Evaluation of ChatGPT and Google Bard Using Prompt Engineering in Cancer Screening Algorithms.

Large language models (LLMs) such as ChatGPT and Bard have emerged as powerful tools in medicine, showcasing strong results in tasks such as radiology report translations and research paper drafting. While their implementation in clinical practice holds promise, their response accuracy remains variable. This study aimed to evaluate the accuracy of ChatGPT and Bard in clinical decision-making based on the American College of Radiology Appropriateness Criteria for various cancers. Both LLMs were evaluated in terms of their responses to open-ended (OE) and select-all-that-apply (SATA) prompts. Furthermore, the study incorporated prompt engineering (PE) techniques to enhance the accuracy of LLM outputs. The results revealed similar performances between ChatGPT and Bard on OE prompts, with ChatGPT exhibiting marginally higher accuracy in SATA scenarios. The introduction of PE also marginally improved LLM outputs in OE prompts but did not enhance SATA responses. The results highlight the potential of LLMs in aiding clinical decision-making processes, especially when guided by optimally engineered prompts. Future studies in diverse clinical situations are imperative to better understand the impact of LLMs in radiology.

Cross-Activation of Regulatory T Cells by Self Antigens Limits Self-Reactive and Activated CD8+ T Cell Responses.

The interaction between regulatory T (Treg) cells and self-reactive T cells is a crucial mechanism for maintaining immune tolerance. In this study, we investigated the cross-activation of Treg cells by self-antigens and its impact on self-reactive CD8+ T cell responses, with a focus on the P53 signaling pathway. We discovered that major histocompatibility complex (MHC) I-restricted self-peptides not only activated CD8+ T cells but also induced the delayed proliferation of Treg cells. Following HLA-A*0201-restricted Melan-A-specific (pMelan) CD8+ T cells, we observed the direct expansion of Treg cells and concurrent suppression of pMelan+CD8+ T cell proliferation upon stimulation with Melan-A peptide. Transcriptome analysis revealed no significant alterations in specific signaling pathways in pMelan+CD8+ T cells that were co-cultured with activated Treg cells. However, there was a noticeable upregulation of genes involved in P53 accumulation, a critical regulator of cell survival and apoptosis. Consistent with such observation, the blockade of P53 induced a continuous proliferation of pMelan+CD8+ T cells. The concurrent stimulation of Treg cells through self-reactive TCRs by self-antigens provides insights into the immune system's ability to control activated self-reactive CD8+ T cells as part of peripheral tolerance, highlighting the intricate interplay between Treg cells and CD8+ T cells and implicating therapeutic interventions in autoimmune diseases and cancer immunotherapy.

Phase 1 trial of 4-1BB-based adoptive T-cell therapy targeting human telomerase reverse transcriptase in patients with advanced refractory solid tumors.

BACKGROUND AIMS: Human telomerase reverse transcriptase (hTERT) is an attractive target for anti-cancer therapies. We developed an effective method for generating hTERT-specific CD8+ T cells (hTERT-induced natural T cells [TERTiNTs]) using peripheral blood mononuclear cells (PBMCs) from patients with solid cancers and investigated their feasibility and safety. METHODS: This was a single-center phase 1 trial using a 3 + 3 dose escalation design to evaluate six dose levels of TERTiNTs. PBMCs from each patient were screened using an hTERT peptide panel to select those that stimulated CD8+ T cells. The four most stimulatory peptides were used to produce autologous CD8+ T cells from patients refractory or intolerant to standard therapies. Eligible patients received a single intravenous infusion of TERTiNTs at different dose levels (4 × 108 cells/m2, 8 × 108 cells/m2 and 16 × 108 cells/m2). Pre-conditioning chemotherapy, including cyclophosphamide alone or in combination with fludarabine, was administered to induce lymphodepletion. RESULTS: From January 2014 to October 2019, a total of 24 patients with a median of three prior lines of therapy were enrolled. The most common adverse events were lymphopenia (79.2%), nausea (58.3%) and neutropenia (54.2%), mostly caused by pre-conditioning chemotherapy. The TERTiNT infusion was well tolerated, and dose-limiting toxicities were not observed. None of the patients showed objective responses. Seven patients (30.4%) achieved stable disease with a median progression-free survival of 3.9 months (range, 3.2-11.3). At the highest dose level (16 × 108 cells/m2), four of five patients showed disease stabilization. CONCLUSIONS: The generation of TERTiNTs was feasible and safe and provided an interesting disease control rate in heavily pre-treated cancer patients.

