Crumbling the Castle: Targeting DNABII Proteins for Collapsing Bacterial Biofilms as a Therapeutic Approach to Treat Disease and Combat Antimicrobial Resistance.

Antimicrobial resistance (AMR) is a concerning global threat that, if not addressed, could lead to increases in morbidity and mortality, coupled with societal and financial burdens. The emergence of AMR bacteria can be attributed, in part, to the decreased development of new antibiotics, increased misuse and overuse of existing antibiotics, and inadequate treatment options for biofilms formed during bacterial infections. Biofilms are complex microbiomes enshrouded in a self-produced extracellular polymeric substance (EPS) that is a primary defense mechanism of the resident microorganisms against antimicrobial agents and the host immune system. In addition to the physical protective EPS barrier, biofilm-resident bacteria exhibit tolerance mechanisms enabling persistence and the establishment of recurrent infections. As current antibiotics and therapeutics are becoming less effective in combating AMR, new innovative technologies are needed to address the growing AMR threat. This perspective article highlights such a product, CMTX-101, a humanized monoclonal antibody that targets a universal component of bacterial biofilms, leading to pathogen-agnostic rapid biofilm collapse and engaging three modes of action-the sensitization of bacteria to antibiotics, host immune enablement, and the suppression of site-specific tissue inflammation. CMTX-101 is a new tool used to enhance the effectiveness of existing, relatively inexpensive first-line antibiotics to fight infections while promoting antimicrobial stewardship.

A propensity score-matched, real-world comparison of ustekinumab vs vedolizumab as a second-line treatment for Crohn's disease. The Cross Pennine study II.

BACKGROUND: The optimal choice of biological agents after failure of anti-tumour-necrosis-factor-(TNF)α agent in Crohn's disease (CD) is yet to be defined. AIMS: To assess the effectiveness and safety of ustekinumab compared to vedolizumab as second-line treatment in CD patients who failed anti-TNFα therapy. METHODS: Retrospective analysis of clinical response and remission at 14 and 52 weeks to ustekinumab by physician global assessment (PGA). A propensity score-matched analysis with a cohort treated with vedolizumab was performed. RESULTS: Of 282 patients (mean age 40 ± 15, F:M ratio 1.7:1) treated with ustekinumab, clinical response or remission was reached by 200/282 patients (70.9%) at 14 weeks, and 162/259 patients (62.5%) at 52 weeks. Overall, 74 adverse events occurred, of which 26 were labelled as serious (8.3 per 100 person-year). After exclusion of patients without prior anti-TNFα exposure and patients previously exposed to vedolizumab or ustekinumab, we analysed 275/282 patients (97.5%) on ustekinumab and 118/135 patients (87.4%) on vedolizumab. Propensity score analysis revealed that at 14 weeks, patients treated with ustekinumab were 38% (95% CI 25%-50%; P < 0.001) more likely to achieve clinical remission, while at 52 weeks, the difference of 9% (95% CI -15% to 33%; P = 0.462) was not significant. CONCLUSIONS: Ustekinumab was effective and well tolerated in this real-world cohort. While ustekinumab proved more effective at 14-weeks, we found no statistically significant differences at 52 weeks compared to vedolizumab.

Standards for the provision of antenatal care for patients with inflammatory bowel disease: guidance endorsed by the British Society of Gastroenterology and the British Maternal and Fetal Medicine Society.

BACKGROUND: Pregnant women with inflammatory bowel disease (IBD) are at increased risk of adverse pregnancy outcomes. Comprehensive guidelines on medical management have been published; yet, there is limited guidance on service set-up and minimum standards of care for pregnant women with IBD. AIM: To develop a position statement on service set-up and minimum standards of care in the UK. METHODS: A working group consisting of 16 gastroenterologists, obstetricians, obstetric physician, IBD specialist nurses and midwives was assembled. Initial draft statements were produced and a modified Delphi process with two rounds of voting applied. Statements were modified according to voters' feedback after each round. Statements with ≥80% agreement were accepted. RESULTS: All 15 statements met criteria for inclusion. To facilitate optimal care, regular and effective communication between IBD and obstetric teams is required. There should be nominated link clinicians for IBD in obstetric units and for pregnancy in IBD units. Preconception counselling should be available for all women with IBD. All pregnant women should be advised on the safety of IBD medication during pregnancy and breast feeding, the optimal mode of delivery, the management of biologics (where applicable) and safety of childhood vaccinations. Regular audit of pregnancy outcomes and documentation of advice given is recommended. CONCLUSION: Position statements have been developed that advise on the importance of joined-up multidisciplinary care, proactive decision-making with clear documentation and communication to the woman and other healthcare practitioners.

IBD in pregnancy: recent advances, practical management.

