1. Oct 2025
    1. eLife Assessment

      This study presents an important finding by identifying OPG as a novel stromal checkpoint influencing T-cell anti-tumor responses, thereby shedding new light on the complex interplay between the tumor microenvironment and immune regulation. The data are robust and the experimental approaches are sound, providing solid support for the study's conclusions; however, there are a number of additional questions raised by the data. Of particular note are the questions raised on the mechanistic effects of TRAIL versus RANKL. In addition, it would broaden the interest in this study to include more translational human data to complement the work presented.

    2. Reviewer #1 (Public review):

      Summary:

      Wang et al. present a compelling study investigating a novel immunosuppressive mechanism within the tumor microenvironment (TME) mediated by a subset of cancer-associated fibroblasts (CAFs)-specifically, inflammatory CAFs (iCAFs) that secrete osteoprotegerin (OPG). Utilizing both genetic and antibody-mediated OPG inhibition in murine breast and pancreatic cancer models, the authors demonstrate that blocking OPG enhances infiltration and effector function of cytotoxic T cells, which leads to significant tumor regression. Their data further show that OPG blockade induces a population of IFN-licensed CAFs characterized by increased expression of antigen presentation genes and immunomodulatory properties that favour T cell infiltration. The manuscript proposes that OPG functions as a "stromal immune checkpoint" and could represent a promising therapeutic target to convert "cold" tumors into "hot," immunotherapy-responsive tumours.

      Strengths:

      (1) Novel role for OPG+ CAF as T-cell immune suppressors:<br /> This study introduces a novel role for OPG+ iCAFs as active suppressors of T cell function and highlights stromal OPG as a critical negative regulator of antitumor immunity.

      (2) Methodological Rigor:<br /> The manuscript is underpinned by a thorough and systematic experimental design, combining genetic mouse models, antibody interventions, in vitro functional assays, single-cell RNA-seq, and human RAN-seq datasets analyses.

      (3) Translational Relevance:<br /> By identifying OPG as a stromal immune checkpoint, the study opens exciting opportunities for developing new immunotherapeutic strategies in stromatogenic tumors.

      (4) Clear and Comprehensive Data Presentation:<br /> The use of high-dimensional single-cell technologies and logical, detailed data presentation supports the study's reproducibility and transparency.

      Weaknesses:

      (1) The manuscript lacks definitive data identifying the cellular origin of OPG, particularly establishing iCAFs as the exclusive functional source.

      (2) There is a paucity of translational evidence directly correlating OPG+ iCAFs with T cell exclusion in human tumors.

      (3) The scope is limited by the reliance on two murine models, including a subcutaneous pancreatic cancer model, which may not fully recapitulate native tumor microenvironments.

      (4) Long-term outcomes and durability of response following OPG blockade, including possible effects on bone homeostasis, are not addressed.

      (5) Mechanistic experiments related to the blockade of TRAIL and RANKL remain incomplete, and alternative pathways are not thoroughly explored.

    3. Reviewer #2 (Public review):

      Summary:

      The work identified a protein called OPD secreted by a particular subtype of cancer-associated fibroblasts and found that it regulated T cell function in the tumor microenvironment. They showed that an antibody that targeted this protein could induce infiltration of immune cells into the tumour and could convert a cold tumor lacking tumour infiltration to a hot tumour with an immune-rich tumour microenvironment. They have supported the conclusion with the data in animal work as well as human tissue data. The authors also stated that it remains unclear whether the IFN-stimulated CAF subset after antibody treatment of OPG is due to reprogramming of existing iCAFs or arises de novo from progenitor populations. Despite their preclinical data suggesting the latter, they rightly suggested that in vivo lineage tracing is needed to further prove the origin and fate of these CAF populations. Overall, this is a well-designed and important study that would benefit from further mechanistic clarification and minor revision.

      Strengths:

      The strength of their data is that they utilized an immunocompetent orthotopic breast cancer model using the GFP-labelled tumor cell line EO771 in C57BL/6J mice, a well-established model for interrogating the role of stromal-immune interactions in carcinogenesis and tumor growth. They also performed scRNA-seq of the sorted stromal cells of the implanted EO771 cells as well as stromal cells from human esophageal carcinoma using tumor samples and matched adjacent non-malignant tissues from patients.

      Weaknesses:

      The key mechanistic aspects remain unclear, in particular the relative contributions of the TRAIL versus RANKL pathways to immunosuppression. The dual inhibition of TRAIL and RANKL by OPG is proposed, but the contribution of each axis to immune suppression was not clearly dissected. It would strengthen the paper to evaluate the effects of TRAIL versus RANKL signalling (e.g., with selective ligands or antagonists), which warrants deeper mechanistic exploration. Moreover, while CD4⁺ T cell cytotoxicity was observed, its functional role was underexplored.

    1. Once a group is singled out in one contrast, it cannot appear in another.  Each contrast compares only two “chunks” of variance.  If you have a control group, the first contrast usually compares the control against all experimental groups.

      dus bepaalde constraten, eerst word de contrast groep vergeleken met alle experimentele groepen en als je iets uit het conntrast haalt kan het niet terugkomen

    2. bootstrapping

      ootstrapping is een statistische techniek die wordt gebruikt om de nauwkeurigheid van schattingen te beoordelen en betrouwbaarheidsintervallen te berekenen zonder aannames te maken over de verdeling van de data. Het is een herhalingsproces waarbij steekproeven worden genomen uit de oorspronkelijke dataset, met vervanging, om een groot aantal nieuwe steekproeven te genereren. Deze steekproeven worden vervolgens gebruikt om de variabiliteit van een statistiek te schatten.) opvallend het neemt herhaaldelijk een steekproef uit een sample met teruglegging wat betekend dat elke observatiie meerdere keren kan worden geslecteerd in een enkele steekproef het vereeist geen aanames over de verdeling van de data waardoor je het kan gebruiken als iets niet normaal verdeeld is.

    1. The liver, while playing important roles in whole body metabolism, is usually considered a part of the gastrointestinal system for two main reasons. First, it provides for excretion from the body of lipid-soluble waste products that cannot enter the urine. Second, the blood flow draining the intestine is arranged such that substances that are absorbed pass first through the liver, allowing for the removal and metabolism of any toxins that have been taken up, as well as clearance of particulates, such as small numbers of enteric bacteria.

      Testing

    1. I put on the body-armor of black rubber the absurd flippers the grave and awkward mask.

      I think it’s interesting how the ordinary diving equipment becomes symbolic, like “black rubber body armor” and “awkward mask,” suggesting emotional or psychological protection.