Systematic comparison of rAAV vectors manufactured using large-scale suspension cultures of Sf9 and HEK293 cells.

Recombinant adeno-associated virus (rAAV) vectors could be manufactured by plasmid transfection into human embryonic kidney 293 (HEK293) cells or baculovirus infection of Spodoptera frugiperda (Sf9) insect cells. However, systematic comparisons between these systems using large-scale, high-quality AAV vectors are lacking. rAAV from Sf9 cells (Sf9-rAAV) at 2-50 L and HEK293 cells (HEK-rAAV) at 2-200 L scales were characterized. HEK-rAAV had ∼40-fold lower yields but ∼10-fold more host cell DNA measured by droplet digital PCR and next-generation sequencing, respectively. The electron microscope observed a lower full/empty capsid ratio in HEK-rAAV (70.8%) than Sf9-rAAV (93.2%), while dynamic light scattering and high-performance liquid chromatography analysis showed that HEK-rAAV had more aggregation. Liquid chromatography tandem mass spectrometry identified different post-translational modification profiles between Sf9-rAAV and HEK-rAAV. Furthermore, Sf9-rAAV had a higher tissue culture infectious dose/viral genome than HEK-rAAV, indicating better infectivity. Additionally, Sf9-rAAV achieved higher in vitro transgene expression, as measured by ELISA. Finally, after intravitreal dosing into a mouse laser choroidal neovascularization model, Sf9-rAAV and HEK-rAAV achieved similar efficacy. Overall, this study detected notable differences in the physiochemical characteristics of HEK-rAAV and Sf9-rAAV. However, the in vitro and in vivo biological functions of the rAAV from these systems were highly comparable. Sf9-rAAV may be preferred over HEK293-rAAV for advantages in yields, full/empty ratio, scalability, and cost.

DNA-PK is activated by SIRT2 deacetylation to promote DNA double-strand break repair by non-homologous end joining.

DNA-dependent protein kinase (DNA-PK) plays a critical role in non-homologous end joining (NHEJ), the predominant pathway that repairs DNA double-strand breaks (DSB) in response to ionizing radiation (IR) to govern genome integrity. The interaction of the catalytic subunit of DNA-PK (DNA-PKcs) with the Ku70/Ku80 heterodimer on DSBs leads to DNA-PK activation; however, it is not known if upstream signaling events govern this activation. Here, we reveal a regulatory step governing DNA-PK activation by SIRT2 deacetylation, which facilitates DNA-PKcs localization to DSBs and interaction with Ku, thereby promoting DSB repair by NHEJ. SIRT2 deacetylase activity governs cellular resistance to DSB-inducing agents and promotes NHEJ. SIRT2 furthermore interacts with and deacetylates DNA-PKcs in response to IR. SIRT2 deacetylase activity facilitates DNA-PKcs interaction with Ku and localization to DSBs and promotes DNA-PK activation and phosphorylation of downstream NHEJ substrates. Moreover, targeting SIRT2 with AGK2, a SIRT2-specific inhibitor, augments the efficacy of IR in cancer cells and tumors. Our findings define a regulatory step for DNA-PK activation by SIRT2-mediated deacetylation, elucidating a critical upstream signaling event initiating the repair of DSBs by NHEJ. Furthermore, our data suggest that SIRT2 inhibition may be a promising rationale-driven therapeutic strategy for increasing the effectiveness of radiation therapy.

The acquired dyad inclination and decreased interpersonal brain communication in the pursuit of collective benefit.

