Genome-wide aggregated trans-effects on risk of type 1 diabetes: A test of the "omnigenic" sparse effector hypothesis of complex trait genetics.

The "omnigenic" hypothesis postulates that the polygenic effects of common SNPs on a typical complex trait are mediated through trans-effects on expression of a relatively sparse set of effector ("core") genes. We tested this hypothesis in a study of 4,964 cases of type 1 diabetes (T1D) and 7,497 controls by using summary statistics to calculate aggregated (excluding the HLA region) trans-scores for gene expression in blood. From associations of T1D with aggregated trans-scores, nine putative core genes were identified, of which three-STAT1, CTLA4 and FOXP3-are genes in which variants cause monogenic forms of autoimmune diabetes. Seven of these genes affect the activity of regulatory T cells, and two are involved in immune responses to microbial lipids. Four T1D-associated genomic regions could be identified as master regulators via trans-effects on gene expression. These results support the sparse effector hypothesis and reshape our understanding of the genetic architecture of T1D.

Impact of Vitamin D deficiency on immunological and metabolic responses in women with recurrent pregnancy loss: focus on VDBP/HLA-G1/CTLA-4/ENTPD1/adenosine-fetal-maternal conflict crosstalk.

BACKGROUND AND AIM: Recurrent pregnancy loss (RPL), also known as recurrent implantation failure (RIF), is a distressing condition affecting women characterized by two or more consecutive miscarriages or the inability to carry a pregnancy beyond 20 weeks. Immunological factors and genetic variations, particularly in Vit D Binding Protein (VDBP), have gained attention as potential contributors to RPL. This study aimed to provide insight into the immunological, genetic, and metabolic networks underlying RPL, placing a particular emphasis on the interactions between VDBP, HLA-G1, CTLA-4, ENTPD1, and adenosine-fetal-maternal conflict crosstalk. METHODS: A retrospective study included 198 women with three or more consecutive spontaneous abortions. Exclusion criteria comprised uterine abnormalities, endocrine disorders, parental chromosomal abnormalities, infectious factors, autoimmune diseases, or connective tissue diseases. Immunological interplay was investigated in 162 female participants, divided into two groups based on their Vit D levels: normal Vit D-RPL and low Vit D-RPL. Various laboratory techniques were employed, including LC/MS/MS for Vit D measurement, ELISA for protein detection, flow cytometry for immune function analysis, and molecular docking for protein-ligand interaction assessment. RESULTS: General characteristics between groups were significant regarding Vit D and glucose levels. Low Vit D levels were associated with decreased NK cell activity and downregulation of HLA-G1 and HLA-G5 proteins, while CTLA-4 revealed upregulation. VDBP was significantly downregulated in the low Vit D group. Our findings highlight the intricate relationship between Vit D status and adenosine metabolism by the downregulation of SGLT1, and NT5E, key components of adenosine metabolism, suggests that Vit D deficiency may disrupt the regulation of adenosine levels, leading to an impaired reproductive outcome. HNF1ß, a negative regulator of VDBP, was upregulated, while HNF1α, a positive regulator, was downregulated in low Vit D women after RPL. Molecular docking analysis revealed crucial residues involved in the interaction between Vit D and HNF1ß. CONCLUSION: Collectively, these findings underscore the importance of Vit D in modulating immune function and molecular pathways relevant to pregnancy maintenance, highlighting the need for further research to elucidate the mechanisms and potential therapeutic interventions for improving pregnancy outcomes in individuals with Vit D deficiency and RPL.

Immunotherapy: Current Status and Future Perspectives.

Hepatocellular carcinoma (HCC) has an increasing incidence and dismal prognosis, with few systemic treatments approved, including several small molecule tyrosine kinase inhibitors. The application of immune checkpoint inhibitors (ICIs) to HCC has resulted in durable activity, and further evaluation is ongoing. In this review, we discuss the immunologic principles and the mechanism of action of the ICIs and present the relevant clinical data. Furthermore, we provide an overview of the current and emerging immunotherapeutic approaches for HCC, such as combination treatments, vaccines, and cellular therapies.

Improved tumor accumulation and therapeutic efficacy of CTLA-4-blocking antibody using liposome-encapsulated antibody: In vitro and in vivo studies.