Screening adequacy of unstained thyroid fine needle aspiration samples using a deep learning-based classifier.

Fine needle aspiration (FNA) biopsy of thyroid nodules is a safe, cost-effective, and accurate diagnostic method for detecting thyroid cancer. However, about 10% of initial FNA biopsy samples from patients are non-diagnostic and require repeated FNA, which delays the diagnosis and appropriate care. On-site evaluation of the FNA sample can be performed to filter out non-diagnostic FNA samples. Unfortunately, it involves a time-consuming staining process, and a cytopathologist has to be present at the time of FNA. To bypass the staining process and expert interpretation of FNA specimens at the clinics, we developed a deep learning-based ensemble model termed FNA-Net that allows in situ screening of adequacy of unstained thyroid FNA samples smeared on a glass slide which can decrease the non-diagnostic rate in thyroid FNA. FNA-Net combines two deep learning models, a patch-based whole slide image classifier and Faster R-CNN, to detect follicular clusters with high precision. Then, FNA-Net classifies sample slides to be non-diagnostic if the total number of detected follicular clusters is less than a predetermined threshold. With bootstrapped sampling, FNA-Net achieved a 0.81 F1 score and 0.84 AUC in the precision-recall curve for detecting the non-diagnostic slides whose follicular clusters are less than six. We expect that FNA-Net can dramatically reduce the diagnostic cost associated with FNA biopsy and improve the quality of patient care.

The value of magnetic resonance imaging and ultrasonography (MRI/US)-fusion biopsy in clinically significant prostate cancer detection in patients with biopsy-naïve men according to PSA levels: A propensity score matching analysis.

Objectives: To evaluate the detection rate of clinically significant prostate cancer (csPCa) in Magnetic resonance imaging and ultrasonography (MRI/US) fusion biopsy in patients with biopsy-naïve men for varying prostate-specific antigen (PSA) levels. Since MRI can efficiently detect csPCa compared to standard transrectal ultrasound (TRUS) guided biopsy; however, the optimal PSA threshold for its use is unclear. Materials and methods: We retrospectively reviewed those who underwent MRI/US-fusion and standard biopsy from January 2016 to June 2018. Patients were divided into three groups: PSA <4, 4-10, >10 ng/mL. Propensity scoring was performed to balance the characteristics of the different biopsy groups, and the detection rate of csPCa was compared. Results: Data from a total of 670 males were included in the analysis (standard TRUS, n = 333; MRI/US fusion, n = 337). Prior to matching, patients who received MRI/US-fusion biopsy had lower prostate volume. Propensity score matching balanced this characteristic and generated a cohort comprising 195 patients from each group. In the matched cohort, patients with PSA 4-10 ng/mL had a significantly increased risk of csPCa by MRI/US-fusion vs. standard biopsy (35.0% vs. 26.6%, P = 0.033). However, patients with PSA <4 ng/mL had csPCa found by MRI/US-fusion versus standard biopsy (12.0% vs. 16.0%, P = 0.342), whereas, patients with PSA >10 ng/mL had csPCa found by MRI/US-fusion versus standard biopsy (78.0% vs. 80.0%, P = 0.596). In multivariate logistic analysis among patients with PSA 4-10 ng/mL, MRI/US-fusion biopsy (odds ratio: 2.46, 95% confidence interval = 1.31-4.60, P = 0.005) were significantly associated with a detection of csPCa. Conclusions: Detection of csPCa by MRI/US-fusion biopsy is more efficient in patients with biopsy-naïve men with PSA 4-10 ng/mL. However, standard TRUS biopsy may identify csPCa in patients with PSA <4 ng/mL and ≥10 ng/mL, emphasizing the importance of performing a standard biopsy in conjunction with MRI/US-fusion biopsy in such populations.

IL7-Fc Enhances the Efficacy of Adoptive T Cell Therapy under Lymphopenic Conditions in a Murine Melanoma Model.

