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Supplementary Table S3. Comprehensive drug sensitivity and resistance testing with 200 cancer drugs. Cell viability in GIST882 and GIST48 cell lines was measured in five different concentrations over a 10,000-fold concentration range.

Supplementary Figure S1. A box-whisker plot showing the relative KIT, ANO1, and PRKCQ (encodes for protein kinase C theta) mRNA expression in soft-tissue sarcoma samples in the MediSapiens database. Supplementary Figure S2. PDE3A (A) and KIT (B) expression in human intestinal smooth muscle tissue (magnification x400, scale bar 100 μm). Supplementary Figure S3 A, Effects of the combinations of PDE3 siRNAs and DMSO on the GIST882 and GIST48 cell lines. siPDE3B and siPDE3A/B reduced GIST882 cell viability, and none of the treatments influenced markedly GIST48 viability. SEM < 0.05 in all data points. B, Effects of PDE3 inhibitors cilostazol, milrinone, and amrinone given with DMSO on GIST882 and GIST48 cell line viability. SE < 0.05 in all data points. Supplementary Figure S4. A Sankey diagram of top SPIA pathways. A Sankey diagram showing differentially expressed miRNAs, their predicted target genes, and the biological pathways estimated to be significantly impacted by the expression change (SPIA pGFDR

Supplementary Table S5. List of genes most affected by anagrelide treatment and protein expression data.

Supplemental legend

Supplementary Table S4. miRNA expression, mRNA expression, and associated pathways perturbed in the GIST882 cell line after anagrelide treatment.

Supplementary Materials and Methods

Supplementary Table S6. The primers used in the quantitative PCR analyses.

Supplementary Table S1. The outlier mRNA expression profiles of the gene tissue index (GTI) outlier analysis. The list of 2415 outlier genes in GIST is ranked by the gene tissue index.

Purpose:Gastrointestinal stromal tumor (GIST) is a common type of soft-tissue sarcoma. Imatinib, an inhibitor of KIT, platelet-derived growth factor receptor alpha (PDGFRA), and a few other tyrosine kinases, is highly effective for GIST, but advanced GISTs frequently progress on imatinib and other approved tyrosine kinase inhibitors. We investigated phosphodiesterase 3 (PDE3) as a potential therapeutic target in GIST cell lines and xenograft models.Experimental Design:The GIST gene expression profile was interrogated in the MediSapiens IST Online transcriptome database comprising human tissue and cancer samples, and PDE3A and PDE3B expression was studied using IHC on tissue microarrays (TMA) consisting of 630 formalin-fixed human tissue samples. GIST cell lines were screened for sensitivity to 217 anticancer compounds, and the efficacy of PDE inhibitors on GIST was further studied in GIST cell lines and patient-derived mouse xenograft models.Results:GISTs expressed PDE3A and PDE3B frequently compared with other human normal or cancerous tissues both in the in silico database and the TMAs. Anagrelide was identified as the most potent of the PDE3 modulators evaluated. It reduced cell viability, promoted cell death, and influenced cell signaling in GIST cell lines. Anagrelide inhibited tumor growth in GIST xenograft mouse models. Anagrelide was also effective in a GIST xenograft mouse model with KIT exon 9 mutation that may pose a therapeutic challenge, as these GISTs require a high daily dose of imatinib.Conclusions:PDE3A and PDE3B are frequently expressed in GIST. Anagrelide had anticancer efficacy in GIST xenograft models and warrants further testing in clinical trials.

Purpose: Gastrointestinal stromal tumor (GIST) is a common type of soft-tissue sarcoma. Imatinib, an inhibitor of KIT, platelet-derived growth factor receptor alpha (PDGFRA), and a few other tyrosine kinases, is highly effective for GIST, but advanced GISTs frequently progress on imatinib and other approved tyrosine kinase inhibitors. We investigated phosphodiesterase 3 (PDE3) as a potential therapeutic target in GIST cell lines and xenograft models. Experimental Design: The GIST gene expression profile was interrogated in the MediSapiens IST Online transcriptome database comprising human tissue and cancer samples, and PDE3A and PDE3B expression was studied using IHC on tissue microarrays (TMA) consisting of 630 formalin-fixed human tissue samples. GIST cell lines were screened for sensitivity to 217 anticancer compounds, and the efficacy of PDE inhibitors on GIST was further studied in GIST cell lines and patient-derived mouse xenograft models. Results: GISTs expressed PDE3A and PDE3B frequently compared with other human normal or cancerous tissues both in the in silico database and the TMAs. Anagrelide was identified as the most potent of the PDE3 modulators evaluated. It reduced cell viability, promoted cell death, and influenced cell signaling in GIST cell lines. Anagrelide inhibited tumor growth in GIST xenograft mouse models. Anagrelide was also effective in a GIST xenograft mouse model with KIT exon 9 mutation that may pose a therapeutic challenge, as these GISTs require a high daily dose of imatinib. Conclusions: PDE3A and PDE3B are frequently expressed in GIST. Anagrelide had anticancer efficacy in GIST xenograft models and warrants further testing in clinical trials.