bims-tumhet Biomed News
on Tumor Heterogeneity
Issue of 2025–02–23
fifteen papers selected by
Sergio Marchini, Humanitas Research



  1. Int J Gynecol Cancer. 2025 Feb;pii: S1048-891X(24)07409-7. [Epub ahead of print]35(2): 100060
       OBJECTIVE: The molecular classification of endometrial cancer into POLE-ultra-mutated, mismatch repair-deficient, p53-mutated, and no specific molecular profile sub-types has significant prognostic value and is recommended in the evaluation of all patients with endometrial cancer. Nonetheless, there has been inconsistent clinical implementation. One possible barrier is the current practice of using several different assays, each with its own result, that subsequently need to be integrated. We developed a single, fully next-generation sequencing (NGS)-based assay that sub-types endometrial samples and evaluated it against an existing algorithm.
    METHODS: Molecular sub-typing was retrospectively performed on 142 formalin-fixed, paraffin-embedded endometrial cancer samples using a clinically validated NGS panel targeting 133 genes and additional loci for micro-satellite instability and tumor mutational burden. In parallel, these same samples were classified by the World Health Organization algorithm using mismatch repair and p53 immunohistochemistry, and POLE sequencing. Concordance between algorithms was assessed, and the prognostic value of each was evaluated. We further explored racial disparities in outcomes and assessed other prognostic and potentially therapeutic biomarkers.
    RESULTS: The sequencing-based method was highly concordant with the World Health Organization algorithm (136/142 patients, Cohen's κ = 0.94) and retained prognostic value, with a significant difference in overall survival and progression-free survival by sub-type, and similar stratification to that initially identified in The Cancer Genome Atlas analysis. In addition, 11 cases had sequence variants in the previously established prognostic biomarker CTNNB1, and 3 cases had potentially targetable sequence variants in ERBB2. Endometrial cancer outcomes in Black individuals were worse, in part owing to the increased incidence of sub-types with a poor prognosis.
    CONCLUSIONS: A fully sequencing-based assay streamlines molecular classification of endometrial cancer and retains the prognostic value of other validated methods, which may aid clinical implementation. The additional genomic information obtained with an NGS panel, beyond the classification markers, can broaden potentially applicable therapies.
    Keywords:  Endometrial Cancer; Molecular Profiling; Next-Generation Sequencing
    DOI:  https://doi.org/10.1016/j.ijgc.2024.100060
  2. J Ovarian Res. 2025 Feb 15. 18(1): 29
       INTRODUCTION: Early diagnosis of ovarian cancer, using cost-effective and non-invasive methods remains an unmet medical need, largely due to unspecific symptoms of the disease.
    OBJECTIVE: Our goal was to identify differentially methylated CpG loci between cervical swabs obtained from patients diagnosed with benign ovarian disease and with malignant pelvic mass.
    METHODOLOGY: Using Infinium EPICv2 array, we interrogated methylation profiles of 77 cervical swabs. The study cohort was then divided into a training and testing set to develop a diagnostic signature. We applied several strategies to pinpoint CpG sites able to differentiate cervical swabs obtained from ovarian cancer patients and patients with benign ovarian disease.
    RESULTS AND CONCLUSIONS: None of the statistical methods applied identified CpG loci capable of diagnosing ovarian cancer with sufficient specificity and sensitivity. We conclude that methylation differences observed do not adequately distinguish between benign and malignant ovarian disease. The variations attributable to clinical conditions are likely obscured by the differences in cell composition, which is the primary source of sample heterogeneity. Therefore, we suggest that diagnostic tools should not rely on local methylation profile of the cervix but rather focus on detecting cancer-specific sequences transferred from the tumor site and present in cervical swabs. Ovarian cancer is difficult to detect early, and we aimed to explore whether DNA methylation in cervical swabs could serve as a diagnostic tool. However, our study found that methylation patterns in these samples do not reliably distinguish between benign and malignant conditions, likely due to variations in cell composition. We recommend future research focus on detecting tumor-specific DNA sequences in cervical swabs instead.
