KLF4 inhibited the senescence-associated secretory phenotype in ox-LDL-treated endothelial cells via PDGFRA/NAMPT/mitochondrial ROS.

BACKGROUND: Inflammation is one of the significant consequences of ox-LDL-induced endothelial cell (EC) dysfunction. The senescence-associated secretory phenotype (SASP) is a critical source of inflammation factors. However, the molecular mechanism by which the SASP is regulated in ECs under ox-LDL conditions remains unknown. RESULTS: The level of SASP was increased in ox-LDL-treated ECs, which could be augmented by KLF4 knockdown whereas restored by KLF4 knock-in. Furthermore, we found that KLF4 directly promoted PDGFRA transcription and confirmed the central role of the NAPMT/mitochondrial ROS pathway in KLF4/PDGFRA-mediated inhibition of SASP. Animal experiments showed a higher SASP HFD-fed mice, compared with normal feed (ND)-fed mice, and the endothelium of EC-specific KLF4-/- mice exhibited a higher proportion of SA-ß-gal-positive cells and lower PDGFRA/NAMPT expression. CONCLUSIONS: Our results revealed that KLF4 inhibits the SASP of endothelial cells under ox-LDL conditions through the PDGFRA/NAMPT/mitochondrial ROS. METHODS: Ox-LDL-treated ECs and HFD-fed mice were used as endothelial senescence models in vitro and in vivo. SA-ß-gal stain, detection of SAHF and the expression of inflammatory factors determined SASP and senescence of ECs. The direct interaction of KLF4 and PDGFRA promotor was analyzed by EMSA and fluorescent dual luciferase reporting analysis.

TAD border deletion at the Kit locus causes tissue-specific ectopic activation of a neighboring gene.

Topologically associated domains (TADs) restrict promoter-enhancer interactions, thereby maintaining the spatiotemporal pattern of gene activity. However, rearrangements of the TADs boundaries do not always lead to significant changes in the activity pattern. Here, we investigated the consequences of the TAD boundaries deletion on the expression of developmentally important genes encoding tyrosine kinase receptors: Kit, Kdr, Pdgfra. We used genome editing in mice to delete the TADs boundaries at the Kit locus and characterized chromatin folding and gene expression in pure cultures of fibroblasts, mast cells, and melanocytes. We found that although Kit is highly active in both mast cells and melanocytes, deletion of the TAD boundary between the Kit and Kdr genes results in ectopic activation only in melanocytes. Thus, the epigenetic landscape, namely the mutual arrangement of enhancers and actively transcribing genes, is important for predicting the consequences of the TAD boundaries removal. We also found that mice without a TAD border between the Kit and Kdr genes have a phenotypic manifestation of the mutation - a lighter coloration. Thus, the data obtained shed light on the principles of interaction between the 3D chromatin organization and epigenetic marks in the regulation of gene activity.

ETV6::NTRK3 Fusion-Positive Wild-Type Gastrointestinal Stromal Tumor (GIST) with Abundant Lymphoid Infiltration (TILs and Tertiary Lymphoid Structures): A Report on a New Case with Therapeutic Implications and a Literature Review.

Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors of the gastrointestinal tract, with proto-oncogene, receptor tyrosine kinase (c-kit), or PDGFRα mutations detected in around 85% of cases. GISTs without c-kit or platelet-derived growth factor receptor alpha (PDGFRα) mutations are considered wild-type (WT), and their diverse molecular alterations and biological behaviors remain uncertain. They are usually not sensitive to tyrosine kinase inhibitors (TKIs). Recently, some molecular alterations, including neurotrophic tyrosine receptor kinase (NTRK) fusions, have been reported in very few cases of WT GISTs. This novel finding opens the window for the use of tropomyosin receptor kinase (TRK) inhibitor therapy in these subtypes of GIST. Herein, we report a new case of NTRK-fused WT high-risk GIST in a female patient with a large pelvic mass (large dimension of 20 cm). The tumor was removed, and the histopathology displayed spindle-predominant morphology with focal epithelioid areas, myxoid stromal tissue, and notable lymphoid infiltration with tertiary lymphoid structures. Ten mitoses were quantified in 50 high-power fields without nuclear pleomorphism. DOG1 showed strong and diffuse positivity, and CD117 showed moderate positivity. Succinate dehydrogenase subunit B (SDHB) was retained, Pan-TRK was focal positive (nuclear pattern), and the proliferation index Ki-67 was 7%. Next-generation sequencing (NGS) detected an ETV6::NTRK3 fusion, and this finding was confirmed by fluorescence in situ hybridization (FISH), which showed NTRK3 rearrangement. In addition, an RB1 mutation was found by NGS. The follow-up CT scan revealed peritoneal nodules suggestive of peritoneal dissemination, and Entrectinib (a TRK inhibitor) was administered. After 3 months of follow-up, a new CT scan showed a complete response. Based on our results and the cases from the literature, GISTs with NTRK fusions are very uncommon so far; hence, further screening studies, including more WT GIST cases, may increase the possibility of finding additional cases. The present case may offer new insights into the potential introduction of TRK inhibitors as treatments for GISTs with NTRK fusions. Additionally, the presence of abundant lymphoid infiltration in the present case may prompt further research into immunotherapy as a possible additional therapeutic option.

Fusion of Platelet Derived Growth Factor Receptor Alpha (PDGFRA) With Ubiquitin Specific Peptidase 8 (USP8) in a Calcified Chondroid Mesenchymal Neoplasm Harboring t(4;15)(q12;q21) as a Sole Aberration.

BACKGROUND/AIM: The term "calcified chondroid mesenchymal neoplasm" was introduced in 2021 to describe a group of tumors characterized by various morphological features, including the formation of cartilage or chondroid matrix. These tumors frequently carry chimeric genes where the 5'-end partner gene is fibronectin 1 and the 3'-end partner gene codes for receptor tyrosine kinase. Our study explores fusion of the genes platelet-derived growth factor receptor alpha (PDGFRA) and ubiquitin-specific peptidase 8 (USP8) in calcified chondroid mesenchymal neoplasm. CASE REPORT: Genetic investigations were conducted on a tumor located in the leg of a 71-year-old woman. G-banding analysis of short-term cultured tumor cells revealed the karyotype 46,XX,t(4;15)(q12;q21)[6]/46,XX[4]. RNA sequencing detected in-frame PDGFRA::USP8 and USP8::PDGFRA chimeric transcripts, which were validated by RT-PCR/Sanger sequencing. The PDGFRA::USP8 chimeric protein is predicted to have cell membrane location and functions as a chimeric ubiquitinyl hydrolase. The USP8::PDGFRA protein was predicted to be nuclear and function as a positive regulator of cellular metabolic process. CONCLUSION: We report, for the first time, a calcified chondroid mesenchymal neoplasm carrying a balanced t(4;15)(q12;q21) chromosomal translocation, resulting in the generation of both PDGFRA::USP8 and USP8::PDGFRA chimeras. The PDGFRA::USP8 protein is located on the cell membrane and functions as a chimeric ubiquitinyl hydrolase, activated by PDGFs. Conversely, USP8::PDGFRA is a nuclear protein regulating metabolic processes.

Single-cell approaches define two groups of mammalian oligodendrocyte precursor cells and their evolution over developmental time.

Here, we used single-cell RNA sequencing (scRNA-seq), single-cell ATAC sequencing (scATAC-seq), and single-cell spatial transcriptomics to characterize murine cortical OPCs throughout postnatal life. During development, we identified two groups of differentially localized PDGFRα+ OPCs that are transcriptionally and epigenetically distinct. One group (active, or actOPCs) is metabolically active and enriched in white matter. The second (homeostatic, or hOPCs) is less active, enriched in gray matter, and predicted to derive from actOPCs. In adulthood, these two groups are transcriptionally but not epigenetically distinct, and relative to developing OPCs are less active metabolically and have less open chromatin. When adult oligodendrogenesis is enhanced during experimentally induced remyelination, adult OPCs do not reacquire a developmental open chromatin state, and the oligodendrogenesis trajectory is distinct from that seen neonatally. These data suggest that there are two OPC groups subserving distinct postnatal functions and that neonatal and adult OPC-mediated oligodendrogenesis are fundamentally different.

