Cluster analysis of multiplex ligation-dependent probe amplification data in choroidal melanoma.

PurposeTo determine underlying correlations in multiplex ligation-dependent probe amplification (MLPA) data and their significance regarding survival following treatment of choroidal melanoma (CM).MethodsMLPA data were available for 31 loci across four chromosomes (1p, 3, 6, and 8) in tumor material obtained from 602 patients with CM treated at the Liverpool Ocular Oncology Center (LOOC) between 1993 and 2012. Data representing chromosomes 3 and 8q were analyzed in depth since their association with CM patient survival is well-known. Unsupervised k-means cluster analysis was performed to detect latent structure in the data set. Principal component analysis (PCA) was also performed to determine the intrinsic dimensionality of the data. Survival analyses of the identified clusters were performed using Kaplan-Meier (KM) and log-rank statistical tests. Correlation with largest basal tumor diameter (LTD) was investigated.ResultsChromosome 3: A two-cluster (bimodal) solution was found in chromosome 3, characterized by centroids at unilaterally normal probe values and unilateral deletion. There was a large, significant difference in the survival characteristics of the two clusters (log-rank, p<0.001; 5-year survival: 80% versus 40%). Both clusters had a broad distribution in LTD, although larger tumors were characteristically in the poorer outcome group (Mann-Whitney, p<0.001). Threshold values of 0.85 for deletion and 1.15 for gain optimized the classification of the clusters. PCA showed that the first principal component (PC1) contained more than 80% of the data set variance and all of the bimodality, with uniform coefficients (0.28±0.03). Chromosome 8q: No clusters were found in chromosome 8q. Using a conventional threshold-based definition of 8q gain, and in conjunction with the chromosome 3 clusters, three prognostic groups were identified: chromosomes 3 and 8q both normal, either chromosome 3 or 8q abnormal, and both chromosomes 3 and 8q abnormal. KM analysis showed 5-year survival figures of approximately 97%, 80%, and 30% for these prognostic groups, respectively (log-rank, p<0.001). All MLPA probes within both chromosomes were significantly correlated with each other (Spearman, p<0.001).ConclusionsWithin chromosome 3, the strong correlation between the MLPA variables and the uniform coefficients from the PCA indicates a lack of evidence for a signature gene that might account for the bimodality we observed. We hypothesize that the two clusters we found correspond to binary underlying states of complete monosomy or disomy 3 and that these states are sampled by the complete ensemble of probes. Consequently, we would expect a similar pattern to emerge in higher-resolution MLPA data sets. LTD may be a significant confounding factor. Considering chromosome 8q, we found that chromosome 3 cluster membership and 8q gain as traditionally defined have an indistinguishable impact on patient outcome

4DCT and VMAT for lung patients with irregular breathing

PurposeIrregular breathing in lung cancer patients is a common contra-indication to 4D computerized tomography (4DCT), which may then limit radiotherapy treatment options. For irregular breathers, we investigated whether 3DCT or 4DCT (1) better represents tumor motion, (2) better represents average tumor densities, and (3) better allows for volumetric modulated arc threarpy (VMAT) plans delivered with acceptable dosimetric accuracy.MethodsTen clinical breathing traces were identified with irregularities in phase and amplitude, and fed to a programmable moving platform incorporating an anthropomorphic lung tumor phantom. 3DCT and 4DCT data resorted by phase (4DCT-P) and amplitude (4DCT-A) were acquired for each trace. Tumors were delineated by Hounsfield unit (HU) thresholding and apparent motion range assessed. HU profiles were extracted from each image and agreement with calculated expected profiles quantified using area-under-curve (AUC) scoring. Clinically representative VMAT plans were created for each image, delivered to the irregularly moving phantom, and measured with a small-volume ion chamber at the tumor center.ResultsMedian difference from expected tumor motion range for 3DCT, 4DCT-P, and 4DCT-A was 2.5 [1.6-3.6] cm, 1.1 [0.1-1.9] cm, and 1.3 [0.4-1.9] cm, respectively (p = 0.005, 4DCT-P vs. 3DCT). Median AUC scores (ideal = 0) for 3DCT, 4DCT-P, and 4DCT-A were 0.25 [0.14-0.49], 0.12 [0.05-0.42], and 0.13 [0.09-0.44], respectively (p = 0.005, 4DCT-P vs. 3DCT). Nine of ten 4DCT-P plans and all 4DCT-A plans measured within 2.5% of expected dose in the treatment planning system (TPS), compared with seven 3DCT plans.ConclusionFor the cases studied tumor motion range and average density was better represented with 4DCT compared with 3DCT, even in the presence of irregular breathing. 4DCT images allowed for delivery of VMAT plans with acceptable dosimetric accuracy. No significant differences were detected between phase and amplitude resorting. In combination with 4D cone beam imaging at treatment, our findings have given us confidence to introduce 4DCT and VMAT for lung radiotherapy patients with irregular breathing