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Background: The increasing prevalence of metabolic dysfunction-associated steatotic liver disease (MASLD) incurs substantial morbidity, mortality and healthcare costs. Detection and clinical intervention at early stages of disease improves prognosis; however, we are currently limited by a lack of reliable diagnostic tests for population screening and monitoring responses to therapy. To address this unmet need, we investigated human invariant Natural Killer T cell (iNKT) activation by fat-loaded hepatocytes, leading to the discovery that circulating soluble CD46 (sCD46) levels accurately predict hepatic steatosis., Methods: sCD46 in plasma was measured using a newly developed immuno-competition assay in two independent cohorts: Prospective living liver donors (n = 156; male = 66, female = 90) and patients with liver tumours (n = 91; male = 58, female = 33). sCD46 levels were statistically evaluated as a predictor of hepatic steatosis., Findings: Interleukin-4-secreting (IL-4 + ) iNKT cells were over-represented amongst intrahepatic lymphocytes isolated from resected human liver samples. IL-4 + iNKT cells preferentially developed in cocultures with a fat-loaded, hepatocyte-like cell line, HepaRG. This was attributed to induction of matrix metalloproteases (MMP) in fat-loaded HepaRG cells and primary human liver organoids, which led to indiscriminate cleavage of immune receptors. Loss of cell-surface CD46 resulted in unrepressed differentiation of IL-4 + iNKT cells. sCD46 levels were elevated in patients with hepatic steatosis. Discriminatory cut-off values for plasma sCD46 were found that accurately classified patients according to histological steatosis grade., Interpretation: sCD46 is a reliable clinical marker of hepatic steatosis, which can be conveniently and non-invasively measured in serum and plasma samples, raising the possibility of using sCD46 levels as a diagnostic method for detecting or grading hepatic steatosis., Funding: F.B. was supported by the Else Kröner Foundation (Award 2016_kolleg.14). G.G. was supported by the Bristol Myers Squibb Foundation for Immuno-Oncology (Award FA-19-009). N.S. was supported by a Wellcome Trust Fellowship (211113/A/18/Z). J.A.H. received funding from the European Union's Horizon 2020 research and innovation programme (Award 860003). J.M.W. received funding from the Else Kröner Foundation (Award 2015_A10)., Competing Interests: Declaration of interests University Hospital Regensburg has filed a not yet published European patent application (Registration Nr. 23 183 382.3) for sCD46 as a clinical biomarker of hepatic steatosis. J.A.H. received in-kind support from Beckman Coulter. The authors have no other conflicts of interest to declare., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Cytoreductive surgery (CRS) combined with hyperthermic intraperitoneal chemotherapy (HIPEC) is a treatment option for peritoneal carcinomatosis (PC) from colorectal cancer (CRC), which is otherwise a terminal stage of disease. Nevertheless, survival outcomes are only marginally superior to other treatments. This fact highlights the need for better strategies to control intra-abdominal disease recurrence after CRS-HIPEC, including the complementary use of immunotherapies. The aim of this study was therefore to investigate the immune phenotype of T cells in patients with PC. Fifty three patients with CRC (34 patients with PC and 19 patients without PC) were enrolled in a prospective study (clinicaltrials.gov: NCT04108936). Peripheral blood and omental fat were collected to isolate peripheral blood mononuclear cells (PBMCs) and adipose tissue mononuclear cells (ATMCs). These cells were analysed by flow cytometry using a panel focused upon T cell memory differentiation and exhaustion markers. We found a more naïve profile for CD8 + T cells in peripheral blood and intra-abdominal fat of PC patients compared to comparator group (CG) patients. Furthermore, there was an over-representation of CD4 + T cells expressing inhibitory receptors in adipose tissue of PC patients, but not in blood. Our description of intraperitoneal T cell subsets gives us a better understanding of how peritoneal carcinomatosis shapes local immune responses., (© 2023. The Author(s).)