Adoptive cell therapy (ACT) using tumor-reactive T cells is a promising form of immunotherapy to specifically target cancer. However, the survival and functional maintenance of adoptively transferred T cells remains a challenge, ultimately limiting their efficacy. Here, we evaluated the use of recombinant IL7-Fc in ACT. In a lymphopenic murine melanoma model, IL7-Fc treatment led to the enhanced inhibition of tumor growth with an increased number of adoptively transferred CD8+ T cells in tumor tissue and tumor-draining lymph nodes. Additionally, IL7-Fc further enhanced anti-tumor responses that were induced by recombinant human IL2 in the same mouse model. In contrast, in an immunocompetent murine melanoma model, IL7-Fc dampened the anti-tumor immunity. Further, IL7-Fc decreased the proliferation of adoptively transferred and immune-activated tumor-reactive CD8+ T cells in immunocompetent mice by inducing the massive expansion of endogenous T cells, thereby limiting the space for adoptively transferred T cells. Our data suggest that IL7-Fc is principally beneficial for enhancing the efficacy of tumor-reactive T-cells in lymphopenic conditions for the ACT.

Chronic activation of 4-1BB signaling induces granuloma development in tumor-draining lymph nodes that is detrimental to subsequent CD8+ T cell responses.

The antitumor capabilities of agonistic anti-4-1BB mAbs have made them an attractive target for tumor immunotherapy. However, the adverse side effects associated with agonist antibodies have hindered their clinical development. Here, we aimed to study the immune-related adverse events of repeated doses and long-term use of agonistic anti-4-1BB mAbs. We show that chronic activation of 4-1BB signals induced the accumulation of IFN-γ-producing PD-1+CD8+ T cells in the secondary lymphoid organs of tumor-bearing mice by increasing the number of dividing CD8+ T cells, which was beneficial for suppressing tumor growth in the early phase of anti-4-1BB induction. However, repeated exposure to anti-4-1BB mAbs led to granuloma development in tumor-draining lymph nodes (TDLNs) of mice due to recruitment and accumulation of macrophages via the CD8+ T cell-IFN-γ axis. This was accompanied by excessive lymph node swelling, which impaired the sequential activation of CD8+ T cells. Our data provide insights into the immune-related adverse events of long-term agonist 4-1BB antibody dosing, which should be considered during the clinical development of immunomodulating therapy.

Polymorphic Region-Specific Antibody for Evaluation of Affinity-Associated Profile of Chimeric Antigen Receptor.

Antibody applications in cancer immunotherapy involve diverse strategies, some of which redirect T cell-mediated immunity via engineered antibodies. Affinity is a trait that is crucial for these strategies, as optimal affinity reduces unwanted side effects while retaining therapeutic function. Antibody-antigen pairs possessing a broad affinity range are required to define optimal affinity and to investigate the affinity-associated functional profiles of T cell-engaging strategies such as bispecific antibodies and chimeric antigen receptor-engineered T cells. Here, we demonstrate the unique binding characteristic of the developed antibody clone MVR, which exhibits robust binding to B-lymphoid cell lines. Intriguingly, MVR specifically recognizes the highly polymorphic human leukocyte antigen (HLA)-DR complex and exhibits varying affinities that are dependent upon the HLA-DRB1 allele type. Remarkably, MVR binds to the conformational epitope that consists of two hypervariable regions. As an application of MVR, we demonstrate an MVR-engineered chimeric antigen receptor (CAR) that elicits affinity-dependent function in response to a panel of target cell lines that express different HLA-DRB1 alleles. This tool evaluates the effect of affinity on cytotoxic killing, polyfunctionality, and activation-induced cell death of CAR-engineered T cells. Collectively, MVR exhibits huge potential for the evaluation of the affinity-associated profile of T cells that are redirected by engineered antibodies.

Replication study: Melanoma exosomes educate bone marrow progenitor cells toward a pro-metastatic phenotype through MET.

