Development and External Validation of the Munich Sorafenib Evaluation Score for Hepatocellular Carcinoma.

INTRODUCTION: In recent years, increasing options for systemic HCC treatment have become available. The development of therapy-specific prognostic scores has been encouraged. Tailoring therapy to individual patients requires prognostic scores for treatment success in addition to the Barcelona-Clinic-Liver-Cancer (BCLC) classification. We have developed and validated a prognostic score for patients treated with sorafenib. METHODS: Prognostic factors identified in a multivariate analysis of 108 sorafenib patients were used to construct the Munich Sorafenib Evaluation (M-SE) score. M-SE and 9 established HCC prognostic systems were ranked according to concordance-index and AIC. External M-SE validation was performed in an independent HCC sorafenib cohort (n = 101) derived from the prospective multicenter randomized controlled SORAMIC trial. RESULTS: Ascites (p < 0.0001; HR 2.923), tumor burden ≥50% of the liver (p = 0.0033; HR 1.946), and GOT (p < 0.0001; HR 1.716) were identified as independent prognostic parameters. All three M-SE stages were characterized by significantly different survival times (p < 0.0001). M-SE stage-A patients had a median OS of 18.7 months (95% CI: 15.6-21.8); patients in stage B and C showed a significantly shorter survival of 5.7 (2.7-8.7) and 2.0 months (1.6-2.4), respectively. M-SE (c-index 0.70; AIC 621) outperformed all other prognostic systems. External validation in a prospective cohort confirmed its superior prognostic performance. CONCLUSION: The M-SE score allows classification of sorafenib patients in three distinct prognostic stages. Provided that M-SE successfully passes prospective validation, it can help to predict the outcome of patients evaluated for sorafenib treatment.

A Simple Prognostic Scoring System for Hepatocellular Carcinoma Treated with Selective Internal Radiation Therapy.

INTRODUCTION: Selective internal radiation therapy (SIRT) is a local treatment option for patients with hepatocellular carcinoma (HCC). Its exact role next to other HCC therapies has yet to be defined. In order to identify patients most suitable for SIRT, a SIRT-specific prognostic score should be developed. METHODS: A cohort of 72 SIRT patients treated at the University Hospital of Munich was retrospectively analyzed. The prognostic performance of 12 HCC staging systems and prognostic scores was assessed. Cox-regression analysis was used to identify independent prognostic factors, which formed the basis of the Munich-SIRT score (M-SIRT). All scores were ranked by calculating the c-Index and Akaike information criterion (AIC). External validation was performed in a cohort of 128 SIRT patients treated at the University Hospital of Pamplona, Spain. RESULTS: median overall survival was 13 months (95% confidence interval 9.9-21.9). AFP (p = 0.005; hazard ratio [HR] 2.38), albumin (p < 0.001; HR 5.87), and alkaline phosphatase (p < 0.001; HR 8.38) were identified as independent prognostic factors. M-SIRT comprises 3 prognostic groups with a median survival of 38.9, 14.6, and 7.7 months, respectively (I vs. II: p = 0.003, II vs. III: p < 0.001). AIC (318) and concordance index (0.711) ranked M-SIRT superior to the established HCC staging systems, and the score successfully passed external validation in an independent SIRT cohort (I vs. II: p = 0.03; II vs. III: p = 0.007). CONCLUSION: Therapy-specific prognostic scores can facilitate treatment decisions and prognostication for HCC patients. Considering its performance in 200 SIRT patients, M-SIRT is a promising prognostic tool for HCC patients evaluated for SIRT.

The Munich-Transarterial Chemoembolisation Score Holds Superior Prognostic Capacities Compared to TACE-Tailored Modifications of 9 Established Staging Systems for Hepatocellular Carcinoma.