Background: The approval of Atezolizumab / Bevacizumab therapy (Atezo/Bev) in 2020 opened up a promising new treatment option for patients with end-stage hepatocellular carcinoma (HCC). However, liver transplant (LTx) patients with HCC are still denied this therapy owing to concerns about ICI-induced organ rejection and lack of regulatory approval., Methods: A prospective observational study at a tertiary liver transplant centre monitored the compassionate, off-label use of Atezo/Bev in a single, stable LTx recipient with non-resectable HCC recurrence. Close clinical, laboratory and immunological monitoring of the patient was performed throughout a four-cycle Atezo/Bev treatment. Measured parameters were selected after a systematic review of the literature on predictive markers for clinical response and risk of graft rejection caused by ICI therapy., Results: 19 articles describing 20 unique predictive biomarkers were identified. The most promising negative prognostic factors were the baseline values and dynamic course of IL-6, alpha-fetoprotein (AFP) and the AFP/CRP ratio. The frequency of regulatory T cells (Treg) reportedly correlates with the success of ICI therapy. PD-L1 and CD28 expression level with the allograft, peripheral blood CD4 + T cell numbers and Torque Teno Virus (TTV) titre may predict risk of LTx rejection following ICI therapy. No relevant side effects or acute rejection occurred during Atezo/Bev therapy; however, treatment did not prevent tumor progression. Absence of PD-L1 expression in pre-treatment liver biopsies, as well as a progressive downregulation of CD28 expression by CD4 + T cells during therapy, correctly predicted absence of rejection. Furthermore, increased IL-6 and AFP levels after starting therapy, as well as a reduction in blood Treg frequency, correctly anticipated a lack of therapeutic response., Conclusion: Atezo/Bev therapy for unresectable HCC in stable LTx patients remains a controversial strategy because it carries a high-risk of rejection and therapeutic response rates are poorly defined. Although previously described biomarkers of rejection risk and therapeutic response agreed with clinical outcomes in the described case, these immunological parameters are difficult to reliably interpret. Clearly, there is an important unmet need for standardized assays and clinically validated cut-offs before we use these biomarkers to guide treatment decisions for our patients., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2023 Yang Zhou, Eder, Weber, Heumann, Kronenberg, Werner, Geissler, Schlitt, Hutchinson and Bitterer.)

Hepatitis E virus (HEV) is a major cause of acute hepatitis globally. Chronic and fulminant courses are observed especially in immunocompromised transplant recipients since administration of ribavirin (RBV) does not always lead to a sustained virologic response. By in vitro stimulation of NK cells through hepatoma cell lines inoculated with a full-length HEV and treatment with RBV, we analyzed the viral replication and cell response to further elucidate the mechanism of action of RBV on immune cells, especially NK cells, in the context of HEV infection. Co-culture of HEV-infected hepatoma cells with PBMCs and treatment with RBV both resulted in a decrease in viral replication, which in combination showed an additive effect. An analysis of NK cell functions after stimulation revealed evidence of reduced cytotoxicity by decreased TRAIL and CD107a degranulation. Simultaneously, IFN-ɣ production was significantly increased through the IL-12R pathway. Although there was no direct effect on the IL-12R subunits, downstream events starting with TYK-2 and subsequently pSTAT4 were upregulated. In conclusion, we showed that RBV has an immunomodulatory effect on the IL-12R pathway of NK cells via TYK-2. This subsequently leads to an enhanced IFN-ɣ response and thus, to an additive antiviral effect in the context of an in vitro HEV infection.

Treatment of advanced melanoma with combined immune checkpoint inhibitor (ICI) therapy is complicated in up to 50% of cases by immune-related adverse events (irAE) that commonly include hepatitis, colitis and skin reactions. We previously reported that pre-therapy expansion of cytomegalovirus (CMV)-reactive CD4 + effector memory T cells (T EM ) predicts ICI-related hepatitis in a subset of patients with Stage IV melanoma given αPD-1 and αCTLA-4. Here, we develop and validate a 10-color flow cytometry panel for reliably quantifying CD4 + T EM cells and other biomarkers of irAE risk in peripheral blood samples. Compared to previous methods, our new panel performs equally well in measuring CD4 + T EM cells (agreement = 98%) and is superior in resolving CD4 + CD197 + CD45RA - central memory T cells (T CM ) from CD4 + CD197 + CD45RA + naive T cells (T naive ). It also enables us to precisely quantify CD14 + monocytes (CV = 6.6%). Our new "monocyte and T cell" (MoT) assay predicts immune-related hepatitis with a positive predictive value (PPV) of 83% and negative predictive value (NPV) of 80%. Our essential improvements open the possibility of sharing our predictive methods with other clinical centers. Furthermore, condensing measurements of monocyte and memory T cell subsets into a single assay simplifies our workflows and facilitates computational analyses., Competing Interests: MK is a Beckman Coulter Life Sciences associate. SH has received consulting fees and speaker’s honoraria from BMS and Merck Sharp & Dohme (MSD). NA is employed by MVZ for Laboratory Medicine Raubling. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Schilling, Glehr, Kapinsky, Ahrens, Riquelme, Cordero, Bitterer, Schlitt, Geissler, Haferkamp, Hutchinson and Kronenberg.)

