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SHP2 phosphatase promotes full activation of the RTK-dependent Ras/MAPK pathway. Its mutations can drive cancer and RASopathies, a group of neurodevelopmental disorders (NDDs). Here we ask how same residue mutations in SHP2 can lead to both cancer and NDD phenotypes, and whether we can predict what the outcome will be. We collected and analyzed mutation data from the literature and cancer databases and performed molecular dynamics simulations of SHP2 mutants. We show that both cancer and Noonan syndrome (NS, a RASopathy) mutations favor catalysis-prone conformations. As to cancer versus RASopathies, we demonstrate that cancer mutations are more likely to accelerate SHP2 activation than the NS mutations at the same genomic loci, in line with NMR data for K-Ras4B more aggressive mutations. The compiled experimental data and dynamic features of SHP2 mutants lead us to propose that different from strong oncogenic mutations, SHP2 activation by NS mutations is less likely to induce a transition of the ensemble from the SHP2 inactive state to the active state. Strong signaling promotes cell proliferation, a hallmark of cancer. Weak, or moderate signals are associated with differentiation. In embryonic neural cells, dysregulated differentiation is connected to NDDs. Our innovative work offers structural guidelines for identifying and correlating mutations with clinical outcomes, and an explanation for why bearers of RASopathy mutations may have a higher probability of cancer. Finally, we propose a drug strategy against SHP2 variants-promoting cancer and RASopathies.

Psoriasis is a complex chronic inflammatory skin disease with unclear molecular mechanisms. We found that the Src homology‐2 domain‐containing protein tyrosine phosphatase‐2 (SHP2) was highly expressed in both psoriatic patients and imiquimod (IMQ)‐induced psoriasis‐like mice. Also, the SHP2 allosteric inhibitor SHP099 reduced pro‐inflammatory cytokine expression in PBMCs taken from psoriatic patients. Consistently, SHP099 significantly ameliorated IMQ‐triggered skin inflammation in mice. Single‐cell RNA sequencing of murine skin demonstrated that SHP2 inhibition impaired skin inflammation in myeloid cells, especially macrophages. Furthermore, IMQ‐induced psoriasis‐like skin inflammation was significantly alleviated in myeloid cells (monocytes, mature macrophages, and granulocytes)—but not dendritic cells conditional SHP2 knockout mice. Mechanistically, SHP2 promoted the trafficking of toll‐like receptor 7 (TLR7) from the Golgi to the endosome in macrophages by dephosphorylating TLR7 at Tyr1024, boosting the ubiquitination of TLR7 and NF‐ κ B‐mediated skin inflammation. Importantly, Tlr7 point‐mutant knock‐in mice showed an attenuated psoriasis‐like phenotype compared to wild‐type littermates following IMQ treatment. Collectively, our findings identify SHP2 as a novel regulator of psoriasis and suggest that SHP2 inhibition may be a promising therapeutic approach for psoriatic patients. Synopsis From the clinical level of psoriasis patients, the whole animal level, and the cellular level, this study reveals that SHP2 promotes TLR7 trafficking to endosomes and activates the downstream NF‐κB pathway through dephosphorylation of TLR7, thus exacerbating the pathogenesis of psoriasis and providing a potential target for the development of therapeutic drugs for psoriasis. SHP2 expression is increased in both human psoriatic patients and IMQ‐induced psoriasis‐like mice. Treatment with SHP2 inhibitor alleviated IMQ‐induced and IL‐23‐induced psoriasis‐like skin inflammation. SHP2 deficiency in macrophages attenuates IMQ‐induced skin inflammation in mice. SHP2 promotes TLR7 trafficking to endosomes in a phosphatase‐dependent manner. Psoriasis‐like skin inflammation is reduced in Tlr7‐Y1025D point mutant mice. Graphical Abstract From the clinical level of psoriasis patients, the whole animal level, and the cellular level, this study reveals that SHP2 promotes TLR7 trafficking to endosomes and activates the downstream NF‐κB pathway through dephosphorylation of TLR7, thus exacerbating the pathogenesis of psoriasis and providing a potential target for the development of therapeutic drugs for psoriasis.