Association between Platelet-Derived Growth Factor Receptor Alpha Gene Polymorphisms and Platelet-Rich Plasma's Efficiency in Treating Lateral Elbow Tendinopathy-A Prospective Cohort Study.

Individual differences in the response to platelet-rich plasma (PRP) therapy can be observed among patients. The genetic background may be the cause of this variability. The current study focused on the impact of genetic variants on the effectiveness of PRP. The aim of the present study was to analyze the impact of single nucleotide polymorphisms (SNP) of the platelet-derived growth factor receptor alpha (PDGFRA) gene on the effectiveness of treating lateral elbow tendinopathy (LET) with PRP. The treatment's efficacy was analyzed over time (2, 4, 8, 12, 24, 52 and 104 weeks after the PRP injection) on 107 patients using patient-reported outcome measures (PROM) and achievement of a minimal clinically important difference (MCID). Four SNPs of the PDGFRA gene (rs7668190, rs6554164, rs869978 and rs1316926) were genotyped using the TaqMan assay method. Patients with the AA genotypes of the rs7668190 and the rs1316926 polymorphisms, as well as carriers of the T allele of rs6554164 showed greater effectiveness of PRP therapy than carriers of other genotypes. Moreover, the studied SNPs influenced the platelets' parameters both in whole blood and in PRP. These results showed that PDGFRA gene polymorphisms affect the effectiveness of PRP treatment. Genotyping the rs6554164 and the rs1316926 SNPs may be considered for use in individualized patient selection for PRP therapy.

PDGFRα+ITGA11+ fibroblasts foster early-stage cancer lymphovascular invasion and lymphatic metastasis via ITGA11-SELE interplay.

Cancer-associated fibroblasts (CAFs) exhibit considerable heterogeneity in advanced cancers; however, the functional annotation and mechanism of CAFs in early-stage cancers remain elusive. Utilizing single-cell RNA sequencing and spatial transcriptomic, we identify a previously unknown PDGFRα+ITGA11+ CAF subset in early-stage bladder cancer (BCa). Multicenter clinical analysis of a 910-case cohort confirms that PDGFRα+ITGA11+ CAFs are associated with lymphovascular invasion (LVI) and poor prognosis in early-stage BCa. These CAFs facilitate LVI and lymph node (LN) metastasis in early-stage BCa, as evidenced in a PDGFRα+ITGA11+ CAFs-specific deficient mouse model. Mechanistically, PDGFRα+ITGA11+ CAFs promote lymphangiogenesis via recognizing ITGA11 surface receptor SELE on lymphatic endothelial cells to activate SRC-p-VEGFR3-MAPK pathway. Further, CHI3L1 from PDGFRα+ITGA11+ CAFs aligns the surrounding matrix to assist cancer cell intravasation, fostering early-stage BCa LVI and LN metastasis. Collectively, our study reveals the crucial role of PDGFRα+ITGA11+ CAFs in shaping metastatic landscape, informing the treatment of early-stage BCa LVI.

Cellular and molecular characteristics of stromal Lkb1 deficiency-induced gastrointestinal polyposis based on single-cell RNA sequencing.

