Nico van Zandwijk

Effects of Concomitant Cisplatin and Radiotherapy on Inoperable Non-Small-Cell Lung Cancer

BACKGROUND AND METHODS Cisplatin (cis-diamminedichloroplatinum) has been reported to enhance the cell-killing effect of radiation, an effect whose intensity varies with the schedule of administration. We randomly assigned 331 patients with nonmetastatic inoperable non-small-cell lung cancer to one of three treatments: radiotherapy for two weeks (3 Gy given 10 times, in five fractions a week), followed by a three-week rest period and then radiotherapy for two more weeks (2.5 Gy given 10 times, five fractions a week); radiotherapy on the same schedule, combined with 30 mg of cisplatin per square meter of body-surface area, given on the first day of each treatment week; or radiotherapy on the same schedule, combined with 6 mg of cisplatin per square meter, given daily before radiotherapy. RESULTS Survival was significantly improved in the radiotherapy-daily-cisplatin group as compared with the radiotherapy group (P = 0.009): survival in the radiotherapy-daily-cisplatin group was 54 percent at one year, 26 percent at two years, and 16 percent at three years, as compared with 46 percent, 13 percent, and 2 percent, respectively, in the radiotherapy group. Survival in the radiotherapy-weekly-cisplatin group was intermediate (44 percent, 19 percent, and 13 percent) and not significantly different from survival in either of the other two groups. The survival benefit of daily combined treatment was due to improved control of local disease (P = 0.003). Survival without local recurrence was 59 percent at one year and 31 percent at two years in the radiotherapy-daily-cisplatin group; 42 percent and 30 percent, respectively, in the radiotherapy-weekly-cisplatin group; and 41 percent and 19 percent, respectively, in the radiotherapy group. Cisplatin induced nausea and vomiting in 86 percent of the patients given it weekly and in 78 percent of those given it daily; these effects were severe in 26 percent and 28 percent, respectively. CONCLUSIONS Cisplatin, given daily in combination with the radiotherapy described here to patients with nonmetastatic but inoperable non-small-cell lung cancer, improved rates of survival and control of local disease at the price of substantial side effects.

K-ras Oncogene Activation as a Prognostic Marker in Adenocarcinoma of the Lung

BACKGROUND The capability of activated oncogenes to induce malignant transformation of immortalized cells in vitro has suggested that they have a similar role in the pathogenesis of human tumors. We previously found that activation of the K-ras oncogene by a point mutation in codon 12 occurs in about one third of human lung adenocarcinomas. METHODS We studied the clinical importance of this oncogene-activation in 69 patients with lung adenocarcinoma in whom complete resection of the tumor was possible. The polymerase chain reaction was used to amplify ras-specific sequences of DNA isolated from frozen or paraffin-embedded tumor samples. Ras point mutations were subsequently detected and classified with the use of mutation-specific oligonucleotide probes. RESULTS Nineteen of the tumors harbored a point mutation in codon 12 of the K-ras oncogene. There was no association between the K-ras point mutation and the age at diagnosis, sex, or presence of previous or concurrent neoplasms. Tumors positive for K-ras point mutations tended to be smaller and less differentiated than those without mutations. The K-ras codon-12 point mutation was a strong (and unfavorable) prognostic factor: 12 of the 19 patients with K-ras point-mutation-positive tumors died during the follow-up period, as compared with 16 of the 50 patients with no mutation in the K-ras oncogene (P = 0.002). This difference in prognosis was also reflected in the duration of disease-free survival (P = 0.038) and in the number of deaths due to cancer (P less than 0.001). CONCLUSIONS The presence of K-ras point mutations defines a subgroup of patients with lung adenocarcinoma in whom the prognosis is very poor and disease-free survival is not usually long despite radical resection and a small tumor load.

Mutational activation of the K-ras oncogene. A possible pathogenetic factor in adenocarcinoma of the lung.

