Downstream Activation Research Articles

CNK2 promotes cancer cell motility by mediating ARF6 activation downstream of AXL signalling

Cell motility is a critical feature of invasive tumour cells that is governed by complex signal transduction events. Particularly, the underlying mechanisms that bridge extracellular stimuli to the molecular machinery driving motility remain partially understood. Here, we show that the scaffold protein CNK2 promotes cancer cell migration by coupling the pro-metastatic receptor tyrosine kinase AXL to downstream activation of ARF6 GTPase. Mechanistically, AXL signalling induces PI3K-dependent recruitment of CNK2 to the plasma membrane. In turn, CNK2 stimulates ARF6 by associating with cytohesin ARF GEFs and with a novel adaptor protein called SAMD12. ARF6-GTP then controls motile forces by coordinating the respective activation and inhibition of RAC1 and RHOA GTPases. Significantly, genetic ablation of CNK2 or SAMD12 reduces metastasis in a mouse xenograft model. Together, this work identifies CNK2 and its partner SAMD12 as key components of a novel pro-motility pathway in cancer cells, which could be targeted in metastasis.

Involvement of oxidative stress in placental dysfunction, the pathophysiology of fetal death and pregnancy disorders.

Placental oxidative stress contributes to both normal and abnormal placentation during pregnancy. This review discusses the potential consequence of oxidative stress-induced placental dysfunction on pregnancies complicated by fetal death and pregnancies with a high risk of fetal death. The placenta is a source of reactive oxygen free radicals due to the oxidative metabolism required to meet the demands of the growing fetus. The placenta has an array of efficient antioxidant defense systems to deal with rising oxidative stress created by free radicals during pregnancy. Properly controlled physiological (low-level) free radical production is a necessary part of cellular signaling pathways and downstream activities during normal placental development; however, poorly controlled oxidative stress can cause aberrant placentation, immune disturbances and placental dysfunction. Abnormal placental function and immune disturbances are linked to many pregnancy-related disorders, including early and recurrent pregnancy loss, fetal death, spontaneous preterm birth, preeclampsia and fetal growth restriction. This review discusses the role of placental oxidative stress in both normal and pathological settings. Finally, based on previously published work, this review presents multiple lines of evidence for the strong association between oxidative stress and adverse pregnancy outcomes, including fetal death and pregnancies with a high risk of fetal death.

Ubiquitination of GRK2 Is Required for the β-Arrestin-Biased Signaling Pathway of Dopamine D2 Receptors to Activate ERK Kinases.

A class-A GPCR dopamine D2 receptor (D2R) plays a critical role in the proper functioning of neuronal circuits through the downstream activation of both G-protein- and β-arrestin-dependent signaling pathways. Understanding the signaling pathways downstream of D2R is critical for developing effective therapies with which to treat dopamine (DA)-related disorders such as Parkinson's disease and schizophrenia. Extensive studies have focused on the regulation of D2R-mediated extracellular-signal-regulated kinase (ERK) 1/2 signaling; however, the manner in which ERKs are activated upon the stimulation of a specific signaling pathway of D2R remains unclear. The present study conducted a variety of experimental techniques, including loss-of-function experiments, site-directed mutagenesis, and the determination of protein interactions, in order to investigate the mechanisms underlying β-arrestin-biased signaling-pathway-mediated ERK activation. We found that the stimulation of the D2R β-arrestin signaling pathway caused Mdm2, an E3 ubiquitin ligase, to move from the nucleus to the cytoplasm and interact with tyrosine phosphorylated G-protein-coupled receptor kinase 2 (GRK2), which was facilitated by Src, a non-receptor tyrosine kinase. This interaction led to the ubiquitination of GRK2, which then moved to the plasma membrane and interacted with activated D2R, followed by the phosphorylation of D2R as well as the mediation of ERK activation. In conclusion, Mdm2-mediated GRK2 ubiquitination, which is selectively triggered by the stimulation of the D2R β-arrestin signaling pathway, is necessary for GRK2 membrane translocation and its interaction with D2R, which in turn mediates downstream ERK signaling. This study is primarily novel and provides essential information with which to better understand the detailed mechanisms of D2R-dependent signaling.