Liver kinase B1 (Lkb1), encoded by serine/threonine kinase (Stk11), is a serine/threonine kinase and tumor suppressor that is strongly implicated in Peutz-Jeghers syndrome (PJS). Numerous studies have shown that mesenchymal-specific Lkb1 is sufficient for the development of PJS-like polyps in mice. However, the cellular origin and components of these Lkb1-associated polyps and underlying mechanisms remain elusive. In this study, we generated tamoxifen-inducible Lkb1flox/flox;Myh11-Cre/ERT2 and Lkb1flox/flox;PDGFRα-Cre/ERT2 mice, performed single-cell RNA sequencing (scRNA-seq) and imaging-based lineage tracing, and aimed to investigate the cellular complexity of gastrointestinal polyps associated with PJS. We found that Lkb1flox/+;Myh11-Cre/ERT2 mice developed gastrointestinal polyps starting at 9 months after tamoxifen treatment. scRNA-seq revealed aberrant stem cell-like characteristics of epithelial cells from polyp tissues of Lkb1flox/+;Myh11-Cre/ERT2 mice. The Lkb1-associated polyps were further characterized by a branching smooth muscle core, abundant extracellular matrix deposition, and high immune cell infiltration. In addition, the Spp1-Cd44 or Spp1-Itga8/Itgb1 axes were identified as important interactions among epithelial, mesenchymal, and immune compartments in Lkb1-associated polyps. These characteristics of gastrointestinal polyps were also demonstrated in another mouse model, tamoxifen-inducible Lkb1flox/flox;PDGFRα-Cre/ERT2 mice, which developed obvious gastrointestinal polyps as early as 2-3 months after tamoxifen treatment. Our findings further confirm the critical role of mesenchymal Lkb1/Stk11 in gastrointestinal polyposis and provide novel insight into the cellular complexity of Lkb1-associated polyp biology. © 2024 The Pathological Society of Great Britain and Ireland.

Sex-determining region Y gene promotes liver fibrosis and accounts for sexual dimorphism in its pathophysiology.

BACKGROUND & AIMS: Men are more prone to develop and die from liver fibrosis than women. In this study, we aim to investigate how sex-determining region Y gene (SRY) in hepatocytes promotes liver fibrosis. METHODS: Hepatocyte-specific Sry knock-in (KI), Sry knockout (KO), and Sry KI with platelet-derived growth factor receptor α (Pdgfrα) KO mice were generated. Liver fibrosis was induced in mice by bile duct ligation for 2 weeks or carbon tetrachloride treatment for 6 weeks. In addition, primary hepatocytes, hepatic stellate cells (HSCs), and immortalized cell lines were used for in vitro studies and mechanistic investigation. RESULTS: Compared to females, the severity of toxin- or cholestasis-induced liver fibrosis is similarly increased in castrated and uncastrated male mice. Among all Y chromosome-encoded genes, SRY was the most significantly upregulated and consistently increased gene in fibrotic/cirrhotic livers in male patients and in mouse models. Sry KI mice developed exacerbated liver fibrosis, whereas Sry KO mice had alleviated liver fibrosis, compared to age- and sex-matched control mice after bile duct ligation or administration of carbon tetrachloride. Mechanistically, both our in vivo and in vitro studies illustrated that SRY in hepatocytes can transcriptionally regulate Pdgfrα expression, and promote HMGB1 (high mobility group box 1) release and subsequent HSC activation. Pdgfrα KO or treatment with the SRY inhibitor DAX1 in Sry KI mice abolished SRY-induced HMGB1 secretion and liver fibrosis. CONCLUSIONS: SRY is a strong pro-fibrotic factor and accounts for the sex disparity observed in liver fibrosis, suggesting its critical role as a potentially sex-specific therapeutic target for prevention and treatment of the disease. IMPACT AND IMPLICATION: We identified that a male-specific gene, sex-determining region Y gene (SRY), is a strong pro-fibrotic gene that accounts for the sex disparity observed in liver fibrosis. As such, SRY might be an appropriate target for surveillance and treatment of liver fibrosis in a sex-specific manner. Additionally, SRY might be a key player in the sexual dimorphism observed in hepatic pathophysiology more generally.

Genomic profiling in GIST: Implications in clinical outcome and future challenges.

Gastrointestinal Stromal Tumors (GIST) are the most frequent mesenchymal neoplasia of the digestive tract. Genomic alterations in KIT, PDFGRA, SDH, and BRAF genes are essential in GIST oncogenesis. Therefore, the mutations in these genes have demonstrated clinical implications. Tumors with deletions in KIT-exon 11 or duplications in exon 9 are associated with a worse prognosis. In contrast, KIT-exon 11 substitutions and duplications are associated with a better clinical outcome. Moreover, mutations in Kit exon 9 and 11 are actionable, due to their response to imatinib, while mutations in PDGFRA respond to sunitinib and/or avapritinib. Although, molecular testing on tissue samples is effective; it is invasive, requires adequate amounts of tissue, and a long experimental process is needed for results. In contrast, liquid biopsy has been proposed as a simple and non-invasive method to test biomarkers in cancer. The most common molecule analyzed by liquid biopsy is circulating tumor DNA (ctDNA). GISTs ctDNA testing has been demonstrated to be effective in identifying known and novel KIT mutations that were not detected using traditional tissue DNA testing and have been useful in determining progression risk and response to TKI therapy. This allows the clinician to have an accurate picture of the genetic changes of the tumor over time. In this work, we aimed to discuss the implications of mutational testing in clinical outcomes, the methods to test ctDNA and the future challenges in the establishment of alternatives of personalized medicine.

