Human embryonic stem cell-derived organoid retinoblastoma reveals a cancerous origin

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Significance

As a genetic malignancy, retinoblastoma (Rb) is caused by RB1 mutations; however, its developmental origin and drug agents for human Rb remain largely unexplored. Here we describe an innovative Rb organoid model derived from human embryonic stem cells with a biallelic mutagenesis of the RB1 gene. We identify tumorigenic growth in the Rb organoids, as well as properties consistent with human primary Rb. We confirm that the Rb cell of origin stemmed from ARR3 ⁺ maturing cone precursor cells and SYK inhibitors displaying a significant therapeutic response. Our elegant in-dish Rb organoid model can be used to efficiently and effectively dissect the origin of Rb and mechanisms of Rb tumorigenesis, as well as screen novel therapies.

Abstract

Retinoblastoma (Rb) is the most prevalent intraocular malignancy in children, with a worldwide survival rate <30%. We have developed a cancerous model of Rb in retinal organoids derived from genetically engineered human embryonic stem cells (hESCs) with a biallelic mutagenesis of the RB1 gene. These organoid Rbs exhibit properties highly consistent with Rb tumorigenesis, transcriptome, and genome-wide methylation. Single-cell sequencing analysis suggests that Rb originated from ARR3-positive maturing cone precursors during development, which was further validated by immunostaining. Notably, we found that the PI3K-Akt pathway was aberrantly deregulated and its activator spleen tyrosine kinase (SYK) was significantly up-regulated. In addition, SYK inhibitors led to remarkable cell apoptosis in cancerous organoids. In conclusion, we have established an organoid Rb model derived from genetically engineered hESCs in a dish that has enabled us to trace the cell of origin and to test novel candidate therapeutic agents for human Rb, shedding light on the development and therapeutics of other malignancies.

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RNA velocity of single cells

Gioele La Manno, Ruslan Soldatov, Amit Zeisel … (2018)

RNA abundance is a powerful indicator of the state of individual cells. Single-cell RNA sequencing can reveal RNA abundance with high quantitative accuracy, sensitivity and throughput1. However, this approach captures only a static snapshot at a point in time, posing a challenge for the analysis of time-resolved phenomena, such as embryogenesis or tissue regeneration. Here we show that RNA velocity—the time derivative of the gene expression state—can be directly estimated by distinguishing unspliced and spliced mRNAs in common single-cell RNA sequencing protocols. RNA velocity is a high-dimensional vector that predicts the future state of individual cells on a timescale of hours. We validate its accuracy in the neural crest lineage, demonstrate its use on multiple published datasets and technical platforms, reveal the branching lineage tree of the developing mouse hippocampus, and examine the kinetics of transcription in human embryonic brain. We expect RNA velocity to greatly aid the analysis of developmental lineages and cellular dynamics, particularly in humans.

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PI3K/Akt/mTOR inhibitors in cancer: At the bench and bedside

Ali Alzahrani (2019)

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Single-cell transcriptomes from human kidneys reveal the cellular identity of renal tumors

Matthew D. Young, Thomas J. Mitchell, Felipe A. Vieira Braga … (2018)

Understanding tumor origins and the similarities and differences between organ-specific cancers is important for determining treatment options. Young et al. generated more than 72,000 single-cell transcriptomes from healthy and cancerous human kidneys. From these data, they determined that Wilms tumor, a pediatric kidney cancer, originates from aberrant fetal cells, whereas adult kidney cancers are likely derived from a specific subtype of proximal convoluted tubular cell. Science , this issue p. [Related article:] 594 Single-cell mRNAs of normal and cancerous kidney cells reveal the cellular identity of childhood and adult tumors. Messenger RNA encodes cellular function and phenotype. In the context of human cancer, it defines the identities of malignant cells and the diversity of tumor tissue. We studied 72,501 single-cell transcriptomes of human renal tumors and normal tissue from fetal, pediatric, and adult kidneys. We matched childhood Wilms tumor with specific fetal cell types, thus providing evidence for the hypothesis that Wilms tumor cells are aberrant fetal cells. In adult renal cell carcinoma, we identified a canonical cancer transcriptome that matched a little-known subtype of proximal convoluted tubular cell. Analyses of the tumor composition defined cancer-associated normal cells and delineated a complex vascular endothelial growth factor (VEGF) signaling circuit. Our findings reveal the precise cellular identities and compositions of human kidney tumors.

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Author and article information

Journal

Journal ID (nlm-ta): Proc Natl Acad Sci U S A

Journal ID (iso-abbrev): Proc Natl Acad Sci U S A

Journal ID (hwp): pnas

Journal ID (pmc): pnas

Journal ID (publisher-id): PNAS

Title: Proceedings of the National Academy of Sciences of the United States of America

Publisher: National Academy of Sciences

ISSN (Print): 0027-8424

ISSN (Electronic): 1091-6490

Publication date (Print): 29 December 2020

Publication date (Electronic): 14 December 2020

Publication date PMC-release: 14 December 2020

Volume: 117

Issue: 52

Pages: 33628-33638

Affiliations

[1] ^aLaboratory of Stem Cell and Retinal Regeneration, The Eye Hospital, Wenzhou Medical University , 325027 Wenzhou, China;

[2] ^bInstitute of Biomedical Big Data, School of Biomedical Engineering, School of Ophthalmology and Optometry, The Eye Hospital, Wenzhou Medical University , 325027 Wenzhou, China;

[3] ^cBeijing Institute of Ophthalmology, Beijing Tongren Eye Center, Beijing Tongren Hospital, Capital Medical University, Beijing Ophthalmology and Visual Sciences Key Laboratory , 100730 Beijing, China;

[4] ^dInstitute of Stem Cell Research, The Eye Hospital, Wenzhou Medical University , 325027 Wenzhou, China

Author notes

²To whom correspondence may be addressed. Email: sujz@ 123456wmu.edu.cn or jinzibing@ 123456foxmail.com .

Edited by Robert Johnston, Johns Hopkins, Baltimore, MD, and accepted by Editorial Board Member Jeremy Nathans November 12, 2020 (received for review June 8, 2020)

Author contributions: Z.-B.J. designed and supervised whole study; H.L., Y.-Y.Z., Y.-P.L., Z.-Q.H., C.-J.Z., and K.-C.W. performed experiments and interpreted the results; J.S. and Z.-B.J. contributed new reagents/analytic tools; Yan Zhang, F.Y., Yaru Zhang, and J.S. analyzed the omics data; H.L. and Z.-B.J. wrote the manuscript; and J.S. and Z.-B.J. revised versions of the manuscript.

¹H.L. and Y.Z. contributed equally to this work.