People perform better collectively than individually, a phenomenon known as the collective benefit. To pursue the benefit, they may learn from previous behaviors, come to know whose initial opinion should be valued, and develop the inclination to take it as the collective one. Such learning may affect interpersonal brain communication. To test these hypotheses, this study recruited participant dyads to conduct a perceptual task on which they made individual decisions first and then the collective one. The enhanced interpersonal brain synchronization (IBS) between participants was explored when individual decisions were in disagreement vs. agreement. Computational modeling revealed that participant dyads developed the dyad inclination of taking the higher-able participants', not the lower-able ones' decisions as their collective ones. Brain analyses unveiled the enhanced IBS at frontopolar areas, premotor areas, supramarginal gyri, and right temporal-parietal junctions. The premotor IBS correlated negatively with dyad inclination and collective benefit in the absence of correction. The Granger causality analyses further supported the negative relation of dyad inclination with inter-brain communication. This study highlights that dyads learn to weigh individuals' decisions, resulting in dyad inclinations, and explores associated inter-brain communication, offering insights into the dynamics of collective decision-making.

Viral communities in a pH>10 serpentinite-like environment: insight into diversity and potential roles in modulating the microbiomes by bioactive vitamin B9 synthesis.

Viral communities exist in a variety of ecosystems and play significant roles in mediating biogeochemical processes, whereas viruses inhabiting strongly alkaline geochemical systems remain underexplored. In this study, the viral diversity, potential functionalities, and virus-host interactions in a strongly alkaline environment (pH = 10.4-12.4) exposed to the leachates derived from the serpentinization-like reactions of smelting slags were investigated. The viral populations (e.g., Herelleviridae, Queuovirinae, and Inoviridae) were closely associated with the dominating prokaryotic hosts (e.g., Meiothermus, Trueperaceae, and Serpentinomonas) in this ultrabasic environment. Auxiliary metabolic genes (AMGs) suggested that viruses may enhance hosts' fitness by facilitating cofactor biosynthesis, hydrogen metabolism, and carbon cycling. To evaluate the activity of synthesis of essential cofactor vitamin B9 by the viruses, a viral folA (vfolA) gene encoding dihydrofolate reductase (DHFR) was introduced into a thymidine-auxotrophic strain Escherichia coli MG1655 ΔfolA mutant, which restored the growth of the latter in the absence of thymidine. Notably, the homologs of the validated vDHFR were globally distributed in the viromes across various ecosystems. The present study sheds new light on the unique viral communities in hyperalkaline ecosystems and their potential beneficial impacts on the coexisting microbial consortia by supplying essential cofactors. IMPORTANCE: This study presents a comprehensive investigation into the diversity, potential functionalities, and virus-microbe interactions in an artificially induced strongly alkaline environment. Functional validation of the detected viral folA genes encoding dihydrofolate reductase substantiated the synthesis of essential cofactors by viruses, which may be ubiquitous, considering the broad distribution of the viral genes associated with folate cycling.

Blue light photoreceptor cryptochrome 1 promotes wood formation and anthocyanin biosynthesis in Populus.

Blue light photoreceptor cryptochrome 1 (CRY1) in herbaceous plants plays crucial roles in various developmental processes, including cotyledon expansion, hypocotyl elongation and anthocyanin biosynthesis. However, the function of CRY1 in perennial trees is unclear. In this study, we identified two ortholog genes of CRY1 (PagCRY1a and PagCRY1b) from Populus, which displayed high sequence similarity to Arabidopsis CRY1. Overexpression of PagCRY1 substantially inhibited plant growth and promoted secondary xylem development in Populus, while CRISPR/Cas9-mediated knockout of PagCRY1 enhanced plant growth and delayed secondary xylem development. Moreover, overexpression of PagCRY1 dramatically increased anthocyanin accumulation. The further analysis supported that PagCRY1 functions specifically in response to blue light. Taken together, our results demonstrated that modulating the expression of blue light photoreceptor CRY1 ortholog gene in Populus could significantly influence plant biomass production and the process of wood formation, laying a foundation for further investigating the light-regulated tree growth.

Field-Induced Non-BEC Transitions in Frustrated Magnets.

