Molecular grading of ductal carcinoma in situ of the breast

Abstract

Purpose: Increased incidence of ductal carcinoma in situ (DCIS) associated with mammographic screening for breast cancer has emphasized the challenges of managing this condition. The aim of this study was to identify informative clinical indicators of DCIS biology by molecular profiling.

Experimental design: Areas of in situ carcinoma, atypical ductal hyperplasia, and benign epithelium were microdissected from 46 invasive breast cancers. Oligonucleotide probes showing differential expression between DCIS associated with grade 1 and 3 invasive cancer were identified by microarray-based gene expression profiling. Expression at these probes was used to define a "molecular grade" subcategorization of all samples. The genomic basis of molecular grade was examined by array-based comparative genomic hybridization. Clinical course was examined in a cohort of 134 patients with DCIS treated by surgery alone.

Results: DCIS samples were designated as low or high molecular grade based on expression at 173 probes. The low molecular grade subgroup included low (n = 10) and intermediate (n = 11) nuclear grade DCIS as well as all samples of atypical ductal hyperplasia (n = 4) and benign epithelium (n = 7). The high molecular grade subgroup included DCIS of intermediate (n = 7) and high (n = 19) nuclear grade. The character and degree of genomic aberration were distinct between molecular grade subgroups. A classification tree model including nuclear grade and Ki67 score accurately predicted molecular grade for 95.7% of samples. In an independent cohort, this showed a pattern of rapid disease recurrence for high molecular grade DCIS.

Conclusions: Molecular profiling indicates a binary grading scheme for DCIS. This practical approach has potential to improve clinical evaluation of DCIS.

Fig. 1.

A, i, hierarchical clustering of 61 samples according to expression of the top 100 differentially expressed oligonucleotide probes (selected by supervised analysis of DCIS lesions associated with grade 1 and 3 invasive cancer). Columns represent samples; rows represent individual probes. Heatmap depicts high (red) and low (green) relative levels of gene expression. ii, grade of associated invasive breast cancer. iii, DCIS nuclear grade or sample type. B, GGI calculated for each sample in the corresponding heatmap column.

Fig. 2.

A, correlation between GGI and DCIS Ki67 score (P < 0.0001, r = 0.79, n = 46). B, Ki67 scores for DCIS samples in low MG and high MG groups (P < 0.0001, n = 46).

Fig. 3.

A, frequency of DNA copy number gains (red) and losses (green) across the genome (plotted from chromosome 1pter to 22qter, X and Y) in in situ carcinoma associated with grade 1 (i) and grade 3 (ii) invasive cancer. Average log 2 ratio copy number compared with normal male reference DNA is shown in blue. iii, representation of random forest algorithm applied to determine the importance measure for each probe in distinguishing lesions associated with grade 1 or grade 3 invasive cancer. Points colored in blue highlight regions that have a much higher copy number in grade 1 cases than in grade 3; points in orange have a higher copy number in grade 3 than grade 1. Higher copy number is defined as a difference of 0.25 mean log 2 ratio or larger. B, correlation between the number of regions of high-level DNA amplification and GGI (P < 0.0001, r = 0.62, n = 50). C, number of regions of high-level DNA amplification in low MG and high MG subgroups of DCIS (P = 0.003, n = 46).

Fig. 4.

A, two-step classification tree model illustrating prediction of DCIS molecular grade by histopathologic and biomarker features. B, local-recurrence -free survival for patients with DCIS of low MG and high MG predicted by nuclear grade and Ki67 score (n = 134).