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Abstract Mutations of receptor tyrosine kinases (RTKs) are associated with the development of many cancers by modifying receptor signaling and contributing to drug resistance in clinical settings. We present enhanced bystander bioluminescence resonance energy transfer-based biosensors providing new insights into RTK biology and pharmacology critical for the development of more effective RTK-targeting drugs. Distinct SH2-specific effector biosensors allow for real-time and spatiotemporal monitoring of signal transduction pathways engaged upon RTK activation. Using EGFR as a model, we demonstrate the capacity of these biosensors to differentiate unique signaling signatures, with EGF and Epiregulin ligands displaying differences in efficacy, potency, and responses within different cellular compartments. We further demonstrate that EGFR single point mutations found in Glioblastoma or non-small cell lung cancer, impact the constitutive activity of EGFR and response to tyrosine kinase inhibitor. The BRET-based biosensors are compatible with microscopy, and more importantly characterize the next generation of therapeutics directed against RTKs.

Partial prostatectomy has been described as an alternative to focal ablation therapy for the management of localized low- to intermediate-risk prostate cancer. This report aims to describe the long-term outcomes in a series of 28 men (2000–2022) who underwent robotic-assisted anterior partial prostatectomy (APP) for anteriorly located tumors entirely or partially within the anterior fibromuscular stroma. The median follow-up is 7 yr (interquartile range [IQR]: 4.2–8). The median prostate-specific antigen (PSA) before APP was 9.6 (6–11). Continence remained uninterrupted in 92% of patients. Erectile function without drug remained uninterrupted in 69%. The median nadir PSA after APP was 0.36 ng/ml (IQR: 0.25–0.60). Cancer recurrence at biopsies at the margins of the primary cancer resected area in case of a PSA elevation was observed in eight patients and led to salvage completion robotic radical prostatectomy at a median time of 3.25 yr (IQR: 2.4–6). Freedom from post-APP cancer recurrence at 7 yr was 62.7% (35.0–81.3%). Pre-APP tumor volume at magnetic resonance imaging (MRI) and volume of grade 4/5 were predictive of recurrence. Freedom from biochemical recurrence after completion radical prostatectomy at 7 yr was 94.7% (68.1–99.3%). All 28 patients are alive. No one had systemic treatment or metastases. These results confirm our initial report of robotic APP with good functional results and acceptable oncological results. The use of the inclusion criteria of pre-APP tumor volume at MRI

Background: The risk of prostate cancer metastatic is correlated with its volume and grade. These parameters are now best estimated preoperatively with magnetic resonance imaging (MRI) and MRI-guided biopsy. Objective: To estimate the risk of metastatic recurrence after radical prostatectomy (RP) in our model versus conventional clinical European Association of Urology (EAU) classification. The secondary objective is biochemical recurrence (BCR). Design, setting, and participants: A retrospective study was conducted of a cohort of 713 patients having undergone MRI-guided biopsies and RP between 2009 and 2018. The preoperative variables included prostate-specific antigen, cT stage, tumor volume (TV) based on the lesion’s largest diameter at MRI, percentage of Gleason pattern 4/5 (%GP4/5) at MRI-guided biopsy, and volume of GP4/5 (VolGP4/5) calculated as TV × %GP4/5. Outcome measurements and statistical analysis: The variables’ ability to predict recurrence was determined in univariable and multivariable Fine-and-Gray models, according to the Akaike information criterion (AIC) and Harrell’s C-index. Results and limitations: Overall, 176 (25%), 430 (60%), and 107 (15%) patients had low, intermediate, and high-risk disease, respectively, according to the EAU classification. During a median follow-up period of 57 mo, metastatic recurrence was observed in 48 patients with a 5-yr probability of 5.6% (95% confidence interval [CI] 3.9–7.7). VolGP4/5 (categories: 3.2 ml) was the parameter with the lowest AIC and the highest C-index for metastatic recurrence of 0.82 (95% CI 0.76–0.88), and for BCR it was 0.73 (95% CI 0.68–0.78). In a multivariable model that included %GP4/5 and TV, C-index values were 0.86 (95% CI 0.79–0.91) for metastatic recurrence and 0.77 (0.72–0.82) for BCR. The same results for EAU classification were 0.74 (0.67–0.80) and 0.67 (0.63–0.72), respectively. Limitations are related to short follow-up and expertise of radiologists and urologists. Conclusions: We developed a preoperative risk tool integrating the VolGP4/5 based on MRI and MRI-guided biopsies to predict metastatic recurrence after RP. Our model showed higher accuracy than conventional clinical risk models. These findings might enable physicians to provide more personalized patient care. Patient summary: Aggressiveness of prostate cancer evaluated before treatment by incorporating magnetic resonance imaging (MRI) and MRI-guided biopsy results gives a better estimate of the risk of metastatic recurrence than previous parameters not based on MRI.