BACKGROUND/AIMS: The recently proposed Munich-transarterial chemoembolisation-score (M-TACE) was tailored to suit hepatocellular Carcinoma (HCC) patients evaluated for TACE. M-TACE outperformed the established HCC-staging-systems and successfully passed external validation. Modifications of staging-systems through the rearrangement of stages or by adding prognostic factors are methods of improving prognostic power. M-TACEs performance compared to scores modified this way should be tested. METHODS: Seven well-known HCC staging-systems (including Cancer of the Liver Italian Program-score [CLIP] and Barcelona Clinic liver cancer [BCLC]) and 2 TACE-specific scores (Selection for Transarterial Chemoembolisation Treatment [STATE] and Hepatoma Arterial embolisation Prognostic [HAP]) were rearranged in a cohort of 186 TACE-patients through score-point-analysis and subsequent linking of non-significant adjacent score-points. Additionally, a new score was constructed by combining the top established staging-system in TACE patients (CLIP-TACE) and the prognostic parameter with the highest hazard ratio for death in the TACE-cohort [C-reactive protein (CRP)]. Additionally, the TACE-tailored-scores were applied to an external TACE-cohort (n = 71). -Results: Rearrangement resulted in optimal stratification and monotonicity. CLIP-TACE demonstrated the best prognostic capability of all rearranged scores (c-index 0.668, AIC 1294) and the addition of CRP yielded further prognostic improvement (c-index 0.680, AIC 1289). However, superiority over M-TACE could not be achieved by any of the new scores in the internal and external cohort. CONCLUSION: M-TACE outperforms TACE-tailored modifications of all relevant HCC-staging-systems. Prospective validation of M-TACE to promote its role as the preferred staging-system for TACE-patients is therefore justified.

Correlation Between Baseline Osteoprotegerin Serum Levels and Prognosis of Advanced-Stage Colorectal Cancer Patients.

BACKGROUND/AIMS: Osteoprotegerin (OPG) is a soluble receptor of the pro-apoptotic cytokine TRAIL which is thought to contribute to tumour development by inhibiting apoptosis or affecting other aspects of tumour biology, including cell proliferation and immune response. Although immunohistochemical studies suggest that OPG correlates with survival in metastatic colorectal cancer (mCRC), only scarce data are available on serum OPG in CRC patients. METHODS: In this pilot study, we assessed the prognostic significance of serum OPG and CEA (Carcinoembryonic antigen) in 81 patients with UICC (Union for International Cancer Control) stage-IV mCRC. OPG was additionally assessed by immunohistochemistry in primary tissue samples from 33 patients of the same cohort. RESULTS: Baseline serum OPG correlated with CEA (r=0.36, p=0.0011), but independently predicted survival of mCRC patients. Life expectancy was poorer in patients with OPG levels above the median concentration of 51ng/ml (median overall survival [95% confidence interval] 1.8 years [1.3-3.0] vs. 1.0 [0.7-1.2] p=0.013). Patients with high levels of both OPG and CEA had an even poorer life expectancy vs. low-OPG/low-CEA patients (0.9 years [0.6-1.5] vs. 3 years [1.2-4.4], p=0.015), indicating that CEA and OPG have additive prognostic significance. Immunohistochemical analysis of OPG failed to show a correlation between OPG staining and survival (p=0.055) or OPG concentration from matched serum samples. CONCLUSIONS: This pilot study provides evidence of independent prognostic significance of serum OPG in patients with advanced mCRC and warrants its further prospective validation.

Methylated free-circulating HPP1 DNA is an early response marker in patients with metastatic colorectal cancer.

Detection of methylated free-circulating DNA (mfcDNA) for hyperplastic polyposis 1 (HPP1) in blood is correlated with a poor prognosis for patients with metastatic colorectal cancers (mCRC). Here, we analyzed the plasma levels of HPP1 mfcDNA in mCRC patients treated with a combination therapy containing a fluoropyrimidine, oxaliplatin and bevacizumab to test whether HPP1 mfcDNA is a suitable prognostic and response biomarker. From 467 patients of the prospective clinical study AIO-KRK-0207, mfcDNA was isolated from plasma samples at different time points and bisulfite-treated mfcDNA was quantified using methylation specific PCR. About 337 of 467 patients had detectable levels for HPP1 mfcDNA before start of treatment. The detection was significantly correlated with poorer overall survival (OS) (HR = 1.86; 95%CI 1.37-2.53). About 2-3 weeks after the first administration of combination chemotherapy, HPP1 mfcDNA was reduced to non-detectable levels in 167 of 337 patients. These patients showed a better OS compared with patients with continued detection of HPP1 mfcDNA (HR HPP1(sample 1: pos/ sample 2: neg) vs. HPP1(neg/neg) = 1.41; 95%CI 1.00-2.01, HPP1(neg,pos/pos) vs. HPP1(neg/neg) = 2.60; 95%CI 1.86-3.64). Receiver operating characteristic analysis demonstrated that HPP1 mfcDNA discriminates well between patients who do (not) respond to therapy according to the radiological staging after 12 or 24 weeks (AUC = 0.77 or 0.71, respectively). Detection of HPP1 mfcDNA can be used as a prognostic marker and an early marker for response (as early as 3-4 weeks after start of treatment compared with radiological staging after 12 or 24 weeks) to identify patients who will likely benefit from a combination chemotherapy with bevacizumab.

