Markus K. A. Herrmann

KRAS and BRAF mutations in patients with rectal cancer treated with preoperative chemoradiotherapy

BACKGROUND AND PURPOSE KRAS and BRAF are mutated in 35% and 10% of colorectal cancers, respectively. However, data specifically for locally advanced rectal cancers are scarce, and the frequency of KRAS mutations in codons 61 and 146 remains to be established. MATERIALS AND METHODS DNA was isolated from pre-therapeutic biopsies of 94 patients who were treated within two phase-III clinical trials receiving preoperative chemoradiotherapy. Mutation status of KRAS exons 1-3 and BRAF exon 15 was established using the ABI PRISM Big Dye Sequencing Kit and subsequently correlated with clinical parameters. RESULTS Overall, KRAS was mutated in 45 patients (48%). Twenty-nine mutations (64%) were located in codon 12, 10 mutations (22%) in codon 13, and 3 mutations (7%) in codons 61 and 146. No V600E BRAF mutation was detected. The presence of KRAS mutations was correlated neither with tumor response or lymph node status after preoperative chemoradiotherapy nor with overall survival or disease-free survival. When KRAS exon 1 mutations were separated based on the amino-acid exchange, we again failed to detect significant correlations (p=0.052). However, G12V mutations appeared to be associated with higher rates of tumor regression than G13D mutations (p=0.012). CONCLUSION We are the first to report the mutation status of KRAS and BRAF in pre-therapeutic biopsies from locally advanced rectal cancers. The high number of KRAS mutations in codons 61 and 146 emphasizes the importance to expand current mutation analyses, whereas BRAF mutations are not relevant for rectal carcinogenesis. Although the KRAS mutation status was not correlated with response, the subtle difference between G12V and G13D mutations warrants analysis of a larger patient population.

The delineation of target volumes for radiotherapy of lung cancer patients.

PURPOSE Differences in the delineation of the gross target volume (GTV) and planning target volume (PTV) in patients with non-small-cell lung cancer are considerable. The focus of this work is on the analysis of observer-related reasons while controlling for other variables. METHODS In three consecutive patients, eighteen physicians from fourteen different departments delineated the GTV and PTV in CT-slices using a detailed instruction for target delineation. Differences in the volumes, the delineated anatomic lymph node compartments and differences in every delineated pixel of the contoured volumes in the CT-slices (pixel-by-pixel-analysis) were evaluated for different groups: ten radiation oncologists from ten departments (ROs), four haematologic oncologists and chest physicians from four departments (HOs) and five radiation oncologists from one department (RO1D). RESULTS Agreement (overlap > or = 70% of the contoured pixels) for the GTV and PTV delineation was found in 16.3% and 23.7% (ROs), 30.4% and 38.6% (HOs) and 32.8% and 35.9% (RO1D), respectively. CONCLUSION A large interobserver variability in the PTV and much more in the GTV delineation were observed in spite of a detailed instruction for delineation. The variability was smallest for group ROID where due to repeated discussions and uniform teaching a better agreement was achieved.

High-Grade Acute Organ Toxicity During Preoperative Radiochemotherapy as Positive Predictor for Complete Histopathologic Tumor Regression in Multimodal Treatment of Locally Advanced Rectal Cancer*

