Jessica Camacho

Full results of GSEA of Hallmark pathway analysis after 90Y microsphere treatment in resistant and sensitive cell lines.

QRT-PCR validation of gene expression representing relevant biological pathways after 90Y microsphere treatment in the most sensitive (PLC/PRF/5), resistant (SK-Hep1), and intermediate (SNU-398) cell lines. Select genes involved in (A) interferon stimulation and antigen presentation were mostly upregulated in both SNU-398 and SK-Hep1 after treatment, with the exception of GBP1 in SK-Hep1 and MX1 and IFI27 in SNU-398 which did not have reliable qPCR readouts. TXNIP was strongly upregulated in SK-Hep1 after treatment, consistent with increased oxidative stress signaling. (B) SNU-398 demonstrated very strong upregulation of inflammatory response genes, in particular CCL5 and TNFAIP2. C, Distinct patterns of DNA damage and cell stress genes were seen across cell lines, with SK-Hep1 showing upregulation of BRCA1 and downregulation of BNIP3. D, ECM genes CD44 and ITGA3 were strongly upregulated in SNU-398 after treatment, suggesting stress-induced EMT acquisition. Although there was slight downregulation of these genes in SK-Hep1, they remained at highest abundance in this line consistent with an EMT-associated expression profile. All experiments were performed in technical triplicate and error bars represent SEM when >1 biological replicate was performed. nd, no reliable qRT-PCR readout.

Primer sequences for genes used for RT-qPCR

Differential expression and GSEA after 90Y treatment in individual cell lines.

Demographics and clinical characteristics of exploratory patient cohort.

Details of liver cancer cell lines

90Y-resistant and -sensitive cell lines demonstrate distinct patterns of biological process enrichment after 90Y microsphere treatment. A, Volcano plot of log2 FC vs. −log10P value of genes upregulated (red) and downregulated (blue) after 90Y microsphere treatment in select 90Y-resistant cell lines (SK-Hep1, SNU-449, and SNU-387). B, GSEA of Hallmark pathways demonstrates upregulation of pathways involved with inflammation and immune response after treatment in the 90Y-resistant group, such as IFNγ response (mean log2FC = 7.4), among others. C, Consistent with Hallmark IFNγ and IFNα enrichment, KEGG and Reactome pathways associated with cytokine signaling and antigen presentation were upregulated after treatment with 90Y. D, Volcano plot of log2 FC vs. −log10P value of genes upregulated (red) and downregulated (blue) after 90Y microsphere treatment in select 90Y-sensitive cell lines (PLC/PRF/5 and HepG2). E, GSEA of Hallmark pathways demonstrates downregulation of many of the inflammatory and immune response–related pathways that were upregulated in the resistant cell lines, including IFNγ response (mean log2 FC = −4.3,) and IFNα response (mean log2 FC = −4.7). No Hallmark pathways demonstrated significant upregulation after treatment in 90Y-sensitive cells. F, KEGG and Reactome pathway analysis corroborates downregulation of interferon- and antigen presentation–associated pathways, along with those involved with growth signaling such as MAPK (mean log FC = −4.01) and TGFβ (mean log FC = −2.62). Numbers next to each gene set bar represent FDR. Significance set at FDR 2. Heterogeneous activation of stress and survival pathways after 90Y microsphere treatment across different liver cancer cell lines. Significant variation in Hallmark gene sets (G) IFNγ, (H) IFNα, (I) TNFα signaling, (J) hypoxia, (K) myogenesis, (L) p53 pathway, (M) oxidative phosphorylation, and (N) DNA repair, showing heterogeneity in stress and survival pathway activation across cell lines after 90Y microsphere treatment.

ANOVA results comparing nAUC between cell lines.

Relative expression of CD44, ITGA3, and Apo genes.

Differential gene expression between 90Y-resistant and sensitive cell lines after 90Y microsphere treatment