Piperacillin concentration in relation to therapeutic range in critically ill patients--a prospective observational study.

BACKGROUND: Piperacillin levels after standard dosing have been shown frequently to be subtherapeutic, especially when renal clearance was augmented. Here, we aimed to determine if piperacillin was in its therapeutic range in a typically heterogeneous intensive care unit patient group, and also to describe target attainment dependent on daily dosage, creatinine clearance, and renal replacement therapy (RRT). METHODS: Sixty patients with severe infections were included in this monocentric prospective observational study. Patients received 4.5 g of piperacillin-tazobactam two to three times daily by intermittent infusion depending on renal function according to clinical guidelines. Over 4 days, multiple serum samples (median per patient, 29; in total, 1627) were obtained to determine total piperacillin concentrations using ultra-high-performance liquid chromatography/tandem mass spectrometry. RESULTS: A high heterogeneity of patient characteristics was observed (e.g., on day 1: creatinine clearance 2-233 mL/min and ten patients on RRT). Piperacillin trough levels showed inter-individual variation from 123 to >1785-fold on different study days. Each day, approximately 50% and 60% of the patients had piperacillin levels below the target ranges 1 and 2, respectively [defined for the calculated unbound piperacillin fraction according to the literature as 100% time above MIC (100%fT > MIC) (target range 1) and ≥ 50%fT > 4 × MIC (target range 2); MIC = 16 mg/L]. Whereas only the minority of patients who received piperacillin-tazobactam three times daily (TID) reached target 1 (38% on day 1), most patients who received piperacillin-tazobactam only twice daily (BID) because of severely impaired renal function reached this target (100% on day 1). Patients with RRT had significant higher percentages of fT > MIC. Zero percent, 55% and 100% of patients without RRT who received antibiotics TID reached target 1 when creatinine clearance was > 65 mL/min, 30-65 mL/min and < 30 mL/min, respectively. In patients with causative strains only sensitive to piperacillin-tazobactam of all antibiotics given to the patient, piperacillin levels negatively correlated with CRP concentrations of day 4 (p < 0.05). CONCLUSIONS: A dosage of 4.5 g piperacillin-tazobactam TID seems to be frequently insufficient in critically ill patients, and also in patients where renal function is mildly to moderately impaired. For these patients, prescription of 4.5 g piperacillin-tazobactam four times daily could be considered. TRIAL REGISTRATION: Clinicaltrials.gov NCT01793012. Registered 24 January 2013.

Preoperative serum markers for individual patient prognosis in stage I-III colon cancer.

Carcinoembryonic antigen (CEA) remains the only recommended biomarker for follow-up care of colorectal cancer (CRC), but besides CEA, several other serological parameters have been proposed as prognostic markers for CRC. The present retrospective analysis investigates a comprehensive set of serum markers with regard to cancer-specific survival (CSS) and disease-free survival (DFS). A total of 472 patients with colon cancer underwent surgery for curative intent between January 1988 and June 2007. Preoperative serum was analyzed for the following parameters: albumin, alkaline phosphatase (aP), beta-human chorionic gonadotropin (ßhCG), bilirubin, cancer antigen 125 (CA 125), cancer antigen 19-9 (CA 19-9), CA 72-4, CEA, C-reactive protein (CRP), cytokeratin-19 soluble fragment (CYFRA 21-1), ferritin, gamma-glutamyltransferase (γGT), glutamate oxaloacetate transaminase (GOT), glutamate pyruvate transaminase (GPT), hemoglobin, haptoglobin, interleukin-6, interleukin-8, creatinine, lactate dehydrogenase (LDH), serum amyloid A (SAA), and 25-hydroxyvitamin D. After a median follow-up period of 5.9 years, the overall 3- and 5-year CSS was 91.7 and 84.9 % and DFS rates were 82.7 % (3 years) and 77.6 % (5 years). Multivariate analyses confirmed preoperative CEA as an independent prognostic factor with regard to CSS and DFS. CA 19-9 and γGT also provided prognostic value for CSS and DFS, respectively. Younger age was negatively associated with DFS. According to UICC stage, CEA provided significant prognostic value with regard to CSS and DFS, while CA 19-9 was only prognostic for CSS. Combined analysis is able to identify patients with favorable prognosis. In addition to tumor baseline parameters, preoperative CEA could be confirmed as prognostic marker in colon cancer. CA 19-9 and γGT also provide additional prognostic value with regard to survival and recurrence in stage III and stage I disease, respectively. The combined use of CEA together with CA 19-9 and γGT improve risk-adapted post-op surveillance.