Phosphorylation of USP20 on Ser334 by IRAK1 promotes IL-1β-evoked signaling in vascular smooth muscle cells and vascular inflammation

Reversible lysine-63 (K63) polyubiquitination regulates proinflammatory signaling in vascular smooth muscle cells (SMCs) and plays an integral role in atherosclerosis. Ubiquitin-specific peptidase 20 (USP20) reduces NFκB activation triggered by proinflammatory stimuli, and USP20 activity attenuates atherosclerosis in mice. The association of USP20 with its substrates triggers deubiquitinase activity; this association is regulated by phosphorylation of USP20 on Ser334 (mouse) or Ser333 (human). USP20 Ser333 phosphorylation was greater in SMCs of atherosclerotic segments of human arteries as compared with nonatherosclerotic segments. To determine whether USP20 Ser334 phosphorylation regulates proinflammatory signaling, we created USP20-S334A mice using CRISPR/Cas9-mediated gene editing. USP20-S334A mice developed ∼50% less neointimal hyperplasia than congenic WT mice after carotid endothelial denudation. WT carotid SMCs showed substantial phosphorylation of USP20 Ser334, and WT carotids demonstrated greater NFκB activation, VCAM-1 expression, and SMC proliferation than USP20-S334A carotids. Concordantly, USP20-S334A primary SMCs invitro proliferated and migrated less than WT SMCs in response to IL-1β. An active site ubiquitin probe bound to USP20-S334A and USP20-WT equivalently, but USP20-S334A associated more avidly with TRAF6 than USP20-WT. IL-1β induced less K63-linked polyubiquitination of TRAF6 and less downstream NFκB activity in USP20-S334A than in WT SMCs. Using invitro phosphorylation with purified IRAK1 and siRNA-mediated gene silencing of IRAK1 in SMCs, we identified IRAK1 as a novel kinase for IL-1β-induced USP20 Ser334 phosphorylation. Our findings reveal novel mechanisms regulating IL-1β-induced proinflammatory signaling: by phosphorylating USP20 Ser334, IRAK1 diminishes the association of USP20 with TRAF6 and thus augments NFκB activation, SMC inflammation, and neointimal hyperplasia.

Activating FcγR function depends on endosomal-signaling platforms

Cell surface receptor internalization can either terminate signaling or activate alternative endosomal signaling pathways. We investigated here whether endosomal signaling is involved in the function of the human receptors for Fc immunoglobulin fragments (FcRs): FcαRI, FcγRIIA, and FcγRI. All these receptors were internalized after their cross-linking with receptor-specific antibodies, but their intracellular trafficking was different. FcαRI was targeted directly to lysosomes, while FcγRIIA and FcγRI were internalized in particular endosomal compartments described by the insulin esponsive minoeptidase (IRAP), where they recruited signaling molecules, such as the active form of the kinase Syk, PLCγ and the adaptor LAT. Destabilization of FcγR endosomal signaling in the absence of IRAP compromised cytokine secretion downstream FcγR activation and macrophage ability to kill tumor cells by antibody-dependent cell-mediated cytotoxicity (ADCC). Our results indicate that FcγR endosomal signaling is required for the FcγR-driven inflammatory reaction and possibly for the therapeutic action of monoclonal antibodies.

Molecular mechanisms of the tyrosine kinase inhibitor pralsetinib activity in in-vitro models of medullary thyroid carcinoma: Aberrant activation of the HH-Gli signaling pathway in acquired resistance