Purpose:To test for a possible correlation between high-grade acute organ toxicity during preoperative radiochemotherapy and complete tumor regression after total mesorectal excision in multimodal treatment of locally advanced rectal cancer.Patients and Methods:From 2001 to 2008, 120 patients were treated. Preoperative treatment consisted of normofractionated radiotherapy at a total dose of 50.4 Gy, and either two cycles of 5-fluorouracil (5-FU) or two cycles of 5-FU and oxaliplatin. Toxicity during treatment was monitored weekly, and any toxicity CTC (Common Toxicity Criteria) ≥ grade 2 of enteritis, proctitis or cystitis was assessed as high-grade organ toxicity for later analysis. Complete histopathologic tumor regression (TRG4) was defined as the absence of any viable tumor cells.Results:A significant coherency between high-grade acute organ toxicity and complete histopathologic tumor regression was found, which was independent of other factors like the preoperative chemotherapy schedule. The probability of patients with acute organ toxicity ≥ grade 2 to achieve TRG4 after neoadjuvant treatment was more than three times higher than for patients without toxicity (odds ratio: 3.29, 95% confidence interval: [1.01, 10.96]).Conclusion:Acute organ toxicity during preoperative radiochemotherapy in rectal cancer could be an early predictor of treatment response in terms of complete tumor regression. Its possible impact on local control and survival is under further prospective evaluation by the authors’ working group.Ziel:Überprüfung einer möglichen Korrelation zwischen höhergradiger akuter Organtoxizität während präoperativer Radiochemotherapie und kompletter Tumorregression nach totaler mesorektaler Exzision in der multimodalen Behandlung von lokal fortgeschrittenen Rektumkarzinomen.Patienten und Methodik:Im Zeitraum von 2001 bis 2008 wurden 120 Patienten behandelt. Die präoperative Behandlung bestand aus einer normofraktionierten Radiotherapie mit einer Gesamtdosis von 50,4 Gy und entweder zwei Zyklen 5-Fluorouracil (5-FU) oder zwei Zyklen 5-FU und Oxaliplatin. Die Toxizität während der Behandlung wurde wöchentlich untersucht. Jede Toxizität ≥ Grad 2 nach CTC (Common Toxicity Criteria) in Form von Enteritis, Proktitis oder Zystitis wurde dabei als höhergradige Organtoxizität gewertet und für spätere Analysen verwendet. Bei vollständigem histopathologischen Tumoransprechen (TRG4) konnten im Operationspräparat nach neoadjuvanter Therapie keine vitalen Tumorzellen mehr identifiziert werden.Ergebnisse:Es fand sich eine signifikante Korrelation zwischen höhergradiger akuter Organtoxizität und kompletter Tumorregression, und zwar in multivariater Analyse unabhängig von anderen Faktoren wie z.B. dem präoperativen Chemotherapieregime. Die Wahrscheinlichkeit für die Patienten mit höhergradigen Nebenwirkungen, eine histopathologische Komplettremission zu entwickeln, war mehr als dreimal höher als für Patienten ohne Toxizität (Odds-Ratio: 3,29, 95%-Konfidenzintervall: [1,01, 10,96]).Schlussfolgerung:Höhergradige akute Organtoxizität während präoperativer Radiochemotherapie bei Patienten mit multimodaler Therapie lokal fortgeschrittener Rektumkarzinome könnte einen frühen Prädiktor für das Ansprechen auf die Therapie in Bezug auf die histopathologische Remission darstellen. Ob dieser Parameter auch für die lokale Kontrolle sowie das Gesamtüberleben prädiktiv sein kann, wird in weiteren prospektiven Untersuchungen durch die Arbeitsgruppe der Autoren untersucht.

Sodium butyrate enemas in the treatment of acute radiation-induced proctitis in patients with prostate cancer and the impact on late proctitis