Comparison of five automated hematology analyzers in a university hospital setting: Abbott Cell-Dyn Sapphire, Beckman Coulter DxH 800, Siemens Advia 2120i, Sysmex XE-5000, and Sysmex XN-2000.

BACKGROUND: Various types of automated hematology analyzers are used in clinical laboratories. Here, we performed a side-by-side comparison of five current top of the range routine hematology analyzers in the setting of a university hospital central laboratory. METHODS: Complete blood counts (CBC), differentials, reticulocyte and nucleated red blood cell (NRBC) counts of 349 patient samples, randomly taken out of routine diagnostics, were analyzed with Cell-Dyn Sapphire (Abbott), DxH 800 (Beckman Coulter), Advia 2120i (Siemens), XE-5000 and XN-2000 (Sysmex). Inter-instrument comparison of CBCs including reticulocyte and NRBC counts and investigation of flagging quality in relation to microscopy were performed with the complete set of samples. Inter-instrument comparison of five-part differential was performed using samples without atypical cells in blood smear (n=292). Automated five-part differentials and NRBCs were additionally compared with microscopy. RESULTS: The five analyzers showed a good concordance for basic blood count parameters. Correlations between instruments were less well for reticulocyte counts, NRBCs, and differentials. The poorest concordance for NRBCs with microscopy was observed for Advia 2120i (Kendall's τb=0.37). The highest flagging sensitivity for blasts was observed for XN-2000 (97% compared to 65%-76% for other analyzers), whereas overall specificity was comparable between different instruments. CONCLUSIONS: To the best of our knowledge, this is the most comprehensive side-by-side comparison of five current top of the range routine hematology analyzers. Variable analyzer quality and parameter specific limitations must be considered in defining laboratory algorithms in clinical practice.

Magnetic resonance colonography for the detection of colorectal neoplasia in asymptomatic adults.

BACKGROUND & AIMS: Colonoscopy is the preferred screening test for colorectal neoplasia; the fecal occult blood test (FOBT) detects neoplasias with low levels of sensitivity. Computed tomographic colonography detects neoplasias with high levels of sensitivity but involves exposure to radiation. We investigated whether magnetic resonance colonography (MRC) can be used to screen for colorectal adenomas and cancers. METHODS: We analyzed data from 286 asymptomatic adults (40-82 years old) who underwent 3 Tesla MRC and colonoscopic examinations on the same day. FOBT was performed before bowel preparation. Colonoscopists were initially blinded to the findings on MRC and unblinded after withdrawal from the respective segments. Sensitivities for adenoma and per-patient sensitivities and specificities were calculated based on the unblinded results of colonoscopy. RESULTS: We detected 133 adenomas and 2 cancers in 86 patients; 37 adenomas were ≥6 mm, and 20 adenomas were advanced. Sensitivities of MRC and colonoscopy for adenomas ≥6 mm were 78.4% (95% confidence interval [CI], 61.8-90.2) and 97.3% (95% CI, 85.8-99.9); for advanced adenomas these values were 75% (95% CI, 50.9-91.3) and 100% (95% CI, 83.2-100.0), respectively. MRC identified 87.1% (95% CI, 70.2-96.4), colonoscopy 96.8% (95% CI, 83.3-99.9), and FOBT 10.0% (95% CI, 2.1-26.5) of individuals with adenomas ≥6 mm and 83.8% (95% CI, 58.6-96.4), 100% (95% CI, 81.5-100.0), and 17.6% (95% CI, 3.8-43.4) of individuals with advanced neoplasia. Specificities of MRC, colonoscopy, and FOBT for individuals with adenomas ≥6 mm were 95.3% (95% CI, 91.9-97.5), 96.9% (95% CI, 93.9-98.6), and 91.8% (95% CI, 87.6-94.9), respectively. CONCLUSIONS: 3 Tesla MRC detects colorectal adenomas ≥6 mm and advanced neoplasia with high levels of sensitivity and specificity. Although MRC detects colorectal neoplasia with lower levels of sensitivity than colonoscopy, it strongly outperforms one-time FOBT.

Diagnostic relevance of plasma DNA and DNA integrity for breast cancer.