To define the role of cellular oncogenes in human cancers, we studied the prevalence of mutational activation of ras oncogenes in untreated non-small-cell lung cancer. Genomic DNA was extracted from 39 tumor specimens obtained by thoracotomy and was examined for activating point mutations in codons 12, 13, and 61 of the H-ras, K-ras, and N-ras genes. A novel, highly sensitive assay based on oligonucleotide hybridization following an in vitro amplification step was employed. The K-ras gene was found to be activated by point mutations in codon 12 in 5 of 10 adenocarcinomas. Two of these tumors were less than 2 cm in size and had not metastasized. No ras gene mutations were observed in 15 squamous-cell carcinomas, 10 large-cell carcinomas, 1 carcinoid, 2 metastatic adenocarcinomas from primary tumors outside the lung, and 1 small-cell carcinoma. An approximately 20-fold amplification of the unmutated K-ras gene was observed in a tumor that proved to be a solitary lung metastasis of a rectal carcinoma. We conclude that mutational K-ras activation may be an important early event in the pathogenesis of adenocarcinoma of the lung but that amplification of ras genes or mutational activation of H-ras or N-ras does not play a major part in non-small-cell lung cancer.

Randomized Controlled Trial of Resection Versus Radiotherapy After Induction Chemotherapy in Stage IIIA-N2 Non–Small-Cell Lung Cancer

BACKGROUND Induction chemotherapy before surgical resection increases survival compared with surgical resection alone in patients with stage IIIA-N2 non-small-cell lung cancer (NSCLC). We hypothesized that, following a response to induction chemotherapy, surgical resection would be superior to thoracic radiotherapy as locoregional therapy. METHODS Selected patients with histologic or cytologic proven stage IIIA-N2 NSCLC were given three cycles of platinum-based induction chemotherapy. Responding patients were subsequently randomly assigned to surgical resection or radiotherapy. Survival curves were estimated using Kaplan-Meier analyses from time of randomization. RESULTS Induction chemotherapy resulted in a response rate of 61% (95% confidence interval [CI] = 57% to 65%) among the 579 eligible patients. A total of 167 patients were allocated to resection and 165 to radiotherapy. Of the 154 (92%) patients who underwent surgery, 14% had an exploratory thoracotomy, 50% a radical resection, 42% a pathologic downstaging, and 5% a pathologic complete response; 4% died after surgery. Postoperative radiotherapy was administered to 62 (40%) of patients in the surgery arm. Among the 154 (93%) irradiated patients, overall compliance to the radiotherapy prescription was 55%, and grade 3/4 acute and late esophageal and pulmonary toxic effects occurred in 4% and 7%; one patient died of radiation pneumonitis. Median and 5-year overall survival for patients randomly assigned to resection versus radiotherapy were 16.4 versus 17.5 months and 15.7% versus 14%, respectively (hazard ratio = 1.06, 95% CI = 0.84 to 1.35). Rates of progression-free survival were also similar in both groups. CONCLUSION In selected patients with pathologically proven stage IIIA-N2 NSCLC and a response to induction chemotherapy, surgical resection did not improve overall or progression-free survival compared with radiotherapy. In view of its low morbidity and mortality, radiotherapy should be considered the preferred locoregional treatment for these patients.

Circulating microRNAs: Association with disease and potential use as biomarkers

The control of gene expression by microRNAs influences many cellular processes and has been implicated in the control of many (patho)physiological states. Recently, microRNAs have been detected in serum and plasma, and circulating microRNA profiles have now been associated with a range of different tumour types, diseases such as stroke and heart disease, as well as altered physiological states such as pregnancy. Here we review the disease-specific profiles of circulating microRNAs, and the methodologies used for their detection and quantification. We also discuss possible functions of circulating microRNAs and their potential as non-invasive biomarkers.