Treatment of advanced melanoma with combined PD-1/CTLA-4 blockade commonly causes serious immune-mediated complications. Here, we identify a subset of patients predisposed to immune checkpoint blockade-related hepatitis who are distinguished by chronic expansion of effector memory CD4 + T cells (T EM cells). Pre-therapy CD4 + T EM cell expansion occurs primarily during autumn or winter in patients with metastatic disease and high cytomegalovirus (CMV)-specific serum antibody titres. These clinical features implicate metastasis-dependent, compartmentalised CMV reactivation as the cause of CD4 + T EM expansion. Pre-therapy CD4 + T EM expansion predicts hepatitis in CMV-seropositive patients, opening possibilities for avoidance or prevention. 3 of 4 patients with pre-treatment CD4 + T EM expansion who received αPD-1 monotherapy instead of αPD-1/αCTLA-4 therapy remained hepatitis-free. 4 of 4 patients with baseline CD4 + T EM expansion given prophylactic valganciclovir and αPD-1/αCTLA-4 therapy remained hepatitis-free. Our findings exemplify how pathogen exposure can shape clinical reactions after cancer therapy and how this insight leads to therapeutic innovations.

Background: Intraoperative ultrasound is a routine procedure in abdominal surgery. Not only detection of focal lesions but also perfusion and elasticity analyses using color-coded Doppler sonography, contrast-enhanced ultrasound and elastography enable highly sensitive and highly specific diagnostics, especially in oncological surgery., Objective: This article provides an overview on current techniques of intraoperative ultrasound and gives an outlook on future possibilities and applications., Material and Methods: Literature-based presentation of evidence-based intraoperative ultrasound, discussion of current basic research and expert recommendations., Results: Modern intraoperative ultrasound is a highly efficient technique for detection and characterization of focal space-occupying lesions in surgery of benign and malignant diseases., Conclusion: Intraoperative ultrasound is a highly sensitive and specific diagnostic tool, providing real-time diagnostics with a manageable amount of technical equipment. Thus, preoperative diagnostics can be refined, partly resulting in a modification of intraoperative treatment management. Future developments in the fields of digital image fusion with preoperative sectional imaging, optical guidance of surgical instruments and molecular targeted tumor therapy will increase the importance of intraoperative sonography even more.

Objective: To investigate the diagnostic accuracy of high-resolution ultrasound (HRU) for the detection of hepatic metastases of breast cancer in a humanized tumour mouse (HTM) using clinical standard technology. In addition, the efficiency of standard contrast-enhanced ultrasound (CEUS) [microbubbles of sulphur hexafluoride] and CEUS using a novel VEGFR2-targeted contrast agent [BR55®] was examined., Methods: A total of 14 HTM were sonographically examined twice. In addition to a human immune system, the animals developed hepatic tumour lesions after intrahepatic injection of BT-474 breast cancer cells. Digital cine loops from the arterial phase (15-35 sec), the portal venous phase (35-90 sec) and the late phase (3-15 min) of the entire liver were analysed. Data were correlated to histopathology., Results: After 9 months, half of the mice (7/14) revealed the development of hepatic breast cancer metastases. The detection limit was 1 mm tumour diameter. In particular, the use of targeted contrast media reduced the needed tumour diameter and helped to precisely classify tumour tissue. In 93% (13/14), the findings of ultrasound could be approved by histological examination by the pathologist., Conclusions: This study in HTM demonstrated the high feasibility of tumour specific contrast media and standard HRU contrast agents to detect early liver metastases.

