Comparison of imaging based single-cell resolution spatial transcriptomics profiling platforms using formalin-fixed paraffin-embedded tumor samples
Nejla Ozirmak Lermi, Max Molina Ayala, Sharia Hernandez, Wei Lu, Khaja Khan, Alejandra G. Serrano, Idania Carolina Lubo Julio, Leticia Hamana, Katarzyna Tomczak, Sean Barnes, Jinzhuang Dou, Qingnan Liang, RTI Team, Ahmed Al-Rawi, Claudio A. Arrechedera, Kimberly S. Ayers, Claudia Bedoya, Elizabeth M. Burton, Connie A. Chon, Randy Chu, Shadarra Crosby, Jonathan Do, Cibelle Lima, Fu Szu-Chin, Andy Futreal, Ana Lineth García, Celia Garica-Prieto, Swati Gite, Curtis Gumbs, Kristin J. Hargraves, Meng He, Chacha Horombe, Heladio P. Ibarguen, Stacy Jackson, Jeena J. Jacob, Isha Khanduri, Walter Kinyua, Mark Knafl, Wenhua Lang, Latasha Little, Saradhi Mallampati, Mary GT Mendoza, Funda Meric‐Bernstam, Mohammad M. Mohammad, Mario Luiz Marques Piubelli, Sabitha Prabhakaran, Kenna Shaw, Xiaofei Song, Sandesh Subramanya, Baohua Sun, Shumaila Virani, Wanlin Wang, Scott E. Woodman, Mingchu Xu, Jianhua Zhang, Qingxiu Zhang, Shanyu Zhang, Maria Gabriela Raso, Ximing Tang, Mei Jiang, Beatriz Sánchez‐Espiridión, Annikka Weissferdt, John V. Heymach, Jianjun Zhang, Boris Sepesi, Tina Cascone, Anne S. Tsao, Mehmet Altan, Reza J. Mehran, Don L. Gibbons, Ignacio Wistuba, Cara Haymaker, Ken Chen, Luisa M. Solis Soto
Abstract
Imaging-based spatial transcriptomics (ST) is evolving as a pivotal technology in studying tumor biology and associated microenvironments. However, the strengths of the commercially available ST platforms in studying spatial biology have not been systematically evaluated using rigorously controlled experiments. We use serial 5 μm sections of formalin-fixed, paraffin-embedded surgically resected lung adenocarcinoma and pleural mesothelioma samples in tissue microarrays to compare the performance of the ST platforms (CosMx, MERFISH, and Xenium (uni/multi-modal)) in reference to bulk RNA sequencing, multiplex immunofluorescence, GeoMx, and hematoxylin and eosin staining data. In addition to an objective assessment of automatic cell segmentation and phenotyping, we perform a manual phenotyping evaluation to assess pathologically meaningful comparisons between ST platforms. Here, we show the intricate differences between the ST platforms, reveal the importance of parameters such as probe design in determining the data quality, and suggest reliable workflows for accurate spatial profiling and molecular discovery. Spatial cell distribution within a tissue microenvironment is a rapidly advancing field. Here, authors assess three commercially available single-cell resolution spatial transcriptomics approaches (CosMx, MERFISH, and Xenium) to inform which technology outperforms for immune profiling of solid tumors using patient samples.