PurposeTo evaluate prospectively the effect of sodium butyrate enemas on the treatment of acute and the potential influence on late radiation-induced proctitis.Patients and Methods31 patients had been treated with sodium butyrate enemas for radiation-induced acute grade II proctitis which had developed after 40 Gy in median. During irradiation the toxicity was evaluated weekly by the Common Toxicity Criteria (CTC) and subsequently yearly by the RTOG (Radiation Therapy Oncology Group) and LENT-SOMA scale.Results23 of 31 patients (74%) experienced a decrease of CTC grade within 8 days on median. A statistical significant difference between the incidence and the severity of proctitis before start of treatment with sodium butyrate enemas compared to 14 days later and compared to the end of irradiation treatment course, respectively, was found. The median follow-up was 50 months. Twenty patients were recorded as suffering from no late proctitis symptom. Eleven patients suffered from grade I and 2 of these patients from grade II toxicity, too. No correlation was seen between the efficacy of butyrate enemas on acute proctitis and prevention or development of late toxicity, respectively.ConclusionSodium butyrate enemas are effective in the treatment of acute radiation-induced proctitis in patients with prostate cancer but have no impact on the incidence and severity of late proctitis.ZusammenfassungZielDiese prospektive Untersuchung wurde durchgeführt, um den Effekt von Natriumbutyrat-Einläufen in der Therapie der akuten Proktitis sowie den potentiellen Einfluss auf radiogene rektale Spätreaktionen zu evaluieren.Patienten und Methodik31 Patienten wurden mit Natriumbutyrat-Einläufen bei radiogen induzierter akuter Grad-II-Proktitis behandelt, die im Mittel nach 40 Gy aufgetreten war. Während der Radiotherapie wurde die Toxizität wöchentlich anhand der Common Toxicity Criteria (CTC) und anschließend jährlich anhand der RTOG- und LENT-SOMA-Skalen erhoben.Ergebnisse23 von 31 Patienten (74%) erfuhren eine Abnahme des CTC-Grades innerhalb von 8 Tagen im Median. Dadurch war der Unterschied in der Häufigkeit und Schwere der Proktitis vor Therapiebeginn und 14 Tage später bzw. am Ende der Radiotherapie statistisch signifikant. Der mediane Follow-up lag bei 50 Monaten. 20 Patienten entwickelten keine späte Proktitis. 11 Patienten entwickelten eine Grad-I- und 2 von diesen Patienten ebenfalls eine Grad-II-Toxizität. Es konnte keine signifikante Korrelation zwischen der Effektivität der Natriumbutyrat-Einläufe und Prävention bzw. Entwicklung einer späten Toxizität entdeckt werden.SchlussfolgerungNatriumbutyrat-Einläufe sind effektiv in der Behandlung der akuten radiotherapieinduzierten Proktitis bei Patienten mit Prostatakarzinom, haben aber keinen Einfluss auf die Häufigkeit und Schwere der späten Proktitis.

A prospective study of faecal calprotectin and lactoferrin in the monitoring of acute radiation proctitis in prostate cancer treatment.

Objective. Acute radiation proctitis is a relevant complication of pelvic radiation. The purpose of this study was to investigate two markers of gut inflammation as non-invasive diagnostic tools to evaluate acute radiation proctitis. Material and methods. Twenty patients who underwent radiotherapy for prostate cancer took part in this prospective study. Radiation-induced toxicity was evaluated weekly during radiotherapy in compliance with the CTC toxicity criteria. Stool samples from patients were examined before treatment, weekly during radiotherapy and 2 weeks after the end of radiotherapy using enzyme-linked immunosorbent assay for calprotectin and lactoferrin and correlated with the CTC toxicity. Results. Calprotectin and lactoferrin faecal values increased significantly during radiation treatment and decreased about 2 weeks after cessation of radiation. Faecal concentrations of calprotectin and lactoferrin correlated with the documented radiation proctitis symptoms (all grades together) in 15/20 patients (75%). With respect to changes in faecal concentrations and correspondence to proctitis symptoms, both markers showed parallel results in 90% of the patients. On comparing calprotectin and lactoferrin concentrations between the 4th week of radiation and the 1st week, it was found that patients with any grade of toxicity exhibited a significantly higher increase in calprotectin (p=0.044) and lactoferrin (p=0.05), respectively, compared with those without toxicity. Conclusions. Calprotectin and lactoferrin faecal values changed during radiation treatment and after cessation of radiation, with correlation to acute proctitis symptoms in most of the patients. Before markers are used to monitor acute radiation proctitis, further experience should be acquired. Patients will be followed to determine the predictive value of the two tested markers for chronic radiation proctitis.

