Mardiyyah/TAPT_data_V2_split · Datasets at Hugging Face

PMCID stringclasses 24 values	Title stringclasses 24 values	Sentences stringlengths 2 40.7k
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Slides with 3.5-µm tissue sections were incubated at 56 °C for 1 h prior to deparaffinization with Tissue-Tek DRS 2000 Automated Slide Stainer (Sakura Finetek Japan, Tokyo, Japan).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	After blocking endogenous peroxidase with hydrogen peroxide, antigen retrieval was performed with EnVision FLEX Target Retrieval Solution Low pH (pH 6, Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. K8005, Agilent) in a decloaking chamber (Biocare Medical, Pacheco, CA, USA) at 110 °C for 3 min.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The slides were then incubated with polyclonal rabbit anti-PDE3A (1:100, Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. HPA014492, Sigma-Aldrich, St. Louis, MO, USA) in WellMed BrightDiluent for 30 min (Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. BD09-500, ImmunoLogic, Duiven, The Netherlands).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	WellMed BrightVision (one component detection system, Goat Anti-Rabbit IgG HRP, Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. DPVR500HRP, ImmunoLogic) was used for antibody detection and the slides were stained with an ImmPACT DAB Peroxidase (HRP) substrate kit (Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. SK-4105, Vector Laboratories, Burlingame, CA, USA).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The tissues were counterstained with hematoxylin (Mayer’s, Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. S3309, Agilent).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The slides were scanned with Pannoramic 250 Flash III (3DHISTECH) in the Biobank Helsinki and analyzed with a slide viewer (3DHISTECH).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Interstitial cells of Cajal from healthy colon tissue were used as a positive control for PDE3A staining.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The intensity of PDE3A staining on TMA was scored as 0 (no staining in tumor cells), 1 (weak), 2 (moderate), or 3 (strong).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Due to a heterogeneous staining pattern in some of the samples, PDE3A expression was evaluated in whole-tissue sections in the LPS series, based on H-scores (staining intensity (0–3) × percentage of positive cells).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	For statistical analysis, the results were further divided into quartiles (0: 0–75; 1: 76–150; 2: 151–225; 3: 226–300) or into two groups (low: 0–150, and high: 151–300), when comparing the differences between LPS subtypes.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	GOT3 (RRID:CVCL_M819), MLS1765-92 (RRID:CVCL_S817), and MLS402-91 (RRID:CVCL_S813) cell lines were established and kindly provided by Prof. Pierre Åman (University of Gothenburg, Sweden); GIST882 (RRID:CVCL_7044) and LPS141 (RRID:CVCL_M823) cell lines were kindly provided by Dr. Jonathan Fletcher (Harvard Medical School, Boston, MA, USA); and the SA4 (RRID:CVCL_8910) cell line was kindly provided by Dr. Kjetil Boye (Oslo University Hospital, Oslo, Norway).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The 93T449 (RRID:CVCL_U614) (Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. CRL-3043), 94T778 (RRID:CVCL_U613) (Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. CRL-3044) and SW872 (RRID:CVCL_1730) (Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. HTB-92) cell lines were purchased from ATCC (Manassas, VA, USA).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	All cell lines used in this study were authenticated using short-tandem-repeat (STR) profiling in the genotyping lab of the FIMM Technology Centre.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	GIST882 was verified as containing a known KIT exon 13 p.K642E mutation.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The SA4 cell line was verified by Western blot as not being HeLa-contaminated, where the HPV18 E7 protein was not detected.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	All cell lines tested negative for mycoplasma and were cultured in a humidified, 5% CO2 atmosphere at 37 °C.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The cell lines were cultured in Gibco™ RPMI Medium 1640 (Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. 11530586, Life Technologies, Carlsbad, CA, USA) with Pen Strep Glut and 5% (MLS402-91 and MLS1765-92), 10% (SA4, GOT3, 93T449, and 94T778), or 20% (GIST882, LPS141 and SW872) FBS.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The cells were lysed in M-PER™ Mammalian Protein Extraction Reagent (Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. 78501, Thermo Fisher Scientific) with HALT™ protease and phosphatase inhibitor cocktails (Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. 78429 and Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. 78420, Thermo Fisher Scientific).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Denatured lysates were subjected to SDS-PAGE, using Mini-PROTEAN TGX™ Precast Gels (Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. 456-1034, Bio-Rad), and blotted to PVDF membranes (Mini Format, Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. 1704156, Bio-Rad).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	After blocking, the membranes were incubated overnight in primary antibodies at 4 °C.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Polyclonal goat anti-rabbit IgG (1:10,000, Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. 111-035-003, Jackson ImmunoResearch, West Grove, PA, USA) was used as a secondary antibody.