Dependencies in heterogeneous, lineage plastic patient-derived prostate cancer organoids revealed through integrated single-cell multiomics and CRISPR screening

  • bioRxiv. 2026 May 8:2026.05.07.723570. doi: 10.64898/2026.05.07.723570.
Samir Zaidi  1  2 ,  Maren Büttner  3 ,  Weiran Feng  4 ,  Caitlin Baxter  1  2 ,  D Henry Kates  5 ,  Tiago Paiva Prudente  1  2 ,  Diana Zakharova  4 ,  Sanjoy Mehta  6 ,  Subhiksha Nandakumar  7 ,  Chen Khuan Wong  5 ,  Hojin Lee  8 ,  Pierre-Jacques Hamard  9 ,  Wenfei Kang  10 ,  Jungmin Choi  8 ,  Ronan Chaligné  11 ,  Robert Cohen  3 ,  Yu Chen  5  12 ,  Ari Firestone  3 ,  Zhenghao Chen  3 ,  Charles L Sawyers  4  13
Affiliations
  • 1. Department of Medicine, Yale School of Medicine, New Haven, CT 06510, USA.
  • 2. Center of Molecular and Cellular Oncology, Yale Cancer Center, New Haven, CT 06510, USA.
  • 3. Calico Life Sciences LLC, South San Francisco, CA USA.
  • 4. Cancer Epigenetics Institute, Nuclear Dynamics & Cancer Program Fox Chase Cancer Center, Philadelphia, PA 19111, USA.
  • 5. Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA.
  • 6. Genome Editing and Screening Core, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA.
  • 7. Department of Epidemiology and Biostatistics, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA.
  • 8. Department of Biomedical Sciences, Korea University College of Medicine, Seoul, Korea.
  • 9. Epigenetic Research Innovation Lab, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA.
  • 10. Molecular Cytology Core Facility, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA.
  • 11. Single Cell Analytics Innovation Lab, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA.
  • 12. Department of Genitourinary Oncology, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA.
  • 13. Howard Hughes Medical Institute, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA.
PMID: 42146571 DOI: 10.64898/2026.05.07.723570
Abstract

Lineage plasticity and tumor heterogeneity limit the effectiveness of targeted therapies, yet the functional dependencies used to nominate therapeutic targets are often derived from homogeneous systems that fail to capture this complexity. Here, we establish a framework to resolve state-specific genetic vulnerabilities by integrating single-cell multiomics (RNA and ATAC) with pooled CRISPR-Cas9 screening across a large panel of patient-derived organoids (PDOs) from castrate-resistant prostate Cancer (CRPC) and neuroendocrine prostate Cancer (NEPC). We generate a single-cell multiome atlas spanning >190,000 cells across 22 PDOs, defining seven lineage states-including intermediate and plastic populations not resolved by bulk profiling-and demonstrate that these lineage programs robustly classify independent transcriptomic datasets from prostate Cancer patient tumors. By systematically coupling this atlas to subtype-resolved CRISPR screens, we construct a functional dependency map linking cell state in heterogeneous 3D human tumor models. We show that intratumoral heterogeneity fundamentally reshapes the interpretation of gene essentiality, whereby gene-level depletion reflects the composite behavior of co-existing subpopulations, and identify a general principle in which resistant "limiting" populations disproportionately determine aggregate fitness effects. This framework reveals both canonical and previously unrecognized lineage-restricted dependencies within highly plastic tumor and NEPC states, including a therapeutically targetable dependency on the Aryl Hydrocarbon Receptor (AHR) in a novel hybrid stem-like/ASCL1 population. Together, these data establish an extensive multi-dimensional prostate Cancer resource, identify novel lineage-resolved biology, and provide a generalizable strategy for interpreting functional genomics in heterogeneous human tumors.

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