High-Grade Acute Organ Toxicity as a Positive Prognostic Factor in Primary Radiochemotherapy for Anal Carcinoma

PURPOSE To test for a possible correlation between high-grade acute organ toxicity during primary radiochemotherapy and treatment outcome for patients with anal carcinoma. METHODS AND MATERIALS From 1991 to 2009, 72 patients with anal carcinoma were treated at our department (10 patients had stage I, 28 patients had stage II, 11 patients had stage IIIA, and 13 patients had stage IIIB cancer [Union Internationale Contre le Cancer criteria]). All patients received normofractionated (1.8 Gy/day, five times/week) whole-pelvis irradiation including iliac and inguinal lymph nodes with a cumulative dose of 50.4 Gy. Concomitant chemotherapy regimen consisted of two cycles of 5-fluorouracil (1,000 mg/m(2)total body surface area (TBSA)/day as continuous intravenous infusion on days 1-4 and 29-32) and mitomycin C (10 mg/m(2)/TBSA, intravenously on days 1 and 29). Toxicity during treatment was monitored weekly, and any incidence of Common Toxicity Criteria (CTC) grade of ≥3 for skin reaction, cystitis, proctitis, or enteritis was assessed as high-grade acute organ toxicity for later analysis. RESULTS We found significant correlation between high-grade acute organ toxicity and overall survival, locoregional control, and stoma-free survival, which was independent in multivariate analysis from other possible prognostic factors: patients with a CTC acute organ toxicity grade of ≥3 had a 5-year overall survival rate of 97% compared to 30% in patients without (p < 0.01, multivariate analysis; 97% vs. 48%, p = 0.03 for locoregional control, and 95% vs. 59%, p = 0.05 for stoma-free survival). CONCLUSIONS Our data indicate that normal tissue and tumor tissue may behave similarly with respect to treatment response, since high-grade acute organ toxicity during radiochemotherapy showed itself to be an independent prognostic marker in our patient population. This hypothesis should be further analyzed by using biomolecular and clinical levels in future clinical trials.

Sodium butyrate enemas in the treatment of acute radiation-induced proctitis in patients with prostate cancer and the impact on late proctitis. A prospective evaluation.

PurposeTo evaluate prospectively the effect of sodium butyrate enemas on the treatment of acute and the potential influence on late radiation-induced proctitis.Patients and Methods31 patients had been treated with sodium butyrate enemas for radiation-induced acute grade II proctitis which had developed after 40 Gy in median. During irradiation the toxicity was evaluated weekly by the Common Toxicity Criteria (CTC) and subsequently yearly by the RTOG (Radiation Therapy Oncology Group) and LENT-SOMA scale.Results23 of 31 patients (74%) experienced a decrease of CTC grade within 8 days on median. A statistical significant difference between the incidence and the severity of proctitis before start of treatment with sodium butyrate enemas compared to 14 days later and compared to the end of irradiation treatment course, respectively, was found. The median follow-up was 50 months. Twenty patients were recorded as suffering from no late proctitis symptom. Eleven patients suffered from grade I and 2 of these patients from grade II toxicity, too. No correlation was seen between the efficacy of butyrate enemas on acute proctitis and prevention or development of late toxicity, respectively.ConclusionSodium butyrate enemas are effective in the treatment of acute radiation-induced proctitis in patients with prostate cancer but have no impact on the incidence and severity of late proctitis.ZusammenfassungZielDiese prospektive Untersuchung wurde durchgeführt, um den Effekt von Natriumbutyrat-Einläufen in der Therapie der akuten Proktitis sowie den potentiellen Einfluss auf radiogene rektale Spätreaktionen zu evaluieren.Patienten und Methodik31 Patienten wurden mit Natriumbutyrat-Einläufen bei radiogen induzierter akuter Grad-II-Proktitis behandelt, die im Mittel nach 40 Gy aufgetreten war. Während der Radiotherapie wurde die Toxizität wöchentlich anhand der Common Toxicity Criteria (CTC) und anschließend jährlich anhand der RTOG- und LENT-SOMA-Skalen erhoben.Ergebnisse23 von 31 Patienten (74%) erfuhren eine Abnahme des CTC-Grades innerhalb von 8 Tagen im Median. Dadurch war der Unterschied in der Häufigkeit und Schwere der Proktitis vor Therapiebeginn und 14 Tage später bzw. am Ende der Radiotherapie statistisch signifikant. Der mediane Follow-up lag bei 50 Monaten. 20 Patienten entwickelten keine späte Proktitis. 11 Patienten entwickelten eine Grad-I- und 2 von diesen Patienten ebenfalls eine Grad-II-Toxizität. Es konnte keine signifikante Korrelation zwischen der Effektivität der Natriumbutyrat-Einläufe und Prävention bzw. Entwicklung einer späten Toxizität entdeckt werden.SchlussfolgerungNatriumbutyrat-Einläufe sind effektiv in der Behandlung der akuten radiotherapieinduzierten Proktitis bei Patienten mit Prostatakarzinom, haben aber keinen Einfluss auf die Häufigkeit und Schwere der späten Proktitis.