Haemolysis during Sample Preparation Alters microRNA Content of Plasma

The presence of cell-free microRNAs (miRNAs) has been detected in a range of body fluids. The miRNA content of plasma/serum in particular has been proposed as a potential source of novel biomarkers for a number of diseases. Nevertheless, the quantification of miRNAs from plasma or serum is made difficult due to inefficient isolation and lack of consensus regarding the optimal reference miRNA. The effect of haemolysis on the quantification and normalisation of miRNAs in plasma has not been investigated in great detail. We found that levels of miR-16, a commonly used reference gene, showed little variation when measured in plasma samples from healthy volunteers or patients with malignant mesothelioma or coronary artery disease. Including samples with evidence of haemolysis led to variation in miR-16 levels and consequently decreased its ability to serve as a reference. The levels of miR-16 and miR-451, both present in significant levels in red blood cells, were proportional to the degree of haemolysis. Measurements of the level of these miRNAs in whole blood, plasma, red blood cells and peripheral blood mononuclear cells revealed that the miRNA content of red blood cells represents the major source of variation in miR-16 and miR-451 levels measured in plasma. Adding lysed red blood cells to non-haemolysed plasma allowed a cut-off level of free haemoglobin to be determined, below which miR-16 and miR-451 levels displayed little variation between individuals. In conclusion, increases in plasma miR-16 and miR-451 are caused by haemolysis. In the absence of haemolysis the levels of both miR-16 and miR-451 are sufficiently constant to serve as normalisers.

Prognostic Relevance of Response Evaluation Using [18F]-2-Fluoro-2-Deoxy-D-Glucose Positron Emission Tomography in Patients With Locally Advanced Non–Small-Cell Lung Cancer

PURPOSE The objective of this study was to determine the accuracy of (early) response measurements using [18F]-2-fluoro-2-deoxy-D-glucose positron emission tomography (18FDG PET) with respect to survival of patients with stage IIIA-N2 non-small-cell lung cancer (NSCLC) undergoing induction chemotherapy (IC), with a comparative analysis of PET methods. PATIENTS AND METHODS In a prospective multicenter study, PET was performed in patients before IC and after one and three cycles. Computed tomography (CT) was performed before and after IC. Glucose consumption (metabolic rate of glucose [MRglu]) was measured using Patlak graphical analysis and correlated with simplified methods. Mediastinal lymph node (MLN) status was assessed visually. Cox proportional hazards analysis was used to determine the prognostic relevance of CT and PET measures of response with respect to survival. RESULTS Complete PET data sets were available in 47 patients. Median survival was 21 months. MLN status after IC by PET predicted survival (hazard ratio [HR], 2.33; 95% CI, 1.04 to 5.22; P = .04) in contrast with CT (HR, 1.87; 95% CI, 0.81 to 4.30; P = .14). Residual MRglu after IC proved to be the best prognostic factor (HR, 1.95; 95% CI, 1.28 to 2.97; P = .002). Multivariate stepwise analysis showed that PET identified prognostically different strata in patients considered responsive according to CT. Residual MRglu after one cycle selected patients with different outcomes (HR, 2.04; 95% CI, 1.18 to 3.52; P = .01). Simplified quantitative 18FDG PET methods were correlated with Patlak graphical analysis during and after therapy (r > or = 0.90). CONCLUSION 18FDG PET has additional value over CT in monitoring response to IC in patients with stage IIIA-N2 NSCLC, and it seems feasible to predict survival early during IC. Simple semiquantitative and complex PET methods perform equally well.

High blood neutrophil-to-lymphocyte ratio is an indicator of poor prognosis in malignant mesothelioma patients undergoing systemic therapy

Purpose: Asbestos-induced chronic inflammation is implicated in the pathogenesis of malignant mesothelioma (MM). We have investigated blood neutrophil-to-lymphocyte ratio (NLR), an index of systemic inflammation, as a prognostic factor in MM patients. Experimental Design: Patients with MM who had systemic therapy at participating institutes were studied. Potential prognostic factors such as age, gender, performance status, histologic subtype, and baseline laboratory parameters, including NLR, were analyzed. Overall survival from commencement of therapy was determined by the Kaplan–Meier method. Multivariate analyses using Cox Regression model were performed with significant factors (P ≤ 0.05) to determine their independent effect. Results: A total of 173 MM patients undergoing systemic therapy including 119 patients receiving first-line therapy and 54 patients receiving second- or third-line therapy were included in this retrospective evaluation. Forty-two percent of patients had an elevated NLR at baseline. The following variables were predictive of survival: female gender (P = 0.044), epithelioid histologic subtype (P < 0.001), baseline white blood cell count less than 8.3 × 109/L (P = 0.008), baseline platelet count 400 × 109/L or less (P = 0.05), and NLR of 5 or less (P < 0.001). After multivariate analysis, histologic epithelioid subtype [hazard ratio (HR) = 2.0; 95% confidence interval (CI) = 1.3–2.9; P = 0.001], and NLR less than 5 (HR = 2.7; 95% CI = 1.8–3.9; P < 0.001) remained independent predictors. The 1-year survival rate was 60% versus 26%, whereas the 2-year survival rate was 34% versus 10% for NLR less than 5 and 5 or greater, respectively. In the separate analyses of chemotherapy-naive and previously treated patient groups, NLR was an independent predictor of survival in both groups. Conclusion: Our results indicate that NLR is an independent predictor of survival for patients with MM undergoing systemic therapy. Clin Cancer Res; 16(23); 5805–13. ©2010 AACR.