As part of the Reproducibility Project: Cancer Biology we published a Registered Report (Lesnik et al., 2016) that described how we intended to replicate selected experiments from the paper 'Melanoma exosomes educate bone marrow progenitor cells toward a pro-metastatic phenotype through MET' (Peinado et al., 2012). Here we report the results. We regenerated tumor cells stably expressing a short hairpin to reduce Met expression (shMet) using the same highly metastatic mouse melanoma cell line (B16-F10) as the original study, which efficiently downregulated Met in B16F10 cells similar to the original study (Supplementary Figure 5A; Peinado et al., 2012). Exosomes from control cells expressed Met, which was reduced in exosomes from shMet cells; however, we were unable to reliably detect phosphorylated Met in exosomes. We tested the effect of exosome-dependent Met signaling on primary tumor growth and metastasis. Similar to the results in the original study, we did not find a statistically significant change in primary tumor growth. Measuring lung and femur metastases, we found a small increase in metastatic burden with exosomes from control cells that was diminished when Met expression was reduced; however, while the effects were in the same direction as the original study (Figure 4E; Peinado et al., 2012), they were not statistically significant. Differences between the original study and this replication attempt, such as level of knockdown efficiency, cell line genetic drift, sample sizes, study endpoints, and variability of observed metastatic burden, are factors that might have influenced the outcomes. Finally, we report meta-analyses for each result.

RELT negatively regulates the early phase of the T-cell response in mice.

RELT (tumor necrosis factor receptor superfamily member 19-like, TNFRSF19L) is a TNFR superfamily member that is primarily expressed in immune cells and lymphoid tissues, but whose immunological function is not well-defined. Here, we show that RELT is expressed by naive T cells and DCs, and their activation or maturation decreases RELT expression. Using RELT knockout (RELT-/- ) mice, we demonstrate that RELT deficiency selectively promotes the homeostatic proliferation of CD4+ T cells but not CD8+ T cells, and enhances anti-tumor CD8+ T-cell responses. We also demonstrate, using an adoptive transfer model in which RELT is knocked-out in either the transferred transgenic CD8+ T cells or the recipient melanoma-bearing mice, that RELT on multiple immune cells limits the hyper-response of tumor-specific CD8+ T cells. Hyper-responsiveness of RELT-deficient T cells was induced by promoting their proliferation. Taken together, our findings suggest that RELT acts as a negative regulator that controls the early phase of T-cell activation probably by promoting T-cell apoptosis.

Desensitized chimeric antigen receptor T cells selectively recognize target cells with enhanced antigen expression.

Chimeric antigen receptor (CAR) T cell therapy is an effective method for treating specific cancers. CARs are normally designed to recognize antigens, which are highly expressed on malignant cells but not on T cells. However, when T cells are engineered with CARs that recognize antigens expressed on the T cell surface, CAR T cells exhibit effector function on other T cells, which results in fratricide, or killing of neighboring T cells. Here, using human leukocyte antigen-DR (HLA-DR)-targeted CAR T cells, we show that weak affinity between CAR and HLA-DR reduces fratricide and induces sustained CAR downregulation, which consequently tunes the avidity of CAR T cells, leading to desensitization. We further demonstrate that desensitized CAR T cells selectively kill Epstein-Barr virus-transformed B cells with enhanced HLA-DR expression, while sparing normal B cells. Our study supports an avidity-tuning strategy that permits sensing of antigen levels by CAR T cells.

Cancer immunotherapy using tumor antigen-reactive T cells.

Studies over the last 30 years have shown the promise of cancer immunotherapy using T cells. In particular, since the report by Rosenberg and colleagues in 2002 that adoptive T-cell therapy (ACT) under lymphopenic conditions substantially increased response rates in melanoma patients, ACT has become a promising immunotherapeutic route to cancer treatment. Here we provide a brief history of ACT and review the characteristics of T-cell therapeutics that are specific to this approach. Since every T-cell treatment has its own unique properties in terms of number and type of target antigens, and number of epitopes and type of T cells, we review the main strategies for designing ACT: how Ag specificity is determined, how is it standardized and the need for lymphodepletion to induce epitope spreading. We also briefly consider the next generation of ACT.

4-1BB signaling activates glucose and fatty acid metabolism to enhance CD8+ T cell proliferation.

