Cytotoxic Programming of CD4+ T Cells Is Regulated by Opposing Actions of the Related Transcription Factors Eos and Aiolos.

In contrast to the "helper" activities of most CD4+ T effector subsets, CD4+ cytotoxic T lymphocytes (CD4-CTLs) perform functions normally associated with CD8+ T and NK cells. Specifically, CD4-CTLs secrete cytotoxic molecules and directly target and kill compromised cells in an MHC class II-restricted fashion. The functions of these cells have been described in diverse immunological contexts, including their ability to provide protection during antiviral and antitumor responses, as well as being implicated in autoimmunity. Despite their significance to human health, the complete mechanisms that govern their programming remain unclear. In this article, we identify the Ikaros zinc finger transcription factor Eos (Ikzf4) as a positive regulator of CD4-CTL differentiation during murine immune responses against influenza virus infection. We find that the frequency of Eos+ cells is elevated in lung CD4-CTL populations and that the cytotoxic gene program is compromised in Eos-deficient CD4+ T cells. Consequently, we observe a reduced frequency and number of lung-residing, influenza virus-responsive CD4-CTLs in the absence of Eos. Mechanistically, we determine that this is due, at least in part, to reduced expression of IL-2 and IL-15 cytokine receptor subunits on the surface of Eos-deficient CD4+ T cells, both of which support the CD4-CTL program. Finally, we find that Aiolos, a related Ikaros family member and known CD4-CTL antagonist, represses Eos expression by antagonizing STAT5-dependent activation of the Ikzf4 promoter. Collectively, our findings reveal a mechanism wherein Eos and Aiolos act in opposition to regulate cytotoxic programming of CD4+ T cells.

Weakened APC/C activity at mitotic exit drives cancer vulnerability to KIF18A inhibition.

The efficacy of current antimitotic cancer drugs is limited by toxicity in highly proliferative healthy tissues. A cancer-specific dependency on the microtubule motor protein KIF18A therefore makes it an attractive therapeutic target. Not all cancers require KIF18A, however, and the determinants underlying this distinction remain unclear. Here, we show that KIF18A inhibition drives a modest and widespread increase in spindle assembly checkpoint (SAC) signaling from kinetochores which can result in lethal mitotic delays. Whether cells arrest in mitosis depends on the robustness of the metaphase-to-anaphase transition, and cells predisposed with weak basal anaphase-promoting complex/cyclosome (APC/C) activity and/or persistent SAC signaling through metaphase are uniquely sensitive to KIF18A inhibition. KIF18A-dependent cancer cells exhibit hallmarks of this SAC:APC/C imbalance, including a long metaphase-to-anaphase transition, and slow mitosis overall. Together, our data reveal vulnerabilities in the cell division apparatus of cancer cells that can be exploited for therapeutic benefit.

Provider Perceptions Regarding Cardiopulmonary Resuscitation in Surgical Patients with Frailty.

OBJECTIVE: To characterize the perceptions of surgeons, anesthesiologists, and geriatricians regarding perioperative CPR in surgical patients with frailty. SUMMARY BACKGROUND DATA: The population of patients undergoing surgery is growing older and more frail. Despite a growing focus on goal-concordant care, frailty assessment, and debate regarding the appropriateness of cardiopulmonary resuscitation (CPR) in patients with frailty, providers' views regarding frailty and perioperative CPR are unknown. METHODS: We performed qualitative thematic analysis of transcripts from semi-structured interviews of anesthesiologists (8), surgeons (10), and geriatricians (9) who care for high-risk surgical patients at two academic medical centers in Boston, MA. The interview guide elicited clinicians' understanding of frailty, approach to decision-making regarding perioperative CPR, and perceptions of perioperative CPR in frail surgical patients. RESULTS: We identified 5 themes: perceptions of perioperative CPR in patients with frailty vary by provider specialty; judgments regarding appropriateness of CPR in surgical patients with frailty are typically multifactorial and include patient goals, age, comorbidities, and arrest etiology; resuscitation in patients with frailty is sometimes associated with moral distress; biases such as ableism and ageism may skew clinicians' perceptions of appropriateness of perioperative CPR in patients with frailty; and evidence to guide risk stratification for patients with frailty undergoing perioperative CPR is inadequate. CONCLUSIONS: Anesthesiologists, surgeons, and geriatricians offer different accounts of frailty's relevance to judgments regarding CPR in surgical patients. Divergent views regarding frailty and perioperative CPR may impede efforts to deliver goal-concordant care and suggest a need for research to inform risk stratification, predict patient-centered outcomes, and understand the role of potential biases such as ageism and ableism.