Levels of ALU 115, ALU 247, DNA integrity ([1, 2]) and of the tumour markers CA 15-3 and CEA were analysed in the blood of 152 patients. Plasma levels of ALU 115 and ALU 247 were significantly higher in patients with locally confined (LBC; N = 65), metastatic breast cancer (MBC; N = 47), and benign diseases (N = 12) than in healthy controls (p < 0.001 for all comparisons). DNA integrity, CEA, and CA 15-3 were significantly higher in MBC than in benign controls and LBC but could not identify LBCs. The best discrimination of LBC from healthy controls was achieved by ALU 115 and ALU 247 (AUC 95.4 and 95.5 %) and of MBC from all control groups by CA 15-3 and CEA (AUC 83.2 and 79.1 %). Plasma DNA is valuable for the detection of LBC, while established tumour markers are most informative in MBC.

Evaluation of preoperative serum markers for individual patient prognosis in stage I-III rectal cancer.

Several independent serum biomarkers have been proposed as prognostic and/or predictive markers for colorectal cancer (CRC). To this date, carcinoembryonic antigen (CEA) remains the only recommended serological CRC biomarker. The present retrospective analysis investigates the prognostic value of several serum markers. A total of 256 patients with rectal cancer underwent surgery for curative intent in a university cancer center between January 1988 and June 2007. Preoperative serum was retrospectively analyzed for albumin, alkaline phosphatase (aP), beta-human chorionic gonadotropin, bilirubin, CA 125, cancer antigen 19-9, cancer antigen 72-4 (CA 72-4), CEA, CRP, CYFRA 21-1, ferritin, gamma-glutamyl transpeptidase, glutamate oxaloacetate transanunase, glutamate pyruvate transaminase, hemoglobin, haptoglobin, interleukin-6, interleukin-8, creatinine, lactate-dehydrogenase, serum amyloid A (SAA), and 25-hydroxyvitamin D. Cancer-specific survival (CSS) and disease-free survival (DFS) were estimated. Median follow-up time was 8.4 years. Overall 3- and 5-year CSS was 88.6 and 78.9 %, respectively. DFS rates were 72.8 % (3 years) and 67.5 % (5 years). Univariate analysis of CSS indicated aP, CA 72-4, CEA, and SAA as prognostic factors, while aP, CEA, and SAA were also prognostic with regard to DFS. Multivariate analysis confirmed SAA together with T and N stage as prognostic factors. According to UICC stage, CEA and SAA add prognostic value in stages II and III with regard to DFS and CSS, respectively. The combined use of CEA and SAA is able to identify patients with favorable and poor prognosis. In addition to tumor baseline parameters, routine analysis of SAA together with CEA provided markedly improved prognostic value on CSS and DFS in resected rectal cancer.

Circulating cell-free methylated DNA and lactate dehydrogenase release in colorectal cancer.

BACKGROUND: Hypermethylation of DNA is an epigenetic alteration commonly found in colorectal cancer (CRC) and can also be detected in blood samples of cancer patients. Methylation of the genes helicase-like transcription factor (HLTF) and hyperplastic polyposis 1 (HPP1) have been proposed as prognostic, and neurogenin 1 (NEUROG1) as diagnostic biomarker. However the underlying mechanisms leading to the release of these genes are unclear. This study aimed at examining the possible correlation of the presence of methylated genes NEUROG1, HLTF and HPP1 in serum with tissue breakdown as a possible mechanism using serum lactate dehydrogenase (LDH) as a surrogate marker. Additionally the prognostic impact of these markers was examined. METHODS: Pretherapeutic serum samples from 259 patients from all cancer stages were analyzed. Presence of hypermethylation of the genes HLTF, HPP1, and NEUROG1 was examined using methylation-specific quantitative PCR (MethyLight). LDH was determined using an UV kinetic test. RESULTS: Hypermethylation of HLTF and HPP1 was detected significantly more often in patients with elevated LDH levels (32% vs. 12% [p = 0.0005], and 68% vs. 11% [p < 0.0001], respectively). Also, higher LDH values correlated with a higher percentage of a fully methylated reference in a linear fashion (Spearman correlation coefficient 0.18 for HLTF [p = 0.004]; 0.49 [p < .0001] for HPP1). No correlation between methylation of NEUROG1 and LDH was found in this study. Concerning the clinical characteristics, high levels of LDH as well as methylation of HLTF and HPP1 were significantly associated with larger and more advanced stages of CRC. Accordingly, these three markers were correlated with significantly shorter survival in the overall population. Moreover, all three identified patients with a worse prognosis in the subgroup of stage IV patients. CONCLUSIONS: We were able to provide evidence that methylation of HLTF and especially HPP1 detected in serum is strongly correlated with cell death in CRC using LDH as surrogate marker. Additionally, we found that prognostic information is given by both HLTF and HPP1 as well as LDH. In sum, determining the methylation of HLTF and HPP1 in serum might be useful in order to identify patients with more aggressive tumors.