Medullary thyroid carcinoma (MTC) is a malignant tumor with challenging management. Multi-targeted kinase inhibitors (MKI) and tyrosine-kinase inhibitors (TKI) with high specificity for RET protein are approved for advanced MTC treatment. However, their efficacy is hindered by evasion mechanisms of tumor cells. Thus, the aim of this study was the identification of an escape mechanism in MTC cells exposed to a highly selective RET TKI. TT cells were treated with TKI, MKI, and/or the HH-Gli inhibitors, GANT61 and Arsenic Trioxide (ATO), in the presence or absence of hypoxia. RET modifications, oncogenic signaling activation, proliferation and apoptosis were assessed. Additionally, cell modifications and HH-Gli activation were also evaluated in pralsetinib-resistant TT cells. Pralsetinib inhibited RET autophosphorylation and RET downstream pathways activation in normoxic and hypoxic conditions. Additionally, pralsetinib impaired proliferation, induced the activation of apoptosis and, in hypoxic cells, downregulated HIF-1α. Focusing on escape molecular mechanisms associated with therapy, we observed increased Gli1 levels in a subset of cells. Indeed, pralsetinib stimulated the re-localization of Gli1 into the cell nuclei. Treatment of TT cells with both pralsetinib and ATO resulted in Gli1 down-regulation and impaired cell viability. Moreover, pralsetinib-resistant cells confirmed Gli1 activation and up-regulation of its transcriptionally regulated target genes. Altogether, we showed that pralsetinib impairs MTC cell growth and induces cell death, also in hypoxic conditions. The HH-Gli pathway is a new molecular mechanism of escape to pralsetinib therapy that can be overcome through combined therapy.

Abstract B021: Examining the role of HER2 signaling in promoting prostate cancer proliferation in an androgen receptor-low environment

Abstract Background: Intense neoadjuvant androgen deprivation therapy (inADT) reduces prostate cancer recurrence in approximately 40% of patients with intermediate/high risk prostate cancer. Using samples from our recent phase 2 study (NCT02430480) from patients undergoing 6 months of inADT prior to radical prostatectomy, we sought to examine molecular features present at baseline that correlate with pathologic response following surgery. Methods: Tumors from 37 patients with locally advanced prostate cancer were subjected to immunohistochemistry (IHC) and laser capture microdissection to obtain 143 histologically distinct foci. RNA was isolated from each focus and used to generate RNA-seq libraries. Differentially-expressed genes were determined per-unit of residual tumor volume using linear mixed-effect models. Histologic features and staining intensities were obtained from IHC of posttreatment specimens. Ingenuity Pathway Analysis (IPA) identified upstream regulators at baseline that interacted with pathologic outcome. Cancer cell lines were treated with a panel of EGFR family inhibitors and androgen receptor (AR) axis inhibitors. Protein and RNA expression were measured to identify the impact of dual inhibition. Results: In pre-treatment targeted biopsies from men with locally advanced prostate cancer, we observed that tumor samples that exhibited worse response to inADT had lower AR activity at baseline, presumably due to a decreased dependency on AR, potentially related to increased dependency on alternative growth pathways. These incomplete/nonresponding tumors displayed a significant increase in expression of EFGR family signaling (EGFR, HER2, and HER3) using IPA, which have previously been shown to serve as bypass mechanisms to AR signaling to drive downstream PI3K activity. HER2 was the most abundantly expressed protein by IHC in posttreatment tumors. Given the multitude of receptor tyrosine kinases (RTKs) regulated by EGFR family signaling and RTK inhibitors targeting this activity, we examined the impact of combining RTK inhibitors with ADT to overcome resistance to ADT. Afatinib, lapatinib, and neratinib showed the greatest anti-tumor efficacy in vitro. Treatment with these agents rapidly and dynamically increased AR expression and activity. We observed synergistic effects on cell viability and proliferation by treating cell lines with combination therapy of antiandrogen therapy and these inhibitors. Conclusions: Our data suggest that elevated HER2 activity defines an AR-low subtype of prostate cancer that is resistant to inADT. Disrupting the HER2-mediated bypass of AR blockade re-sensitized resistant PC cells to AR-targeted therapy. These effects were observed in an untreated setting and represent a divergent path in tumorigenesis where prostate tumors develop with intrinsically lower AR activity. These findings will be validated using genetic abrogation of HER2 in additional clinical samples, organoids, and patient-derived xenograft models, and represent new opportunities for combination therapy in newly-diagnosed high-risk disease. Citation Format: Scott Wilkinson, Anson Ku, Isaiah King, Anna Baj, Daniel Low, Huihui Ye, Stephanie A. Harmon, Nicholas T. Terrigino, John R. Bright, Rayann Atway, Shana Y. Trostel, Rosina T. Lis, Baris Turkbey, William L. Dahut, Fatima Karzai, Adam G. Sowalsky. Examining the role of HER2 signaling in promoting prostate cancer proliferation in an androgen receptor-low environment [abstract]. In: Proceedings of the AACR Special Conference: Advances in Prostate Cancer Research; 2023 Mar 15-18; Denver, Colorado. Philadelphia (PA): AACR; Cancer Res 2023;83(11 Suppl):Abstract nr B021.

