Abstract Background: Invasive lobular carcinoma (ILC) is typically characterized by lobular morphology, CDH1 inactivation and reduced E-cadherin expression. However, discordance between morphologic and immunohistochemical (IHC) findings often complicates diagnosis. To address this issue, we performed whole transcriptome sequencing (WTS) on selected breast cancer cases exhibiting variable E-cadherin IHC patterns and morphology, including cases with and without CDH1 mutations identified by prior targeted sequencing. Methods: Whole transcriptome sequencing (WTS) was performed on 11 FFPE tissues (8 tumors and 3 normal breast tissue) using poly(A)-selected RNA libraries and 100 bp paired-end reads on the Illumina NovaSeq 6000 platform (Macrogen, Seoul, Korea). All 8 tumor cases had previously undergone targeted DNA sequencing using the TruSight Oncology 500 assay (TSO500; illumina). Based on morphologic, IHC, and molecular features, samples were stratified into three groups: Group 1 (n=4) consisted of tumors lacking lobular morphology, despite harboring CDH1 mutations identified by prior targeted sequencing. Group 2 (n=4) comprised tumor with classic lobular morphology and complete loss of E-cadherin expression, but no detectable CDH1 mutations on targeted sequencing. Group 3 (n=3) consisted of normal breast tissue samples serving as controls. Results: In Group 1, WTS confirmed the presence of CDH1 mutations previously identified by targeted sequencing (E880K in three cases, L630V in one case). Additionally, a novel transcript-derived variant, p.D756Y, was detected in one case. In Group 2, all four cases harbored CDH1 mutations that had not been detected by prior targeted sequencing. These included a stop-gained variant (Q610*), a frameshift mutation (D476fs), and several missense mutations (F69Y, S70P, I584V, I707T). Additionally, an intronic alteration near a canonical splice site (c.163+19330CG) was identified, suggesting potential splicing disruption. No pathogenic CDH1 mutations were detected in any of the three normal breast tissue samples in Group 3. Conclusion: WTS confirmed the presence of CDH1 mutations in breast tumors lacking lobular morphology, suggesting that molecular alterations can precede or occur independently of classic morphologic features. This finding suggests a diagnostic gray zone, where tumors with CDH1 mutations may exhibit non-lobular morphology and thus risk misclassification. Furthermore, WTS detected pathogenic CDH1 variants in tumors with classic lobular morphology, despite the absence of detectable mutations by targeted sequencing. This finding underscores the added sensitivity of transcriptome-based analysis. Together, these findings reveal the molecular and diagnostic complexity of lobular carcinoma and support an expanded role for RNA-level profiling in its classification. Citation Format: Y. Sung, J. Sim, J. Gim, T. Jeon, J. Lee, J. Oh, S. Jung, K. Park, A. Kim. Whole transcriptome sequencing reveals diagnostic and molecular complexity of lobular carcinoma associated with CDH1 alteration abstract. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(4 Suppl):Abstract nr PD9-05.
Sung et al. (Tue,) studied this question.