Frustrated spin systems have traditionally proven challenging to understand, owing to a scarcity of controlled methods for their analyses. By contrast, under strong magnetic fields, certain aspects of spin systems admit simpler and universal description in terms of hardcore bosons. The bosonic formalism is anchored by the phenomenon of Bose-Einstein condensation (BEC), which has helped explain the behaviors of a wide range of magnetic compounds under applied magnetic fields. Here, we focus on the interplay between frustration and externally applied magnetic field to identify instances where the BEC paradigm is no longer applicable. As a representative example, we consider the antiferromagnetic J_{1}-J_{2}-J_{3} model on the square lattice in the presence of a uniform external magnetic field, and demonstrate that the frustration-driven suppression of the Néel order leads to a Lifshitz transition for the hardcore bosons. In the vicinity of the Lifshitz point, the physics becomes unmoored from the BEC paradigm, and the behavior of the system, both at and below the saturation field, is controlled by a Lifshitz multicritical point. We obtain the resultant universal scaling behaviors, and provide strong evidence for the existence of a frustration and magnetic-field driven correlated bosonic liquid state along the entire phase boundary separating the Néel phase from other magnetically ordered states.

Tepidibacillus marianensis sp. nov., a novel heterotrophic iron-reducing bacterium isolated from Mariana Trench sediment.

A novel chemoheterotrophic iron-reducing micro-organism, designated as strain LSZ-M11000T, was isolated from sediment of the Marianas Trench. Phylogenetic analysis based on the 16S rRNA gene revealed that strain LSZ-M11000T belonged to genus Tepidibacillus, with 97 % identity to that of Tepidibacillus fermentans STGHT, a mesophilic bacterium isolated from the Severo-Stavropolskoye underground gas storage facility in Russia. The polar lipid profile of strain LSZ-M11000T consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, as well as other unidentified phospholipids and lipids. The major fatty acids were C16 : 0 (28.4 %), C18 : 0 (15.8 %), iso-C15 : 0 (12.9 %), and anteiso-C15 : 0 (12.0 %). Strain LSZ-M11000T had no menaquinone. Genome sequencing revealed that the genome size of strain LSZ-M11000T was 2.97 Mb and the DNA G+C content was 37.9 mol%. The average nucleotide identity values between strain LSZ-M11000T and its close phylogenetic relatives, Tepidibacillus fermentans STGHT and Tepidibacillus decaturensis Z9T, were 76.4 and 72.6 %, respectively. The corresponding DNA-DNA hybridization estimates were 20.9 and 23.4 %, respectively. Cells of strain LSZ-M11000T were rod-shaped (1.0-1.5×0.3-0.5 µm). Using pyruvate as an electron donor, it was capable of reducing KMnO4, MnO2, As(V), NaNO3, NaNO2, Na2SO4, Na2S2O3, and K2Cr2O7. Based on phenotypic, genotypic, and phylogenetic evidence, strain LSZ-M11000T is proposed to be a novel strain of the genus Tepidibacillus, for which the name Tepdibacillus marianensis is proposed. The type strain is LSZ-M11000T (=CCAM 1008T=JCM 39431T).

The m6A reader IGF2BP1 contributes to the activation of hepatic stellate cells through facilitating TUBB4B mRNA stabilization.

The m6A reader insulin-like growth factor-2 mRNA-binding protein 1 (IGF2BP1) is involved in multiple pathophysiological processes through enhanced expression of the proteins encoded by their target mRNAs. However, the functional role of IGF2BP1-mediated m6A in liver fibrosis remains elusive. Here, we report that IGF2BP1 is highly expressed in activated hepatic stellate cells (HSCs), the major driver of fibrogenesis, and TUBB4B is identified as a potential target of IGF2BP1 by re-analysis of the RNA-seq, RIP-seq, and m6A-seq data. The relevant findings were subsequently demonstrated by a series of molecular and cellular evidences. The knockdown of IGF2BP1 or TUBB4B and pharmacological inhibition of TUBB4B by mebendazole treatments significantly suppress the proliferation, migration, and activation of HSCs. Mechanistically, IGF2BP1 upregulates TUBB4B expression through stabilizing TUBB4B in an m6A-dependent manner, and TUBB4B induces liver fibrosis by activating the FAK signaling pathway. Collectively, our results indicate that targeting IGF2BP1/TUBB4B/FAK axis in HSCs could be a promising therapeutic approach for liver fibrosis.