The recognition that individual GPCRs can activate multiple signaling pathways has raised the possibility of developing drugs selectively targeting therapeutically relevant ones. This requires tools to determine which G proteins and βarrestins are activated by a given receptor. Here, we present a set of BRET sensors monitoring the activation of the 12 G protein subtypes based on the translocation of their effectors to the plasma membrane (EMTA). Unlike most of the existing detection systems, EMTA does not require modification of receptors or G proteins (except for Gs). EMTA was found to be suitable for the detection of constitutive activity, inverse agonism, biased signaling and polypharmacology. Profiling of 100 therapeutically relevant human GPCRs resulted in 1500 pathway-specific concentration-response curves and revealed a great diversity of coupling profiles ranging from exquisite selectivity to broad promiscuity. Overall, this work describes unique resources for studying the complexities underlying GPCR signaling and pharmacology.

Abstract Glycogen storage disease type Ia (GSD Ia) is a rare metabolic disease due to glucose‐6‐phosphatase deficiency. Chronic kidney disease is a frequent complication that may manifest itself by glomerular lesions and tubular dysfunction from the second decade of life. We report two young GSDIa patients with malignant renal tumor. The first patient was a 25‐year‐old man. He had chronic metabolic imbalance without kidney involvement. The tumor, a type 2 papillary renal carcinoma, was accidentally discovered during follow‐up. The second patient was a 27‐year‐old woman with chronic metabolic imbalance and chronic kidney involvement. The tumor, a grade 2 papillary carcinoma, was accidentally discovered during follow‐up. These two observations are, to date, the first to be reported. We suggest that annual monitoring of kidney imaging in GSDI patients should be systematic to detect renal cancer, from the second decade of life.

Objective: Recently, we observed that the specialized proresolving mediator (SPM) entity resolvin D1 activates lipoxin A4/formyl peptide receptor 2 (ALX/FPR2), which facilitates cardiac healing and persistent inflammation is a hallmark of impaired cardiac repair in aging. Splenic leukocyte-directed SPMs are essential for the safe clearance of inflammation and cardiac repair after injury; however, the target of SPMs remains undefined in cardiac healing and repair. Methods: To define the mechanistic basis of ALX/FPR2 as a resolvin D1 target, ALX/FPR2-null mice were examined extensively. The systolic-diastolic heart function was assessed using echocardiography, leukocytes were phenotyped using flow cytometry, and SPMs were quantitated using mass spectrometry. The presence of cardiorenal syndrome was validated using histology and renal markers. Results: Lack of ALX/FPR2 led to the development of spontaneous obesity and diastolic dysfunction with reduced survival with aging. After cardiac injury, ALX/FPR2−/− mice showed lower expression of lipoxygenases (−5, −12, −15) and a reduction in SPMs in the infarcted left ventricle and spleen, indicating nonresolving inflammation. Reduced SPM levels in the infarcted heart and spleen are suggestive of impaired cross-talk between the injured heart and splenic leukocytes, which are required for the resolution of inflammation. In contrast, cyclooxygenases (−1 and −2) were over amplified in the infarcted heart. Together, these results suggest interorgan signaling in which the spleen acts as both an SPM biosynthesizer and supplier in acute heart failure. ALX/FPR2 dysfunction magnified obesogenic cardiomyopathy and renal inflammation (↑NGAL, ↑TNF-α, ↑CCL2, ↑IL-1β) with elevated plasma creatinine levels in aging mice. At the cellular level, ALX/FPR2−/− mice showed impairment of macrophage phagocytic function ex-vivo with expansion of neutrophils after myocardial infarction. Conclusions: Lack of ALX/FPR2 induced obesity, reduced the life span, amplified leukocyte dysfunction, and facilitated profound interorgan nonresolving inflammation. Our study shows the integrative and indispensable role of ALX/FPR2 in lipid metabolism, cardiac inflammation–resolution processes, obesogenic aging, and renal homeostasis. Keywords: Inflammation, Leukocytes, Myocardial infarction, Nonresolving inflammation, Obesogenic aging, Resolution of inflammation