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Antigens were detected with SuperSignal™ West Pico PLUS chemiluminescent substrate (Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. 34580, Thermo Fisher Scientific) on autoradiography films and quantitated using Fiji ImageJ 1.53 (64-bit).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Antibodies used for Western blotting were polyclonal rabbit anti-PDE3A (1:1000, Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. HPA014492, Sigma-Aldrich) and monoclonal rabbit anti-SLFN12 (1:500, Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. ab234418, Abcam, Cambridge, UK).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Target protein expression was normalized to polyclonal rabbit anti-cytoskeletal actin (1:150 000, Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. A300-491A, Bethyl Laboratories, Montgomery, TX, USA).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The following drugs were used in the experiments: anagrelide hydrochloride (CAS No. 58579-51-4, Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. 735554, Lancrix, Shanghai, China), BAY 2666605 (CAS No. 2275774-60-0, Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. HY-145924, MedChemExpress, Monmouth Junction, NJ, USA), cilostazol (CAS No. 73963-72-1, Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. C0737, Sigma–Aldrich), and DNMDP (CAS No. 328104-79-6, Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. HY-W028690, MedChemExpress).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	All of the drugs were dissolved in dimethyl sulfoxide (DMSO).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	A corresponding DMSO concentration in cell media was used as the control treatment.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	A total of 10,000 GIST882 cells, 3000 SA4 cells, and 1800 GOT3 cells were seeded per well on a Falcon 96-Well White/Clear bottom microplate (Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. 353377, CORNING, Corning, NY, USA) and allowed to attach for 24 h before the addition of drugs.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The number of cells plated in wells differed between cell lines to attain 70–80% confluency in the control treatment on the final measuring day.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Cell viabilities were measured with CellTiter-Glo Luminescent Cell Viability Assay (Cat.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No. G7572, Promega, Madison, WI, USA), following the manufacturer’s instructions.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Luminescence was recorded with a Hidex Sense microplate reader (Hidex Oy, Turku, Finland).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Background values from media were excluded from the results.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Eight well replicates per treatment were always performed.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The results presented were obtained from two separate experiments.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Luminescence measurements were divided by the average of control treatment, indicating a relative cell viability change percentage.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Statistical analyses were performed using IBM SPSS Statistics 28.0.0.0 (Chicago, IL, USA).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	p values < 0.05 were considered statistically significant.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Cross tabulations were analyzed using a Pearson χ test or a Fisher–Freeman–Halton test.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Correlations between PDE3A staining with age at time of diagnosis and tumor size were examined using the Mann–Whitney U Test.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Correlations between PDE3A expression and overall survival or metastasis-free survival were estimated with the Kaplan–Meier method and compared using the univariate Cox proportional hazard model.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Survival was calculated from the date of diagnosis to the date of death or the date of the first detected metastases, excluding patients who were without any events at the end of follow-up.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The correlation between the PDE3A H-score and RT-qPCR expression was analyzed using Spearman’s rank-order correlation, and the correlation between PDE3A and SLFN12 mRNA expression was analyzed with Pearson correlation.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Statistical significance was analyzed using the Kruskall–Wallis test to compare PDE3A and SLFN12 mRNA expression levels between the LPS subtypes and gender.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Differences in cell viability across treatments were analyzed using a two-tailed unpaired t-test.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	A comparison between transcriptomes of 53 DDLPS samples, 54 MLPS samples, and 24 PLPS samples and the IST transcriptome database identified 97 highly expressed genes specific in DDLPS, 247 highly expressed genes specific in MLPS, and 37 highly expressed genes specific in PLPS (Table S2).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The top 25 upregulated genes in each subtype, with the corresponding logFC (fold change) and p values, are shown in Table 1.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Genes related to CDK4 and MDM2 co-amplification, B4GALNT1, TSPAN31, YEATS4, HMGA2, FRS2, and GLI1 have been described in WDLPS/DDLPS and emerged in our DDLPS gene list .