α4 nicotinic receptors on GABAergic neurons mediate a cholinergic analgesic circuit in the substantia nigra pars reticulata.

Nicotinic acetylcholine receptors (nAChRs) regulate pain pathways with various outcomes depending on receptor subtypes, neuron types, and locations. But it remains unknown whether α4ß2 nAChRs abundantly expressed in the substantia nigra pars reticulata (SNr) have potential to mitigate hyperalgesia in pain states. We observed that injection of nAChR antagonists into the SNr reduced pain thresholds in naïve mice, whereas injection of nAChR agonists into the SNr relieved hyperalgesia in mice, subjected to capsaicin injection into the lower hind leg, spinal nerve injury, chronic constriction injury, or chronic nicotine exposure. The analgesic effects of nAChR agonists were mimicked by optogenetic stimulation of cholinergic inputs from the pedunculopontine nucleus (PPN) to the SNr, but attenuated upon downregulation of α4 nAChRs on SNr GABAergic neurons and injection of dihydro-ß-erythroidine into the SNr. Chronic nicotine-induced hyperalgesia depended on α4 nAChRs in SNr GABAergic neurons and was associated with the reduction of ACh release in the SNr. Either activation of α4 nAChRs in the SNr or optogenetic stimulation of the PPN-SNr cholinergic projection mitigated chronic nicotine-induced hyperalgesia. Interestingly, mechanical stimulation-induced ACh release was significantly attenuated in mice subjected to either capsaicin injection into the lower hind leg or SNI. These results suggest that α4 nAChRs on GABAergic neurons mediate a cholinergic analgesic circuit in the SNr, and these receptors may be effective therapeutic targets to relieve hyperalgesia in acute and chronic pain, and chronic nicotine exposure.

New mitogenomes from the genus Cricotopus (Diptera: Chironomidae, Orthocladiinae): Characterization and phylogenetic implications.

Cricotopus is a large and diverse genus of non-biting midges composed of several subgenera. Complete mitogenome sequences are available for very few Cricotopus species. The subgenus Pseudocricotopus unites species with unusual morphological structures in adult male and pupal stages, however, molecular methods are needed to verify the placement of this subgenus within Cricotopus. We obtained mitogenomes of C. (Pseudocricotopus) cf. montanus and nine other Cricotopus species for phylogenetic analysis, coupled with two Rheocricotopus species and one Synorthocladius species as outgroups. The structure of the mitogenome was similar among these Cricotopus species, exhibiting A+T bias and retaining ancestral gene order. Mutation rate, estimated as Ka/Ks, varied among genes, and was highest for ATP8 and lowest for COI. The phylogenetic relationships among species of Cricotopus, Rheocricotopus and Synorthocladius was reconstructed using Bayesian inference and maximum likelihood estimation. The phylogenetic trees confirmed placement of subgenus Pseudocricotopus, represented by Cricotopus cf. montanus, within Cricotopus. Our study increases the library of chironomid mitogenomes and provides insight into the properties of their constituent genes.

Correlation between serum trimethylamine-N-oxide and body fat distribution in middle-aged and older adults: a prospective cohort study.