Avapritinib-based SAR studies unveil a binding pocket in KIT and PDGFRA.

Avapritinib is the only potent and selective inhibitor approved for the treatment of D842V-mutant gastrointestinal stromal tumors (GIST), the most common primary mutation of the platelet-derived growth factor receptor α (PDGFRA). The approval was based on the NAVIGATOR trial, which revealed overall response rates of more than 90%. Despite this transformational activity, patients eventually progress, mostly due to acquired resistance mutations or following discontinuation due to neuro-cognitive side effects. These patients have no therapeutic alternative and face a dismal prognosis. Notable, little is known about this drug's binding mode and its medicinal chemistry development, which is instrumental for the development of the next generation of drugs. Against this background, we solve the crystal structures of avapritinib in complex with wild-type and mutant PDGFRA and stem cell factor receptor (KIT), which provide evidence and understanding of inhibitor binding and lead to the identification of a sub-pocket (Gα-pocket). We utilize this information to design, synthesize and characterize avapritinib derivatives for the determination of key pharmacophoric features to overcome drug resistance and limit potential blood-brain barrier penetration.

[Reappraisals of biological behaviors of PDGFRA mutant gastrointestinal stromal tumor].

Objective: To investigate the biological behavior spectrum of platelet-derived growth factor alpha receptor (PDGFRA)-mutant gastrointestinal stromal tumor (GIST), and to compare the clinical values of the Zhongshan method of benign and malignant evaluation with the modified National Institutes of Health (NIH) risk stratification. Methods: A total of 119 cases of GIST with PDGFRA mutation who underwent surgical resection at Zhongshan Hospital, Fudan University from 2009 to 2020 were collected. The clinicopathological data, follow-up records, and subsequent treatment were reviewed and analyzed statistically. Results: There were 79 males and 40 females. The patients ranged in age from 25 to 80 years, with a median age of 60 years. Among them, 115 patients were followed up for 1-154 months, and 13 patients progressed to disease. The 5-year disease-free survival (DFS) and overall survival (OS) were 90.1% and 94.1%, respectively. According to the modified NIH risk stratification, 8 cases, 32 cases, 38 cases, and 35 cases were very-low risk, low risk, intermediate risk, and high risk, and 5-year DFS were 100.0%, 95.6%, 94.3%, and 80.5%, respectively. There was no significant difference in prognosis among the non-high risk groups, only the difference between high risk and non-high risk groups was significant (P=0.029). However, the 5-year OS was 100.0%, 100.0%, 95.0% and 89.0%, and there was no difference (P=0.221). According to the benign and malignant evaluation Zhongshan method, 43 cases were non-malignant (37.4%), 56 cases were low-grade malignant (48.7%), 9 cases were moderately malignant (7.8%), and 7 cases were highly malignant (6.1%). The 5-year DFS were 100.0%, 91.7%, 77.8%, 38.1%, and the difference was significant (P<0.001). The 5-year OS were 100.0%, 97.5%, 77.8%, 66.7%, the difference was significant (P<0.001). Conclusions: GIST with PDGFRA gene mutation shows a broad range of biological behavior, ranging from benign to highly malignant. According to the Zhongshan method, non-malignant and low-grade malignant tumors are common, the prognosis after surgery is good, while the fewer medium-high malignant tumors showed poor prognosis after surgical resection. The overall biological behavior of this type of GIST is relatively inert, which is due to the low proportion of medium-high malignant GIST. The modified NIH risk stratification may not be effective in risk stratification for PDGFRA mutant GIST.