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	SHANK2 and CTAG2 emerged in the MLPS-specific list and AXL emerged in the PLPS-specific list; these genes have previously been described as upregulated in these subtypes .
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	For emerging oncogenic genes that have been characterized in other cancers, we identified PAPPA , FBN2 , and METTL1 in DDLPS, POTEE and LBX1 in MLPS, and CDCA2 and TROAP in PLPS as differentially expressed genes.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Figure 1 shows the top 10 enriched pathways, with their overlapping genes, in three LPS subtypes.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The most important pathways in DDLPS were the hedgehog signaling pathway and the calcium-activated potassium channel, which were previously described as being involved in many cancers .
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The two most upregulated genes identified in DDLPS, GLI1 and HHIP, are related to the hedgehog signaling pathways.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The MLPS-specific DEGs displayed upregulated phospholipase C-mediated cascade and insulin signaling pathways, which are also recognized pathways that are altered in cancer .
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The pathways identified in PLPS formed an apparent cluster and are involved in M phase, DNA replication, kinesins, and cell cycle.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Phosphodiesterase 3A (PDE3A) emerged in the MLPS gene list (Table S2) and is related to upregulated insulin signaling (Figure 1).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	PDE3A protein is highly expressed in cardiovascular tissues and has been identified as a promising drug target in GISTs .
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Therefore, we decided to further investigate its role in LPS.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	IHC staining for PDE3A in a TMA series consisting of nine different STS types revealed expression in nearly half of the samples (Table 2).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Examples illustrating the staining intensities in different STS types are shown in Figure 2.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Consistent with previous studies, PDE3A was most frequently expressed in GISTs .
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	GIST samples also exhibited PDE3A expression that was significantly higher than it was in other STS subtypes.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Other STS types that most often exhibited moderate or high PDE3A expression (staining intensity ≥2) were leiomyosarcoma (39.0%), malignant peripheral nerve sheath tumors (25.0%), not-otherwise-specified sarcoma (21.2%), and LPS (19.5%).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The transcriptome data (Table S2) indicated that PDE3A expression was higher in the MLPS subtype than in various other tissue types or the two other LPS subtypes.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Therefore, we investigated 181 LPS tissue sections with specified histology, using IHC.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Consistent with the transcriptome data, elevated PDE3A expression was associated with the MLPS subtype, but also with male gender (Table 3) (p < 0.05).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Associations between PDE3A expression and tumor site, age at time of diagnosis, tumor size, metastasis at diagnosis, overall survival (hazard ratio = 0.853, 95% confidence interval = 0.479–1.519, p = 0.588), or metastasis-free survival (hazard ratio = 0.905, 95% confidence interval = 0.374–2.190, p = 0.824) were not observed.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	PDE3A expression was also analyzed by dividing the samples into quartiles, based on H-scores (Figure 3A).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Examples of PDE3A immunostaining intensity levels in LPS tissues are shown in Figure 3B.
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Interestingly, in comparison to low-grade MLPS, high PDE3A expression was particularly associated with high-grade MLPS (p = 0.032) (Figure 3A).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	Higher PDE3A expression in MLPS was also seen in mRNA levels (Figure S1A), and a positive correlation between mRNA and protein expression was observed (Spearman’s ρ = 0.423, p < 0.001) (Figure S1B).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	The cytotoxic effect of PDE3A modulators is based on their ability to induce complex formation between PDE3A and SLFN12, and drug sensitivity positively correlates with expression levels of the two proteins .
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	We examined SLFN12 mRNA expression in 62 LPS samples and found it to be expressed at higher levels in MLPS samples than in DDLPS and PLPS samples (Figure 3C) (Kruskal–Wallis test, p = 0.012).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	In addition, there was a correlation between SLFN12 and PDE3A mRNA expression levels (Pearson correlation coefficient = 0.362, p = 0.004); almost all of the samples (11/12) that exhibited equal or greater PDE3A and SLFN12 mRNA expression than a PDE3A modulator-sensitive cell line were MLPS samples (Figure 3D).
PMC10669966	PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma	No association between male gender and PDE3A was observed in the mRNA levels (Kruskal–Wallis test, p = 0.389).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Slides with 3.5-µm tissue sections were incubated at 56 °C for 1 h prior to deparaffinization with Tissue-Tek DRS 2000 Automated Slide Stainer (Sakura Finetek Japan, Tokyo, Japan).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