Inflammatory bowel disease (IBD) poses complex issues in pregnancy, but with high-quality care excellent pregnancy outcomes are achievable. In this article, we review the current evidence and recommendations for pregnant women with IBD and aim to provide guidance for clinicians involved in their care. Many women with IBD have poor knowledge about pregnancy-related issues and a substantial minority remains voluntarily childless. Active IBD is associated with an increased risk of preterm birth, low for gestation weight and fetal loss. With the exception of methotrexate and tofacitinib the risk of a flare outweighs the risk of IBD medication and maintenance of remission from IBD should be the main of care. Most women with IBD will experience a normal pregnancy and can have a vaginal delivery. Active perianal Crohn's disease is an absolute and ileal pouch surgery a relative indication for a caesarean section. Breast feeding is beneficial to the infant and the risk from most IBD medications is negligible.

Impact of the coronavirus infectious disease (COVID-19) pandemic on the provision of inflammatory bowel disease (IBD) antenatal care and outcomes of pregnancies in women with IBD.

BACKGROUND: The impact of COVID-19 on pregnant inflammatory bowel disease (IBD) patients is currently unknown. Reconfiguration of services during the pandemic may negatively affect medical and obstetric care. We aimed to examine the impacts on IBD antenatal care and pregnancy outcomes. METHODS: Retrospective data were recorded in consecutive patients attending for IBD antenatal care including outpatient appointments, infusion unit visits and advice line encounters. RESULTS: We included 244 pregnant women with IBD, of which 75 (30.7%) were on biologics in whom the treatment was stopped in 29.3% at a median 28 weeks gestation. In addition, 9% of patients were on corticosteroids and 21.5% continued on thiopurines. The care provided during 460 patient encounters was not affected by the pandemic in 94.1% but 68.2% were performed via telephone (compared with 3% prepandemic practice; p<0.0001). One-hundred-ten women delivered 111 alive babies (mean 38.2 weeks gestation, mean birth weight 3324 g) with 12 (11.0%) giving birth before week 37. Birth occurred by vaginal delivery in 72 (56.4%) and by caesarean section in 48 (43.6%) cases. Thirty-three were elective (12 for IBD indications) and 15 emergency caesarean sections. Breast feeding rates were low (38.6%). Among 244 pregnant women with IBD, 1 suspected COVID-19 infection was recorded. CONCLUSION: IBD antenatal care adjustments during the COVID-19 pandemic have not negatively affected patient care. Despite high levels of immunosuppression, only a single COVID-19 infection occurred. Adverse pregnancy outcomes were infrequent.

Patients' perceptions of faecal calprotectin testing in inflammatory bowel disease: results from a prospective multicentre patient-based survey.

INTRODUCTION: Despite its success, there appears to be practical issues with Faecal Calprotectin (FC) testing in Inflammatory Bowel Diseases (IBD), including sample collection, delivery and processing delays. Patients' perception and barriers to FC testing are yet to be explored in clinical practice. METHOD: A prospective patient survey was undertaken at IBD units in UK, Europe and Australia. A 9-point patient-based questionnaire was completed in clinic and included demographics, previous FC testing and FC sample difficulty rating score. Predictors of testing difficulty were derived using multivariable logistic regression analysis. RESULTS: A total of 585 patients with IBD completed the survey; 306 males with a median age of 43 years (IQR: 31-54). There were 446 patients (76%) who had prior FC testing experience. Of these, 37% (n = 165) rated FC testing difficult; 'sample collection' (n = 106; 67%) being the most common reason reported. Multivariable regression analysis identified age <49 years (odds ratio (OR): 2.5, CI:1.6-4.0), disease duration <35 months (OR 1.4, CI:0.9-2.1) and testing location (UK centre: OR 1.9, CI:1.2-3.1) as predictors of a difficult FC rating score. CONCLUSIONS: A total of 37% of patients find FC testing challenging, in particular those aged <49 years, disease duration <35 months. Further studies understanding and addressing these practical issues may aid higher FC uptake in clinic.

Assessing and supporting patients who have IBD.

This two-part series gives general nurses information about inflammatory bowel disease. Part 1 explored the aetiology, pathophysiology, signs and symptoms. This second part looks at how patients are assessed and treated, and the nurse's role. Although treatment of this condition may be complex, nurses can embed the 6Cs into practice to ensure they provide care and support of a high quality.

Assessment and management of diarrhoea.

All nurses are likely to care for many patients with diarrhoea. A new Nursing Times Learning unit has been launched to complement guidance from the Royal College of Nursing to assist them in approaching the difficult, sometimes embarrassing issues related to planning care for people with diarrhoea. This article summarises the resources and gives guidance on identifying types and causes of diarrhoea, and advice on management.

