spOT-NMF

Optimal Transport-Based Matrix Factorization for Accurate Deconvolution of Spatial Transcriptomics Abdelkareem, A.O. et al.(2025)

spOT-NMF is a Python package for unsupervised deconvolution and discovery of gene programs in spatial transcriptomics. It integrates Optimal Transport (OT) into a non-negative matrix factorization (NMF) framework, enabling robust topic modeling, high-resolution spatial deconvolution, and rich biological annotation.

This package supports the analyses in: spOT-NMF: Optimal Transport-Based Matrix Factorization for Accurate Deconvolution of Spatial Transcriptomics — bioRxiv (2025). DOI: 10.1101/2025.08.02.668292

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✨ Key Features

• OT-NMF Deconvolution: Reference-free topic modeling with OT-regularized NMF.
• HVG Selection: Flexible, batch-aware highly variable gene selection.
• Biological Annotation: Automated enrichment and gene-set overlap of inferred programs.
• Spatial Visualization: Publication-quality spatial plots for topic/program usage.
• Scalable & Modular: Built for large datasets and multi-sample workflows.
• CLI & Python API: Run from the command line or import in notebooks.

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📦 Installation

spOT-NMF requires Python ≥ 3.12. We recommend uv for a fast, reproducible setup. PyTorch is installed separately so you can pick the build (CPU or CUDA) for your platform.

Recommended: uv

# 1. Create and activate an isolated environment (uv fetches Python 3.12 if needed)
uv venv --python 3.12
# Linux/macOS:  source .venv/bin/activate
# Windows:      .venv\Scripts\activate

# 2. Install PyTorch for your platform (see pytorch.org)
#    CPU-only:
uv pip install torch --index-url https://download.pytorch.org/whl/cpu
#    CUDA 12.x (Linux/Windows with NVIDIA GPUs):
#    uv pip install torch torchvision torchaudio --index-url https://download.pytorch.org/whl/cu121

# 3. Install spOT-NMF
uv pip install spot-nmf

Alternative: pip

python -m venv .venv && source .venv/bin/activate   # Windows: .venv\Scripts\activate
pip install torch --index-url https://download.pytorch.org/whl/cpu
pip install spot-nmf

From source (development)

git clone https://github.com/MorrissyLab/spOT-NMF.git
cd spOT-NMF
uv venv --python 3.12
uv pip install torch --index-url https://download.pytorch.org/whl/cpu
uv pip install -e ".[dev]"     # editable install with test dependencies
uv run pytest -q               # run the test suite

Verify the install

spotnmf --help

If no GPU is available, spOT-NMF automatically runs on CPU.

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🚀 Quick Start

Command Line

Full pipeline (deconvolution → annotation → spatial plots → networks):

spotnmf spotnmf \
  --sample_name SAMPLE1 \
  --adata_path ./data/sample1.h5ad \
  --data_mode h5ad \
  --results_dir ./results \
  --k 5 \
  --genome GRCh38

--data_mode selects how the input is read: h5ad for a single AnnData .h5ad file, visium (the default) for a Space Ranger output directory, or visium_hd for Visium HD. Pass --data_mode h5ad whenever --adata_path points to a .h5ad file.

Other commands:

spotnmf deconvolve --sample_name SAMPLE1 --adata_path ./data/sample1.h5ad --data_mode h5ad --results_dir ./results --k 5
spotnmf plot       --sample_name SAMPLE1 --adata_path ./data/sample1.h5ad --data_mode h5ad --results_dir ./results
spotnmf annotate   --sample_name SAMPLE1 --results_dir ./results --genome GRCh38
spotnmf network    --sample_name SAMPLE1 --results_dir ./results --usage_threshold 0 --n_bins 1000 --edge_threshold 0.199

The network command reuses the per-spot usages written by deconvolve. On small datasets no topic pairs may pass --n_bins / --edge_threshold; in that case it prints a notice and skips plotting — lower the thresholds to force a graph.