Molecular and clinicopathological features of KIT/PDGFRA wild-type gastrointestinal stromal tumors.

Approximately 10% of gastrointestinal stromal tumors (GISTs) harbor reportedly no KIT and PDGFRA mutations (wild-type GISTs). The clinicopathological features and oncologic outcomes of wild-type GISTs based on molecular profiles are unknown. We recruited 35 wild-type GIST patients from the two registry studies of high-risk GISTs between 2012 and 2015 and primary GISTs between 2003 and 2014. Molecular profiling of wild-type GISTs was performed by targeted next-generation sequencing (NGS) using formalin-fixed paraffin-embedded tumor samples. Among 35 wild-type GISTs, targeted NGS analysis detected NF1, SDH, or BRAF mutation: 16 NF1-GISTs with various NF1 mutations, 12 SDH-GISTs (4 with SDHA mutations, 4 with SDHB mutations, and 4 with SDHB-negative staining), and 5 BRAF-GISTs with the V600E mutation. Two GISTs showed no mutations based on our targeted NGS analysis. Additional gene mutations were infrequent in primary wild-type GISTs and found in TP53, CREBBP, CDKN2A, and CHEK2. Most NF1-GISTs were located in the small intestine (N = 12; 75%) and showed spindle cell features (N = 15; 94%) and multiple tumors (N = 6, 38%) with modest proliferation activities. In contrast, SDH-GISTs were predominantly found in the stomach (N = 11; 92%), exhibiting epithelioid cell (N = 6; 50%) and multiple (N = 6, 50%) features. The overall survival of patients with SDH-GISTs appeared to be better than that of BRAF-GISTs (p = 0.0107) or NF1-GISTs (p = 0.0754), respectively. In conclusion, major molecular changes in wild-type GISTs include NF1, SDH, and BRAF. NF1-GISTs involved multifocal spindle cell tumors in the small intestine. SDH-GISTs occurred in young patients and were multifocal in the stomach and clinically indolent.

Clinical Benefit of Avapritinib in KIT-Mutant Gastrointestinal Stromal Tumors: A Post Hoc Analysis of the Phase I NAVIGATOR and Phase I/II CS3007-001 Studies.

PURPOSE: The efficacy of the selective KIT/PDGFRA inhibitor avapritinib (300 mg once daily) was explored in patients with non-PDGFRA-mutant gastrointestinal stromal tumors (GISTs) from the phase I NAVIGATOR and phase I/II CS3007-001 trials. PATIENTS AND METHODS: Adults with unresectable/metastatic, KIT-only-mutant GISTs and progression following ≥1 tyrosine kinase inhibitors (TKIs) were included in this post hoc analysis. Baseline mutational status was identified in tumor and plasma. Primary endpoints were objective response rate (ORR) and progression-free survival (PFS) by blinded independent radiology review per modified RECIST v1.1 in patients harboring KIT activation-loop mutations (KIT exons 17 or 18) without ATP binding-pocket mutations (KIT exons 13 or 14; ALposABPneg), and other KIT mutations (OTHERS). RESULTS: Sixty KIT ALposABPneg and 100 KIT OTHERS predominantly heavily pretreated patients (61.3% with ≥3 prior TKIs) were included. ORR was significantly higher in KIT ALposABPneg than KIT OTHERS patients (unadjusted: 26.7% vs. 12.0%; P = 0.0852; adjusted: 31.4% vs. 12.1%; P = 0.0047). Median PFS (mPFS) was significantly longer in KIT ALposABPneg patients compared with KIT OTHERS patients (unadjusted: 9.1 vs. 3.5 months; P = 0.0002; adjusted: 9.1 vs. 3.4 months; P < 0.0001), and longer in second- versus later-line settings (19.3 vs. 5.6-10.6 months). Benefit with avapritinib was observed in patients with KIT exon 9 mutations in the ≥4 line settings (mPFS: 5.6 and 3.7 months for 4 line and >4 line, respectively). CONCLUSIONS: Avapritinib showed greater antitumor activity in patients with GISTs harboring KIT ALposABPneg mutations versus KIT OTHERS, and may be considered in the former subpopulation. Patients with KIT exon 9 mutations may also benefit in ≥4 line settings.

