Abstract
The tumor microenvironment in colorectal cancer (CRC) is a heterogeneous ecosystem in which host cells and microbial communities interact dynamically, influencing disease progression. However, the clinical utility is limited by the lack of a scalable spatial host-microbiome technique and by insufficient integration of artificial intelligence for the interpretation of high-dimensional multi-omics data. To overcome the barriers, we present AlphaFISH, a platform technology integrating both technical and computational innovations for multi-omics spatial analysis of clinical biopsies at subcellular resolution. The system uses a sequencing-free, high-throughput, spatial profiling technique to construct, to date, the largest clinical spatial transcriptomics and spatial microbiome datasets acquired from 149 colorectal biopsies from 68 human subjects, supported by a comprehensive scRNA-seq atlas covering 4.27 million cells across 650 patients for robust cell annotation. Deep learning of the cohort-scale dual-omics data, consisting of more than 10 million subcellular sampling vectors, enables the development of a transformer model with joint embeddings of gene expression, spatial architecture, and the microbial microenvironment in colon tissues, achieving nearly 90% accuracy in predicting CRC-associated pathological features using unseen spatial omics inputs. The AI interrogation further predicts tumour recurrence at 81% accuracy in 28 patients followed within 1 year post tumor resection period. AlphaFISH reveals that spatial interactions between Fusobacterium and cellular niche consisting of tumor and T cells serve as key markers of CRC malignancy, progression, and recurrence, indicating the critical role of spatial bacterial-immune crosstalks.