BACKGROUND: Trimethylamine-N-oxide (TMAO) is linked with obesity, while limited evidence on its relationship with body fat distribution. Herein, we investigated the associations between serum TMAO and longitudinal change of fat distribution in this prospective cohort study. METHODS: Data of 1964 participants (40-75y old) from Guangzhou Nutrition and Health Study (GNHS) during 2008-2014 was analyzed. Serum TMAO concentration was quantified by HPLC-MS/MS at baseline. The body composition was assessed by dual-energy X-ray absorptiometry at each 3-y follow-up. Fat distribution parameters were fat-to-lean mass ratio (FLR) and trunk-to-leg fat ratio (TLR). Fat distribution changes were derived from the coefficient of linear regression between their parameters and follow-up duration. RESULTS: After an average of 6.2-y follow-up, analysis of covariance (ANCOVA) and linear regression displayed women with higher serum TMAO level had greater increments in trunk FLR (mean ± SD: 1.47 ± 4.39, P-trend = 0.006) and TLR (mean ± SD: 0.06 ± 0.24, P-trend = 0.011). Meanwhile, for women in the highest TMAO tertile, linear mixed-effects model (LMEM) analysis demonstrated the annual estimated increments (95% CI) were 0.03 (95% CI: 0.003 - 0.06, P = 0.032) in trunk FLR and 1.28 (95% CI: -0.17 - 2.73, P = 0.083) in TLR, respectively. In men, there were no similar significant observations. Sensitivity analysis yielded consistent results. CONCLUSION: Serum TMAO displayed a more profound correlation with increment of FLR and TLR in middle-aged and older community-dwelling women in current study. More and further studies are still warranted in the future. TRIAL REGISTRATION: NCT03179657.

Enhanced cartilage regeneration by icariin and mesenchymal stem cell-derived extracellular vesicles combined in alginate-hyaluronic acid hydrogel.

OBJECTIVE: Osteoarthritis (OA) is characterized by progressive cartilage degeneration and absence of curative therapies. Therefore, more efficient therapies are compellingly needed. Both mesenchymal stem cells (MSCs)-derived extracellular vesicles (EVs) and Icariin (ICA) are promising for repair of cartilage defect. This study proposes that ICA may be combined to potentiate the cartilage repair capacity of MSC-EVs. MATERIALS AND METHODS: MSC-EVs were isolated from sodium alginate (SA) and hyaluronic acid (HA) composite hydrogel (SA-HA) cell spheroid culture. EVs and ICA were combined in SA-HA hydrogel to test therapeutic efficacy on cartilage defect in vivo. RESULTS: EVs and ICA were synergistic for promoting both proliferation and migration of MSCs and inflammatory chondrocytes. The combination therapy led to strikingly enhanced repair on cartilage defect in rats, with mechanisms involved in the concomitant modulation of both cartilage degradation and synthesis makers. CONCLUSION: The MSC-EVs-ICA/SA-HA hydrogel potentially constitutes a novel therapy for cartilage defect in OA.

Evidence for Topological Magnon-Phonon Hybridization in a 2D Antiferromagnet down to the Monolayer Limit.

Topological phonons and magnons potentially enable low-loss, quantum coherent, and chiral transport of information and energy at the atomic scale. Van der Waals magnetic materials are promising to realize such states due to their recently discovered strong interactions among the electronic, spin, and lattice degrees of freedom. Here, we report the first observation of coherent hybridization of magnons and phonons in monolayer antiferromagnet FePSe3 by cavity-enhanced magneto-Raman spectroscopy. The robust magnon-phonon cooperativity in the 2D limit occurs even in zero magnetic field, which enables nontrivial band inversion between longitudinal and transverse optical phonons caused by the strong coupling with magnons. The spin and lattice symmetry theoretically guarantee magnetic-field-controlled topological phase transition, verified by nonzero Chern numbers calculated from the coupled spin-lattice model. The 2D topological magnon-phonon hybridization potentially offers a new route toward quantum phononics and magnonics with an ultrasmall footprint.

Source identification and migration fate of heavy metals of soil-groundwater system in a thousand-year cultivation region.