Summary: Esthesioneuroblastoma (ENB) is a rare cancer of the olfactory mucosa, with no established molecular stratification to date. We report similarities of ENB with tumors arising in the neural crest and perform integrative analysis of these tumors. We propose a molecular-based subtype classification of ENB as basal or neural, both of which have distinct pathological, transcriptomic, proteomic, and immune features. Among the basal subtype, we uncovered an IDH2 R172 mutant-enriched subgroup (∼35%) harboring a CpG island methylator phenotype reminiscent of IDH2 mutant gliomas. Compared with the basal ENB methylome, the neural ENB methylome shows genome-wide reprogramming with loss of DNA methylation at the enhancers of axonal guidance genes. Our study reveals insights into the molecular pathogenesis of ENB and provides classification information of potential therapeutic relevance. : Classe et al. report an integrative multi-omics analysis of esthesioneuroblastomas (ENBs) and identify two subgroups of ENBs: neural-like and basal-like. These subgroups are linked to cell ontogeny and are associated with distinct clinicopathological features and patient outcomes. Notably, one-third of basal ENBs harbor an IDH2 R172 mutation with a CpG island methylator phenotype. Keywords: esthesioneuroblastomas, IDH2, neural, basal, DNA methylation, sequencing, genetics, epigenetics, survival, ki67

Renal cell carcinoma, the most common neoplasm of adult kidney, accounts for about 3% of adult malignancies and is usually highly resistant to conventional therapy. MicroRNAs are a class of small non-coding RNAs, which have been previously shown to promote malignant initiation and progression. In this study, we focused our attention on miR-21, a well described oncomiR commonly upregulated in cancer. Using a cohort of 99 primary renal cell carcinoma samples, we showed that miR-21 expression in cancer tissues was higher than in adjacent non-tumor tissues whereas no significant difference was observed with stages, grades, and metastatic outcome. In vitro, miR-21 was also overexpressed in renal carcinoma cell lines compared to HK-2 human proximal tubule epithelial cell line. Moreover, using Boyden chambers and western blot techniques, we also showed that miR-21 overexpression increased migratory, invasive, proliferative, and anti-apoptotic signaling pathways whereas opposite results were observed using an anti-miR-21-based silencing strategy. Finally, we assessed the role of miR-21 in mediating renal cell carcinoma chemoresistance and further showed that miR-21 silencing significantly (1) increased chemosensitivity of paclitaxel, 5-fluorouracil, oxaliplatin, and dovitinib; (2) decreased expression of multi-drug resistance genes; and (4) increased SLC22A1/OCT1, SLC22A2/OCT2, and SLC31A1/CTR1 platinum influx transporter expression. In conclusion, our results showed that miR-21 is a key actor of renal cancer progression and plays an important role in the resistance to chemotherapeutic drugs. In renal cell carcinoma, targeting miR-21 is a potential new therapeutic strategy to improve chemotherapy efficacy and consequently patient outcome.

Renal proximal tubular epithelial cells play a central role in renal physiology and are among the cell types most sensitive to ischemia and xenobiotic nephrotoxicity. In order to investigate the molecular and cellular mechanisms underlying the pathophysiology of kidney injuries, a stable and well-characterized primary culture model of proximal tubular cells is required. An existing model of proximal tubular cells is hampered by the cellular heterogeneity of kidney; a method based on cell sorting for specific markers must therefore be developed. In this study, we present a primary culture model based on the mechanical and enzymatic dissociation of healthy tissue obtained from nephrectomy specimens. Renal epithelial cells were sorted using co-labeling for CD10 and CD13, two renal proximal tubular epithelial markers, by flow cytometry. Their purity, phenotypic stability and functional properties were evaluated over several passages. Our results demonstrate that CD10/CD13 double-positive cells constitute a pure, functional and stable proximal tubular epithelial cell population that displays proximal tubule markers and epithelial characteristics over the long term, whereas cells positive for either CD10 or CD13 alone appear to be heterogeneous. In conclusion, this study describes a method for establishing a robust renal proximal tubular epithelial cell model suitable for further experimentation.

Abstract  This article describes the development and formal verification (proof of semantic preservation) of a compiler back-end from Cminor (a simple imperative intermediate language) to PowerPC assembly code, using the Coq proof assistant both for programming the compiler and for proving its soundness. Such a verified compiler is useful in the context of formal methods applied to the certification of critical software: the verification of the compiler guarantees that the safety properties proved on the source code hold for the executable compiled code as well. [ABSTRACT FROM AUTHOR]

Abstract  This article presents the formal semantics of a large subset of the C language called Clight. Clight includes pointer arithmetic, struct and union types, C loops and structured switch statements. Clight is the source language of the CompCert verified compiler. The formal semantics of Clight is a big-step operational semantics that observes both terminating and diverging executions and produces traces of input/output events. The formal semantics of Clight is mechanized using the Coq proof assistant. In addition to the semantics of Clight, this article describes its integration in the CompCert verified compiler and several ways by which the semantics was validated. [ABSTRACT FROM AUTHOR]