Cancer immunotherapies using monoclonal antibodies including cytotoxic T-lymphocyte-associated protein-4 (CTLA-4) blocking monoclonal antibody have several drawbacks including lack of appropriate penetration to the tumors, and organ toxicity. To address these obstacles, PEGylated and non-PEGylated liposomes containing CTLA-4 was prepared and characterized and the anti-tumor therapeutic responses were studied on the mice bearing C26 colon cancer tumors. The biodistribution study showed that the PEGylated liposomes had prolonged blood half-lives and accumulated remarkably more than non-PEGylated liposomes and free CTLA-4 antibody in tumor area. The lowest tumor volumes, highest time to reach end points (TTE: 34.29±3.09 days) and tumor growth delay percent (TGD: 29.37%) were seen in mice that received PEGylated liposomes than free CTLA-4 blocking antibody treatment (TTE: 31.16±4.13 days, TGD: 17.57%). In conclusion, PEGylated liposomes containing anti CTLA-4 antibody are delivered to tumor sites more efficiently and have a greater effect on anti-tumor immune responses than free antibodies and merits further investigation.

[Immunotherapy of cancer with checkpoint inhibitors : Not only in malignant melanoma]. / Immuntherapie von Tumorerkrankungen mit Checkpoint-Inhibitoren : Nicht nur beim malignen Melanom.

The newest weapon in cancer therapy is checkpoint inhibition, which is the result of basic immunology research. The success of this therapy is based on the fact that upon light microscopy, many solid tumors harbor lymphocytic cells infiltrating the tumor (TILs), and in many solid tumors, the presence of these TILs are prognostic. Ipilimumab was the first monoclonal antibody developed against a target present on T cells after becoming activated, CTLA-4. In malignant melanoma, ipilimumab showed its beneficial effect as compared to a placebo peptide. However, the therapy with this antibody harbors significant toxicity. Meanwhile, other targets such as PD-1, also expressed on (late) activated T cells, were identified, and therapies with antibodies inhibiting PD-1/PD-L1 are less toxic. Although these antibodies show response only in a minority of patients, the benefit seems durable in some of these patients. In solid tumors such as melanoma or non-small cell lung cancer (NSCLC), treatment with PD-1 inhibitors has resulted in a significant prolongation of survival, even in first-line treatment. As these drugs have been approved in many indications, it is important to know the drugs and side effects. Resistance towards these drugs are caused by low expression of the natural ligand, PD-L1, in the tumor tissue, as well as acquired loss of signal transduction of interferon-related genes such as JAK1 or JAK2, respectively. Also new in cancer therapy are bispecific T cell engager monoclonal antibodies (BiTEs) such as blinatumomab, and autologous chimeric antigen receptor-modified T cells (CAR-Ts). The later have proven their efficacy mainly in hematological neoplasias such as precursor-B-ALL. The dramatic costs of all these new drugs will have an enormous impact on the health care systems in the near future.

[Immunotherapeutic studies of head and neck tumors : Highlights of the 2016 ASCO Annual Meeting]. / Immuntherapeutische Studien bei Kopf-Hals-Tumoren : Highlights der ASCO-Jahrestagung 2016.

Immunotherapeutic drugs in the form of novel immune checkpoint inhibitors have had a significant impact on and revival of the treatment standards for head and neck tumors. Recently, at the annual meeting of the American Society of Clinical Oncology (ASCO) several innovative immunotherapies in head and neck cancer were presented, which might lead to a paradigm shift in the palliative as well as curative setting in the near future.The most common approaches are antibodies targeting the programmed cell death 1 (PD-1) axis. These therapies seem to be effective in a significant proportion of patients (independent of human papillomavirus) and show an extended duration of response.In a phase III trial for palliative second-line therapy, the PD-1 antibody nivolumab demonstrated a significant improvement in survival in patients with head and neck squamous cell carcinoma (HNSCC) who were experiencing disease progression after platinum-based therapy; therefore, the Food and Drug Administration gave it a breakthrough therapy designation.Further clinical trials focusing on first-line palliative treatment (compared with the EXTREME protocol) as well as neoadjuvant therapies using immune checkpoint-inhibitors are ongoing. However, valid testing systems (e. g., PD-L1 testing) as well as reliable predictive markers for patient selection are necessary to avoid increasing public health costs and to protect patients from potentially serious adverse events.

Management of infliximab refractory immune checkpoint inhibitor gastrointestinal toxicity: a multicenter case series.