Pollution of farmland by heavy metals threatens food security and human health. In addition, heavy metals in soil could infiltrate into groundwater to influence the water quality and safety of drinking water. However, the relationship between heavy metal pollution in soil and groundwater is still not clear. In this study, we investigated the soil and groundwater in the Guanzhong Plain region, which is a significant grain production base in China, and determined the spatial distributions, ecological risk, sources, and migration fates of heavy metals (As, Cd, Cr, Cu, Ni, Pb, and Zn). The results showed that the mean values (0-20 cm) in the soil were 19.57 mg kg-1 for As, 0.71 mg kg-1 for Cd, 69.65 mg kg-1 for Cr, 21.97 mg kg-1 for Cu, 28.67 mg kg-1 for Ni, 17.54 mg kg-1 for Pb, and 73.77 mg kg-1 for Zn, and the corresponding mean values in groundwater were 1.2, 0.04, 4.69, 0.15, 0.07, 0.3, and 3.6 µg L-1, respectively. The mean values for As, Cd, Cr, Pb, and Zn in soil exceeded the background values, and the mean values for As, Cd and Pb exceeded those in groundwater. Positive matrix factorization models identified five sources (fertilizers and organic fertilizers, natural sources, pesticides and herbicides, industrial activities, and sedimentation caused by transportation) for heavy metal pollution in soil and four sources (industry activity, atmospheric sedimentation caused by transportation, natural sources, and agriculture) for heavy metal pollution in groundwater. The soil particle composition and soil organic carbon content were important factors that affected the vertical distribution of heavy metals in the soil. The migration modes (convection and diffusion) were not found for all heavy metals. These results help to understand the relationships between heavy metals in soil and groundwater in farmland ecosystems regionally.

Effects of Cuisine-Based Chinese Heart-Healthy Diet in Lowering Blood Pressure Among Adults in China: Multicenter, Single-Blind, Randomized, Parallel Controlled Feeding Trial.

BACKGROUND: More than one-fifth of the world's population consumes Chinese cuisines regularly, but no evidence-based healthy diets fitting the Chinese food culture are available for implementation. METHODS: A multicenter, patient- and outcome assessor-blind, randomized feeding trial was conducted among 265 participants with 130 to 159 mm Hg baseline systolic blood pressure (SBP) for 4 major Chinese cuisines (Shangdong, Huaiyang, Cantonese, Szechuan). After a 7-day run-in period on a control diet matching the usual local diets, participants were randomized to continue with the control diet or the cuisine-based Chinese heart-healthy diet for another 28 days. The primary outcome was SBP, and secondary outcomes included diastolic blood pressure and food preference score. Linear regression models were used to estimate the intervention effects and adjustments for the center. The incremental cost per 1 mm Hg reduction in SBP was also calculated. RESULTS: A total of 265 participants were randomized (135 on the Chinese heart-healthy diet and 130 on the control diet), with 52% women, mean age of 56.5±9.8 years, and mean SBP and diastolic blood pressure of 139.4±8.3 and 88.1±8.0 mm Hg, respectively, at baseline. The change in SBP and diastolic blood pressure from baseline to the end of the study in the control group was -5.0 (95% CI, -6.5 to -3.5) mm Hg and -2.8 (95% CI, -3.7 to -1.9) mm Hg, respectively. The net difference of change between the 2 groups in SBP and diastolic blood pressure were -10.0 (95% CI, -12.1 to -7.9) mm Hg and -3.8 (95% CI, -5.0 to -2.5) mm Hg, respectively. The effect size did not differ among cuisines (P for interaction=0.173). The mean food preference score was 9.5 (with 10 the best preferred) at baseline, and the net change during intervention was 0.1 (95% CI, -0.1 to 0.2; P=0.558). The incremental cost-effectiveness ratio per 1 mm Hg SBP reduction was CNY 0.4 (USD 0.06) per day. No difference in the number of adverse events was found between the 2 groups (P=0.259), and none of the adverse events was associated with the intervention. CONCLUSIONS: The Chinese heart-healthy diet is effective, palatable, and cost-effective in reducing blood pressure in Chinese adults with high blood pressure, with a clinically significant effect applicable across major Chinese cuisine cultures. REGISTRATION: URL: https://www. CLINICALTRIALS: gov; Unique identifier: NCT03882645.

Selective CDK9 knockdown sensitizes TRAIL response by suppression of antiapoptotic factors and NF-kappaB pathway.

