technology is useful for research and clinical applications including disease modeling, regenerative medicine, and drug discovery/drug cytotoxicity studies

An inflammation based score can optimize the selection of patients with advanced cancer considered for early phase clinical trials.

Background: Adequate organ function and good performance status (PS) are common eligibility criteria for phase I trials. As inflammation is pathogenic and prognostic in cancer we investigated the prognostic performance of inflammation-based indices including the neutrophil (NLR) and platelet to lymphocyte ratio (PLR). Methods: We studied inflammatory scores in 118 unselected referrals. NLR normalization was recalculated at disease reassessment. Each variable was assessed for progression-free (PFS) and overall survival (OS) on uni- and multivariate analyses and tested for 90 days survival (90DS) prediction using receiving operator curves (ROC). Results: We included 118 patients with median OS 4.4 months, 23% PS>1. LDH 65450 and NLR 655 were multivariate predictors of OS (p<0.001). NLR normalization predicted for longer OS (p<0.001) and PFS (p<0.05). PS and NLR ranked as most accurate predictors of both 90DS with area under ROC values of 0.66 and 0.64, and OS with c-score of 0.69 and 0.60. The combination of NLR+PS increased prognostic accuracy to 0.72. The NLR was externally validated in a cohort of 126 subjects. Conclusions: We identified the NLR as a validated and objective index to improve patient selection for experimental therapies, with its normalization following treatment predicting for a survival benefit of 7 months. Prospective validation of the NLR is warranted. Copyright: \ua9 2014 Pinato et al

The ALBI grade provides objective hepatic reserve phenotyping across each BCLC stage of hepatocellular carcinoma

BACKGROUND & AIMS: Overall survival (OS) is a composite clinical endpoint in hepatocellular carcinoma (HCC) due to the mutual influence of cirrhosis and active malignancy in dictating patient's mortality. The ALBI grade is a recently described index of liver dysfunction in hepatocellular carcinoma, based solely on albumin and bilirubin levels. Whilst accurate, this score lacks cross-validation, especially in intermediate stage HCC, where OS is highly heterogeneous. METHODS: We evaluated the prognostic accuracy of the ALBI grade in estimating OS in a large, multi-centre study of 2426 patients, including a large proportion of intermediate stage patients treated with chemoembolization (n=1461) accrued from Europe, the United States and Asia. RESULTS: Analysis of survival by primary treatment modality confirmed the ALBI grade as a significant predictor of patient OS after surgical resection (p<0.001), transarterial chemoembolization (p<0.001) and sorafenib (p<0.001). Stratification by Barcelona Clinic Liver Cancer stage confirmed the independent prognostic value of the ALBI across the diverse stages of the disease, geographical regions of origin and time of recruitment to the study (p<0.001). CONCLUSIONS: In this large, multi-centre retrospective study, the ALBI grade satisfied the criteria for accuracy and reproducibility following statistical validation in Eastern and Western HCC patients, including those treated with chemoembolization. Consideration should be given to the ALBI grade as a stratifying biomarker of liver reserve in routine clinical practice. LAY SUMMARY: Liver failure is a key determinant influencing the natural history of hepatocellular carcinoma (HCC). In this large multi-centre study we externally validate a novel biomarker of liver functional reserve, the ALBI grade, across all the stages of HCC

Prognostic Utility of Albumin-Bilirubin Grade for Short- and Long-Term Outcomes Following Hepatic Resection for Intrahepatic Cholangiocarcinoma: a Multi-Institutional Analysis of 706 Patients

Background: The objective of the current study was to define the impact of albumin-bilirubin (ALBI) grade on short- as well as long-term outcomes among patients with intrahepatic cholangiocarcinoma (ICC). Methods: Patients who underwent hepatectomy for ICC between 1990 and 2016 were identified using an international multi-institutional database. Clinicopathologic factors including ALBI score were assessed using bivariate and multivariable analyses, as well as standard survival analyses. Results: Among 706 patients, 453 (64.2%) patients had ALBI grade 1, 231 (32.7%) ALBI grade 2, and 22 (3.1%) had ALBI grade 3. After adjusting for all competing factors, patients with ALBI grade 2/3 had higher odds of a prolonged length-of-stay (>10 days, odds ratio [OR] = 2.37, 95% confidence interval [CI]:1.47-3.80), perioperative transfusion (OR = 2.15, 95% CI:1.45-3.18) and 90-day mortality (OR = 2.50, 95% CI:1.16-5.38). Median and 5-year overall survival (OS) for the entire cohort was 41.5 months (IQR:15.7-107.8) and 39.8%, respectively. Of note, median OS incrementally worsened with increased ALBI grade: grade 1, 49.6 months (IQR:18.3-NR) vs grade 2, 29.6 months (IQR:12.6-98.4) vs grade 3, 16.9 months (IQR:6.5-32.4; P < 0.001). On multivariable analysis, higher ALBI grade remained associated with higher hazards of death (grade 2/3: hazard ratio = 1.36, 95% CI:1.04-1.78). Conclusion: The ALBI score was associated with both short- and long-term outcomes following resection for ICC and could prove a useful surrogate marker to identify patients at risk for adverse outcomes.info:eu-repo/semantics/publishedVersio

Less than 5 Netrin-1 molecules initiate attraction but 200 Sema3A molecules are necessary for repulsion

Guidance molecules, such as Sema3A or Netrin-1, induce growth cone (GC) repulsion or attraction. In order to determine the speed of action and efficiency of these guidance cues we developed an experimental procedure to deliver controlled amounts of these molecules. Lipid vesicles encapsulating 10-10 4 molecules of Sema3A or Netrin-1 were manipulated with high spatial and temporal resolution by optical tweezers and their photolysis triggered by laser pulses. Guidance molecules released from the vesicles diffused and reached the GC membrane in a few seconds. Following their arrival, GCs retracted or grew in 20-120 s. By determining the number of guidance molecules trapped inside vesicles and estimating the fraction of guidance molecules reaching the GC, we show that the arrival of less than 5 Netrin-1 molecules on the GC membrane is sufficient to induce growth. In contrast, the arrival of about 200 Sema3A molecules is necessary to induce filopodia repulsion