Variability of linezolid concentrations after standard dosing in critically ill patients: a prospective observational study.

INTRODUCTION: Severe infections in intensive care patients show high morbidity and mortality rates. Linezolid is an antimicrobial drug frequently used in critically ill patients. Recent data indicates that there might be high variability of linezolid serum concentrations in intensive care patients receiving standard doses. This study was aimed to evaluate whether standard dosing of linezolid leads to therapeutic serum concentrations in critically ill patients. METHODS: In this prospective observational study, 30 critically ill adult patients with suspected infections received standard dosing of 600 mg linezolid intravenously twice a day. Over 4 days, multiple serum samples were obtained from each patient, in order to determine the linezolid concentrations by liquid chromatography tandem mass spectrometry. RESULTS: A high variability of serum linezolid concentrations was observed (range of area under the linezolid concentration time curve over 24 hours (AUC24) 50.1 to 453.9 mg/L, median 143.3 mg*h/L; range of trough concentrations (Cmin) < 0.13 to 14.49 mg/L, median 2.06 mg/L). Furthermore, potentially subtherapeutic linezolid concentrations over 24 hours and at single time points (defined according to the literature as AUC24 < 200 mg*h/L and Cmin < 2 mg/L) were observed for 63% and 50% of the patients, respectively. Finally, potentially toxic levels (defined as AUC24 > 400 mg*h/L and Cmin > 10 mg/L) were observed for 7 of the patients. CONCLUSIONS: A high variability of linezolid serum concentrations with a substantial percentage of potentially subtherapeutic levels was observed in intensive care patients. The findings suggest that therapeutic drug monitoring of linezolid might be helpful for adequate dosing of linezolid in critically ill patients. TRIAL REGISTRATION: Clinicaltrials.gov NCT01793012. Registered 24 January 2013.

Immunogenic cell death biomarkers HMGB1, RAGE, and DNAse indicate response to radioembolization therapy and prognosis in colorectal cancer patients.

Radioembolization therapy (RE) is an efficient locoregional treatment for liver metastases from colorectal cancer. Serum biomarkers involved in immunogenic cell death are potentially valuable for early predicting therapy response and estimating prognosis. In a prospective observation study, blood samples were taken from 49 consecutive colorectal cancer patients with extensive hepatic metastases before, 24 and 48 hr after RE. Serum levels of high mobility group box 1 (HMGB1), receptor of glycation end products (RAGE) and activity of desoxyribonuclease were compared with response to therapy regularly determined radiologically 3 months after therapy and with overall survival. Serum levels of HMGB1 were increased already 24 hr after RE, while RAGE levels were decreased and DNAse remained unchanged. In radiological staging, 35 patients demonstrated disease progression while 14 patients had stable disease or remission. Serum HMGB1 levels 24 hr after RE were significantly higher in progressive than in nonprogressive patients while for RAGE and DNAse no difference was observed between the response groups. Concerning overall survival, high pretherapeutic (0 hr) and 24 hr levels of HMGB1 were associated with poor outcome. Multivariate analysis including HMGB1, tumor, liver and inflammation markers revealed HMGB1 and CRP as independent prognostic parameters. HMGB1 is a valuable serum biomarker for early estimation of therapy response and prognosis in colorectal cancer patients with liver metastases undergoing RE therapy.

Circulating nucleosomes and immunogenic cell death markers HMGB1, sRAGE and DNAse in patients with advanced pancreatic cancer undergoing chemotherapy.