BACKGROUND: Immune checkpoint inhibitor (ICI) gastrointestinal toxicity (gastritis, enteritis, colitis) is a major cause of morbidity and treatment-related death. Guidelines agree steroid-refractory cases warrant infliximab, however best management of infliximab-refractory ICI gastrointestinal toxicity (IRIGItox) is unknown. METHODS: We conducted an international multicenter retrospective case series. IRIGItox was defined as failure of symptom resolution ≤grade 1 (Common Terminology Criteria for Adverse Events V.5.0) following ≥2 infliximab doses or failure of symptom resolution ≤grade 2 after one dose. Data were extracted regarding demographics, steroid use, response to treatment, and survival outcomes. Toxicity was graded at symptom onset and time of infliximab failure. Efficacy of infliximab refractory therapy was assessed by symptom resolution, time to resolution and steroid wean duration. Survival outcomes were examined based on immunosuppressive therapy received. RESULTS: 78 patients were identified: median age 60 years; 56% men; majority melanoma (N=70, 90%); 60 (77%) received anti-cytotoxic T-lymphocyte-associated protein 4 alone or in combination with anti-programmed cell death protein-1 and most had colitis (N=74, 95%). 106 post-infliximab treatments were given: 31 calcineurin inhibitors (CNIs); 27 antimetabolites (mycophenolate, azathioprine); 16 non-systemic immunomodulatory agents (eg, mesalazine or budesonide); 15 vedolizumab; 5 other biologics (anti-interleukin-12/23, 16, Janus kinase inhibitors) and 7 interventional procedures (including colectomy); 5 did not receive post-infliximab therapy. Symptom resolution was achieved in most (N=23/31, 74%) patients treated with CNIs; 12/27 (44%) with antimetabolites; 7/16 (44%) with non-systemic immunomodulation, 8/15 (53%) with vedolizumab and 5/7 (71%) with interventional procedures. No non-vedolizumab biologics resulted in toxicity resolution. CNIs had the shortest time to symptom resolution (12 days) and steroid wean (43 days); however, were associated with poorer event-free survival (6.3 months) and overall survival (26.8 months) than other agents. Conversely, vedolizumab had the longest time to toxicity resolution and steroid wean, 66 and 124 days, but most favorable survival data: EFS 24.5 months; median OS not reached. Six death occurred (three due to IRIGItox or management of toxicity; three with persisting IRIGItox and progressive disease). CONCLUSIONS: IRIGItox causes major morbidity and mortality. Management is heterogeneous. CNIs appear most likely to result in toxicity resolution in the shortest time period, however, are associated with poorer oncological outcomes in contrast to vedolizumab.

Transforming growth factor-ß1 and soluble co-inhibitory immune checkpoints as putative drivers of immune suppression in patients with basal cell carcinoma.

The current study compared the levels and possible associations between systemic soluble immune checkpoints (sICPs, n = 17) and a group of humoral modulators of immune suppressor cells (n = 7) in a cohort of patients with basal cell carcinoma (BCC, n = 40) and a group of healthy control subjects (n = 20). The seven humoral modulators of immunosuppressor cells were represented by the enzymes, arginase 1 and fibroblast activation protein (FAP), the chemokine, RANTES (CCL5) and the cytokines, interleukin-10 and transforming growth factor-ß1 (TGF-ß1), as well as the M2-type macrophage markers, soluble CD163 (sCD163) and sCD206. The plasma levels of six co-inhibitory sICPs, sCTLA-4, sLAG-3, sPD-1, sPD-L1, sTIM-3 and sPD-L2 were significantly elevated in the cohort of BCC patients (p<0.001-p<0.00001), while that of sBTLA was significantly decreased (p<0.006). Of the co-stimulatory sICPs, sCD27 and sGITR were significantly increased (p<0.0002 and p<0.0538) in the cohort of BCC patients, while the others were essentially comparable with those of the control participants; of the dual active sICPs, sHVEM was significantly elevated (p<0.00001) and TLR2 comparable with the control group. A correlation heat map revealed selective, strong associations of TGF-ß1 with seven co-stimulatory (z = 0.618468-0.768131) and four co-inhibitory (z = 0.674040-0.808365) sICPs, as well as with sTLR2 (z = 0.696431). Notwithstanding the association of BCC with selective elevations in the levels of a large group of co-inhibitory sICPs, our novel findings also imply the probable involvement of TGF-ß1 in driving immunosuppression in this malignancy, possibly via activation of regulatory T cells. Notably, these abnormalities were present in patients with either newly diagnosed or recurrent disease.

In silico engineering a CD80 variant with increased affinity to CTLA-4 and decreased affinity to CD28 for optimized cancer immunotherapy.