Abstract  This paper formalizes and proves correct a compilation scheme for mutually-recursive definitions in call-by-value functional languages. This scheme supports a wider range of recursive definitions than previous methods. We formalize our technique as a translation scheme to a lambda-calculus featuring in-place update of memory blocks, and prove the translation to be correct. [ABSTRACT FROM AUTHOR]

Abstract  Ezrin is a cytoskeleton linker protein that is actively involved in the metastatic process of cancer cells. We have recently reported that ezrin expression in conventional osteosarcoma was an independent prognostic factor for event-free survival and overall survival. In this work, ezrin expression was found in all histological subtypes. Especially cartilaginous areas in chondroblastic osteosarcomas were immunopositive for ezrin. We wanted to know if ezrin could be a useful diagnostic marker in bone pathology. We have searched for ezrin expression in 208 cartilaginous tumours by immunohistochemistry and in 16 chondroblastic osteosarcomas. All conventional chondrosarcomas, whatever their grade, were negative, while ten of 16 chondroblastic osteosarcomas were positive. In contrast, dedifferentiated (five of 14) and mesenchymal chondrosarcomas (five of ten) showed ezrin positivity. Some chondroblastomas and more rarely chondromyxoid fibromas also exhibited ezrin expression. These data suggest that ezrin is a useful immunohistochemical marker for differential diagnosis between chondroblastic osteosarcomas and conventional chondrosarcomas with a specificity of 100%. Ezrin expression in dedifferentiated and mesenchymal chondrosarcomas which are aggressive neoplasms resistant to conventional treatment means that ezrin could be a therapeutic target. Ezrin expression in chondroblastomas is more intriguing and requires further study to assess prognostic value. [ABSTRACT FROM AUTHOR]

Abstract  The helicase-like transcription factor (HLTF) belongs to the SWI/SNF family of proteins that use the energy from adenosine triphosphate hydrolysis to remodel chromatin during a variety of cellular processes. HLTF is also involved in DNA repair. Using computer-assisted microscopy, the immunohistochemical expression of HLTF was determined using a series of 100 hypopharyngeal and 56 laryngeal squamous cell carcinomas (SCCs) compared to tumor-free epithelia (60 cases) and dysplasias (92 cases). In hypopharyngeal SCC tumor progression, increased HLTF expression was associated with the percentage of immunopositive epithelial tissue areas (p = 0.02) and the staining intensity of the positive area (p = 0.0005). In the cases of laryngeal lesions, the immunolabeling intensity of HLTF significantly decreased with malignancy (p = 0.01). We also observed a significant shift of HLTF expression from the cytoplasm toward the nuclear compartment (p = 0.0007). Our data reveal an association between the presence of HLTF and neoplastic progression of hypopharyngeal and laryngeal SCCs. [ABSTRACT FROM AUTHOR]

Abstract  This article presents the formal verification, using the Coq proof assistant, of a memory model for low-level imperative languages such as C and compiler intermediate languages. Beyond giving semantics to pointer-based programs, this model supports reasoning over transformations of such programs. We show how the properties of the memory model are used to prove semantic preservation for three passes of the Compcert verified compiler. [ABSTRACT FROM AUTHOR]

Abstract  This article describes the formal verification of a compilation algorithm that transforms parallel moves (parallel assignments between variables) into a semantically-equivalent sequence of elementary moves. Two different specifications of the algorithm are given: an inductive specification and a functional one, each with its correctness proofs. A functional program can then be extracted and integrated in the Compcert verified compiler. [ABSTRACT FROM AUTHOR]

pT1G3 bladder tumors have a high tendency to recur and progress. We evaluated the prognostic values of the depth of submucosal invasion and immunostaining with survivin and p53 in 30 pT1G3 urothelial carcinomas at the first endoscopic resection. The depth of invasion was evaluated toward the muscularis mucosa and measured using a micrometer. Survivin and p53 immunostaining were performed using an automated immunostainer. Of the patients, 19 (63%) had tumor recurrence, 11 (37%) had tumor progression, 10 (33%) had metastatic spread, and 10 (33%) died of the disease. Infiltration of deep lamina propria (pT1b) and a micrometric measure of 1.5 mm or more were associated with an increased risk of tumor local and/or metastatic progression (P = .03 and P = .02, respectively). A combined high expression of survivin (

This issue of the Journal of Functional Programming (JFP)marks a point of transition. After serving since 1991 as Editor, then since 2004 as co-Editor in Chief (along with Greg Morrisett from 2004 to 2006 and Xavier Leroy since 2007), Paul Hudak is stepping down. [ABSTRACT FROM AUTHOR]