After blocking endogenous peroxidase with hydrogen peroxide, antigen retrieval was performed with EnVision FLEX Target Retrieval Solution Low pH (pH 6, Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. K8005, Agilent) in a decloaking chamber (Biocare Medical, Pacheco, CA, USA) at 110 °C for 3 min.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The slides were then incubated with polyclonal rabbit anti-PDE3A (1:100, Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. HPA014492, Sigma-Aldrich, St. Louis, MO, USA) in WellMed BrightDiluent for 30 min (Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. BD09-500, ImmunoLogic, Duiven, The Netherlands).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

WellMed BrightVision (one component detection system, Goat Anti-Rabbit IgG HRP, Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. DPVR500HRP, ImmunoLogic) was used for antibody detection and the slides were stained with an ImmPACT DAB Peroxidase (HRP) substrate kit (Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. SK-4105, Vector Laboratories, Burlingame, CA, USA).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The tissues were counterstained with hematoxylin (Mayer’s, Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. S3309, Agilent).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The slides were scanned with Pannoramic 250 Flash III (3DHISTECH) in the Biobank Helsinki and analyzed with a slide viewer (3DHISTECH).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Interstitial cells of Cajal from healthy colon tissue were used as a positive control for PDE3A staining.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The intensity of PDE3A staining on TMA was scored as 0 (no staining in tumor cells), 1 (weak), 2 (moderate), or 3 (strong).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Due to a heterogeneous staining pattern in some of the samples, PDE3A expression was evaluated in whole-tissue sections in the LPS series, based on H-scores (staining intensity (0–3) × percentage of positive cells).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

For statistical analysis, the results were further divided into quartiles (0: 0–75; 1: 76–150; 2: 151–225; 3: 226–300) or into two groups (low: 0–150, and high: 151–300), when comparing the differences between LPS subtypes.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

GOT3 (RRID:CVCL_M819), MLS1765-92 (RRID:CVCL_S817), and MLS402-91 (RRID:CVCL_S813) cell lines were established and kindly provided by Prof. Pierre Åman (University of Gothenburg, Sweden); GIST882 (RRID:CVCL_7044) and LPS141 (RRID:CVCL_M823) cell lines were kindly provided by Dr. Jonathan Fletcher (Harvard Medical School, Boston, MA, USA); and the SA4 (RRID:CVCL_8910) cell line was kindly provided by Dr. Kjetil Boye (Oslo University Hospital, Oslo, Norway).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The 93T449 (RRID:CVCL_U614) (Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. CRL-3043), 94T778 (RRID:CVCL_U613) (Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. CRL-3044) and SW872 (RRID:CVCL_1730) (Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. HTB-92) cell lines were purchased from ATCC (Manassas, VA, USA).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

All cell lines used in this study were authenticated using short-tandem-repeat (STR) profiling in the genotyping lab of the FIMM Technology Centre.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

GIST882 was verified as containing a known KIT exon 13 p.K642E mutation.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The SA4 cell line was verified by Western blot as not being HeLa-contaminated, where the HPV18 E7 protein was not detected.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

All cell lines tested negative for mycoplasma and were cultured in a humidified, 5% CO2 atmosphere at 37 °C.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The cell lines were cultured in Gibco™ RPMI Medium 1640 (Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. 11530586, Life Technologies, Carlsbad, CA, USA) with Pen Strep Glut and 5% (MLS402-91 and MLS1765-92), 10% (SA4, GOT3, 93T449, and 94T778), or 20% (GIST882, LPS141 and SW872) FBS.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The cells were lysed in M-PER™ Mammalian Protein Extraction Reagent (Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. 78501, Thermo Fisher Scientific) with HALT™ protease and phosphatase inhibitor cocktails (Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. 78429 and Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. 78420, Thermo Fisher Scientific).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Denatured lysates were subjected to SDS-PAGE, using Mini-PROTEAN TGX™ Precast Gels (Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. 456-1034, Bio-Rad), and blotted to PVDF membranes (Mini Format, Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. 1704156, Bio-Rad).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