CD80 or cluster of differentiation 80, also known as B7-1, is a member of the immunoglobulin super family, which binds to CTLA-4 and CD28 T cell receptors and induces inhibitory and inductive signals respectively. Although CTLA-4 and CD28 receptors belong to the same protein family, slight differences in their structures leads to CD80 having a higher binding affinity to CTLA-4 (-14.55 kcal/mol) compared with CD28(-12.51 kcal/mol). In this study, we constructed a variant of CD80 protein with increased binding affinity to CTLA-4 and decreased binding affinity to CD28. This variant has no signaling capability, and can act as a cap for these receptors to protect them from natural CD80 proteins existing in the body. The first step was the evolutionary and alanine scanning analysis of CD80 protein to determine conserved regions in this protein. Next, complex alanine scanning technique was employed to determine CD80 protein hotspots in CD80-CTLA-4 and CD80-CD28 protein complexes. This information was fed into a computational model developed in R for in silico mutagenesis and CD80 variant library construction. The 3D structures of variants were modeled using the Swiss model webserver. After modeling the 3D structures, HADDOCK server was employed to build all protein-protein complexes, which contain CTLA-4-CD80 variant complexes, Wild type CD80-CD28 complexes and CD28-CD80 variant complexes. Protein-protein binding free energy was determined using FoldX and the variant number 316 with mutations at 29, 31, 33 positions showed increased binding affinity to CTLA-4 (-21.43 kcal/mol) and decreased binding affinity to CD28 (- 9.54 kcal/mol). Finally, molecular dynamics (MD) simulations confirmed the stability of variant 316. In conclusion, we designed a new CD80 protein variant with potential immunotherapeutic applications.

Targeting Cbl-b in cancer immunotherapy.

Cancer immunotherapy with immune-checkpoint blockade has improved the outcomes of patients with various malignancies, yet a majority do not benefit or develop resistance. To address this unmet need, efforts across the field are targeting additional coinhibitory receptors, costimulatory proteins, and intracellular mediators that could prevent or bypass anti-PD1 resistance mechanisms. The CD28 costimulatory pathway is necessary for antigen-specific T cell activation, though prior CD28 agonists did not translate successfully to clinic due to toxicity. Casitas B lymphoma-b (Cbl-b) is a downstream, master regulator of both CD28 and CTLA-4 signaling. This E3 ubiquitin ligase regulates both innate and adaptive immune cells, ultimately promoting an immunosuppressive tumor microenvironment (TME) in the absence of CD28 costimulation. Recent advances in pharmaceutical screening and computational biology have enabled the development of novel platforms to target this once 'undruggable' protein. These platforms include DNA encoded library screening, allosteric drug targeting, small-interfering RNA inhibition, CRISPR genome editing, and adoptive cell therapy. Both genetic knock-out models and Cbl-b inhibitors have been shown to reverse immunosuppression in the TME, stimulate cytotoxic T cell activity, and promote tumor regression, findings augmented with PD1 blockade in experimental models. In translating Cbl-b inhibitors to clinic, we propose specific gene expression profiles that may identify patient populations most likely to benefit. Overall, novel Cbl-b inhibitors provide antigen-specific immune stimulation and are a promising therapeutic tool in the field of immuno-oncology.

Local and distant response to intratumoral immunotherapy assessed by immunoPET in mice.

BACKGROUND: Despite the promising efficacy of immune checkpoint blockers (ICB), tumor resistance and immune-related adverse events hinder their success in cancer treatment. To address these challenges, intratumoral delivery of immunotherapies has emerged as a potential solution, aiming to mitigate side effects through reduced systemic exposure while increasing effectiveness by enhancing local bioavailability. However, a comprehensive understanding of the local and systemic distribution of ICBs following intratumoral administration, as well as their impact on distant tumors, remains crucial for optimizing their therapeutic potential.To comprehensively investigate the distribution patterns following the intratumoral and intravenous administration of radiolabeled anti-cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) and to assess its corresponding efficacy in both injected and non-injected tumors, we conducted an immunoPET imaging study. METHODS: CT26 and MC38 syngeneic colorectal tumor cells were implanted subcutaneously on both flanks of Balb/c and C57Bl/6 mice, respectively. Hamster anti-mouse CTLA-4 antibody (9H10) labeled with zirconium-89 ([89Zr]9H10) was intratumorally or intravenously administered. Whole-body distribution of the antibody was monitored by immunoPET imaging (n=12 CT26 Balb/c mice, n=10 MC38 C57Bl/6 mice). Tumorous responses to injected doses (1-10 mg/kg) were correlated with specific uptake of [89Zr]9H10 (n=24). Impacts on the tumor microenvironment were assessed by immunofluorescence and flow cytometry. RESULTS: Half of the dose was cleared into the blood 1 hour after intratumoral administration. Despite this, 7 days post-injection, 6-8% of the dose remained in the intratumoral-injected tumors. CT26 tumors with prolonged ICB exposure demonstrated complete responses. Seven days post-injection, the contralateral non-injected tumor uptake of the ICB was comparable to the one achieved through intravenous administration (7.5±1.7% ID.cm-3 and 7.6±2.1% ID.cm-3, respectively) at the same dose in the CT26 model. This observation was confirmed in the MC38 model. Consistent intratumoral pharmacodynamic effects were observed in both intratumoral and intravenous treatment groups, as evidenced by a notable increase in CD8+T cells within the CT26 tumors following treatment. CONCLUSIONS: ImmunoPET-derived pharmacokinetics supports intratumoral injection of ICBs to decrease systemic exposure while maintaining efficacy compared with intravenous. Intratumoral-ICBs lead to high local drug exposure while maintaining significant therapeutic exposure in non-injected tumors. This immunoPET approach is applicable for clinical practice to support evidence-based drug development.