Individuals with Noonan syndrome (NS) are predisposed to hematologic cancers, solid tumors, and low-grade gliomas. We report an 8-year-old girl originally referred at age 14 months for short stature, developmental delay, and failure to thrive who was subsequently found to have pathogenetic variants both in MECP2 and PTPN11. Family history included a maternal half-sister with NS and a mother carrying the PTPN11 mutation. Familial single-gene testing showed a heterozygous pathogenic variant in PTPN11 (c.417G > C p.Glu139Asp) suggesting NS, prompting initiation of growth hormone (GH) treatment at 26 months. Due to associated language delays, gross motor delays, microcephaly, and seizures, exome sequencing (ES) was pursued. ES identified a heterozygous de novo pathogenic variant (c.763C > T p.Arg255Ter) in MECP2 and led to the additional diagnosis of Rett syndrome (RTT). Seizure onset prompted neuroimaging, which demonstrated hydrocephalus due to aqueductal stenosis secondary to a tectal neoplasm. GH treatment was discontinued. The co-occurrence of NS and RTT is rare. ES enabled the additional diagnosis of RTT in our patient with NS, who presented with atypical features and developmental regression.

Germline mutations of NF1 cause neurofibromatosis type 1 (NF1) through the activation of the RAS signaling pathway, and some NF1 patients develop malignant peripheral nerve sheath tumors (MPNSTs). Here, we established subclones of the human NF1-MPNST cell line sNF96.2 that manifest increased tumorigenic activity and increased phosphorylation of the protein kinases MEK and Akt relative to the parental cells. Genomic DNA sequencing identified 14 additional heterozygous mutations within the coding regions of 13 cancer- and other disease-related genes in these subclones. One of these genes, PTPN11, encodes SHP-2, and the forced expression of the identified G503V mutant of SHP-2 increased both tumorigenic activity and MEK phosphorylation in parental sNF96.2 cells, suggesting that the combination of PTPN11 and NF1 mutations induces the pathological activation of the RAS pathway. These effects of SHP-2 (G503V) were inhibited by the coexpression of the G370A mutant of BRAP, which was also detected in the highly malignant subclones, and this inhibition was accompanied by the calpain-dependent cleavage of SHP-2 (G503V). The cleavage of SHP-2 (G503V) and suppression of MEK phosphorylation mediated by BRAP (G370A) were not detected in NF1-intact (HeLa) cells. Tumor promotion by SHP-2 (G503V) and its suppression by BRAP (G370A) may serve as a basis for the development of new treatment strategies for NF1.

Noonan syndrome (NS) is a relatively common genetic disorder, characterized by typical facies, short stature, developmental delay, and cardiac abnormalities. Known causative genes account for 70–80% of clinically diagnosed NS patients, but the genetic basis for the remaining 20–30% of cases is unknown. We performed next-generation sequencing on germ-line DNA from 27 NS patients lacking a mutation in the known NS genes. We identified gain-of-function alleles in Ras-like without CAAX 1 (RIT1) and mitogen-activated protein kinase kinase 1 (MAP2K1) and previously unseen loss-of-function variants in RAS p21 protein activator 2 (RASA2) that are likely to cause NS in these patients. Expression of the mutant RASA2 , MAP2K1 , or RIT1 alleles in heterologous cells increased RAS-ERK pathway activation, supporting a causative role in NS pathogenesis. Two patients had more than one disease-associated variant. Moreover, the diagnosis of an individual initially thought to have NS was revised to neurofibromatosis type 1 based on an NF1 nonsense mutation detected in this patient. Another patient harbored a missense mutation in NF1 that resulted in decreased protein stability and impaired ability to suppress RAS-ERK activation; however, this patient continues to exhibit a NS-like phenotype. In addition, a nonsense mutation in RPS6KA3 was found in one patient initially diagnosed with NS whose diagnosis was later revised to Coffin–Lowry syndrome. Finally, we identified other potential candidates for new NS genes, as well as potential carrier alleles for unrelated syndromes. Taken together, our data suggest that next-generation sequencing can provide a useful adjunct to RASopathy diagnosis and emphasize that the standard clinical categories for RASopathies might not be adequate to describe all patients.