[Unusual gluteal localization of unicentric Castleman's disease: A case report and review of the literature]. / Localisation intramusculaire d'une maladie de Castleman unicentrique : cas clinique et revue de la littérature.

BACKGROUND: Castleman's disease is a rare and benign lymphoproliferative disorder which can be unicentric (UCD) or multicentric (MCD). UCD usually involves a single lymph node or less frequently a group of lymph nodes. The most common sites of nodal UCD presentation are the mediastinum, neck, abdomen and retroperitoneum. Rarely extranodal involvement has been reported. The intramuscular location is very unusual with only about 10 cases described in medical literature so far. CASE REPORT: We present a case of atypical localization of Castleman's disease occurring in the right gluteal area in a 40-years-old female patient. The patient was asymptomatic and clinical examination was unremarkable except for a right gluteal palpable mass. The CT scanner-guided needle core biopsy was inconclusive. A surgical excision was then performed that revealed a hyaline-vascular type of Castleman's disease. The patient has an uneventful post-operative course. CONCLUSION: The present case is instructive in the work-up of primary soft tissue tumors, for which Castleman's disease is extremely rare and not considered in the differential diagnosis of clinicians. Pathologists must be aware of its existence so that it can be evoked in the presence of a lymphoid population on histological examination.

BAP1 promotes osteoclast function by metabolic reprogramming.

Treatment of osteoporosis commonly diminishes osteoclast number which suppresses bone formation thus compromising fracture prevention. Bone formation is not suppressed, however, when bone degradation is reduced by retarding osteoclast functional resorptive capacity, rather than differentiation. We find deletion of deubiquitinase, BRCA1-associated protein 1 (Bap1), in myeloid cells (Bap1∆LysM), arrests osteoclast function but not formation. Bap1∆LysM osteoclasts fail to organize their cytoskeleton which is essential for bone degradation consequently increasing bone mass in both male and female mice. The deubiquitinase activity of BAP1 modifies osteoclast function by metabolic reprogramming. Bap1 deficient osteoclast upregulate the cystine transporter, Slc7a11, by enhanced H2Aub occupancy of its promoter. SLC7A11 controls cellular reactive oxygen species levels and redirects the mitochondrial metabolites away from the tricarboxylic acid cycle, both being necessary for osteoclast function. Thus, in osteoclasts BAP1 appears to regulate the epigenetic-metabolic axis and is a potential target to reduce bone degradation while maintaining osteogenesis in osteoporotic patients.

Causes of Death in Cutaneous T-Cell Lymphoma Patients.

BACKGROUND: The advancing evolution toward a Th2 immune environment confers a progressive immunosuppression in patients with longstanding cutaneous T-cell lymphoma (CTCL). The conjunction of the disease-related immunosuppression as well as the immunosuppressive character of some CTCL treatments increase the risk of infectious and neoplastic diseases, sometimes with fatal outcomes. OBJECTIVES: The aim of the study was to prospectively study the causes of death in a cohort of CTCL patients, in a tertiary university skin cancer center. METHODS: All CTCL patients who died between 2008 and 2020 were included. The cause of the death was classified as directly or indirectly related or unrelated to CTCL. RESULTS: Over the study period, 31 (13F/18m) patients with CTCL died (mean age: 75.2 years), mean delay between diagnosis and death: 3.2 years (min: 1, max: 12 years), 58.1% of death causes were classified as indirect (infection), 12.9% directly related (blastic transformation), 22.5% unrelated, and 6.5% of unknown cause. 51.6% of mycosis fungoides (MF) patients who died had early-stage disease (1A-2A) or were on remission. 45.2% of dead patients had advanced-stage MF (2B-4B). Mean CRP level is increased in patients who died from infection whereas LDH level increased in patients with blastosis. A tertiary center is expected to manage of a higher proportion of CTCL patients with advanced-stage disease. CONCLUSIONS: As infection represented more than 50% of the causes of death in CTCL patients, particular attention should be given to preventive measures such as anti-infective vaccination. Regular surveillance of CRP and LDH levels could be helpful for follow-up of MF patients, respectively, with regards to infection and blastosis.

Deletion of Glycogen Synthase Kinase 3 Beta Reprograms NK Cell Metabolism.