XTX101, a tumor-activated, Fc-enhanced anti-CTLA-4 monoclonal antibody, demonstrates tumor-growth inhibition and tumor-selective pharmacodynamics in mouse models of cancer.

INTRODUCTION: The clinical benefit of the anti-CTLA-4 monoclonal antibody (mAb) ipilimumab has been well established but limited by immune-related adverse events, especially when ipilimumab is used in combination with anti-PD-(L)1 mAb therapy. To overcome these limitations, we have developed XTX101, a tumor-activated, Fc-enhanced anti-CTLA-4 mAb. METHODS: XTX101 consists of an anti-human CTLA-4 mAb covalently linked to masking peptides that block the complementarity-determining regions, thereby minimizing the mAb binding to CTLA-4. The masking peptides are designed to be released by proteases that are typically dysregulated within the tumor microenvironment (TME), resulting in activation of XTX101 intratumorally. Mutations within the Fc region of XTX101 were included to enhance affinity for FcγRIII, which is expected to enhance potency through antibody-dependent cellular cytotoxicity. RESULTS: Biophysical, biochemical, and cell-based assays demonstrate that the function of XTX101 depends on proteolytic activation. In human CTLA-4 transgenic mice, XTX101 monotherapy demonstrated significant tumor growth inhibition (TGI) including complete responses, increased intratumoral CD8+T cells, and regulatory T cell depletion within the TME while maintaining minimal pharmacodynamic effects in the periphery. XTX101 in combination with anti-PD-1 mAb treatment resulted in significant TGI and was well tolerated in mice. XTX101 was activated in primary human tumors across a range of tumor types including melanoma, renal cell carcinoma, colon cancer and lung cancer in an ex vivo assay system. CONCLUSIONS: These data demonstrate that XTX101 retains the full potency of an Fc-enhanced CTLA-4 antagonist within the TME while minimizing the activity in non-tumor tissue, supporting the further evaluation of XTX101 in clinical studies.

Outcomes and a prognostic classifier in patients with microsatellite instability-high metastatic gastric cancer receiving PD-1 blockade.

BACKGROUND: Subgroup analyses of randomized trials suggest the superiority of immune checkpoint inhibitor-based therapy over chemotherapy in patients with mismatch-repair deficient (dMMR) and/or microsatellite instability-high (MSI-high) advanced gastric or gastroesophageal junction adenocarcinoma. However, these subgroups are small and studies examining prognostic features within dMMR/MSI-high patients are lacking. METHODS: We conducted an international cohort study at tertiary cancer centers and collected baseline clinicopathologic features of patients with dMMR/MSI-high metastatic or unresectable gastric cancer treated with anti-programmed cell death protein-1 (PD-1)-based therapies. The adjusted HRs of variables significantly associated with overall survival (OS) were used to develop a prognostic score. RESULTS: One hundred and thirty patients were included. At a median follow-up of 25.1 months, the median progression-free survival (PFS) was 30.3 months (95% CI: 20.4 to NA) and 2-year PFS rate was 56% (95% CI: 48% to 66%). Median OS was of 62.5 months (95% CI: 28.4 to NA) and 2-year OS rate was 63% (95% CI: 55% to 73%). Among the 103 Response Evaluation Criteria in Solid Tumors-evaluable patients, objective response rate was 66% and disease control rate 87% across lines of therapy. In the multivariable models, Eastern Cooperative Oncology Group Performance Status of 1 or 2, non-resected primary tumor, presence of bone metastases and malignant ascites were independently associated with poorer PFS and OS. These four clinical variables were used to build a three-category (ie, good, intermediate, and poor risk) prognostic score. Compared with patients with good risk, patients with intermediate risk score had numerically inferior PFS and OS (2-year PFS rate: 54.3% versus 74.5%, HR 1.90, 95% CI: 0.99 to 3.66; 2-year OS rate: 66.8% versus 81.2%, HR 1.86, 95% CI: 0.87 to 3.98), whereas patients with poor risk score had significantly inferior PFS and OS (2-year PFS rate: 10.6%, HR 9.65, 95% CI: 4.67 to 19.92; 2-year OS rate: 13.3%, HR 11.93, 95% CI: 5.42 to 26.23). CONCLUSIONS: Overall outcomes with anti-PD-1-based therapies are favorable in MSI-high gastroesophageal adenocarcinomas. However, within this overall favorable subgroup a more accurate prognostication using baseline clinical characteristics might identify patients at higher risk of rapid disease progression who may deserve intensified immunotherapy combination strategies.

