Abstract
BackgroundThe acquisition of oncogenic drivers is a critical feature of cancer progression. For some carcinomas, it is clear that certain genetic drivers occur early in neoplasia and others late. Why these drivers are selected and how these changes alter the neoplasiaās fitness is less understood.MethodsHere we use spatially oriented genomic approaches to identify transcriptomic and genetic changes at the single-duct level within precursor neoplasia associated with invasive breast cancer. We study HER2 amplification in ductal carcinoma in situ (DCIS) as an event that can be both quantified and spatially located via fluorescence in situ hybridization (FISH) and immunohistochemistry on fixed paraffin-embedded tissue.ResultsBy combining the HER2-FISH with the laser capture microdissection (LCM) Smart-3SEQ method, we found that HER2 amplification in DCIS alters the transcriptomic profiles and increases diversity of copy number variations (CNVs). Particularly, interferon signaling pathway is activated by HER2 amplification in DCIS, which may provide a prolonged interferon signaling activation in HER2-positive breast cancer. Multiple subclones of HER2-amplified DCIS with distinct CNV profiles are observed, suggesting that multiple events occurred for the acquisition of HER2 amplification. Notably, DCIS acquires key transcriptomic changes and CNV events prior to HER2 amplification, suggesting that pre-amplified DCIS may create a cellular state primed to gain HER2 amplification for growth advantage.ConclusionBy using genomic methods that are spatially oriented, this study identifies several features that appear to generate insights into neoplastic progression in precancer lesions at a single-duct level.
Highlights
The acquisition of oncogenic drivers is a critical feature of cancer progression
Our study found that HER2 amplification altered the transcriptomic profiles in ductal carcinoma in situ (DCIS), and interferon signaling pathway was activated by HER2 amplification in DCIS, which may provide a prolonged interferon signaling activation in HER2-positive breast cancer
HER2-fluorescence in situ hybridization (FISH) analysis in DCIS Here we employed spatial genomic methods that can quantify small amounts of RNA and DNA from single breast ducts isolated by laser capture microdissection (LCM)-collected formalin-fixed paraffin-embedded (FFPE) tissue [11] (Fig. 1a)
Summary
The acquisition of oncogenic drivers is a critical feature of cancer progression. For some carcinomas, it is clear that certain genetic drivers occur early in neoplasia and others late. We sub-sampled DCIS ducts at different regions within a large lesion, aiming to identify the changes by HER2 amplification and uncover the evolutionary path during HER2 amplification in neoplasia progression These in situ ducts, which can only be identified in clinical archival tissues and are difficult to categorize by morphology, are challenging to capture by the conventional single-cell RNA sequencing techniques. The Smart-3SEQ data can be combined with other in situ profiles such as FISH and immunohistochemistry on the same ducts using adjacent tissue sections This new approach enables us to profile multiple microscopic ducts that have different HER2 status within a single lesion, allowing us to identify the changes by HER2 amplification and uncover the evolutionary path during HER2 amplification in neoplasia progression
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