Reappraisals of biological behaviors of PDGFRA mutant gastrointestinal stromal tumor / 中华病理学杂志

Objective: To investigate the biological behavior spectrum of platelet-derived growth factor alpha receptor (PDGFRA)-mutant gastrointestinal stromal tumor (GIST), and to compare the clinical values of the Zhongshan method of benign and malignant evaluation with the modified National Institutes of Health (NIH) risk stratification. Methods: A total of 119 cases of GIST with PDGFRA mutation who underwent surgical resection at Zhongshan Hospital, Fudan University from 2009 to 2020 were collected. The clinicopathological data, follow-up records, and subsequent treatment were reviewed and analyzed statistically. Results: There were 79 males and 40 females. The patients ranged in age from 25 to 80 years, with a median age of 60 years. Among them, 115 patients were followed up for 1-154 months, and 13 patients progressed to disease. The 5-year disease-free survival (DFS) and overall survival (OS) were 90.1% and 94.1%, respectively. According to the modified NIH risk stratification, 8 cases, 32 cases, 38 cases, and 35 cases were very-low risk, low risk, intermediate risk, and high risk, and 5-year DFS were 100.0%, 95.6%, 94.3%, and 80.5%, respectively. There was no significant difference in prognosis among the non-high risk groups, only the difference between high risk and non-high risk groups was significant (P=0.029). However, the 5-year OS was 100.0%, 100.0%, 95.0% and 89.0%, and there was no difference (P=0.221). According to the benign and malignant evaluation Zhongshan method, 43 cases were non-malignant (37.4%), 56 cases were low-grade malignant (48.7%), 9 cases were moderately malignant (7.8%), and 7 cases were highly malignant (6.1%). The 5-year DFS were 100.0%, 91.7%, 77.8%, 38.1%, and the difference was significant (P<0.001). The 5-year OS were 100.0%, 97.5%, 77.8%, 66.7%, the difference was significant (P<0.001). Conclusions: GIST with PDGFRA gene mutation shows a broad range of biological behavior, ranging from benign to highly malignant. According to the Zhongshan method, non-malignant and low-grade malignant tumors are common, the prognosis after surgery is good, while the fewer medium-high malignant tumors showed poor prognosis after surgical resection. The overall biological behavior of this type of GIST is relatively inert, which is due to the low proportion of medium-high malignant GIST. The modified NIH risk stratification may not be effective in risk stratification for PDGFRA mutant GIST.

PDGFRß Activation Induced the Bovine Embryonic Genome Activation via Enhanced NFYA Nuclear Localization.

Embryonic genome activation (EGA) is a critical step during embryonic development. Several transcription factors have been identified that play major roles in initiating EGA; however, this gradual and complex mechanism still needs to be explored. In this study, we investigated the role of nuclear transcription factor Y subunit A (NFYA) in bovine EGA and bovine embryonic development and its relationship with the platelet-derived growth factor receptor-ß (PDGFRß) by using a potent selective activator (PDGF-BB) and inhibitor (CP-673451) of PDGF receptors. Activation and inhibition of PDGFRß using PDGF-BB and CP-673451 revealed that NFYA expression is significantly (p < 0.05) affected by the PDGFRß. In addition, PDGFRß mRNA expression was significantly increased (p < 0.05) in the activator group and significantly decreased (p < 0.05) in the inhibitor group when compared with PDGFRα. Downregulation of NFYA following PDGFRß inhibition was associated with the expression of critical EGA-related genes, bovine embryo development rate, and implantation potential. Moreover, ROS and mitochondrial apoptosis levels and expression of pluripotency-related markers necessary for inner cell mass development were also significantly (p < 0.05) affected by the downregulation of NFYA while interrupting trophoblast cell (CDX2) differentiation. In conclusion, the PDGFRß-NFYA axis is critical for bovine embryonic genome activation and embryonic development.

Dipeptidyl Peptidase 4 Stimulation Induces Adipogenesis-Related Gene Expression of Adipose Stromal Cells.