Loss of cytotoxicity and defective metabolism are linked to glycogen synthase kinase 3 beta (GSK3ß) overexpression in natural killer (NK) cells from patients with acute myeloid leukemia or from healthy donors after expansion ex vivo with IL-15. Drug inhibition of GSK3ß in these NK cells improves their maturation and cytotoxic activity, but the mechanisms of GSK3ß-mediated dysfunction have not been well studied. Here, we show that expansion of NK cells with feeder cells expressing membrane-bound IL-21 maintained normal GSK3ß levels, allowing us to study GSK3ß function using CRISPR gene editing. We deleted GSK3B and expanded paired-donor knockout and wild-type (WT) NK cells and then assessed transcriptional and functional alterations induced by loss of GSK3ß. Surprisingly, our data showed that deletion of GSK3B did not alter cytotoxicity, cytokine production, or maturation (as determined by CD57 expression). However, GSK3B-KO cells demonstrated significant changes in expression of genes related to rRNA processing, cell proliferation, and metabolic function, suggesting possible metabolic reprogramming. Next, we found that key genes downregulated in GSK3B-KO NK cells were upregulated in GSK3ß-overexpressing NK cells from AML patients, confirming this correlation in a clinical setting. Lastly, we measured cellular energetics and observed that GSK3B-KO NK cells exhibited 150% higher spare respiratory capacity, a marker of metabolic fitness. These findings suggest a role for GSK3ß in regulating NK cell metabolism.

RAB18 is a key regulator of GalNAc-conjugated siRNA-induced silencing in Hep3B cells.

Small interfering RNA (siRNA) therapeutics have developed rapidly in recent years, despite the challenges associated with delivery of large, highly charged nucleic acids. Delivery of siRNA therapeutics to the liver has been established, with conjugation of siRNA to N-acetylgalactosamine (GalNAc) providing durable gene knockdown in hepatocytes following subcutaneous injection. GalNAc binds the asialoglycoprotein receptor (ASGPR) that is highly expressed on hepatocytes and exploits this scavenger receptor to deliver siRNA across the plasma membrane by endocytosis. However, siRNA needs to access the RNA-induced silencing complex (RISC) in the cytoplasm to provide effective gene knockdown, and the entire siRNA delivery process is very inefficient, likely because of steps required for endosomal escape, intracellular trafficking, and stability of siRNA. To reveal the cellular factors limiting delivery of siRNA therapeutics, we performed a genome-wide pooled knockout screen on the basis of delivery of GalNAc-conjugated siRNA targeting the HPRT1 gene in the human hepatocellular carcinoma line Hep3B. Our primary genome-wide pooled knockout screen identified candidate genes that when knocked out significantly enhanced siRNA efficacy in Hep3B cells. Follow-up studies indicate that knockout of RAB18 improved the efficacy of siRNA delivered by GalNAc, cholesterol, or antibodies, but not siRNA delivered by Lipofectamine transfection, suggesting a role for RAB18 in siRNA delivery and intracellular trafficking.

Novel Selective Galectin-3 Antagonists Are Cytotoxic to Acute Lymphoblastic Leukemia.

Galectin-3 is a ß-galactoside-specific, carbohydrate-recognizing protein (lectin) that is strongly implicated in cancer development, metastasis, and drug resistance. Galectin-3 promotes migration and ability to withstand drug treatment of B-cell precursor acute lymphoblastic leukemia (BCP-ALL) cells. Due to high amino acid conservation among galectins and the shallow nature of their glycan-binding site, the design of selective potent antagonists targeting galectin-3 is challenging. Herein, we report the design and synthesis of novel taloside-based antagonists of galectin-3 with enhanced affinity and selectivity. The molecules were optimized by in silico docking, selectivity was established against four galectins, and the binding modes were confirmed by elucidation of X-ray crystal structures. Critically, the specific inhibition of galectin-3-induced BCP-ALL cell agglutination was demonstrated. The compounds decreased the viability of ALL cells even when grown in the presence of protective stromal cells. We conclude that these compounds are promising leads for therapeutics, targeting the tumor-supportive activities of galectin-3 in cancer.

The Relevance of Skin Biopsies in General Internal Medicine: Facts and Myths.