Efficacy of abatacept treatment in a patient with enteropathy carrying a variant of unsignificance in CTLA4 gene: A case report.

BACKGROUND: Cytotoxic T Lymphocyte Antigen-4 (CTLA4) deficiency is a genetic defect that causes a common variable immunodeficiency (CVID) clinical phenotype. Several studies have reported an association between CTLA mutations or variants and various autoimmune diseases. Targeted therapy models, which have become increasingly popular in recent years, have been successful in treating CTLA4 deficiency. In this article, we discuss the clinical outcomes of abatacept treatment in a patient with CTLA4 and lipopolysaccharide-responsive beige-like anchor (LRBA) variants that was previously diagnosed with CVID. CASE SUMMARY: A 25-year-old female patient, who was visibly cachectic, visited our clinic over the course of five years, complaining of diarrhea. The patient was diagnosed with ulcerative colitis in the centers she had visited previously, and various treatments were administered; however, clinical improvement could not be achieved. Severe hypokalemia was detected during an examination. Her serum immunoglobulin levels, CD19+ B-cell percentage, and CD4/CD8 ratio were low. An endoscopic examination revealed erosive gastritis, nodular duodenitis, and pancolitis. Histopathological findings supported the presence of immune mediated enteropathy. When the patient was examined carefully, she was diagnosed with CVID, and intravenous immunoglobulin treatment was initiated. Peroral and rectal therapeutic drugs including steroid therapy episodes were administered to treat the immune mediated enteropathy. Strict follow-ups and treatment were performed due to the hypokalemia. After conducting genetic analyses, the CTLA4 and LRBA variants were identified and abatacept treatment was initiated. With targeted therapy, the patient's clinical and laboratory findings rapidly regressed, and there was an increase in weight. CONCLUSION: The heterozygous CTLA4 variant identified in the patient has been previously shown to be associated with various autoimmune diseases. The successful clinical outcome of abatacept treatment in this patient supports the idea that this variant plays a role in the immunopathogenesis of the disease. In the presence of severe disease, abatacept therapy should be considered until further testing can be conducted.

First-line nivolumab plus ipilimumab for metastatic non-small cell lung cancer, including patients with ECOG performance status 2 and other special populations: CheckMate 817.

BACKGROUND: CheckMate 817, a phase 3B study, evaluated flat-dose nivolumab plus weight-based ipilimumab in patients with metastatic non-small cell lung cancer (NSCLC). Here, in this research, we report on first-line treatment in patients with Eastern Cooperative Oncology Group (ECOG) performance status (PS) 0-1 (cohort A) and special populations (cohort A1: ECOG PS 2; or ECOG PS 0-1 with untreated brain metastases, renal impairment, hepatic impairment, or controlled HIV infection). METHODS: Cohorts A and A1 received nivolumab 240 mg every 2 weeks plus ipilimumab 1 mg/kg every 6 weeks. The primary endpoint was the incidence of grade 3-4 and grade 5 immune-mediated adverse events (IMAEs; adverse events (AEs) deemed potentially immune-related, occurring <100 days of last dose, and treated with immune-modulating medication (except endocrine events)) and treatment-related select AEs (treatment-related AEs with potential immunological etiology requiring frequent monitoring/intervention, reported between first dose and 30 days after the last dose) in cohort A; efficacy endpoints were secondary/exploratory. In cohort A1, safety/efficacy assessment was exploratory. RESULTS: The most common grade 3-4 IMAEs were pneumonitis (5.1%), diarrhea/colitis (4.9%), and hepatitis (4.6%) in cohort A (N=391) and diarrhea/colitis (3.5%), hepatitis (3.5%), and rash (3.0%) in cohort A1 (N=198). The most common grade 3-4 treatment-related select AEs were hepatic (5.9%), gastrointestinal (4.9%), and pulmonary (4.6%) events in cohort A and gastrointestinal (4.0%), skin (3.5%), and endocrine (3.0%) events in cohort A1. No grade 5 IMAEs or treatment-related select AEs occurred. Treatment-related deaths occurred in 4 (1.0%) and 3 (1.5%) patients in cohorts A and A1, respectively. Three-year overall survival (OS) rates were 33.7% and 20.5%, respectively. CONCLUSIONS: Flat-dose nivolumab plus weight-based ipilimumab was associated with manageable safety and durable efficacy in cohort A, consistent with data from phase 3 metastatic NSCLC studies. Special populations of cohort A1 including patients with ECOG PS 2 or ECOG PS 0-1 with untreated brain metastases had manageable treatment-related toxicity and clinically meaningful 3-year OS rate. TRIAL REGISTRATION NUMBER: NCT02869789.