Serum biomarkers are urgently needed for patient stratification and efficient treatment monitoring in pancreatic cancer (PC). Within a prospective diagnostic observation study, blood samples were obtained from 78 patients with advanced PC before and weekly during the course of palliative chemotherapy. Circulating nucleosomes and immunogenic cell death markers, high-mobility group box 1 (HMGB1), soluble receptors of advanced glycation end products (sRAGE) and DNAse activity, were measured by enzyme-linked immunosorbent assay and correlated with results of radiological staging after 2 months of treatment, with time to progression (TTP) and overall survival (OS). Median TTP and OS of PC patients were 3.9 and 7.7 months, respectively. Pretherapeutic baseline biomarker levels did not correlate with objective response; however, nucleosome levels on day (d) 28 were higher (p = 0.048) and sRAGE levels at time of staging (d56) were lower in progressive patients (p = 0.046). Concerning estimation of prognosis, high nucleosome levels (d7, d14, d21 and d56), low sRAGE levels (d56) and DNAse activity courses (d0-d7) correlated with TTP, whereas high nucleosomes (d7, d14 and d56), high HMGB1 (d21 and d56) and DNAse (d0-d7) were associated with OS. After adjustment to Karnofsky performance score, nucleosomes and HMGB1 (both d56) and DNAse (d0-d7) remained independent prognostic factors. Thus, courses of circulating nucleosomes and immunogenic cell death markers HMGB1 and sRAGE show prognostic relevance in PC patients undergoing chemotherapy.

Circulating immunogenic cell death biomarkers HMGB1 and RAGE in breast cancer patients during neoadjuvant chemotherapy.

Neoadjuvant chemotherapy in breast cancer patients aims at preoperative reduction of tumor volume for better resection results and prognosis. As not all patients respond to neoadjuvant therapy, predictive biomarkers are needed for more efficient individual management. In prospectively collected sera of 51 consecutive locally confined breast cancer (LBC) patients receiving preoperative, neoadjuvant chemotherapy, value level kinetics of soluble high mobility group box 1 (HMGB1), soluble receptor for advanced glycation end products (sRAGE) as well as the established breast cancer biomarkers CA 15-3 and carcinoembryonic antigen (CEA) were investigated and correlated with therapy response objectified by pathological staging at surgery. In addition, biomarkers were measured in sera of 30 healthy controls (HC), 13 patients with benign breast diseases, and 28 metastatic breast cancer (MBC) patients. Pretherapeutic levels of soluble HMGB1 were decreased in MBC, while sRAGE was already decreased in LBC. In contrast, CA 15-3 and CEA were strongly elevated in MBC, but not in LBC. Combination of sRAGE and CA 15-3 enabled best discrimination of LBC from HC (AUC 78.2 %; sens 58 % at 95 % spec), while CA15-3 and CEA discriminated best between MBC and all controls (AUC 90.9 %; sens 70 % at 95 % spec). In LBC patients undergoing neoadjuvant chemotherapy, nine patients achieved complete remission (CR), 29 achieved partial remission (PR), while 13 had no change of disease (NC). NC patients tended to have higher HMGB1 and lower sRAGE levels before therapy onset (p = 0.056 and p = 0.054), while CA 15-3 and CEA did not predict therapeutic outcome. Furthermore, kinetics of HMGB1 during therapy correlated with efficacy of the treatment (p = 0.053). Markers of immunogenic cell death are valuable for the diagnosis of MBC and early estimation of response to neoadjuvant therapy in LBC patients.

Cell-free circulating ALU repeats in serum have a prognostic value for colorectal cancer patients.

BACKGROUND: Carcinoembryonic antigen (CEA) is the only established serum biomarker for colorectal cancer (CRC). To facilitate therapy decisions and improve the overall survival of CRC patients, prognostic biomarkers are required. OBJECTIVE: We studied the prognostic value of five different cell free circulating DNA (fcDNA) fragments. The potential markers were ALU115, ALU247, LINE1-79, LINE1-300 and ND1-mt. METHODS: The copy numbers of the DNA fragments were measured in the peripheral blood serum of 268 CRC patients using qPCR, the results were compared to common and previously described markers. RESULTS: We found that ALU115 and ALU247 fcDNA levels correlate significantly with several clinicopathological parameters. An increased amount of ALU115 and ALU247 fcDNA fragments coincides with methylation of HPP1 (P< 0.001; P< 0.01), which proved to be a prognostic marker itself in former studies and also with increased CEA level (both P< 0.001). ALU115 and ALU247 can define patients with poor survival in UICC stage IV (ALU115: HR = 2.9; 95% Cl 1.8-4.8, P< 0.001; ALU247: HR = 2.2; 95% Cl 1.3-3.6; P= 0.001). Combining ALU115 and HPP1, the prognostic value in UICC stage IV is highly significant (P< 0.001). CONCLUSIONS: This study shows that an increased level of ALU fcDNA is an independent prognostic biomarker for advanced colorectal cancer disease.