4-1BB (CD137) is a strong enhancer of the proliferation of CD8+ T cells. Since these cells require increased production of energy and biomass to support their proliferation, we hypothesized that 4-1BB signaling activated glucose and fatty acid metabolism. We found that treatment with agonistic anti-4-1BB mAb promoted the proliferation of CD8+ T cells in vitro, increasing their size and granularity. Studies with a glycolysis inhibitor and a fatty acid oxidation inhibitor revealed that CD8+ T cell proliferation required both glucose and fatty acid metabolism. Anti-4-1BB treatment increased glucose transporter 1 expression and activated the liver kinase B1 (LKB1)-AMP-activated protein kinase (AMPK)-acetyl-CoA carboxylase (ACC) signaling pathway, which may be responsible for activating the metabolism of glucose and fatty acids. We also examined whether blocking glucose or fatty acid metabolism affected cell cycle progression and the anti-apoptotic effect of 4-1BB signaling. The increase of anti-apoptotic factors and cyclins in response to anti-4-1BB treatment was completely prevented by treating CD8+ T cells with the fatty acid oxidation inhibitor, etomoxir, but not with the glycolysis inhibitor, 2-deoxy-D-glucose. We conclude that anti-4-1BB treatment activates glucose and fatty acid metabolism thus supporting the increased demand for energy and biomass, and that fatty acid metabolism plays a crucial role in enhancing the cell cycle progression of anti-CD3-activated CD8+ T cells in vitro and the anti-apoptotic effects of 4-1BB signaling on these cells.

The repopulating cancer cells in melanoma are characterized by increased mitochondrial membrane potential.

Although considerable effort has been expended in identifying definitive markers for cancer stem cells (CSCs) or cancer-initiating cells (CICs), the phenotypic plasticity of these cells obviates simple characterization using cell surface markers. We hypothesized that these cells could be characterized by their metabolic properties because they are in a quiescent state with low energy needs. We examined whether cancer cells differ in mitochondrial membrane potential (Δψm) when they are under stress. The Δψm of B16-F10 melanoma cells increased when they were exposed in vitro to serum starvation and chemotherapeutic agents, but not when exposed to hypoxia. Such TMREhigh cells were also present in tumor tissue. They primarily used glucose and/or lactate, and were superior to TMRElow B16-F10 cells in their ability to drive tumor growth. These findings suggest that CSCs or CICs could be identified in heterogeneous melanoma populations by measuring Δψm.

Extracellular stimulation of VSIG4/complement receptor Ig suppresses intracellular bacterial infection by inducing autophagy.

VSIG4/CRIg (V-set and immunoglobulin domain containing 4) is a transmembrane receptor of the immunoglobulin superfamily that is expressed specifically on macrophages and mature dendritic cells. VSIG4 signaling accelerates phagocytosis of C3-opsonized bacteria, thereby efficiently clearing pathogens within macrophages. We found that VSIG4 signaling triggered by C3-opsonized Listeria (opLM) or by agonistic anti-VSIG4 monoclonal antibody (mAb) induced macrophages to form autophagosomes. VSIG4-induced autophagosomes were selectively colocalized with the intracellular LM while starvation-induced autophagosomes were not. Consistent with these results, the frequency of autophagosomes induced by infection with opLM was lower in VSIG4-deficient bone marrow-derived macrophages (BMDMs) than in WT BMDMs. Furthermore, when VSIG4 molecules were overexpressed in HeLa cells, which are non-macrophage cells, VSIG4 triggering led to efficient uptake of LM, autophagosome formation, and killing of the infected LM. These findings suggest that VSIG4 signaling not only promotes rapid phagocytosis and killing of C3-opsonized intracellular bacteria, as previously reported, but also induces autophagosome formation, eliminating the LM that have escaped from phagosomes. We conclude that VSIG4 signaling provides an anti-immune evasion mechanism that prevents the outgrowth of intracellular bacteria in macrophages.

Redefining Budd-Chiari syndrome: A systematic review.