The aberrantly up-regulated CDK9 can be targeted for cancer therapy. The CDK inhibitor dinaciclib (Dina) has been found to drastically sensitizes cancer response to TRAIL-expressing extracellular vesicle (EV-T). However, the low selectivity of Dina has limited its application for cancer. We propose that CDK9-targeted siRNA (siCDK9) may be a good alternative to Dina. The siCDK9 molecules were encapsulated into EV-Ts to prepare a complexed nanodrug (siEV-T). It was shown to efficiently suppress CDK9 expression and overcome TRAIL resistance to induce strikingly augmented apoptosis in lung cancer both in vitro and in vivo, with a mechanism related to suppression of both anti-apoptotic factors and nuclear factor-kappa B pathway. Therefore, siEV-T potentially constitutes a novel, highly effective and safe therapy for cancers.

Dynamic peripheral blood immune cell markers for predicting the response of patients with metastatic cancer to immune checkpoint inhibitors.

PURPOSE: Immune checkpoint inhibitors (ICIs) have shown durable responses in various malignancies. However, the response to ICI therapy is unpredictable, and investigation of predictive biomarkers needs to be improved. EXPERIMENTAL DESIGN: In total, 120 patients receiving ICI therapy and 40 patients receiving non-ICI therapy were enrolled. Peripheral blood immune cell markers (PBIMs), as liquid biopsy biomarkers, were analyzed by flow cytometry before ICI therapy, and before the first evaluation. In the ICI cohort, patients were randomly divided into training (n = 91) and validation (n = 29) cohorts. Machine learning algorithms were applied to construct the prognostic and predictive immune-related models. RESULTS: Using the training cohort, a peripheral blood immune cell-based signature (BICS) based on four hub PBIMs was developed. In both the training and the validation cohorts, and the whole cohort, the BICS achieved a high accuracy for predicting overall survival (OS) benefit. The high-BICS group had significantly shorter progression-free survival and OS than the low-BICS group. The BICS demonstrated the predictive ability of patients to achieve durable clinical outcomes. By integrating these PBIMs, we further constructed and validated the support vector machine-recursive and feature elimination classifier model, which robustly predicts patients who will achieve optimal clinical benefit. CONCLUSIONS: Dynamic PBIM-based monitoring as a noninvasive, cost-effective, highly specific and sensitive biomarker has broad potential for prognostic and predictive utility in patients receiving ICI therapy.

Nras Q61R/+ and Kras-/- cooperate to downregulate Rasgrp1 and promote lympho-myeloid leukemia in early T-cell precursors.

Early T-cell precursor acute lymphoblastic leukemia (ETP-ALL) is an aggressive subtype of T-cell ALL. Although genetic mutations hyperactivating cytokine receptor/Ras signaling are prevalent in ETP-ALL, it remains unknown how activated Ras signaling contributes to ETP-ALL. Here, we find that in addition to the frequent oncogenic RAS mutations, wild-type (WT) KRAS transcript level was significantly downregulated in human ETP-ALL cells. Similarly, loss of WT Kras in NrasQ61R/+ mice promoted hyperactivation of extracellular signal-regulated kinase (ERK) signaling, thymocyte hyperproliferation, and expansion of the ETP compartment. Kras-/-; NrasQ61R/+ mice developed early onset of T-cell malignancy that recapitulates many biological and molecular features of human ETP-ALL. Mechanistically, RNA-sequencing analysis and quantitative proteomics study identified that Rasgrp1, a Ras guanine nucleotide exchange factor, was greatly downregulated in mouse and human ETP-ALL. Unexpectedly, hyperactivated Nras/ERK signaling suppressed Rasgrp1 expression and reduced Rasgrp1 level led to increased ERK signaling, thereby establishing a positive feedback loop to augment Nras/ERK signaling and promote cell proliferation. Corroborating our cell line data, Rasgrp1 haploinsufficiency induced Rasgrp1 downregulation and increased phosphorylated ERK level and ETP expansion in NrasQ61R/+ mice. Our study identifies Rasgrp1 as a negative regulator of Ras/ERK signaling in oncogenic Nras-driven ETP-like leukemia.