Prognostic role of methylated free circulating DNA in colorectal cancer.

DNA hypermethylation is frequently found in colorectal cancer (CRC). Methylation of helicase-like transcription factor (HLTF) and hyperplastic polyposis 1 (HPP1) are potential and carcinoembryonic antigen (CEA) is an established prognostic factor in serum of patients with CRC. The aim of this study was to perform a direct comparison of the prognostic roles of these markers. Methylation status of HLTF and HPP1 was examined in pretherapeutic sera of 311 patients with CRC and matched primary tissues of 54 stage IV patients using methylation-specific quantitative PCR. CEA was determined using an immunoenzymometric assay. Methylation of HLTF and HPP1 DNA in serum significantly correlated with tumor size, stage, grade and metastatic disease. HPP1 methylation correlated with nodal status. Overall survival was shortened in case of methylation of HLTF or HPP1 or elevated levels of CEA (p < 0.0001 for all). In stage IV, patients survival was impaired if HLTF (p = 0.0005) or HPP1 (p = 0.0003) were methylated or CEA was above the median of 27 ng/ml (p = 0.002). Multivariate analysis revealed that methylation of HLTF [hazard ratio (HR) 1.8, p = 0.0438], HPP1 (HR 1.6, p = 0.0495) and CEA >27 ng/ml (HR 1.7, p = 0.0317) were independent prognostic factors in stage IV. The combination of any two or all three of these factors outperformed each marker on its own. In conclusion, the presence of methylated DNA of the genes HLTF or HPP1 in serum are independent prognostic factors in metastasized CRC. Prospective validation is required to determine their usefulness in clinical routine along with the established marker CEA.

Predictive value of immunogenic cell death biomarkers HMGB1, sRAGE, and DNase in liver cancer patients receiving transarterial chemoembolization therapy.

Transarterial chemoembolization (TACE) therapy is an effective locoregional anticancer treatment for liver cancer patients. Serum biomarkers involved in immunogenic cell death may be valuable for early predicting therapy response and estimating prognosis. Sera of 50 prospectively and consecutively included hepatocellular carcinoma (HCC) patients, undergoing TACE therapy, were taken before and 24 h after TACE application. In these samples, soluble biomarkers involved in immunogenic cell death, and among them, high-mobility group box 1 (HMGB1), soluble receptor of advanced glycation end products (sRAGE), and DNase activity were measured. They were compared with radiological response to therapy. A total of 71 TACE therapies were evaluated, of which 32 were classified as "no progression," and 39, as "progression." While HMGB1 levels increased already 24 h after TACE, there was an early decrease of sRAGE and DNase activity. Pretherapeutic and 24-h values of sRAGE were significantly higher in the no progression group than those in the progression group. There was no difference with respect to treatment response for DNase and HMGB1. Soluble RAGE is a new parameter with predictive relevance in primary liver cancer patients undergoing TACE therapy.

Prognostic relevance of oncological serum biomarkers in liver cancer patients undergoing transarterial chemoembolization therapy.

As transarterial chemoembolization (TACE) therapy is an effective locoregional treatment for patients with advanced liver cancer, prognostic biomarkers are highly needed for pretherapeutic stratification of patients to TACE therapy. Sera of 50 prospectively and consecutively included patients with hepatocellular carcinoma (HCC) undergoing TACE were taken before and 24 h after TACE application. Levels of liver-specific, tumor-related, and cell death biomarkers were analyzed and correlated with overall patient survival. The study was particularly focused on patients treated by TACE with palliative intention (N = 38). Sixteen of 38 patients died within 1 year after TACE, 22 were still alive. In univariate analysis, high levels of cytokeratin 19-fragments (CYFRA 21-1), alpha fetoprotein (AFP), and low choline esterase (CHE) levels measured before and 24 h after TACE were correlated with unfavorable outcome. Further high pretherapeutic lactate dehydrogenase (LDH), aspartate-aminotransferase, and bilirubin levels as well as high 24 h C-reactive protein values were associated with poor survival. In multivariate analysis of clinical and only pretherapeutic biomarkers, AFP, CHE, and LDH showed to be independent prognostic parameters. When additionally 24 h values were included, CHE (24 h) and AFP (24 h) were the strongest independent prognostic biomarkers with a slightly higher prognostic power (Akaike's information criterion 90.3 vs. 92.7). The combination of AFP, CHE, and LDH enables efficient pretherapeutic stratification of HCC patients in advanced tumor stage for TACE therapy.