Spatial distribution of heavy metal contaminants: The effects of water-sediment regulation in the Henan section of the Yellow River

Water-sediment regulation (WSR) affects sediment transport in the middle and lower reaches of the Yellow River through periodic interception of sediment by dams and alteration of downstream flow rates, which leads to redistribution and recycling of elements. In this study, based on the evaluation of seven heavy metals (V, Cr, Ni, Co, Cu, Zn, and Cd) in water, sediment and suspended particulate matter (SPM) in the Henan section of the Yellow River, the effects of WSR on the redistribution and risk of release of metals to the downstream aquatic environment were investigated, and the range of impact and pollution sources were determined. Dissolved metals were well below guideline values for water quality, but Cd, Cr, and Cu were enriched in the sediments, especially in the reservoir, and the pollution load index (PLI) results indicated that the contamination in the study area remained very low. The level of contamination was decreased, and metals migrated downstream after WSR. The chemical speciation of metals indicates that anthropogenic input of metals occurs in upstream tributaries, and the risk of release of metals in the sediments increases after WSR, but the spatial distribution of the risk is more homogeneous. The distribution coefficient explains the distribution pattern of pollution, where SPM carries pollutants to the downstream while formed surface sediments through natural deposition during the sand-discharge stage. Based on geological and field investigations, upstream gold mining and downstream riverbank cultivation activities in the study area are potential sources of Cu and Cd pollution. These findings are crucial to the better understanding of patterns of metal release from artificial impoundments in river systems and provide a theoretical basis for ecological development in the Yellow River Basin.

Preliminary results from ERAS-007 plus palbociclib (palbo) in patients (pts) with KRAS/NRAS mutant (m) colorectal cancer (CRC) or KRASm pancreatic ductal adenocarcinoma (PDAC) in HERKULES-3 study: A phase 1b/2 study of agents targeting the mitogen-activated protein kinase (MAPK) pathway in pts with advanced gastrointestinal malignancies (GI cancers).

3558 Background: The RAS/MAPK pathway is dysregulated in a broad range of cancers including CRC and PDAC, resulting in downstream activation of ERK1/2. ERAS-007 is a novel, orally bioavailable inhibitor of ERK. Palbo is an oral CDK4/6 inhibitor that inhibits cellular proliferation, an essential feature of tumor growth. Both in vitro & in vivo data exploring the combination of ERAS-007 and palbo in a panel of CRC and pancreatic CDX and/or PDX models have shown promising activity to support the potential combinatorial clinical benefit in RASm CRC and PDAC pts. Methods: HERKULES-3 is a Phase 1b/2 study to assess safety, tolerability, PK, and preliminary clinical activity of ERAS-007 combinations targeting the MAPK pathway in pts with advanced GI cancers. Within this study, we are currently evaluating the safety, tolerability, and PK of escalating doses of ERAS-007 twice daily-once a week (BID-QW) (75, 100 mg) in combination with palbo once daily (QD) (75, 100, 125 mg) in pts with KRASm/NRASm CRC or KRASm PDAC. Results: As of 30 November 2022, 30 pts were dosed with the combination of palbo and ERAS-007. Treatment emergent AEs (TEAEs) occurring in ≥20% pts were diarrhea (40%), nausea (40%), anemia (33%), vision blurred (27%), fatigue (23%), and neutrophil count decreased (20%). ERAS-007 treatment related AEs (TRAEs) were reported in 19 pts (63%), with the most frequently reported as diarrhea (40%), nausea (33%), and vision blurred (27%). Grade (Gr) ≥3 TEAEs were reported in 12 pts (40%), including 3 related to ERAS-007 (neutrophil count decreased, diarrhea and dermatitis acneiform). Neutrophil count decreased and anemia were the only Gr 3 events reported in ≥2 pts. No Gr 4 events were reported. Two Gr 5 events unrelated to any drugs (hemorrhage intracranial and malignant pleural effusion) and one Gr 5 event (anemia) related to palbo were reported. Two pts discontinued ERAS-007 due to TEAEs (Gr 5 malignant pleural effusion and Gr 2 neutrophil count decreased). Three pts reported 4 DLTs: 1 pt at 75mg ERAS-007 & 125mg palbo (Gr 3 maculopapular rash & Gr 4 sepsis), 1 pt at 100mg ERAS-007 & 100mg palbo (Gr 3 dermatitis acneiform), and 1 pt at 100mg ERAS-007 & 125mg palbo (Gr 3 thrombocytopenia). Based on preliminary PK, no clinically relevant PK interactions were identified between ERAS-007 and palbo. The evaluation of clinical activity is ongoing. Conclusions: ERAS-007 in combination with palbo in pts with KRASm/NRASm CRC or KRASm PDAC shows expected preliminary safety with reversible and manageable AEs. The highest dose evaluated and cleared by the safety review committee to date is ERAS-007 100 mg BID-QW in combination with the approved monotherapy dose of palbo 125 mg QD. Clinical trial information: NCT05039177 .

Preliminary results from ERAS-007 plus encorafenib and cetuximab (EC) in patients (pts) with metastatic BRAF V600E mutated colorectal cancer (CRC) in HERKULES-3 study: A phase 1b/2 study of agents targeting the mitogen-activated protein kinase (MAPK) pathway in pts with advanced gastrointestinal malignancies (GI cancers).

3557 Background: The RAS/MAPK pathway (including BRAF) is dysregulated in a broad range of cancers including CRC, resulting in downstream activation of ERK1/2. Metastatic CRC with BRAF V600E mutation (BRAF V600E mCRC) has dramatically worse survival than non-BRAF V600E mutated CRCs, and novel therapies are needed. ERAS-007 is a novel, potent, and orally bioavailable inhibitor of ERK. The combination of a BRAF plus EGFR inhibitor (EC) is approved for the treatment of pts with BRAF V600E mCRC; however, only 20% of pts experience an objective response. ERAS-007 alone or in combination with EC showed promising in vitro and in vivo activity in BRAF V600E CRC models to support the combinatorial clinical benefit of ERAS-007+EC in BRAF V600E mCRC. Methods: HERKULES-3 is a Phase 1b/2 study to assess the safety, tolerability, PK, and preliminary clinical activity of ERAS-007 combinations targeting the MAPK pathway in pts with advanced GI cancers. Within this study, we are currently evaluating the safety and tolerability of escalating doses of ERAS-007 + EC in pts with BRAF V600E CRC. Prior BRAF inhibitor and EGFR inhibitor treatment is neither required nor excluded to be enrolled in this study. Results: As of 30 November 2022, 12 patients were dosed with ERAS-007 twice daily-once a week (BID-QW) (75 and 100 mg; n = 10) or once daily once a week (QW) (150 mg; n = 2) in combination with EC (300 mg oral daily + 500 mg/m2 intravenous infusion once every 2 weeks). The treatment-emergent AEs (TEAEs) occurring in ≥20% of pts were fatigue (50%), headache (42%), constipation, diarrhea, nausea, dermatitis acneiform (33% each), and vomiting (25%). No TEAEs led to ERAS-007 discontinuation or death. Grade ≥3 TEAEs were reported in 3 pts (25%). Grade ≥3 treatment-related AEs reported in ≥ 2 patients (17%) include hypertension, headache, confusional state, and skin toxicity. Three pts (25%) died due to disease progression. No DLTs were reported. Out of 4 efficacy evaluable EC naïve pts, one confirmed partial response (PR) and one unconfirmed PR were observed. Evaluation of PK is ongoing and preliminary data will be presented. Conclusions: ERAS-007+EC in pts with BRAF V600E CRC shows acceptable preliminary safety/tolerability and evidence of clinical activity. The highest dose of ERAS-007 evaluated and cleared by the safety review committee to date is 100 mg BID-QW when combined with EC. Observed PK, toxicity, and preliminary activity support continued evaluation of this combination in pts with BRAF V600E CRC. Clinical trial information: NCT05039177 .

