weri rb1 cell line  (ATCC)

Journal: International Journal of Molecular Sciences

Article Title: Human Retinal Organoid Modeling Defines Developmental Window and Therapeutic Vulnerabilities in MYCN-Amplified Retinoblastoma

doi: 10.3390/ijms26178675

Figure Lengend Snippet: Establishment and in vivo tumorigenicity of MYCN-overexpressing cell lines derived from retinal organoids (MYCN O/E -cells). ( A ) MYCN O/E -cell lines derived from retinal organoid tumors were cultured under suspension conditions and underwent single-cell dissociation at each passage. Dissociated cells consistently formed tumor spheres over multiple passages, exhibiting morphology similar to established human retinoblastoma cell lines (e.g., Y79, WERI-Rb1). Scale bar: 100 μm. ( B ) Schematic illustration of the subretinal xenograft procedure performed using MYCN O/E -cells. ( C ) Representative images of immunodeficient mice two months post-subretinal injection of MYCN O/E -cells. Injected eyes exhibited clinical leukocoria and significant intraocular tumor formation, while control eyes injected with normal retinal organoid (nRO) remained morphologically normal. ( D , E ) Histopathological characterization of xenograft tumors at 2 months ( D ) and 9 months ( E ) post-injection revealed highly aggressive and poorly differentiated tumors, composed of densely packed, pleomorphic, hyperchromatic cells with enlarged nuclei, prominent nucleoli, and high nuclear-to-cytoplasmic ratios, closely recapitulating clinical MYCN-amplified retinoblastoma. Rosette structures were notably absent. Scale bar: 100 μm ( D ); 50 μm ( E ). ( F ) Immunohistochemical analyses of xenograft tumors at 9 months post-injection demonstrated robust proliferation (Ki-67 positivity), strong retinal progenitor identity (SOX2 positivity), and lack of photoreceptor differentiation (absence of CRX expression), further confirming the undifferentiated state of tumors. Scale bars: 100 μm.

Article Snippet: Retinoblastoma cell lines (Y79 and WERI-Rb1; ATCC, Manassas, VA, USA) were grown in suspension cultures in RPMI-1640 medium (GibcoTM, cat. no. A1049101, Carlsbad, CA, USA) supplemented with 20% fetal bovine serum (FBS; Rd Tech, cat. no. A1500m, Namyangju, Republic of Korea) and 1% penicillin/streptomycin (Thermo Fisher Scientific, cat. no. 15140-122, Waltham, MA, USA).

Techniques: In Vivo, Derivative Assay, Cell Culture, Suspension, Injection, Control, Amplification, Immunohistochemical staining, Expressing

Journal: International Journal of Molecular Sciences

Article Title: Human Retinal Organoid Modeling Defines Developmental Window and Therapeutic Vulnerabilities in MYCN-Amplified Retinoblastoma

doi: 10.3390/ijms26178675

Figure Lengend Snippet: Transcriptomic profiling reveals distinct molecular signatures and oncogenic pathways in MYCN-overexpressing retinoblastoma organoids (MYCN O/E -RBOs) and similarity to patient-derived tumors. ( A ) Principal component analysis (PCA) comparing transcriptomic profiles among MYCN-overexpressing retinal organoids (MYCN O/E -RBOs), MYCN O/E -RBOs derived cell lines (MYCN O/E -cells), normal retinal organoids (nROs), and Y79 retinoblastoma cells (RB1 -/- /MYCN A ). MYCN O/E samples formed distinct clusters separate from nROs and Y79, indicating unique transcriptional programs driven by MYCN overexpression. ( B ) Volcano plot showing significantly upregulated (1533 genes, yellow) and downregulated (892 genes, blue) differentially expressed genes (DEGs) between MYCN O/E -RBOs and nROs (adjusted p < 0.05, |Log2FC| > 1). ( C ) Gene Ontology (GO) enrichment analysis highlighting significantly downregulated biological processes (photoreceptor differentiation, phototransduction, sensory perception of light stimulus) and significantly upregulated biological processes (cell cycle phase transitions, mitotic cell cycle regulation, DNA repair pathways) in MYCN O/E -RBOs compared to nROs. ( D , E ) Gene Set Enrichment Analysis (GSEA) demonstrated significant enrichment of MYC target genes, unfolded protein response, mTORC1 signaling, and hypoxia-related pathways in MYCN O/E -RBOs ( D ) and independent MYCN-amplified patient tumor datasets ( E ). Concordant pathway activation between MYCN O/E -RBOs and patient-derived tumors supports the clinical relevance of this organoid model. ( F ) Schematic summary highlighting key molecular signatures identified from transcriptomic analyses. MYCN overexpression in retinoblastoma drives increased expression of MYC target genes, enhanced mTORC1 signaling, activation of cell-cycle-related and neural/retinal ganglion cell (RGC)-related genes, and downregulation of photoreceptor differentiation-related genes, collectively promoting a proliferative, undifferentiated retinal progenitor-like phenotype.

Article Snippet: Retinoblastoma cell lines (Y79 and WERI-Rb1; ATCC, Manassas, VA, USA) were grown in suspension cultures in RPMI-1640 medium (GibcoTM, cat. no. A1049101, Carlsbad, CA, USA) supplemented with 20% fetal bovine serum (FBS; Rd Tech, cat. no. A1500m, Namyangju, Republic of Korea) and 1% penicillin/streptomycin (Thermo Fisher Scientific, cat. no. 15140-122, Waltham, MA, USA).

Techniques: Derivative Assay, Over Expression, Amplification, Activation Assay, Expressing

Journal: International Journal of Molecular Sciences

Article Title: Human Retinal Organoid Modeling Defines Developmental Window and Therapeutic Vulnerabilities in MYCN-Amplified Retinoblastoma

doi: 10.3390/ijms26178675

Figure Lengend Snippet: Selective Sensitivity of MYCN-Overexpressing Retinoblastoma Cells to MYC-Targeted Agents. Dose–response curves for MYCN O/E -cells and RB1-deficient Y79 cells following 48 h treatment with various small-molecule inhibitors. Data represent mean ± SEM ( n = 3), with IC 50 values indicated where applicable (N/A: not applicable due to insufficient efficacy). ( A ) Inhibitors targeting MYC transcription. MYCN O/E -cells exhibited marked sensitivity to the CDK inhibitors THZ1 (IC 50 = 21 nM) and Flavopiridol (IC 50 = 50.46 nM), whereas Y79 cells were largely resistant. ( B ) Inhibitors targeting MYC translation. The dual PI3K/mTOR inhibitor Dactolisib showed moderate efficacy against MYCN O/E -cells (IC 50 = 370.8 nM), whereas the selective mTORC1 inhibitor Rapamycin was ineffective. In contrast, both inhibitors demonstrated negligible efficacy in Y79 cells (IC 50 = N/A). ( C ) Inhibitors targeting MYC stability. The PLK1 inhibitor Volasertib demonstrated potency in the MYCN-driven subtype, effectively inhibiting MYCN O/E -cell viability (IC 50 = 273.7 nM) with minimal impact on Y79 cells.

Article Snippet: Retinoblastoma cell lines (Y79 and WERI-Rb1; ATCC, Manassas, VA, USA) were grown in suspension cultures in RPMI-1640 medium (GibcoTM, cat. no. A1049101, Carlsbad, CA, USA) supplemented with 20% fetal bovine serum (FBS; Rd Tech, cat. no. A1500m, Namyangju, Republic of Korea) and 1% penicillin/streptomycin (Thermo Fisher Scientific, cat. no. 15140-122, Waltham, MA, USA).

Techniques: