Single-nucleus and Spatial Transcriptome Profiling of Pancreatic Cancer Identifies Multicellular Dynamics Associated with Neoadjuvant Treatment

Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal and treatment-refractory cancer. Molecular stratification in pancreatic cancer remains rudimentary and does not yet inform clinical management or therapeutic development. Here, we construct a high-resolution molecular landscape of the cellular subtypes and spatial communities that compose PDAC using single-nucleus RNA sequencing and whole-transcriptome digital spatial profiling (DSP) of 43 primary PDAC tumor specimens that either received neoadjuvant therapy or were treatment naive. We uncovered recurrent expression programs across malignant cells and fibroblasts, including a newly identified neural-like progenitor malignant cell program that was enriched after chemotherapy and radiotherapy and associated with poor prognosis in independent cohorts. Integrating spatial and cellular profiles revealed three multicellular communities with distinct contributions from malignant, fibroblast and immune subtypes: classical, squamoid-basaloid and treatment enriched. Our refined molecular and cellular taxonomy can provide a framework for stratification in clinical trials and serve as a roadmap for therapeutic targeting of specific cellular phenotypes and multicellular interactions.

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References

Versteijne E, van Dam J, Suker M, Janssen Q, Groothuis K, Akkermans-Vogelaar J . Neoadjuvant Chemoradiotherapy Versus Upfront Surgery for Resectable and Borderline Resectable Pancreatic Cancer: Long-Term Results of the Dutch Randomized PREOPANC Trial. J Clin Oncol. 2022; 40(11):1220-1230. DOI: 10.1200/JCO.21.02233. View

Buttner M, Ostner J, Muller C, Theis F, Schubert B . scCODA is a Bayesian model for compositional single-cell data analysis. Nat Commun. 2021; 12(1):6876. PMC: 8616929. DOI: 10.1038/s41467-021-27150-6. View

Dardenne E, Beltran H, Benelli M, Gayvert K, Berger A, Puca L . N-Myc Induces an EZH2-Mediated Transcriptional Program Driving Neuroendocrine Prostate Cancer. Cancer Cell. 2016; 30(4):563-577. PMC: 5540451. DOI: 10.1016/j.ccell.2016.09.005. View

Harder J, Ihorst G, Heinemann V, Hofheinz R, Moehler M, Buechler P . Multicentre phase II trial of trastuzumab and capecitabine in patients with HER2 overexpressing metastatic pancreatic cancer. Br J Cancer. 2012; 106(6):1033-8. PMC: 3304403. DOI: 10.1038/bjc.2012.18. View

VanDussen K, Sonnek N, Stappenbeck T . L-WRN conditioned medium for gastrointestinal epithelial stem cell culture shows replicable batch-to-batch activity levels across multiple research teams. Stem Cell Res. 2019; 37:101430. PMC: 6579736. DOI: 10.1016/j.scr.2019.101430. View

Muus C, Luecken M, Eraslan G, Sikkema L, Waghray A, Heimberg G . Single-cell meta-analysis of SARS-CoV-2 entry genes across tissues and demographics. Nat Med. 2021; 27(3):546-559. PMC: 9469728. DOI: 10.1038/s41591-020-01227-z. View

Grunwald B, Devisme A, Andrieux G, Vyas F, Aliar K, McCloskey C . Spatially confined sub-tumor microenvironments in pancreatic cancer. Cell. 2021; 184(22):5577-5592.e18. DOI: 10.1016/j.cell.2021.09.022. View

Kotliar D, Veres A, Nagy M, Tabrizi S, Hodis E, Melton D . Identifying gene expression programs of cell-type identity and cellular activity with single-cell RNA-Seq. Elife. 2019; 8. PMC: 6639075. DOI: 10.7554/eLife.43803. View

Rorke E, Adhikary G, Young C, Rice R, Elias P, Crumrine D . Structural and biochemical changes underlying a keratoderma-like phenotype in mice lacking suprabasal AP1 transcription factor function. Cell Death Dis. 2015; 6:e1647. PMC: 4669787. DOI: 10.1038/cddis.2015.21. View

10.

de Man F, Goey A, van Schaik R, Mathijssen R, Bins S . Individualization of Irinotecan Treatment: A Review of Pharmacokinetics, Pharmacodynamics, and Pharmacogenetics. Clin Pharmacokinet. 2018; 57(10):1229-1254. PMC: 6132501. DOI: 10.1007/s40262-018-0644-7. View

11.

Collisson E, Bailey P, Chang D, Biankin A . Molecular subtypes of pancreatic cancer. Nat Rev Gastroenterol Hepatol. 2019; 16(4):207-220. DOI: 10.1038/s41575-019-0109-y. View

12.

Denny S, Yang D, Chuang C, Brady J, Lim J, Gruner B . Nfib Promotes Metastasis through a Widespread Increase in Chromatin Accessibility. Cell. 2016; 166(2):328-342. PMC: 5004630. DOI: 10.1016/j.cell.2016.05.052. View

13.

Zhang M, Liu T, Xia B, Yang C, Hou S, Xie W . Platelet-Derived Growth Factor D Is a Prognostic Biomarker and Is Associated With Platinum Resistance in Epithelial Ovarian Cancer. Int J Gynecol Cancer. 2017; 28(2):323-331. DOI: 10.1097/IGC.0000000000001171. View

14.

Guo J, Hoffman H, Weekes C, Zheng L, Ting D, Hwang W . Refining the Molecular Framework for Pancreatic Cancer with Single-cell and Spatial Technologies. Clin Cancer Res. 2021; 27(14):3825-3833. PMC: 8282742. DOI: 10.1158/1078-0432.CCR-20-4712. View

15.

Andre P, Song H, Kim W, Kispert A, Yang Y . Wnt5a and Wnt11 regulate mammalian anterior-posterior axis elongation. Development. 2015; 142(8):1516-27. PMC: 4392599. DOI: 10.1242/dev.119065. View

16.

Lau H, Ng N, Loo L, Jasmen J, Teo A . The molecular functions of hepatocyte nuclear factors - In and beyond the liver. J Hepatol. 2017; 68(5):1033-1048. DOI: 10.1016/j.jhep.2017.11.026. View

17.

Roa-Pena L, Leiton C, Babu S, Pan C, Vanner E, Akalin A . Keratin 17 identifies the most lethal molecular subtype of pancreatic cancer. Sci Rep. 2019; 9(1):11239. PMC: 6677817. DOI: 10.1038/s41598-019-47519-4. View

18.

Biasci D, Smoragiewicz M, Connell C, Wang Z, Gao Y, Thaventhiran J . CXCR4 inhibition in human pancreatic and colorectal cancers induces an integrated immune response. Proc Natl Acad Sci U S A. 2020; 117(46):28960-28970. PMC: 7682333. DOI: 10.1073/pnas.2013644117. View

19.

Shi D, Guo J, Hoffman H, Su J, Mino-Kenudson M, Barth J . Therapeutic avenues for cancer neuroscience: translational frontiers and clinical opportunities. Lancet Oncol. 2022; 23(2):e62-e74. PMC: 9516432. DOI: 10.1016/S1470-2045(21)00596-9. View

20.

Di Donato V, De Santis F, Albadri S, Auer T, Duroure K, Charpentier M . An Attractive Reelin Gradient Establishes Synaptic Lamination in the Vertebrate Visual System. Neuron. 2018; 97(5):1049-1062.e6. DOI: 10.1016/j.neuron.2018.01.030. View