Acute Toxicity of Radiochemotherapy in Rectal Cancer Patients: A Risk Particularly for Carriers of the TGFB1 Pro25 variant

PURPOSE Transforming growth factor-beta1 is related to adverse events in radiochemotherapy. We investigated TGFB1 genetic variability in relation to quality of life-impairing acute organ toxicity (QAOT) of neoadjuvant radiochemotherapy under clinical trial conditions. METHODS AND MATERIALS Two independent patient cohorts (n = 88 and n = 75) diagnosed with International Union Against Cancer stage II/III rectal cancer received neoadjuvant radiation doses of 50.4 Gy combined with 5-fluorouracil-based chemotherapy. Toxicity was monitored according to Common Terminology Criteria for Adverse Events. QAOT was defined as a CTCAE grade ≥2 for at least one case of enteritis, proctitis, cystitis, or dermatitis. Nine germline polymorphisms covering the common genetic diversity in the TGFB1 gene were genotyped. RESULTS In both cohorts, all patients carrying the TGFB1 Pro25 variant experienced QAOT (positive predictive value of 100%, adjusted p = 0.0006). In a multivariate logistic regression model, gender, age, body mass index, type of chemotherapy, or disease state had no significant impact on QAOT. CONCLUSION The TGFB1 Pro25 variant could be a relevant marker for individual treatment stratification and carriers may benefit from adaptive clinical care or specific radiation techniques.

Comparison of the Micronucleus and Chromosome Aberration Techniques for the Documentationof Cytogenetic Damage in Radiochemotherapy-Treated Patients with Rectal Cancer