After blocking, the membranes were incubated overnight in primary antibodies at 4 °C.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Polyclonal goat anti-rabbit IgG (1:10,000, Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. 111-035-003, Jackson ImmunoResearch, West Grove, PA, USA) was used as a secondary antibody.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Antigens were detected with SuperSignal™ West Pico PLUS chemiluminescent substrate (Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. 34580, Thermo Fisher Scientific) on autoradiography films and quantitated using Fiji ImageJ 1.53 (64-bit).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Antibodies used for Western blotting were polyclonal rabbit anti-PDE3A (1:1000, Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. HPA014492, Sigma-Aldrich) and monoclonal rabbit anti-SLFN12 (1:500, Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. ab234418, Abcam, Cambridge, UK).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Target protein expression was normalized to polyclonal rabbit anti-cytoskeletal actin (1:150 000, Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. A300-491A, Bethyl Laboratories, Montgomery, TX, USA).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The following drugs were used in the experiments: anagrelide hydrochloride (CAS No. 58579-51-4, Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. 735554, Lancrix, Shanghai, China), BAY 2666605 (CAS No. 2275774-60-0, Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. HY-145924, MedChemExpress, Monmouth Junction, NJ, USA), cilostazol (CAS No. 73963-72-1, Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. C0737, Sigma–Aldrich), and DNMDP (CAS No. 328104-79-6, Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. HY-W028690, MedChemExpress).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

All of the drugs were dissolved in dimethyl sulfoxide (DMSO).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

A corresponding DMSO concentration in cell media was used as the control treatment.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

A total of 10,000 GIST882 cells, 3000 SA4 cells, and 1800 GOT3 cells were seeded per well on a Falcon 96-Well White/Clear bottom microplate (Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. 353377, CORNING, Corning, NY, USA) and allowed to attach for 24 h before the addition of drugs.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The number of cells plated in wells differed between cell lines to attain 70–80% confluency in the control treatment on the final measuring day.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Cell viabilities were measured with CellTiter-Glo Luminescent Cell Viability Assay (Cat.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No. G7572, Promega, Madison, WI, USA), following the manufacturer’s instructions.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Luminescence was recorded with a Hidex Sense microplate reader (Hidex Oy, Turku, Finland).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Background values from media were excluded from the results.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Eight well replicates per treatment were always performed.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The results presented were obtained from two separate experiments.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Luminescence measurements were divided by the average of control treatment, indicating a relative cell viability change percentage.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Statistical analyses were performed using IBM SPSS Statistics 28.0.0.0 (Chicago, IL, USA).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

p values < 0.05 were considered statistically significant.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Cross tabulations were analyzed using a Pearson χ test or a Fisher–Freeman–Halton test.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Correlations between PDE3A staining with age at time of diagnosis and tumor size were examined using the Mann–Whitney U Test.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Correlations between PDE3A expression and overall survival or metastasis-free survival were estimated with the Kaplan–Meier method and compared using the univariate Cox proportional hazard model.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Survival was calculated from the date of diagnosis to the date of death or the date of the first detected metastases, excluding patients who were without any events at the end of follow-up.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The correlation between the PDE3A H-score and RT-qPCR expression was analyzed using Spearman’s rank-order correlation, and the correlation between PDE3A and SLFN12 mRNA expression was analyzed with Pearson correlation.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Statistical significance was analyzed using the Kruskall–Wallis test to compare PDE3A and SLFN12 mRNA expression levels between the LPS subtypes and gender.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Differences in cell viability across treatments were analyzed using a two-tailed unpaired t-test.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

A comparison between transcriptomes of 53 DDLPS samples, 54 MLPS samples, and 24 PLPS samples and the IST transcriptome database identified 97 highly expressed genes specific in DDLPS, 247 highly expressed genes specific in MLPS, and 37 highly expressed genes specific in PLPS (Table S2).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The top 25 upregulated genes in each subtype, with the corresponding logFC (fold change) and p values, are shown in Table 1.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Genes related to CDK4 and MDM2 co-amplification, B4GALNT1, TSPAN31, YEATS4, HMGA2, FRS2, and GLI1 have been described in WDLPS/DDLPS and emerged in our DDLPS gene list .