AIM: To re-examine whether hepatic vein thrombosis (HVT) (classical Budd-Chiari syndrome) and hepatic vena cava-Budd Chiari syndrome (HVC-BCS) are the same disorder. METHODS: A systematic review of observational studies conducted in adult subjects with primary BCS, hepatic vein outflow tract obstruction, membranous obstruction of the inferior vena cava (IVC), obliterative hepatocavopathy, or HVT during the period of January 2000 until February 2015 was conducted using the following databases: Cochrane Library, CINAHL, MEDLINE, PubMed and Scopus. RESULTS: Of 1299 articles identified, 26 were included in this study. Classical BCS is more common in women with a pure hepatic vein obstruction (49%-74%). HVC-BCS is more common in men with the obstruction often located in both the inferior vena cava and hepatic veins (14%-84%). Classical BCS presents with acute abdominal pain, ascites, and hepatomegaly. HVC-BCS presents with chronic abdominal pain and abdominal wall varices. Myeloproliferative neoplasms (MPN) are the most common etiology of classical BCS (16%-62%) with the JAK2V617-F mutation found in 26%-52%. In HVC-BCS, MPN are found in 4%-5%, and the JAK2V617-F mutation in 2%-5%. Classical BCS responds well to medical management alone and 1(st) line management of HVC-BCS involves percutaneous recanalization, with few managed with medical management alone. CONCLUSION: Systematic review of recent data suggests that classical BCS and HVC-BCS may be two clinically different disorders that involve the disruption of hepatic venous outflow.

Phase I Clinical Trial of 4-1BB-based Adoptive T-Cell Therapy for Epstein-Barr Virus (EBV)-positive Tumors.

Although adoptive cell therapy using Ag-specific T cells has been tested successfully in the clinic, the production of these T cells has been challenging. By applying our simple and practical 4-1BB-based method for the generation of Ag-specific CD8 T cells, here we determined the maximum tolerated dose, toxicity profile, immunologic responses, and clinical efficacy of autologous Epstein-Barr virus (EBV)/LMP2A-specific CD8 T cells (EBV-induced Natural T cell; EBViNT) in patients with relapsed/refractory EBV-positive tumors. This was a single-center, phase I, dose-escalation trial study evaluating 4 escalating dosing schedules of single injected EBViNT. CD8 T-cell responses against different LMP2A peptides in each patient were determined, and the most effective peptides were used to produce EBViNT. The produced autologous EBViNTs were single infused to patients with EBV-associated malignancy who had failed to standard treatments and were of HLA-A02 or A24 type. Of 11 patients enrolled, 8 patients received a single infusion of EBViNT: 4 with nasopharyngeal carcinomas, 1 with Hodgkin lymphoma, 2 with extranodal NK/T lymphomas, and 1 with diffuse large B-cell lymphoma. Single infusion of EBViNT was well tolerated by all the patients and generated objective antitumor responses in 3 of them. EBViNT infusion induced 2 waves of interferon-γ response: 1 approximately 1 week and the other 4-8 weeks after the treatment. The strength of the second wave was related to the efficacy of the treatment. The current trial shows that EBViNT therapy is safe and may provide a new option for treating EBV-positive recurrent cancer patients resistant to conventional therapy.

Prognostic value of fluorine-18 fludeoxyglucose positron emission tomography parameters differs according to primary tumour location in small-cell lung cancer.

OBJECTIVE: To investigate the prognostic value of fluorine-18 fludeoxyglucose (FDG) positron emission tomography (PET) parameters for small-cell lung cancer (SCLC), according to the primary tumour location, adjusted by conventional prognostic factors. METHODS: From 2008 to 2013, we enrolled consecutive patients with histologically proven SCLC, who had undergone FDG-PET/CT prior to initial therapy. The primary tumour location was categorized into central or peripheral types. PET parameters and clinical variables were evaluated using univariate and multivariate analysis. RESULTS: A total of 69 patients were enrolled in this study; 28 of these patients were categorized as having the central type and 41 patients as having the peripheral type. In univariate analysis, stage, serum neuron-specific enolase, whole-body metabolic tumour volume (WB-MTV) and whole-body total lesion glycolysis (WB-TLG) were found to be significant in both types of patients. In multivariate analysis, the independent prognostic factor was found to be stage in the central type, but WB-MTV and WB-TLG in the peripheral type. Kaplan-Meier analysis demonstrated that patients with peripheral type with limited disease and low WB-MTV or WB-TLG showed significantly better overall survival than all of the other groups (p < 0.0083). CONCLUSION: The FDG-PET volumetric parameters were demonstrated to be significant and independent prognostic factors in patients with peripheral type of SCLC, while stage was the only independent prognostic factor in patients with central type of SCLC. ADVANCES IN KNOWLEDGE: FDG-PET is a non-invasive method that could potentially be used to estimate the prognosis of patients, especially those with peripheral-type SCLC.