Transcriptomic Hallmarks of Tumor Plasticity and Stromal Interactions in Brain Metastasis
Authors: Wingrove Emily , Liu Zongzhi Z , Patel Kiran D , Arnal‐Estape Anna , Melnick Mary‐Ann , Politi Katerina , Monteiro Cátia , Zhu Lucía , Valiente Manuel , Kluger Harriet M , Chiang Veronica L , Nguyen Don X 
Published: June 2021

Purpose

Brain metastasis remains a major site of relapse for several cancer types. However, the molecular mechanisms of brain metastasis are largely unknown given the complex relationship between tumor cells and the surrounding brain tumor microenvironment (TME). We have developed a Brain Metastatic RNA‐Sequencing (BMX‐Seq) approach, which leverages xenograft models and can distinguish between the transcriptomes of human tumor and mouse stroma gene expression in vivo. These data provide an extensive molecular portrait of the co‐adaptation of brain metastasis with the brain TME.

Methods

Following intra‐arterial injection of the human, lung adenocarcinoma (LUAD), H2030‐BrM3 cell model into athymic mice, metastatic brain tumors were macrodisasected, flash frozen and RNA extracted. Lung tumors, healthy brain regions, subcutaneous lesions and H2030‐BrM3 cells grown in monolayer were harvested in parallel. Our BMX‐Seq pipeline was engineered to analyze bulk xenograft tissue, which includes tumor lesions as well as surrounding stromal tissue. Species‐specific Taqman primers and immunofluorescent (IF) staining were used to validate our BMX‐Seq results including both human tumor and mouse stroma gene expression profiles. Models of melanoma and breast cancer brain metastasis were also included in this study.

In comparing the transcriptomic profiles of tumors grown in the brain as compared to other sites, we identify shifts in WNT signaling pathway components and epithelial and neuronal‐like gene expression programs in malignant cells (Figure 1). Further work revealed such trends are reversible once tumor cells are removed from the brain TME. In describing the neuroinflammatory response of stromal regions surrounding brain lesions, our BMX‐Seq analysis revealed increased expression of astrocyte and microglial enriched transcripts. We then utilized IF to confirm the elevated density of both of these cell types in areas directly surrounding and infiltrating tumor regions. We also detect significantly induced expression of stromal TIM3 (or HAVCR2) in areas surrounding tumor lesions and in a model system devoid of T‐Cells. Follow‐up work revealed that expression of TIM3 is induced on tumor‐associated macrophages (including microglia) within the tumor region, and we further validate such results in a syngeneic model system as well as in human, brain tissue (Figure 2).

Conclusions

We have developed a highly sensitive RNA‐sequencing based approach that can accurately map the transcriptomic adaptation of brain metastatic tumor cells and the surrounding brain TME. We believe these data may serve as a powerful resource to guide future discovery in CNS metastases.

Support or Funding Information

R01CA166376, R01CA191489, W81XWH‐15‐1‐0117 (D.X.N.). Yale Lung Cancer SPORE (P50CA196530) (K.P and D.X.N); Yale SPORE Skin Cancer (P50 CA121974) K24CA172123, R01 CA216846 (H.K.M); MINECO‐Europa Excelencia SAF2015‐62547‐ERC (M.V.).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.