OT-knn: A neighborhood-aware optimal transport framework for aligning spatial transcriptomics data.

Spatial transcriptomics (ST) measures gene expression while preserving spatial context within tissues, enabling detailed characterization of tissue organization. As ST technologies advance, aligning datasets across tissue sections, individuals, platforms, and developmental stages has become increasingly important but remains challenging due to sparse expression, biological heterogeneity, and geometric distortions between slices. We introduce OT-knn, a method for ST alignment that integrates local neighborhood information within an optimal transport framework. Rather than relying solely on single-spot expression, OT-knn reconstructs each spot using its spatial k-nearest neighbors, capturing microenvironment context that is more robust to noise and variability. These representations are then used to derive probabilistic correspondences between slices. We evaluate OT-knn using simulated data with known ground-truth alignment and real datasets from multiple ST platforms, including human dorsolateral prefrontal cortex data (10x Genomics Visium), mouse brain aging data with both within-donor and cross-donor comparisons (MERFISH), a multi-stage axolotl brain dataset (Stereo-seq), and a cross-platform analysis using mouse embryo datasets profiled by Stereo-seq and seqFISH. Across these settings, OT-knn achieves accurate and robust alignment, particularly in the presence of spatial deformation, donor heterogeneity, developmental variation and technological differences.