Randomized Phase II and Pharmacogenetic Study of Pemetrexed Compared With Pemetrexed Plus Carboplatin in Pretreated Patients With Advanced Non–Small-Cell Lung Cancer

PURPOSE We performed a randomized phase II trial comparing pemetrexed with pemetrexed plus carboplatin (PC) in patients experiencing relapse after platinum-based chemotherapy. PATIENTS AND METHODS Main eligibility criteria were histologic or cytologic proof of advanced non-small-cell lung cancer (NSCLC), relapse more than 3 months after platinum-based chemotherapy, normal organ function, and Eastern Cooperative Oncology Group performance status 0 to 2. Patients were randomly assigned to pemetrexed 500 mg/m(2) (arm A) or carboplatin area under the curve 5 and pemetrexed 500 mg/m(2) (arm B), both administered intravenously every 3 weeks. Response assessment was performed every 6 weeks; toxicity assessment was performed every 3 weeks. Primary end point was time to progression (TTP); secondary end points were objective response rate (ORR), overall survival (OS), and toxicity. The study was designed to detect a 33% decrease in the hazard of disease progression in the combination arm (alpha = 0.05, two-sided log-rank test). Polymorphisms of thymidylate synthase, the reduced folate carrier, gamma-glutamyl hydrolase, and methylenetetrahydrofolate reductase (MTHF) were investigated in peripheral WBCs of consenting patients. RESULTS Two hundred forty patients were enrolled. Median TTP was 2.8 months for arm A versus 4.2 months for arm B (hazard ratio, 0.67; 95% CI, 0.51 to 0.89; P = .005). Median OS was 7.6 months and 8.0 months and ORR was 4% and 9% for arms A and B, respectively. Subgroup analyses found adenocarcinoma to be associated with favorable outcome. Toxicities in both arms was negligible, with one potential toxic death in arm A. Patients with MTHFR C677T homozygous mutation had increased progression-free survival compared with patients with wild-type or heterozygous mutations (P = .03). CONCLUSION PC as second-line treatment for relapsed NSCLC resulted in a significant 33% reduction of the hazard of disease progression as compared with pemetrexed alone.

An Immune Response Enriched 72-Gene Prognostic Profile for Early-Stage Non–Small-Cell Lung Cancer

Purpose: Current staging methods are imprecise for predicting prognosis of early-stage non–small-cell lung cancer (NSCLC). We aimed to develop a gene expression profile for stage I and stage II NSCLC, allowing identification of patients with a high risk of disease recurrence within 2 to 3 years after initial diagnosis. Experimental Design: We used whole-genome gene expression microarrays to analyze frozen tumor samples from 172 NSCLC patients (pT1-2, N0-1, M0) from five European institutions, who had undergone complete surgical resection. Median follow-up was 89 months (range, 1.2-389) and 64 patients developed a recurrence. A random two thirds of the samples were assigned as the training cohort with the remaining samples set aside for independent validation. Cox proportional hazards models were used to evaluate the association between expression levels of individual genes and patient recurrence-free survival. A nearest mean analysis was used to develop a gene-expression classifier for disease recurrence. Results: We have developed a 72-gene expression prognostic NSCLC classifier. Based on the classifier score, patients were classified as either high or low risk of disease recurrence. Patients classified as low risk showed a significantly better recurrence-free survival both in the training set (P < 0.001; n = 103) and in the independent validation set (P < 0.01; n = 69). Genes in our prognostic signature were strongly enriched for genes associated with immune response. Conclusions: Our 72-gene signature is closely associated with recurrence-free and overall survival in early-stage NSCLC patients and may become a tool for patient selection for adjuvant therapy.