ID: 203843 A Rare Case of Ibrutinib-Induced Polyneuropathy

The B-cell-receptor (BCR) pathway, which includes Bruton’s-Tyrosine-Kinase (Btk) protein, regulates proliferation, differentiation, and apoptosis of B-cells. In B-cell malignancies such as chronic lymphocytic leukemia (CLL), aberrant BCR signaling plays critical role in pathogenesis of disease. Ibrutinab is irreversible Btk inhibitor that prevents downstream activation of the BCR pathway and subsequent proliferation of malignant B-cells. In this case report, patient with history of Ibrutinab use for several years for CLL treatment had rare adverse effect (AE) of Ibrutinib-inducted-polyneuropathy, which was only reported once in literature.

A structural blueprint for interleukin-21 signal modulation

Interleukin-21 (IL-21) plays a critical role in generating immunological memory by promoting the germinal center reaction, yet clinical use of IL-21 remains challenging because of its pleiotropy and association with autoimmune disease. To better understand the structural basis of IL-21 signaling, we determine the structure of the IL-21-IL-21R-γc ternary signaling complex by X-ray crystallography and a structure of a dimer of trimeric complexes using cryo-electron microscopy. Guided by the structure, we design analogs of IL-21 by introducing substitutions to the IL-21-γc interface. These IL-21 analogs act as partial agonists that modulate downstream activation of pS6, pSTAT3, and pSTAT1. These analogs exhibit differential activity on T and B cell subsets and modulate antibody production in human tonsil organoids. These results clarify the structural basis of IL-21 signaling and offer a potential strategy for tunable manipulation of humoral immunity.

Modulating retinoid-X-receptor alpha (RXRA) expression sensitizes chronic myeloid leukemia cells to imatinib in vitro and reduces disease burden in vivo.

Introduction: The ligand-activated transcription factors, nuclear hormone receptors (NHRs), remain unexplored in hematological malignancies except for retinoic acid receptor alpha (RARA). Methods: Here we profiled the expression of various NHRs and their coregulators in Chronic myeloid leukemia (CML) cell lines and identified a significant differential expression pattern between inherently imatinib mesylate (IM)-sensitive and resistant cell lines. Results: Retinoid-X-receptor alpha (RXRA) was downregulated in CML cell lines inherently resistant to IM and in primary CML CD34+ cells. Pre-treatment with clinically relevant RXRA ligands improved sensitivity to IM in-vitro in both CML cell lines and primary CML cells. This combination effectively reduced the viability and colony-forming capacity of CML CD34+ cells in-vitro. In-vivo, this combination reduced leukemic burden and prolonged survival. Overexpression (OE) of RXRA inhibited proliferation and improved sensitivity to IM in-vitro. In-vivo, RXRA OE cells showed reduced engraftment of cells in the bone marrow, improved sensitivity to IM, and prolonged survival. Both RXRA OE and ligand treatment markedly reduced BCR::ABL1 downstream kinase activation, activating apoptotic cascades and improving sensitivity to IM. Importantly, RXRA OE also led to the disruption of the oxidative capacity of these cells. Conclusion: Combining IM with clinically available RXRA ligands could form an alternative treatment strategy in CML patients with suboptimal response to IM.

Endosomal trafficking of two-pore K+ efflux channel TWIK2 to plasmalemma mediates NLRP3 inflammasome activation and inflammatory injury.