AbstractPurpose:The goal of the interdisciplinary Clinical Research Unit KFO179 (Biological Basis of Individual Tumor Response in Patients with Rectal Cancer) is to develop an individual Response and Toxicity Score for patients with locally advanced rectal cancer treated with neoadjuvant radiochemotherapy. The aim of the present study was to find a reliable and sensitive method with easy scoring criteria and high numbers of cell counts in a short period of time in order to analyze DNA damage in peripheral blood lymphocytes. Thus, the cytokinesis-block micronucleus (CBMN) assay and the chromosome aberration technique (CAT) were tested.Materials and Methods:Peripheral blood lymphocytes obtained from 22 patients with rectal cancer before (0 Gy), during (21.6 Gy), and after (50.4 Gy) radiochemotherapy were stimulated in vitro by phytohemagglutinin (PHA); the cultures were then processed for the CBMN assay and the CAT to compare the two methods.Results:A significant increase of chromosomal damage was observed in the course of radiochemotherapy parallel to increasing radiation doses, but independent of the chemotherapy applied. The equivalence of both methods was shown by Westlake’s equivalence test.Conclusion:The results show that the CBMN assay and the CAT are equivalent. For further investigations, we prefer the CBMN assay, because it is simpler through easy scoring criteria, allows high numbers of cell counts in less time, is reliable, sensitive, and has higher statistical power. In the future, we plan to integrate cytogenetic damage during radiochemotherapy into the planned Response and Toxicity Score within our interdisciplinary Clinical Research Unit.ZusammenfassungZiel:Ziel der interdisziplinären Klinischen Forschergruppe KFO179 (Biological Basis of Individual Tumor Response in Patients with Rectal Cancer) ist es, einen individuellen Response-/Toxizitätsscore für Patienten zu entwickeln, die bei diagnostiziertem lokal fortgeschrittenem Rektumkarzinom mit neoadjuvanter Radiochemotherapie behandelt werden. Ziel der vorliegenden Arbeit war, eine einfache und zuverlässige Methode zur Detektion des individuellen zytogenetischen Schadens durch die Radiochemotherapie herauszuarbeiten, die im weiteren Verlauf der Arbeit der Forschergruppe Anwendung finden soll. Wir verglichen dabei den Mikronukleustest (MN) mit der Chromosomenaberrationsanalyse (CAA).Patienten und Methodik:Periphere Blutlymphozyten von 22 Patienten wurden vor (0 Gy), während (21,6 Gy) und nach (50,4 Gy) Radiochemotherapie untersucht. Der zytogenetische Schaden wurde mittels MN und CAA analysiert und die Äquivalenz beider Methoden geprüft.Ergebnisse:Eine signifikante Zunahme chromosomaler Schädigungen durch die Bestrahlung in Abhängigkeit von der applizierten Dosis konnte bei beiden Techniken, unabhängig von der applizierten Chemotherapie, beobachtet werden. Die Gleichwertigkeit beider Methoden konnte durch den Äquivalenztest nach Westlake gezeigt werden.Schlussfolgerung:Es zeigte sich eine Äquivalenz der angewandten Methoden, was uns nun die Möglichkeit bietet, den MN gleichwertig gegenüber der CAA anzuwenden und für die geplanten Analysen bezüglich des individuellen Response-/Toxizitätsscores zu verwenden. Die Mikronukleustechnik ermöglicht durch leichtere Zählkriterien in kürzerer Zeit eine größere Anzahl von Zellen zu zählen, was zu einem valideren statistischen Endergebnis führt.

Faecal calprotectin and lactoferrin values during irradiation of prostate cancer correlate with chronic radiation proctitis: results of a prospective study.

Objective. Acute proctitis and chronic radiation proctitis are relevant complications of pelvic radiation. The purpose of this study was to investigate two markers of gut inflammation during and after irradiation for prostate cancer to evaluate a correlation between acute and chronic proctitis. Material and methods. Two patient groups were analysed. In group 1, stool samples from 20 patients were collected before therapy, every week during therapy, at the end of therapy, and 13 and 27 months after therapy. Group 2 comprised 47 patients who had undergone irradiation 40 months earlier. Toxicity was determined by common toxicity criteria (CTC) and the LENT soma scale. Calprotectin and lactoferrin values were determined by ELISA. Results. In group 1, acute values for both faecal markers were significantly correlated with chronic proctitis symptoms and all patients with chronic toxicity had acute proctitis symptoms with elevated faecal values. In group 2, where stool samples were solely collected 40 months after irradiation, the Pearson square test showed both a significant correlation between calprotectin and lactoferrin values and toxicity after 40 months. Conclusions. Within a group of 19 patients followed for two years after irradiation for prostate cancer, and 47 patients tested 40 months after irradiation, increased faecal values of calprotectin and lactoferrin were significantly correlated with the occurrence of chronic proctitis. This observation should be confirmed in an expanded study.