Non-invasive PD-L1 quantification using [18F]DK222-PET imaging in cancer immunotherapy.

BACKGROUND: Combination therapies that aim to improve the clinical efficacy to immune checkpoint inhibitors have led to the need for non-invasive and early pharmacodynamic biomarkers. Positron emission tomography (PET) is a promising non-invasive approach to monitoring target dynamics, and programmed death-ligand 1 (PD-L1) expression is a central component in cancer immunotherapy strategies. [18F]DK222, a peptide-based PD-L1 imaging agent, was investigated in this study using humanized mouse models to explore the relationship between PD-L1 expression and therapy-induced changes in cancer. METHODS: Cell lines and xenografts derived from three non-small cell lung cancers (NSCLCs) and three urothelial carcinomas (UCs) were used to validate the specificity of [18F]DK222 for PD-L1. PET was used to quantify anti-programmed cell death protein-1 (PD-1) therapy-induced changes in PD-L1 expression in tumors with and without microsatellite instability (MSI) in humanized mice. Furthermore, [18F]DK222-PET was used to validate PD-L1 pharmacodynamics in the context of monotherapy and combination immunotherapy in humanized mice bearing A375 melanoma xenografts. PET measures of PD-L1 expression were used to establish a relationship between pathological and immunological changes. Lastly, spatial distribution analysis of [18F]DK222-PET was developed to assess the effects of different immunotherapy regimens on tumor heterogeneity. RESULTS: [18F]DK222-PET and biodistribution studies in mice with NSCLC and UC xenografts revealed high but variable tumor uptake at 60 min that correlated with PD-L1 expression. In MSI tumors treated with anti-PD-1, [18F]DK222 uptake was higher than in control tumors. Moreover, [18F]DK222 uptake was higher in A375 tumors treated with combination therapy compared with monotherapy, and negatively correlated with final tumor volumes. In addition, a higher number of PD-L1+ cells and higher CD8+-to-CD4+ cell ratio was observed with combination therapy compared with monotherapy, and positively correlated with PET. Furthermore, spatial distribution analysis showed higher [18F]DK222 uptake towards the core of the tumors in combination therapy, indicating a more robust and distinct pattern of immune cell infiltration. CONCLUSION: [18F]DK222-PET has potential as a non-invasive tool for monitoring the effects of immunotherapy on tumors. It was able to detect variable PD-L1 expression in tumors of different cancer types and quantify therapy-induced changes in tumors. Moreover, [18F]DK222-PET was able to differentiate the impact of different therapies on tumors.

Immune checkpoint inhibitors-induced systemic capillary leak syndrome: A report of two cases.

INTRODUCTION: The occurrence of systemic capillary leak syndrome under immune checkpoint inhibitors has seldom been reported in the literature. OBSERVATION: We report two cases of systemic capillary leak syndrome that occurred with nivolumab (anti-PD-1 antibody) for one, and with an anti-PD-1/CTLA-4 bi-specific antibody for the other. Patients presented with anasarca, hypoalbuminemia, acute kidney injury and, in one case, circulatory collapse. Immune checkpoint inhibitor causality was retained in the lack of evidence for other causes of secondary capillary leak syndrome or for an idiopathic form. The symptoms resolved after a few days of supportive measures (associated with glucocorticoids in one case). DISCUSSION: A high index of suspicion is required for the diagnosis of immune checkpoint inhibitors-induced systemic capillary leak syndrome because its presentation may differ from that of the idiopathic form. Activated CD8+ T-cells play a prominent role in the occurrence of immune checkpoint inhibitors-induced capillary leakage via their cytolytic action on the vascular endothelium. Treatment relies on supportive measures and discontinuation of the immune checkpoint inhibitor while the place of immunomodulatory drugs remains to be defined.