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

SHANK2 and CTAG2 emerged in the MLPS-specific list and AXL emerged in the PLPS-specific list; these genes have previously been described as upregulated in these subtypes .

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

For emerging oncogenic genes that have been characterized in other cancers, we identified PAPPA , FBN2 , and METTL1 in DDLPS, POTEE and LBX1 in MLPS, and CDCA2 and TROAP in PLPS as differentially expressed genes.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Figure 1 shows the top 10 enriched pathways, with their overlapping genes, in three LPS subtypes.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The most important pathways in DDLPS were the hedgehog signaling pathway and the calcium-activated potassium channel, which were previously described as being involved in many cancers .

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The two most upregulated genes identified in DDLPS, GLI1 and HHIP, are related to the hedgehog signaling pathways.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The MLPS-specific DEGs displayed upregulated phospholipase C-mediated cascade and insulin signaling pathways, which are also recognized pathways that are altered in cancer .

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The pathways identified in PLPS formed an apparent cluster and are involved in M phase, DNA replication, kinesins, and cell cycle.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Phosphodiesterase 3A (PDE3A) emerged in the MLPS gene list (Table S2) and is related to upregulated insulin signaling (Figure 1).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

PDE3A protein is highly expressed in cardiovascular tissues and has been identified as a promising drug target in GISTs .

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Therefore, we decided to further investigate its role in LPS.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

IHC staining for PDE3A in a TMA series consisting of nine different STS types revealed expression in nearly half of the samples (Table 2).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Examples illustrating the staining intensities in different STS types are shown in Figure 2.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Consistent with previous studies, PDE3A was most frequently expressed in GISTs .

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

GIST samples also exhibited PDE3A expression that was significantly higher than it was in other STS subtypes.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Other STS types that most often exhibited moderate or high PDE3A expression (staining intensity ≥2) were leiomyosarcoma (39.0%), malignant peripheral nerve sheath tumors (25.0%), not-otherwise-specified sarcoma (21.2%), and LPS (19.5%).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The transcriptome data (Table S2) indicated that PDE3A expression was higher in the MLPS subtype than in various other tissue types or the two other LPS subtypes.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Therefore, we investigated 181 LPS tissue sections with specified histology, using IHC.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Consistent with the transcriptome data, elevated PDE3A expression was associated with the MLPS subtype, but also with male gender (Table 3) (p < 0.05).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Associations between PDE3A expression and tumor site, age at time of diagnosis, tumor size, metastasis at diagnosis, overall survival (hazard ratio = 0.853, 95% confidence interval = 0.479–1.519, p = 0.588), or metastasis-free survival (hazard ratio = 0.905, 95% confidence interval = 0.374–2.190, p = 0.824) were not observed.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

PDE3A expression was also analyzed by dividing the samples into quartiles, based on H-scores (Figure 3A).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Examples of PDE3A immunostaining intensity levels in LPS tissues are shown in Figure 3B.

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Interestingly, in comparison to low-grade MLPS, high PDE3A expression was particularly associated with high-grade MLPS (p = 0.032) (Figure 3A).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

Higher PDE3A expression in MLPS was also seen in mRNA levels (Figure S1A), and a positive correlation between mRNA and protein expression was observed (Spearman’s ρ = 0.423, p < 0.001) (Figure S1B).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

The cytotoxic effect of PDE3A modulators is based on their ability to induce complex formation between PDE3A and SLFN12, and drug sensitivity positively correlates with expression levels of the two proteins .

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

We examined SLFN12 mRNA expression in 62 LPS samples and found it to be expressed at higher levels in MLPS samples than in DDLPS and PLPS samples (Figure 3C) (Kruskal–Wallis test, p = 0.012).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

In addition, there was a correlation between SLFN12 and PDE3A mRNA expression levels (Pearson correlation coefficient = 0.362, p = 0.004); almost all of the samples (11/12) that exhibited equal or greater PDE3A and SLFN12 mRNA expression than a PDE3A modulator-sensitive cell line were MLPS samples (Figure 3D).

PMC10669966

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma

No association between male gender and PDE3A was observed in the mRNA levels (Kruskal–Wallis test, p = 0.389).