Adipogenesis has emerged as a new therapeutic target for regulating metabolism and achieving anti-inflammatory and anti-atherosclerotic effects via the release of adiponectin. However, at present, the effects and mechanism of action of dipeptidyl peptidase 4 (DPP4) stimulation on adiponectin production and adipogenesis have not been clarified. Here, we investigated the effects of DPP4 stimulation with monocyte chemoattractant protein-1 (MCP-1) on platelet-derived growth factor receptor alpha (PDGFRα) expression in adipose tissue and blood adiponectin levels. Stromal vascular fractions (SVFs) purified from human subcutaneous adipose tissue and inguinal adipose tissue of obese and diabetic (Leprdb/db) mice were treated with 50 ng of MCP-1 and plasma from control (Lepr+/+) mice supplemented with 10 ng or 50 ng of MCP-1. Treatment of SVFs from human subcutaneous adipose tissues with 50 ng of MCP-1 significantly increased AdipoQ, DPP4, peroxisome proliferator-activated receptor gamma (PPARγ), fatty-acid-binding protein (FABP4), and SERBF1 mRNA expression. MCP-1-supplemented plasma increased adiponectin, CCAAT-Enhancer-binding protein alpha (C/EBPα), DPP4, IL-33, and PDGFRα mRNA expression and adiponectin and DPP4 protein expression, while decreasing the expression of IL-10 mRNA in SVFs compared with the levels in the plasma treatment group. MCP-1-supplemented plasma was shown to increase PPARγ, PPARγ2, adiponectin, DPP4, and FABP4 and decrease IL-10 mRNA expression in PDGFRα cells from adipose tissue. Meanwhile, MCP-1-supplemented plasma increased MCP-1, PDGFRα, TNFα, adiponectin, and IL-1ß and decreased IL-10 and FOXP3 mRNA expression in DPP4 cells. Moreover, the injection of MCP-1-supplemented plasma into adipose tissue increased the proportion of DPP4+ cells among PDGFRα+ cells from adipose tissue and plasma adiponectin levels of Leprdb/db mice compared with the levels in the plasma injection group. Our results demonstrate that DPP4+ cells are important adipose progenitor cells. Stimulation of DPP4 with MCP-1 increases adipogenesis-related gene expression and the population of DPP4+ cells among PDGFRα+ cells in SVFs and blood adiponectin levels. DPP4 stimulation could be a novel therapy to increase local adipogenesis and systemic adiponectin levels.

Genetic alterations that deregulate RB and PDGFRA signaling pathways drive tumor progression in IDH2-mutant astrocytoma.

In IDH-mutant astrocytoma, IDH2 mutation is quite rare and biological mechanisms underlying tumor progression in IDH2-mutant astrocytoma remain elusive. Here, we report a unique case of IDH2 mutant astrocytoma, CNS WHO grade 3 that developed tumor progression. We performed a comprehensive genomic and epigenomic analysis for primary and recurrent tumors and found that both tumors harbored recurrent IDH2R172K and TP53R248W mutation with CDKN2A/B hemizygous deletion. We also found amplifications of CDK4 and MDM2 with PDGFRA gain in the recurrent tumor and upregulated protein expressions of these genes. We further developed, for the first time, a xenograft mouse model of IDH2R172K and TP53R248W mutant astrocytoma from the recurrent tumor, but not from the primary tumor. Consistent with parent recurrent tumor cells, amplifications of CDK4 and MDM2 and PDGFRA gain were found, while CDKN2A/B was identified as homozygous deletion in the xenografts, qualifying for integrated diagnosis of astrocytoma, IDH2-mutant, CNS WHO grade 4. Cell viability assay found that CDK4/6 inhibitor and PDGFR inhibitor potently decreased cell viability in recurrent tumor cells, as compared to primary tumor cells. These findings suggest that gene alterations that activate retinoblastoma (RB) signaling pathways and PDGFR may drive tumor progression and xenograft formation in IDH2-mutant astrocytoma, which is equivalent to progressive IDH1-mutant astrocytoma. Also, our findings suggest that these genomic alterations may represent therapeutic targets in IDH2-mutant astrocytoma.