Python / Notebooks

from pathlib import Path
import spotnmf as spot

# === Configuration === #
DATA_PATH = Path("data/test_data/dataset10_adata_spatial.h5ad")
RESULTS_DIR = Path(r"/data/test_results/")
SAMPLE_NAME = "TestSample"
GENOME = "mm10"

# === Read Data === #
adata = spot.io.read_adata(
    data_path=DATA_PATH,
    data_mode="h5ad"
)

# === Model Parameters === #
model_params = {
    "lr": 0.001,         # Learning rate
    "h": 0.01,           # H regularization
    "w": 0.01,           # W regularization
    "eps": 0.05,         # Epsilon
    "normalize_rows": True,
}

# === Run Factorization === #
results = spot.cli.run_experiment(
    adata_spatial=adata,
    k=5,                        # Number of ranks
    sample_name=SAMPLE_NAME,
    results_dir=str(RESULTS_DIR),
    genome=GENOME,
    annotate=False,
    plot=False,
    network=False,
    is_visium=True,
    model_params=model_params,
)

# === Annotate Programs === #
spot.cli.annotate_programs(
    results_dir=str(RESULTS_DIR),
    sample_name=SAMPLE_NAME,
    genome=GENOME,
)

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📓 Tutorials

A fully worked, well-commented notebook runs the entire pipeline end-to-end on the small example dataset that ships with the repo (CPU-only, ~1 minute) — loading data, selecting HVGs, running the OT-NMF deconvolution, mapping programs spatially, extracting marker genes, and validating the recovered programs against ground-truth cell types. All figures are pre-rendered in the notebook.

• Full pipeline tutorial → (docs/source/tutorials/full_pipeline.ipynb)

GitHub renders the notebook (with figures) directly in the browser — just click the link.

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⚙️ CLI Overview

Command	Description
spotnmf	Full pipeline: deconvolution → annotation → spatial plotting
deconvolve	Run OT-NMF and save results
plot	Visualize spatial topic/program usage
annotate	Enrich and annotate gene programs
network	Visualize niche networks based on topic interactions

Run spotnmf <command> --help for per-command options.

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📁 Outputs

• topics_per_spot_{sample}.csv — topic/program usage per spot
• genescores_per_topic_{sample}.csv — gene scores per topic
• ranked_genescores_{sample}.csv — ranked marker genes per topic
• Pathway enrichment and gene-set overlap tables
• Spatial plots & QC visualizations
• Network plots of topic–topic interactions

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🔬 Reproducibility (Manuscript Notebooks)

The main branch provides the reusable software package. The original Jupyter notebooks used to reproduce manuscript figures are maintained in the manuscript branch:

git fetch origin
git checkout manuscript

Notebooks are in:

scripts/manuscript_notebooks/

Use manuscript to regenerate paper figures; use main for running the package on your data.

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🧾 Citation

Please cite:

Abdelkareem, A.O., Gill, G.S., Manoharan, V.T., Verhey, T.B., & Morrissy, A.S. spOT-NMF: Optimal Transport-Based Matrix Factorization for Accurate Deconvolution of Spatial Transcriptomics. bioRxiv (2025). https://doi.org/10.1101/2025.08.02.668292

@article{abdelkareem2025spotnmf,
  title   = {spOT-NMF: Optimal Transport-Based Matrix Factorization for Accurate Deconvolution of Spatial Transcriptomics},
  author  = {Abdelkareem, Aly O. and Gill, Gurveer S. and Manoharan, Varsha Thoppey and Verhey, Theodore B. and Morrissy, A. Sorana},
  journal = {bioRxiv},
  year    = {2025},
  doi     = {10.1101/2025.08.02.668292},
  url     = {https://www.biorxiv.org/content/10.1101/2025.08.02.668292v1},
  note    = {Preprint}
}

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🤝 Contributing

We welcome ideas, bug reports, and feature requests—please open a GitHub Issue: https://github.com/MorrissyLab/spOT-NMF/issues

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📜 License

GPL-3.0. See LICENSE for details.

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💬 Support

Questions or need help? Open an Issue: https://github.com/MorrissyLab/spOT-NMF/issues