Vectorized Treg-depleting αCTLA-4 elicits antigen cross-presentation and CD8+ T cell immunity to reject 'cold' tumors.

BACKGROUND: Immune checkpoint blockade (ICB) is a clinically proven concept to treat cancer. Still, a majority of patients with cancer including those with poorly immune infiltrated 'cold' tumors are resistant to currently available ICB therapies. Cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) is one of few clinically validated targets for ICB, but toxicities linked to efficacy in approved αCTLA-4 regimens have restricted their use and precluded full therapeutic dosing. At a mechanistic level, accumulating preclinical and clinical data indicate dual mechanisms for αCTLA-4; ICB and regulatory T cell (Treg) depletion are both thought to contribute efficacy and toxicity in available, systemic, αCTLA-4 regimens. Accordingly, strategies to deliver highly effective, yet safe αCTLA-4 therapies have been lacking. Here we assess and identify spatially restricted exposure to a novel strongly Treg-depleting, checkpoint-blocking, vectorized αCTLA-4, as a highly efficacious and potentially safe strategy to target CTLA-4. METHODS: A novel human IgG1 CTLA-4 antibody (4-E03) was identified using function-first screening for monoclonal antibodies (mAbs) and targets associated with superior Treg-depleting activity. A tumor-selective oncolytic vaccinia vector was then engineered to encode this novel, strongly Treg-depleting, checkpoint-blocking, αCTLA-4 antibody or a matching surrogate antibody, and Granulocyte-macrophage colony-stimulating factor (GM-CSF) (VVGM-αCTLA-4). RESULTS: The identified 4-E03 antibody showed significantly stronger Treg depletion, but equipotent checkpoint blockade, compared with clinically validated αCTLA-4 ipilimumab against CTLA-4-expressing Treg cells in a humanized mouse model in vivo. Intratumoral administration of VVGM-αCTLA-4 achieved tumor-restricted CTLA-4 receptor saturation and Treg depletion, which elicited antigen cross-presentation and stronger systemic expansion of tumor-specific CD8+ T cells and antitumor immunity compared with systemic αCTLA-4 antibody therapy. Efficacy correlated with FcγR-mediated intratumoral Treg depletion. Remarkably, in a clinically relevant mouse model resistant to systemic ICB, intratumoral VVGM-αCTLA-4 synergized with αPD-1 to reject cold tumors. CONCLUSION: Our findings demonstrate in vivo proof of concept for spatial restriction of Treg depletion-optimized immune checkpoint blocking, vectorized αCTLA-4 as a highly effective and safe strategy to target CTLA-4. A clinical trial evaluating intratumoral VVGM-αhCTLA-4 (BT-001) alone and in combination with αPD-1 in metastatic or advanced solid tumors has commenced.

Immunosuppressive Effects and Potent Anti-tumor Efficacy of mTOR Inhibitor Everolimus in Breast Tumor-bearing Mice.

To investigate the effects of everolimus, a mechanistic/mammalian target of rapamycin (mTOR) inhibitor, on tumor growth and immune response in a mouse model of breast cancer. Human hormone receptor-positive (HR+)/human epidermal growth receptor 2-negative (HER2-) MC4-L2 cell line was used to establish a mouse model of breast cancer. The inhibitory effects of high (10 mg/kg) and low (5 mg/kg) doses of everolimus were investigated on tumor growth. Additionally, the frequency of CD4+Foxp3+ regulatory T cells (Tregs), CD8+Foxp3+ Tregs, and CD4+ and CD8+ T cells expressing cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) was explored by flow cytometry in bone marrow, lymph nodes, and spleen. Our results showed that both 10 mg/kg and 5 mg/kg doses of everolimus efficiently inhibited tumor growth, resulting in reduced breast tumor volume. In addition, it was revealed that everolimus-treated mice induced a higher frequency of CD4+Foxp3+ Tregs, CD8+Foxp3+ Tregs, and CD4+Foxp3+CTLA-4+ Tregs as well as CD4+ and CD8+ T cells expressing CTLA-4 in their bone marrow, lymph nodes, and spleen compared with standard control (vehicle-treated) in a dose-dependent manner. Furthermore, we found that everolimus treatment with 10 mg/kg and 5 mg/kg increased the frequency of Helios+Foxp3+ Tregs in the bone marrow of treated mice compared with the control group. Our results indicate that treatment with everolimus not only inhibits tumor growth but also exerts an immunomodulatory effect by inducing Tregs in the lymphoid organs of breast cancer-bearing mice. The combination of therapy with other anti-cancer agents may negate immune suppression and improve the efficacy of mTOR-targeted breast cancer therapy.