TCR-dependent and -independent activation underlie liver-specific regulation of NKT cells.

The fate of invariant NKT (iNKT) cells following activation remains controversial and unclear. We systemically examined how iNKT cells are regulated following TCR-dependent and -independent activation with α-galactosylceramide (αGC) or IL-18 plus IL-12, respectively. Our studies reveal activation by αGC or IL-18 plus IL-12 induced transient depletion of iNKT cells exclusively in the liver that was independent of caspase 3-mediated apoptosis. The loss of iNKT cells was followed by repopulation and expansion of phenotypically distinct cells via different mechanisms. Liver iNKT cell expansion following αGC, but not IL-18 plus IL-12, treatment required an intact spleen and IFN-γ. Additionally, IL-18 plus IL-12 induced a more prolonged expansion of liver iNKT cells compared with αGC. iNKT cells that repopulate the liver following αGC had higher levels of suppressive receptors PD-1 and Lag3, whereas those that repopulate the liver following IL-18 plus IL-12 had increased levels of TCR and ICOS. In contrast to acute treatment that caused a transient loss of iNKT cells, chronic αGC or IL-18 plus IL-12 treatment caused long-term systemic loss requiring an intact thymus for repopulation of the liver. This report reveals a previously undefined role for the liver in the depletion of activated iNKT cells. Additionally, TCR-dependent and -independent activation differentially regulate iNKT cell distribution and phenotype. These results provide new insights for understanding how iNKT cells are systemically regulated following activation.

Resistance to IFN-alpha-induced apoptosis is linked to a loss of STAT2.

Type I IFNs (IFN-alpha/beta) are pleitropic cytokines widely used in the treatment of certain malignancies, hepatitis B and C, and multiple sclerosis. IFN resistance is a challenging clinical problem to overcome. Hence, understanding the molecular mechanism by which IFN immunotherapy ceases to be effective is of translational importance. In this study, we report that continuous IFN-alpha stimulation of the human Jurkat variant H123 led to resistance to type I IFN-induced apoptosis due to a loss of signal transducers and activators of transcription 2 (STAT2) expression. The apoptotic effects of IFN-alpha were hampered as STAT2-deficient cells were defective in activating the mitochondrial-dependent death pathway and ISGF3-mediated gene activation. Reconstitution of STAT2 restored the apoptotic effects of IFN-alpha as measured by the loss of mitochondrial membrane potential, cytochrome c release from mitochondria, caspase activation, and ultimately cell death. Nuclear localization of STAT2 was a critical event as retention of tyrosine-phosphorylated STAT2 in the cytosol was not sufficient to activate apoptosis. Furthermore, silencing STAT2 gene expression in Saos2 and A375S.2 tumor cell lines significantly reduced the apoptotic capacity of IFN-alpha. Altogether, we show that STAT2 is a critical mediator in the activation of type I IFN-induced apoptosis. More importantly, defects in the expression or nuclear localization of STAT2 could lessen the efficacy of type I IFN immunotherapy.

Successful immunotherapy with IL-2/anti-CD40 induces the chemokine-mediated mitigation of an immunosuppressive tumor microenvironment.

Treatment of mice bearing orthotopic, metastatic tumors with anti-CD40 antibody resulted in only partial, transient anti-tumor effects whereas combined treatment with IL-2/anti-CD40, induced tumor regression. The mechanisms for these divergent anti-tumor responses were examined by profiling tumor-infiltrating leukocyte subsets and chemokine expression within the tumor microenvironment after immunotherapy. IL-2/anti-CD40, but not anti-CD40 alone, induced significant infiltration of established tumors by NK and CD8(+) T cells. To further define the role of chemokines in leukocyte recruitment into tumors, we evaluated anti-tumor responses in mice lacking the chemokine receptor, CCR2. The anti-tumor effects and leukocyte recruitment mediated by anti-CD40 alone, were completely abolished in CCR2(-/-) mice. In contrast, IL-2/anti-CD40-mediated leukocyte recruitment and reductions in primary tumors and metastases were maintained in CCR2(-/-) mice. Treatment of mice with IL-2/anti-CD40, but not anti-CD40 alone, also caused an IFN-gamma-dependent increase in the expression of multiple Th1 chemokines within the tumor microenvironment. Interestingly, although IL-2/anti-CD40 treatment increased Tregs in the spleen, it also caused a coincident IFN-gamma-dependent reduction in CD4(+)/FoxP3(+) Tregs, myeloid-derived suppressor cells and Th2 chemokine expression specifically within the tumor microenvironment that was not observed after treatment with anti-CD40 alone. Similar effects were observed using IL-15 in combination with anti-CD40. Taken together, our data demonstrate that IL-2/anti-CD40, but not anti-CD40 alone, can preferentially reduce the overall immunosuppressive milieu within the tumor microenvironment. These results suggest that the use of anti-CD40 in combination with IL-2 or IL-15 may hold substantially more promise for clinical cancer treatment than anti-CD40 alone.