Purpose: A. fumigatus infections represent a major threat for patients with a suppressed immune system. Early diagnosis is of importance for a favorable outcome but appears to be difficult due to limited diagnostic procedures. Here we investigated the sensitivity of high-resolution ultrasound (HRU) for the detection of A. fumigatus infection in the liver., Materials and Methods: BALB/c mice were intravenously infected with A. fumigatus and monitored by HRU, Doppler sonography (CCDS), contrast-enhanced ultrasound (CEUS), and real-time strain color-coded elastography (CCE) using a multi-frequency probe (6 - 15 MHz). Contrast media bolus injection of sulfur-hexafluoride micro-bubbles was applied and digital cine-loops from the arterial phase, as well as the portal venous phase up to the late phase of the whole liver were analyzed. All data were correlated to the histopathological findings., Results: Using HRU and CEUS, a sonic shadow was detected in all infected animals. All Aspergillus-infected nodes from 3 - 6 mm in the liver showed a shadow with rim enhancement and no intranodal enhancement when using CEUS. A. fumigatus infection was confirmed by CFU assessment and histopathological analysis. Granulomas were not associated with shadowing on B-mode. In contrast, granulomas with a diameter above 5 mm and a higher stiffness in CCE generated particularly an arterial rim enhancement and portal venous washout without contrast media uptake in the late phase. In addition, CEUS was able to define dynamic capillary microvascularization of infected liver areas., Conclusion: Liver lesions associated with A. fumigatus infection can be detected in mice when combined with CEUS and CCE in vivo., (© Georg Thieme Verlag KG Stuttgart · New York.)

The first PH-functional phosphines (1, 3, and 5) containing the 1,1'-binaphthyl-2,2'-bis(methylene) or 1,1'-biphenyl-2,2'-bis(methylene) backbone have been obtained by two-phase phosphination of 2,2'-bis(halomethyl)-1,1'-binaphthyls with PH(3) or in a protected-group synthesis using P(SiMe(3))(3) as the starting material. The 4,5-dihydro-3H-dinaphtho[2,1-c:1',2'-e]phosphepine (1) is configurationally stable, as indicated by the inequivalence of the two CH(2) and naphthyl substituents in the (13)C{(1)H} NMR spectra. The X-ray crystal structure of 1.0.5C(6)H(5)CH(3) shows an intracyclic C-P-C angle of 99.5(2) degrees, the interplanar angle of the phosphepine ring system being 67.6(5) degrees. The borane adduct 7 of the secondary phosphine 1 has been employed for the syntheses of atropisomeric mono- and bidentate ligands (8-14) with the bulky 1,1'-binaphthyl moieties. Results of force field calculations on the conformations of 1, 3, and 14 are presented. The ability of these phosphines to form mononuclear and polynuclear complexes with transition-metal centers is discussed. Compound 14 exhibits a large variety of low-energy conformations, and some of them seem to be capable of forming mononuclear transition-metal complexes.

Chiral water-soluble secondary phosphines (2-6) were obtained by nucleophilic phosphination of FC(6)H(4)-4-SO(3)K (1a), FC(6)H(3)-2,4-(SO(3)K)(2) (1b), and FC(6)H(4)-2-SO(3)K (1c) with RPH(2) (R = Ph, 2,4,6-Me(3)C(6)H(2), 2,4,6-iPr(3)C(6)H(2)) in the superbasic medium DMSO/KOH by employing steric control of substitution at phosphorus by bulky substituents R and sulfonic groups in the ortho position of the aromatic ring systems in 1b or 1c. The secondary phosphines may be deprotonated in DMSO/KOH to give phosphido anions which on reaction with alkyl halides (PhCH(2)Cl, Br(CH(2))(3)Br, and C(12)H(25)Br) yield mono- or bidentate tertiary phosphines (7-10). Ligands of this type are alternatively accessible by nucleophilic arylation of secondary phosphines, e.g. Ph(Me)PH or Ph(H)P(CH(2))(3)P(H)Ph with 1a or 1b, respectively. The crystal structure of the starting material 1b.H(2)O (space group P2(1)/m) has been determined. In the solid state of 1b.H(2)O the individual molecules are interconnected by ionic interactions between the potassium cations and the SO(3)(-) anions. The C-F bond (C(1)-F 1.347(4) Å) is shorter than that in C(6)H(5)F (1.356(4) Å). The unit cell of 7a.0.5H(2)O (space group P&onemacr;), the first structurally characterized chiral phosphine with a sulfonated phenyl substituent, contains the two enantiomers. Due to the asymmetrical substitution at phosphorus the PC(3) skeletons are significantly distorted (P(1)-C(1,11,31) 1.864(10), 1.825(8), 1.841(7) Å). The electronic structure of sulfonated fluorobenzenes FC(6)H(5)(-)(n)()(SO(3)M)(n)() (M = K, NH(4), n = 1-3) is discussed on the basis of quantum chemical calculations. In particular, the reactivity difference toward nucleophilic phosphination within the series is rationalized in terms of steric factors and of the -I effect of the sulfonic groups.