Oncode Institute

Neoadjuvant immunotherapy is increasingly used in resectable stage III melanoma, with major pathological response (MPR) closely linked to improved outcomes. Predictive biomarkers are needed to identify responders and potentially allow surgical de-escalation. [¹⁸F]FDG-PET/CT is a non-invasive method to assess metabolic response, and the index lymph node (ILN) may reflect response of the entire nodal bain. This retrospective analysis of largely prospectively collected data included patients with resectable stage III melanoma treated with two cycles of neoadjuvant pembrolizumab at two Dutch melanoma centres. [¹⁸F]FDG-PET/CT was performed before and after treatment, with metabolic response classified using EORTC criteria. Volumetric PET parameters, metabolic tumour volume (MTV) and total lesion glycolysis (TLG), were analysed retrospectively. Pathological response was assessed according to INMC criteria. In patients undergoing therapeutic lymph node dissection, ILN response was compared with the entire nodal basin. Forty-seven patients initiated neoadjuvant pembrolizumab, and 39 underwent surgery. Twenty-four patients had evaluable PET/CT scans: complete metabolic response (CMR) in 12.5%, partial metabolic response (PMR) in 37.5%, stable disease in 16.7%, and progressive metabolic disease (PMD) in 33%. Overall, 59% achieved MPR. All patients with CMR achieved MPR; 64% with PMR achieved MPR, compared with only 12.5% with PMD. All patients with pathological complete or near-complete response showed reductions in both ΔTLG and ΔMTV. ILN response was fully concordant with total nodal basin response in all evaluable cases. Three patients who declined surgery after CMR remained recurrence-free during follow-up. Metabolic response on [¹⁸F]FDG-PET/CT, particularly CMR, strongly correlates with pathological response after neoadjuvant pembrolizumab. ILN response accurately reflects total nodal basin response, supporting response-adapted surgical strategies.

Cell competition is an evolutionarily conserved quality control mechanism that eliminates less-fit cells to ensure optimal tissue integrity during development, homeostasis, and regeneration. Beyond these physiological roles, recent evidence implicates a role for cell competition in disease, particularly in cancer, where it can function by either suppressing or promoting malignant progression. In this review, we provide an overview of the different molecular mechanisms that drive cell competition and their impact on cancer development and progression. We will evaluate the current state-of-the-art in vitro experimental systems that can be employed to study these processes. Ranging from classical 2D co-culture systems to advanced organoid and organ-on-chip platforms, these model systems collectively enhance our understanding of the complex cellular interactions that underlie the competitive differences between cells. By integrating insights from diverse model systems, we highlight how cell competition shapes tumor dynamics and discuss how this knowledge could inspire novel therapeutic strategies to prevent or control tumor growth.

Abstract Transforming growth factor-beta (TGF-β), a potent promoter of extracellular matrix deposition and suppressor of infiltrating immunity, has arisen as an attractive target for improving outcomes in tissue fibrosis and cancer immune therapy. Despite the promise of TGF-β inhibitors for attenuating the progression of fibrotic disorders or as adjuncts for cancer immunotherapy, current systemically administered inhibitors that target the ligand or receptors have significant on-target liabilities, including cardiotoxicity and development of pre-malignant cutaneous squamous lesions. Recently, an engineered mini monomer of TGF-β (mmTGF-β), which potently and specifically inhibits TGF-β activity, was shown to strongly synergize with checkpoint inhibitors to suppress cancer progression in an aggressive model of melanoma when genetically delivered using an engineered form of vaccinia virus that preferentially infects cancer cells. Despite these promising results, however, a significant fraction of the mmTGF-β was found to misfold, likely due to mispairing of the cysteines that comprise its cystine knot. Here, we demonstrate that inclusion of a modified form of the TGF-β pro-domain that lacks its dimerization motif, the bowtie knot, dramatically improves both the folding and inhibitory activity upon secretion by mammalian cells, thus overcoming one of the major limitations of genetically delivering mmTGF-β. Furthermore, we show that fusion of mmTGF-β to a CD44 binding domain enhances the inhibitory potential of mmTGF-β on immune cells, and on other cell types which express CD44, by more than 30-fold compared to cells negative for CD44. Together, these modifications provide a framework for further enhancing the efficacy and safety of mmTGF-β for cancer immune therapy, and possibly also tissue fibrosis, when delivered genetically using vaccinia, or other related approaches.

Targeted therapies are important for invasive breast cancer (IBC) treatment but are generally not standard of care in the neoadjuvant setting. To identify therapies with the potential to improve neoadjuvant treatment response, it is essential to develop patient-derived preclinical models that faithfully reflect the diversity of primary IBC subtypes in patients. Here, we generated and characterized a large-scale cohort of 60 mouse-intraductal patient-derived xenograft (MIND-PDX) models representing all subtypes of primary IBC, as well as seven matched PDX-derived organoids. We showed that our IBC-MIND cohort can serve as a platform for preclinical evaluation of experimental neoadjuvant treatments. For triple-negative IBC, we demonstrated that neoadjuvant treatment does not benefit from addition of a PARP inhibitor, whereas for estrogen receptor-positive IBC, the combination of a CDK4/6 inhibitor and fulvestrant improved neoadjuvant treatment response. Our work provides a valuable resource of primary IBC models to study breast cancer biology and develop neoadjuvant treatments.