The platelet receptors, glycoprotein VI (GPVI) and integrin α2β1 jointly control collagen-dependent thrombus formation via protein tyrosine kinases. It is unresolved to which extent the ITIM (immunoreceptor tyrosine-based inhibitory motif) receptor PECAM1 and its downstream acting protein tyrosine phosphatase PTPN11 interfere in this process. Here, we hypothesized that integrin α2β1 has a co-regulatory role in the PECAM1- and PTPN11-dependent restraint of thrombus formation. We investigated platelet activation under flow on collagens with a different GPVI dependency and using integrin α2β1 blockage. Blood was obtained from healthy subjects and from patients with Noonan syndrome with a gain-of-function mutation of PTPN11 and variable bleeding phenotype. On collagens with decreasing GPVI activity (types I, III, IV), the surface-dependent inhibition of PECAM1 did not alter thrombus parameters using control blood. Blockage of α2β1 generally reduced thrombus parameters, most effectively on collagen IV. Strikingly, simultaneous inhibition of PECAM1 and α2β1 led to a restoration of thrombus formation, indicating that the suppressing signaling effect of PECAM1 is masked by the platelet-adhesive receptor α2β1. Blood from 4 out of 6 Noonan patients showed subnormal thrombus formation on collagen IV. In these patients, effects of α2β1 blockage were counterbalanced by PECAM1 inhibition to a normal phenotype. In summary, we conclude that the suppression of GPVI-dependent thrombus formation by either PECAM1 or a gain-of-function of PTPN11 can be overruled by α2β1 engagement.

Psoriasis is a chronic inflammatory skin disease, often accompanied by increased infiltration of immune cells, especially neutrophils. However, the detailed mechanism of the neutrophil function in psoriasis progression remains unclear. Here, we found that both Src homology‐2 domain‐containing protein tyrosine phosphatase‐2 (SHP2) and neutrophils were highly correlated to developing psoriasis by single‐cell ribonucleic acid (RNA) sequencing and experiment verification. The deficiency of SHP2 in neutrophils significantly alleviated psoriasis‐like phenotype in an imiquimod‐induced murine model. Interestingly, high levels of neutrophil extracellular traps (NETs) were produced in the inflamed lesions of psoriatic patients. In addition, imiquimod‐induced psoriasis‐like symptoms were remarkably ameliorated in peptidyl arginine deiminase 4 (PAD4) knockout mice, which cannot form NETs. Mechanistically, RNA‐seq analysis revealed that SHP2 promoted the formation of NETs in neutrophils via the ERK5 pathway. Functionally, this mechanism resulted in the infiltration of pro‐inflammatory cytokines such as TNF‐α, IL‐1β, IL‐6, IL‐17A, and CXCL‐15, which enhances the inflammatory response in skin lesions and reinforces the cross‐talk between neutrophils and keratinocytes, ultimately aggravating psoriasis. Our findings uncover a role for SHP2 in NET release and subsequent cell death known as NETosis in the progression of psoriasis and suggest that SHP2 may be a promising therapeutic target for psoriasis. Single‐cell RNA sequencing and experimental verification were combined to announce that SHP2 aggravates psoriasis‐like skin inflammation in mice via ERK5‐dependent NETosis. During this process, inflammatory cytokines: TNF‐α, IL‐1β, IL‐6, IL‐17a, and CXCL‐15 were infiltrated in skin and contributed to psoriasis. Our study provides evidence for the role of SHP2 in NETosis and psoriasis. SHP2 may be a potential therapeutic target for the treatment of psoriasis.