Development of regulatory T cells requires IL-7Ralpha stimulation by IL-7 or TSLP.

Interleukin-7 (IL-7), a cytokine produced by stromal cells, is required for thymic development and peripheral homeostasis of most major subsets of T cells. We examined whether regulatory T (Treg) cells also required the IL-7 pathway by analyzing IL-7Ralpha(-/-) mice. We observed a striking reduction in cells with the Treg surface phenotype (CD4, CD25, GITR (glucocorticoid-induced tumor necrosis factor [TNF]-like receptor), CD45RB, CD62L, CD103) or intracellular markers (cytotoxic T-lymphocyte-associated antigen-4, CTLA-4, and forkhead box transcription factor 3, Foxp3). Foxp3 transcripts were virtually absent in IL-7Ralpha(-/-) lymphoid tissues, and no Treg cell suppressive activity could be detected. There are 2 known ligands for IL-7Ralpha: IL-7 itself and thymic stromal lymphopoietin (TSLP). Surprisingly, mice deficient in IL-7 or the other chain of the TSLP receptor (TSLPR) developed relatively normal numbers of Treg cells. Combined deletion of IL-7 and TSLP receptor greatly reduced Treg cell development in the thymus but was not required for survival of mature peripheral Treg cells. We conclude that Treg cells, like other T cells, require signals from the IL-7 receptor, but unlike other T cells, do not require IL-7 itself because of at least partially overlapping actions of IL-7 and TSLP for development of Treg cells.

Phosphatidylinositol 3-kinase and Akt nonautonomously promote perineurial glial growth in Drosophila peripheral nerves.

Drosophila peripheral nerves, structured similarly to their mammalian counterparts, comprise a layer of motor and sensory axons wrapped by an inner peripheral glia (analogous to the mammalian Schwann cell) and an outer perineurial glia (analogous to the mammalian perineurium). Growth and proliferation within mammalian peripheral nerves are increased by Ras pathway activation: loss-of-function mutations in Nf1, which encodes the Ras inhibitor neurofibromin, cause the human genetic disorder neurofibromatosis, which is characterized by formation of neurofibromas (tumors of peripheral nerves). However, the signaling pathways that control nerve growth downstream of Ras remain incompletely characterized. Here we show that expression specifically within the Drosophila peripheral glia of the constitutively active Ras(V12) increases perineurial glial thickness. Using chromosomal loss-of-function mutations and transgenes encoding dominant-negative and constitutively active proteins, we show that this nonautonomous effect of Ras(V12) is mediated by the Ras effector phosphatidylinositol 3-kinase (PI3K) and its downstream kinase Akt. We also show that the nonautonomous, growth-promoting effects of activated PI3K are suppressed by coexpression within the peripheral glia of FOXO+ (forkhead box O) a transcription factor inhibited by Akt-dependent phosphorylation. We suggest that Ras-PI3K-Akt activity in the peripheral glia promotes growth of the perineurial glia by inhibiting FOXO. In mammalian peripheral nerves, the Schwann cell releases several growth factors that affect the proliferative properties of neighbors. Some of these factors are oversecreted in Nf1 mutants. Our results raise the possibility that neurofibroma formation in individuals with neurofibromatosis might result in part from a Ras-PI3K-Akt-dependent inhibition of FOXO within Schwann cells.

Enhanced antitumor response by divergent modulation of natural killer and natural killer T cells in the liver.

The use of interleukin-18 (IL-18) together with IL-12 induced high levels of IFN-gamma in tumor-bearing mice and regression of liver tumors that was abolished in IFN-gamma((-/-)) mice. Natural killer (NK) and NKT cells were the major producers of IFN-gamma in the livers of mice treated with IL-18 and/or IL-12. Liver NK cells were significantly increased by treatment with IL-18/IL-12, whereas the degree of liver NKT cell TCR detection was diminished by this treatment. Reduction of NK cells with anti-asGM1 decreased the antitumor activity of IL-18/IL-12 therapy and revealed NK cells to be an important component for tumor regression in the liver. In contrast, the antitumor effects of both IL-18 and IL-12 were further increased in CD1d((-/-)) mice, which lack NKT cells. Our data, therefore, show that the antitumor activity induced in mice by IL-18/IL-12 is NK and IFN-gamma dependent and is able to overcome an endogenous immunosuppressive effect of NKT cells in the liver microenvironment. These results suggest that immunotherapeutic approaches that enhance NK cell function while eliminating or altering NKT cells could be effective in the treatment of cancer in the liver.