Potassium efflux via the two-pore K+ channel TWIK2 is a requisite step for the activation of NLRP3 inflammasome, however, it remains unclear how K+ efflux is activated in response to select cues. Here, we report that during homeostasis, TWIK2 resides in endosomal compartments. TWIK2 is transported by endosomal fusion to the plasmalemma in response to increased extracellular ATP resulting in the extrusion of K+. We showed that ATP-induced endosomal TWIK2 plasmalemma translocation is regulated by Rab11a. Deleting Rab11a or ATP-ligated purinergic receptor P2X7 each prevented endosomal fusion with the plasmalemma and K+ efflux as well as NLRP3 inflammasome activation in macrophages. Adoptive transfer of Rab11a-depleted macrophages into mouse lungs prevented NLRP3 inflammasome activation and inflammatory lung injury. We conclude that Rab11a-mediated endosomal trafficking in macrophages thus regulates TWIK2 localization and activity at the cell surface and the downstream activation of the NLRP3 inflammasome. Results show that endosomal trafficking of TWIK2 to the plasmalemma is a potential therapeutic target in acute or chronic inflammatory states.

Neglecting complex network structures underestimates delays in a large-capital project

Completing large-scale projects on time is a daunting challenge, partly due to the intricate network of dependencies between a project’s activities. To support this challenge, existing theory focuses on predicting whether a delay in completing a single activity is likely to spread and impact downstream activities. Using fine-grained information from 68 546 activities and 84 934 pairs, associated with the delivery of a $1.86Bn infrastructure project, we show that the core mechanism that underpins existing theory underestimates delay propagation. To elucidate the mechanisms that drive delay, we generated null models that destroyed the structural and temporal correlations of the original project activity network. By doing so, we argue that this underestimation is the result of neglecting endogenous structural features within the project’s activity network. Formulating a new mechanism that utilizes both temporal and structural features may help improve our capacity to predict how delays spread within projects.

Rethinking Oncologic Treatment Strategies with Interleukin-2

High-dose recombinant human IL-2 (rhIL-2, aldesleukin) emerged as an important treatment option for selected patients with metastatic melanoma and metastatic renal cell carcinoma, producing durable and long-lasting antitumor responses in a small fraction of patients and heralding the potential of cancer immunotherapy. However, the adoption of high-dose rhIL-2 has been restricted by its severe treatment-related adverse event (TRAE) profile, which necessitates highly experienced clinical providers familiar with rhIL-2 administration and readily accessible critical care medicine support. Given the comparatively wide-ranging successes of immune checkpoint inhibitors and chimeric antigen receptor T cell therapies, there have been concerted efforts to significantly improve the efficacy and toxicities of IL-2-based immunotherapeutic approaches. In this review, we highlight novel drug development strategies, including biochemical modifications and engineered IL-2 variants, to expand the narrow therapeutic window of IL-2 by leveraging downstream activation of the IL-2 receptor to selectively expand anti-tumor CD8-positive T cells and natural killer cells. These modified IL-2 cytokines improve single-agent activity in solid tumor malignancies beyond the established United States Food and Drug Administration (FDA) indications of metastatic melanoma and renal cell carcinoma, and may also be safer in rational combinations with established treatment modalities, including anti-PD-(L)1 and anti-CTLA-4 immunotherapy, chemotherapies, and targeted therapy approaches.

Post-injury Inhibition of Endothelin-1 Dependent Renal Vasoregulation Mitigates Rhabdomyolysis-Induced Acute Kidney Injury.

In patients with rhabdomyolysis, the overwhelming release of myoglobin into the circulation is the primary cause of kidney injury. Myoglobin causes direct kidney injury as well as severe renal vasoconstriction. An increase in renal vascular resistance (RVR) results in renal blood flow (RBF) and glomerular filtration rate (GFR) reduction, tubular injury, and acute kidney injury (AKI). The mechanisms that underlie rhabdomyolysis-induced AKI are not fully understood but may involve the local production of vasoactive mediators in the kidney. Studies have shown that myoglobin stimulates endothelin-1 (ET-1) production in glomerular mesangial cells. Circulating ET-1 is also increased in rats subjected to glycerol-induced rhabdomyolysis. However, the upstream mechanisms of ET-1 production and downstream effectors of ET-1 actions in rhabdomyolysis-induced AKI remain unclear. Vasoactive ET-1 is generated by ET converting enzyme 1 (ECE-1)-induced proteolytic processing of inactive big ET to biologically active peptides. The downstream ion channel effectors of ET-1-induced vasoregulation include the transient receptor potential cation channel, subfamily C member 3 (TRPC3). This study demonstrates that glycerol-induced rhabdomyolysis in Wistar rats promotes ECE-1-dependent ET-1 production, RVR increase, GFR decrease, and AKI. Rhabdomyolysis-induced increases in RVR and AKI in the rats were attenuated by post-injury pharmacological inhibition of ECE-1, ET receptors, and TRPC3 channels. CRISPR/Cas9-mediated knockout of TRPC3 channels attenuated ET-1-induced renal vascular reactivity and rhabdomyolysis-induced AKI. These findings suggest that ECE-1-driven ET-1 production and downstream activation of TRPC3-dependent renal vasoconstriction contribute to rhabdomyolysis-induced AKI. Hence, post-injury inhibition of ET-1-mediated renal vasoregulation may provide therapeutic targets for rhabdomyolysis-induced AKI.