Expression of NrasQ61R and MYC transgene in germinal center B cells induces a highly malignant multiple myeloma in mice.

NRAS Q61 mutations are prevalent in advanced/relapsed multiple myeloma (MM) and correlate with poor patient outcomes. Thus, we generated a novel MM model by conditionally activating expression of endogenous NrasQ61R and an MYC transgene in germinal center (GC) B cells (VQ mice). VQ mice developed a highly malignant MM characterized by a high proliferation index, hyperactivation of extracellular signal-regulated kinase and AKT signaling, impaired hematopoiesis, widespread extramedullary disease, bone lesions, kidney abnormalities, preserved programmed cell death protein 1 and T-cell immunoreceptor with immunoglobulin and immunoreceptor tyrosine-based inhibition motif domain immune-checkpoint pathways, and expression of human high-risk MM gene signatures. VQ MM mice recapitulate most of the biological and clinical features of human advanced/high-risk MM. These MM phenotypes are serially transplantable in syngeneic recipients. Two MM cell lines were also derived to facilitate future genetic manipulations. Combination therapies based on MEK inhibition significantly prolonged the survival of VQ mice with advanced-stage MM. Our study provides a strong rationale to develop MEK inhibition-based therapies for treating advanced/relapsed MM.

[18F]FAPI PET/CT in the evaluation of focal liver lesions with [18F]FDG non-avidity.

PURPOSE: This prospective study was aimed to investigate the potential utility of [18F]fibroblast activation protein inhibitor (FAPI) PET/CT for evaluating focal liver lesions (FLLs) with [18F]FDG non-avidity. METHODS: From January 2021 to March 2022, this prospective study included 80 FLLs that were not avid on [18F]FDG PET/CT from 37 patients, then underwent [18F]FAPI PET/CT. All patients with FLL(s) with biopsy-proof or follow-up confirmation were categorized into four subgroups (20 hepatocellular carcinomas [HCCs]/5 non-HCC malignancies/4 inflammatory FLLs/8 benign noninflammatory FLLs). The diagnostic value of [18F]FAPI for detecting liver malignancy was determined by visual evaluation. Differences in the maximum standardized uptake value (SUVmax) and lesion-to-background ratio (LBR) obtained from [18F]FAPI PET/CT among the four subgroups were analyzed by semiquantitative analysis. RESULTS: Among the thirty-seven enrolled participants (34 males; median age 57 years, range 48-67 years), on visual evaluation, the sensitivity, specificity, and accuracy of [18F]FAPI PET for detecting liver malignancy in the patient-based analysis were 96.0% (24/25), 58.3% (7/12), and 83.8% (31/37), respectively. On semiquantitative analysis, the SUVmax and LBR of [18F]FAPI PET in liver malignancy (33 HCC lesions; 19 non-HCC malignant lesions) were significantly higher than those in 11 benign noninflammatory FLLs [HCC: SUVmax: 6.4 vs. 4.5, P = 0.017; LBR: 5.1 vs. 1.5, P = 0.003; non-HCC: SUVmax: 5.5 vs. 4.5, P = 0.008; LBR: 4.4 vs. 1.5, P = 0.042]. Notably, there was no significant difference in the SUVmax of [18F]FAPI PET between 33 HCC lesions and 17 inflammatory FLLs (6.4 vs. 8.2, P = 0.37), but the LBR of [18F]FAPI PET in HCC were significantly lower than that in inflammatory FLLs (5.1 vs. 9.1, P = 0.003). CONCLUSIONS: [18F]FAPI PET/CT shows high sensitivity in detecting HCC and non-HCC malignancy with [18F]FDG non-avidity. [18F]FAPI might be a promising radiopharmaceutical for the differential diagnosis of benign noninflammatory FLLs and liver malignancy with [18F]FDG non-avidity.