Activating epidermal growth factor receptor (EGFR) mutations are key drivers in non-small cell lung cancer (NSCLC) and other solid tumours, predicting responses to tyrosine kinase inhibitors (TKIs). Tumour heterogeneity alongside sampling and technical factors may contribute to discordant EGFR status across biopsies, complicating treatment decisions. However, systematic evidence on prevalence and drivers of discordance remains limited. This systematic review and meta-analysis followed Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines and was registered in PROSPERO (CRD42024615727). MEDLINE, Embase, and Scopus (2004-2024) were searched for studies reporting EGFR mutation discordance in adult solid tumours. Eligible studies compared primary and metastatic tumours (tissue-tissue), tissue and liquid biopsies (tissue-liquid), or different liquid biopsies. Data extraction and QUADAS-2 risk of bias assessment were performed independently. Discordance proportions were analysed on the logit scale with Haldane-Anscombe correction when needed. Random-effects meta-analysis was conducted using the Paule-Mandel estimator with Hartung-Knapp-Sidik-Jonkman confidence intervals. Subgroup analyses and meta-regression were used to estimate pooled discordance and explore potential predictors. A total of 154 studies (15,560 patients) predominantly involving NSCLC were included. The pooled discordance rate was 16.1% (95% confidence interval 14.2-18.2). Rates were similar for tissue-tissue (16.8%) and tissue-liquid (15.5%), but higher for liquid-liquid (34.0%). Plasma was the most-studied liquid source (16.5%), while cerebrospinal fluid showed the highest discordance (35.5%). Prior TKI exposure was associated with higher discordance (25.8%) compared with treatment-naive patients (14.6%; p = 0.003). Patients who later developed resistance also had higher baseline discordance (21.0% vs 15.0%; p = 0.042). Discordance varied by metastatic site, from 15.1% in lymph nodes to 17.9% in brain/central nervous system. Meta-regression identified TKI exposure, resistance, and mutation prevalence as predictors. EGFR mutation discordance is common and clinically relevant, particularly in NSCLC, but varies substantially by sampling strategy, biofluid, treatment context, and metastatic site. Given the high between-study heterogeneity and the predominance of NSCLC and Asian cohorts, pooled estimates should be interpreted as descriptive summaries rather than universally generalisable benchmarks. These findings support integrated and context-aware sampling strategies for EGFR-targeted therapy and resistance monitoring.

Intracellular signaling pathways form networks through which information is transmitted, often in the form of kinase-mediated phosphorylation events, to interpret extracellular signals and elicit appropriate cellular responses. Yet, even isogenic cells in a homogenous environment show heterogeneity in their intracellular "cell-state", as well as their response to extracellular signals. Here, we aimed to better understand this relation between these phenomena by investigating how information flows through the EGF-receptor centered network upon targeted drug treatment, and how this is affected by cell-to-cell-state differences. Using single-cell ID-seq, we profiled the cell-state and signaling activity in primary human epidermal stem cells by measuring 69 (phospho-)proteins upon inhibition of the Erk/MAPK (p90RSK) and Akt/mTOR (p70S6K) routes downstream of the EGF pathway. We found that the effects of drug treatment propagated from the EGF-signaling pathway to other connected parts of the cellular signaling network, indicating altered signaling flow. We identified nine distinct cell-states that show pervasive state-dependent drug-responses for many (phospho-)proteins. Computational modeling of the signaling network using single-cell Comparative Network Reconstruction showed that many interactions between phospho-proteins (i.e. network wiring) were quantitatively different between cell-states. Furthermore, (phospho-)proteins with a cell-state dependent drug response, were more likely to be involved in interactions that showed a cell-state dependent strength. Overall, our results indicate that drug treatment response and signaling interactions between proteins are closely related and modulated by cell-state.

Ribosomes are responsible for protein synthesis in all living systems. Determining their cellular organization, movement, and translational activity is crucial for dissecting ribosomes' complex functions. In this study, we describe the development of a selective fluorescent probe for eukaryotic ribosomes - RiboBright. Using C-H activation, the natural product cycloheximide was aminated at the C13-position and fluorescently modified to afford RiboBright. We employ RiboBright for the quantification of ribosome content in 10 cell lines through microscopy and flow cytometry. RiboBright is applicable in live cells for tracking and quantification of ribosome movement and in fixed cells for visualization of sub-micrometer-sized spots, at the single-cell level. RiboBright reveals lineage-specific ribosome content, organization, and movement upon differentiation into either extraembryonic endoderm or ectoderm-like cells. Thus, RiboBright provides a versatile and convenient approach for imaging the cellular dynamics of ribosomes.

In the era of immune checkpoint inhibitors for cancers, the need for prognostic biomarkers to identify patients most likely to achieve a durable response has become increasingly more relevant. Tumour‐infiltrating lymphocytes (TILs) have gained significant interest, as they can be evaluated using standard haematoxylin and eosin‐stained slides, making it a widely accessible and cost‐effective biomarker. In addition to their practicality, TILs provide prognostic insights into the interplay between the immune system and tumour cells. While the morphological assessment of TILs has been standardised in breast cancer, comprehensive guidelines for their evaluation in gastro‐oesophageal carcinomas (GEC) are still lacking. This narrative review examines the current literature on the composition, clinical implications and therapeutic utility of TILs in GEC. These insights are used to propose a framework with recommendations for standardised evaluation and reporting of TILs in GEC, while also highlighting pitfalls specific to GEC pathology. These recommendations serve as a vital first step towards the widespread use and validation of TILs as a biomarker.