The RASopathies are a group of genetic rare diseases caused by mutations affecting genes involved in the RAS/MAPK (RAS–mitogen activated protein kinase) pathway. Among them, PTPN11 pathogenic variants are responsible for approximately 50% of Noonan syndrome (NS) cases and, albeit to a lesser extent, of Leopard syndrome (LPRD1), which present a few overlapping clinical features, such as facial dysmorphism, developmental delay, cardiac defects, and skeletal deformities. Motor impairment and decreased muscle strength have been recently reported. The etiology of the muscle involvement in these disorders is still not clear but probably multifactorial, considering the role of the RAS/MAPK pathway in skeletal muscle development and Acetylcholine Receptors (AChR) clustering at the neuromuscular junction (NMJ). We report, herein, four unrelated children carrying three different heterozygous mutations in the PTPN11 gene. Intriguingly, their phenotypic features first led to a clinical suspicion of congenital myasthenic syndrome (CMS), due to exercise-induced fatigability with a variable degree of muscle weakness, and serum proteomic profiling compatible with a NMJ defect. Moreover, muscle fatigue improved after treatment with CMS-specific medication. Although the link between PTPN11 gene and neuromuscular transmission is unconfirmed, an increasing number of patients with RASopathies are affected by muscle weakness and fatigability. Hence, NS or LPDR1 should be considered in children with suspected CMS but negative genetic workup for known CMS genes or additional symptoms indicative of NS, such as facial dysmorphism or intellectual disability.

Characterization of spermatogonial stem cells (SSCs) has been hampered by their low frequency and lack of features that distinguish them from committed spermatogonia. Few conserved SSC markers have been discovered. To identify a new SSC marker, we evaluated SIRPA expression in mouse and rat SSCs. SIRPA was expressed in a small population of undifferentiated spermatogonia. SIRPA, and its ligand CD47 were expressed in cultured SSCs. Expression of both SIRPA and CD47 was upregulated by supplementation of GDNF and FGF2, which promoted SSC self-renewal. Sirpa depletion by short hairpin RNA impaired the proliferation of cultured SSCs, and these cells showed decreased MAP2K1 activation and PTPN11 phosphorylation. Immunoprecipitation experiments showed that SIRPA associates with PTPN11. Ptpn11 depletion impaired SSC activity in a manner similar to Sirpa depletion. SIRPA was expressed in undifferentiated spermatogonia in rat and monkey testes. Xenogenic transplantation experiments demonstrated that SIRPA is expressed in rat SSCs. These results suggest that SIRPA is a conserved SSC marker that promotes SSC self-renewal division by activating the MAP2K1 pathway via PTPN11. Summary Sentence SIRPA is expressed on mouse and rat SSCs.

BRAF V600 mutations have been found in 1–2% of non-small-cell lung cancer (NSCLC) patients, with Food and Drug Administration (FDA) approved treatment of dabrafenib plus trametinib and progression free survival (PFS) of 10.9 months. However, 50–80% of BRAF mutations in lung cancer are non-V600, and can be class II, with intermediate to high kinase activity and RAS independence, or class III, with impaired kinase activity, upstream signaling dependence, and consequently, sensitivity to receptor tyrosine kinase (RTK) inhibitors. Plasma cell-free DNA (cfDNA) of 185 newly diagnosed advanced lung adenocarcinoma patients (Spanish Lung Liquid versus Invasive Biopsy Program, SLLIP, NCT03248089) was examined for BRAF and other alterations with a targeted cfDNA next-generation sequencing (NGS) assay (Guardant360®, Guardant Health Inc., CA, USA), and results were correlated with patient outcome. Cell viability with single or combined RAF, MEK, and SHP2 inhibitors was assessed in cell lines with BRAF class I, II, and III mutations. Out of 185 patients, 22 had BRAF alterations (12%) of which seven patients harbored amplifications (32%) and 17 had BRAF mutations (77%). Of the BRAF mutations, four out of 22 (18%) were V600E and 18/22 (82%) were non-V600. In vitro results confirmed sensitivity of class III and resistance of class I and II BRAF mutations, and BRAF wild type cells to SHP2 inhibition. Concomitant MEK or RAF and SHP2 inhibition showed synergistic effects, especially in the class III BRAF-mutant cell line. Our study indicates that the class of the BRAF mutation may have clinical implications and therefore should be defined in the clinical practice and used to guide therapeutic decisions.