INTRODUCTION: Non-dermatology medical specialties may refer patients for skin biopsies, searching for a particular diagnosis. However, the diagnostic impact of the skin biopsy is not clearly established. This article aims to assess the indications for, and evaluate the clinical relevance of, skin biopsies in non-dermatology medical specialties. METHODS: A questionnaire was sent to 23 non-dermatology specialty departments in a university medical center, requesting a list of indications for skin biopsies, as well as to 10 staff dermatologists to collect the indications of skin biopsies requested by non-dermatology specialties. Once the indications were collected, a literature search was performed to evaluate their clinical value and relevance. RESULTS: Eleven non-dermatology specialties provided a list of skin biopsy indications, to which staff dermatologists added seven more indications. A literature search revealed evidence-based medicine data for six diseases, that is, amyloidosis, peripheral autonomic neuropathy, Sneddon's syndrome, intravascular lymphoma, sarcoidosis, and chronic graft-versus-host disease. Results were questionable concerning infectious endocarditis, acute graft-versus-host-disease, and the lupus band test. Skin biopsy were not evidenced as useful for the diagnosis of calciphylaxis, systemic scleroderma, Behçet's disease, or hypermobile Ehlers-Danlos syndrome. For the diagnosis of Alport's syndrome, pseudoxanthoma elasticum, and vascular Ehlers-Danlos syndrome, skin biopsy is currently outperformed by genetic analyses. For diagnoses such as Henoch-Schönlein purpura and Sjögren's syndrome, skin biopsy represents an additional item among other diagnostic criteria. CONCLUSION: The usefulness of skin biopsy as requested by non-dermatology specialties is only evidenced for amyloidosis, peripheral autonomic neuropathy, Sneddon's syndrome, intravascular lymphoma, sarcoidosis, chronic graft-versus-host-disease, Henoch-Schönlein purpura, and Sjögren's syndrome.

Cancer cell-intrinsic resistance to BiTE therapy is mediated by loss of CD58 costimulation and modulation of the extrinsic apoptotic pathway.

BACKGROUND: Bispecific T-cell engager (BiTE) molecules induce redirected lysis of cancer cells by T cells and are an emerging modality for solid tumor immunotherapy. While signs of clinical activity have been demonstrated, efficacy of T-cell engagers (TCEs) in solid tumors settings, molecular determinants of response, and underlying mechanisms of resistance to BiTE therapy require more investigation. METHODS: To uncover cancer cell-intrinsic genetic modifiers of TCE-mediated cytotoxicity, we performed genome-wide CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) loss-of-function and CRISPRa (CRISPR activation) gain-of-function screens using TCEs against two distinct tumor-associated antigens (TAAs). By using in vitro T-cell cytotoxicity assays and in vivo efficacy studies, we validated the roles of two common pathways identified in our screen, T-cell costimulation pathway and apoptosis pathway, as key modifiers of BiTE activity. RESULTS: Our genetic screens uncovered TAAs-independent cancer cell-intrinsic genes with functions in autophagy, T-cell costimulation, the apoptosis pathway, chromatin remodeling, and cytokine signaling that altered responsiveness to BiTE-mediated killing. Notably, loss of CD58 (the ligand of the CD2 T-cell costimulatory receptor), a gene frequently altered in cancer, led to decreased TCE-mediated cytotoxicity, T-cell activation and antitumor efficacy in vitro and in vivo. Moreover, the effects of CD58 loss were synergistically compounded by concurrent loss of CD80/CD86 (ligands for the CD28 T-cell costimulatory receptor), whereas joint CD2 and CD28 costimulation additively enhanced TCE-mediated killing, indicating non-redundant costimulatory mechanisms between the two pathways. Additionally, loss of CFLAR (Caspase-8 and FADD Like Apoptosis Regulator), BCL2L1, and BID (BH3 Interacting Domain Death Agonist) induced profound changes in sensitivity to TCEs, indicating that key regulators of apoptosis, which are frequently altered in cancer, impact tumor responsiveness to BiTE therapy. CONCLUSIONS: This study demonstrates that genetic alterations central to carcinogenesis and commonly detected in cancer samples lead to significant modulation of BiTE antitumor activity in vitro and in vivo, findings with relevance for a better understanding of patient responses to BiTE therapy and novel combinations that enhance TCE efficacy.

Gene Regulatory Circuits in Innate and Adaptive Immune Cells.

Cell identity and function largely rely on the programming of transcriptomes during development and differentiation. Signature gene expression programs are orchestrated by regulatory circuits consisting of cis-acting promoters and enhancers, which respond to a plethora of cues via the action of transcription factors. In turn, transcription factors direct epigenetic modifications to revise chromatin landscapes, and drive contacts between distal promoter-enhancer combinations. In immune cells, regulatory circuits for effector genes are especially complex and flexible, utilizing distinct sets of transcription factors and enhancers, depending on the cues each cell type receives during an infection, after sensing cellular damage, or upon encountering a tumor. Here, we review major players in the coordination of gene regulatory programs within innate and adaptive immune cells, as well as integrative omics approaches that can be leveraged to decipher their underlying circuitry.

Re-amputation and survival following toe amputation: outcome data from a tertiary referral centre.