Downstream Space Activities in the New Space Era: Paradigm Shift and Evaluation Challenges

New Space refers to the recent opening-up of the space sector to private companies. The liberalization of space activities, which coincides with the digital evolution of the economy, is associated with the rapid expansion of the downstream space segment, i.e., space-related commercial products and services. In this paper, we address the issue of evaluating the size of the downstream space sector in light of New Space characteristics. We describe New Space as a structural transformation of the space sector associated with the entry of private companies and investors, the adoption of new business models, and the reorientation of space agencies towards market-oriented policies. In this specific context, we claim that assessing the economic weight of downstream space activities is of prime interest. Our review of space sector evaluations shows limitations of the existing methodological tools when identifying and evaluating downstream activities. We conclude this work by outlining theoretical issues regarding New Space and the expansion of space-based services markets to be considered for a new evaluation methodology.

Regulation of cGAS-STING signalling in cancer: Approach for combination therapy.

Innate immunity plays an important role not only during infection but also homeostatic role during stress conditions. Activation of the immune system including innate immune response plays a critical role in the initiation and progression of tumorigenesis. The innate immune sensor recognizes pathogen-associated molecular patterns (PAMPs) and activates cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) (cGAS-STING) and induces type-1 immune response during viral and bacterial infection. cGAS-STING is regulated differently in conditions like cellular senescence and DNA damage in normal and tumor cells and is implicated in the progression of tumors from different origins. cGAS binds to cytoplasmic dsDNA and synthesize cyclic GMP-AMP (2'3'-cGAMP), which selectively activates STING and downstream IFN and NF-κB activation. We here reviewed the cGAS-STING signalling pathway and its cross-talk with other pathways to modulate tumorigenesis. Further, the review also focused on emerging studies that targeted the cGAS-STING pathway for developing targeted therapeutics and combinatorial regimens for cancer of different origins.

MTHFD2 reprograms macrophage polarization by inhibiting PTEN

The one-carbon metabolism enzyme methylenetetrahydrofolate dehydrogenase 2 (MTHFD2) is involved in the regulation of tumor oncogenesis and immune cell functions, but whether it can contribute to macrophage polarization remains elusive. Here, we show that MTHFD2 suppresses polarization of interferon-γ-activated macrophages (M(IFN-γ)) but enhances that of interleukin-4-activated macrophages (M(IL-4)) both invitro and invivo. Mechanistically, MTHFD2 interacts with phosphatase and tensin homolog (PTEN) to suppress PTEN's phosphatidylinositol3,4,5-trisphosphate (PIP3) phosphatase activity and enhance downstream Akt activation, independent of the N-terminal mitochondria-targeting signal of MTHFD2. MTHFD2-PTEN interaction is promoted by IL-4 but not IFN-γ. Furthermore, amino acid residues (aa 215-225) of MTHFD2 directly target PTEN catalytic center (aa 118-141). Residue D168 of MTHFD2 is also critical for regulating PTEN's PIP3 phosphatase activity by affecting MTHFD2-PTEN interaction. Our study suggests a non-metabolic function of MTHFD2 by which MTHFD2 inhibits PTEN activity, orchestrates macrophage polarization, and alters macrophage-mediated immune responses.