PURPOSE: Toe amputation is a commonly performed procedure for irreversible foot sepsis. However, outcome and predictors of outcome are poorly understood. Our aim was to determine survival and rate of progression to further amputation following index toe amputation. METHODS: Consecutive patients between 2010 and 2015 were included. Progression to further minor amputation, major amputation or death was recorded. Multivariable Cox regression analyses were undertaken to determine independent predictors of outcome and survival. RESULTS: One hundred forty-six patients were included, with mean age of 65 years. Fifty-five (37.7%) patients underwent hallux amputation, while 91 (62.3%) underwent amputation of non-hallux digit(s). Following index toe amputation, 63 (43.2%) patients progressed to further minor or major ipsilateral amputation, median time to which was 36 months. Twenty-one patients (14.4%) progressed to major ipsilateral amputation. Patients undergoing index non-hallux amputation were significantly more likely to require further minor amputation (P = 0.050); however, the rate of major amputation between hallux (14.5%) and non-hallux (14.3%) groups was similar. Overall, 5-year ipsilateral amputation-free (iAFS) was 39.6 ± 4.1%, ipsilateral major amputation-free (iMAFS) was 55.9 ± 4.1% and overall survival (OS) was 64.3 ± 4.0% and did not differ between index amputation sites. CONCLUSION: Almost half of patients undergoing toe amputation required further digital amputation. However, limb preservation rates are high, and a majority of patients are alive at 5-year follow-up. There was no significant difference in outcome between patients undergoing hallux and non-hallux primary procedures. Overall, increasing age remains the only independent predictor of iMAFS and OS.

The Utility of Preoperative HbA1c as a Standardized Protocol in Elective Carpal Tunnel Release: A Retrospective Review of Clinical Outcomes.

Background: In an effort to reduce surgical complications, some institutions have implemented universal hemoglobin A1c (HbA1c) screening for all preoperative patients. However, the value of HbA1c screening for predicting clinically meaningful complications after elective carpal tunnel release (CTR) remains unclear. The purpose of this study was to investigate the clinically meaningful predicative value of HbA1c screening on postoperative complications following elective CTR. Methods: A retrospective cohort study of 790 patients who underwent CTR was performed. All patients had an HbA1c screening performed, regardless of whether they underwent the diagnosis for diabetes or not. Primary outcomes were overall complication rate, rates of major complications (readmission or reoperation), and rates of minor complications (surgical site infection and wound dehiscence). Patients were stratified into 3 groups based on HbA1c: HbA1c <7, HbA1c 7-8, and HbA1c >8. Results: The overall complication rate for all groups was 4.8%. Rates of major complications were 0.4% for readmission and 0.1% for reoperation. For minor complications, the odds ratio (OR) for the HbA1c 7-8 group was 0.6 (95% confidence interval [CI], 0.14-1.77), and for the HbA1c >8 group, the OR was 1.6 (95% CI, 0.66-3.60). All minor complications resolved with outpatient treatment. There were no statistically significant differences between the groups for any comparisons. Conclusions: Elective CTR has a low complication rate. Routine preoperative screening of HbA1c is of little value in predicting clinically meaningful complications.

A Comprehensive Update of the Atypical, Rare and Mimicking Presentations of Mycosis Fungoides.

INTRODUCTION: Mycosis fungoides (MF) is the most frequent subtype of primary cutaneous T cell lymphomas (pCTCL). The diagnosis may be particularly difficult in the early stages as well as in atypical and rare clinical presentations. Furthermore, MF may simulate a large variety of common dermatologic disorders and patterns, both histopathologically and clinically. METHODS: A literature search was performed to provide a comprehensive update on the rare and atypical MF manifestations as well as the dermatoses and dermatological patterns that could be imitated by MF. RESULTS: A total of 114 publications were found describing a series of different dermatoses and dermatological patterns mimicked by MF, as well as some particular localizations of MF lesions and dermatoses that occur in preexisting MF lesions. CONCLUSIONS: The number of dermatoses that can be imitated by MF is ever-increasing. Patients with common dermatologic conditions that prove to be treatment refractory should be biopsied without delay, and sequentially as necessary, to prevent delay in diagnosis and progression of disease. Clinicopathologic correlation is the best way of diagnosis.

Necrobiosis Lipoidica following Breast Reduction.

Necrobiosis lipoidica (NL) is a rare granulomatous skin disorder of unknown physiopathology that is frequently associated with diabetes mellitus. The typical skin lesions of NL present as chronic, bilateral, well-defined red to yellow-brown plaques with telangectasias, a violaceous border and a waxy atrophic center. The lower legs are the most often involved areas, but NL may exceptionally develop on scar tissue following surgery. The treatment is very challenging and notoriously difficult. We report a 60-year-old diabetic woman who developed NL all along the surgical scars following breast reduction, without presenting NL on the lower legs. NL should be considered among the rare but possible skin healing complications of surgery.