    Keywords:  Cervical swabs; DNA methylation; Early diagnosis; Ovarian cancer
    DOI:  https://doi.org/10.1186/s13048-025-01609-2
  3. Int J Gynecol Cancer. 2025 Jan 23. pii: S1048-891X(25)00180-X. [Epub ahead of print] 101657
      In the era of the serous tubal intraepithelial carcinoma hypothesis, investigation continues as to what proportions of high-grade serous tubo-ovarian carcinomas originate in the distal fallopian tube versus in the ovary. In this retrospective cohort study of 118,619 patients with high-grade serous tubo-ovarian carcinoma identified in the Commission-on-Cancer's National Cancer Database from 2004 to 2021, a diagnosis shift from high-grade serous ovarian carcinoma to high-grade serous fallopian tubal carcinoma occurred from 2004 to 2018 that the proportional distribution of high-grade serous fallopian tubal carcinoma increased 6.1-fold from 4.5% in 2004 to 27.6% in 2018 (p-trend < .001). This rapid diagnosis shift from high-grade serous ovarian carcinoma to high-grade serous fallopian tubal carcinoma reached a plateau at 2018, followed by steady proportional distribution of high-grade serous fallopian tubal carcinoma among the high-grade serous tubo-ovarian carcinomas for 4 consecutive years (27.6% in 2018 to 28.0% in 2021, p-trend = .801). The average rate of tubal carcinomas during this post-plateau period was 27.7%. In conclusion, the diagnosis shift in the primary site of high-grade serous tubo-ovarian carcinoma from the ovary to the fallopian tube may have ended in the late 2010s. After the implementation of College of American Pathologists diagnosis criteria, 1 in 3 to 4 high-grade serous tubo-ovarian carcinomas were classified as of fallopian tube origin.
    Keywords:  Diagnosis Shift; High-Grade Serous Fallopian Tubal Carcinoma; High-Grade Serous Ovarian Carcinoma; Serous Tubal Intraepithelial Carcinoma
    DOI:  https://doi.org/10.1016/j.ijgc.2025.101657
  4. Nat Commun. 2025 Feb 21. 16(1): 1838
      Inferring cell spatial profiles from histology images is critical for cancer diagnosis and treatment in clinical settings. In this study, we report a weakly-supervised deep-learning method, HistoCell, to directly infer super-resolution cell spatial profiles consisting of cell types, cell states and their spatial network from histology images at the single-nucleus-level. Benchmark analysis demonstrates that HistoCell robustly achieves state-of-the-art performance in terms of cell type/states prediction solely from histology images across multiple cancer tissues. HistoCell can significantly enhance the deconvolution accuracy for the spatial transcriptomics data and enable accurate annotation of subtle cancer tissue architectures. Moreover, HistoCell is applied to de novo discovery of clinically relevant spatial organization indicators, including prognosis and drug response biomarkers, across diverse cancer types. HistoCell also enable image-based screening of cell populations that drives phenotype of interest, and is applied to discover the cell population and corresponding spatial organization indicators associated with gastric malignant transformation risk. Overall, HistoCell emerges as a powerful and versatile tool for cancer studies in histology image-only cohorts.
    DOI:  https://doi.org/10.1038/s41467-025-57072-6
  5. Epigenomics. 2025 Feb 21. 1-13
      Cardiac development is a precisely regulated process governed by both genetic and epigenetic mechanisms. Among these, DNA methylation is one mode of epigenetic regulation that plays a crucial role in controlling gene expression at various stages of heart development and maturation. Understanding stage-specific DNA methylation dynamics is critical for unraveling the molecular processes underlying heart development from specification of early progenitors, formation of a primitive and growing heart tube from heart fields, heart morphogenesis, organ function, and response to developmental and physiological signals. This review highlights research that has explored profiles of DNA methylation that are highly dynamic during cardiac development and maturation, exploring stage-specific roles and the key molecular players involved. By exploring recent insights into the changing methylation landscape, we aim to highlight the complex interplay between DNA methylation and stage-specific cardiac gene expression, differentiation, and maturation.
    Keywords:  Heart development; cardiac progenitors; cardiogenesis; cell differentiation; epigenetics; epigenomics; pluripotent stem cells
    DOI:  https://doi.org/10.1080/17501911.2025.2467024
  6. Acta Biochim Pol. 2025 ;72 13922
      In recent years, significant advancements in biochemistry, materials science, engineering, and computer-aided testing have driven the development of high-throughput tools for profiling genetic information. Single-cell RNA sequencing (scRNA-seq) technologies have established themselves as key tools for dissecting genetic sequences at the level of single cells. These technologies reveal cellular diversity and allow for the exploration of cell states and transformations with exceptional resolution. Unlike bulk sequencing, which provides population-averaged data, scRNA-seq can detect cell subtypes or gene expression variations that would otherwise be overlooked. However, a key limitation of scRNA-seq is its inability to preserve spatial information about the RNA transcriptome, as the process requires tissue dissociation and cell isolation. Spatial transcriptomics is a pivotal advancement in medical biotechnology, facilitating the identification of molecules such as RNA in their original spatial context within tissue sections at the single-cell level. This capability offers a substantial advantage over traditional single-cell sequencing techniques. Spatial transcriptomics offers valuable insights into a wide range of biomedical fields, including neurology, embryology, cancer research, immunology, and histology. This review highlights single-cell sequencing approaches, recent technological developments, associated challenges, various techniques for expression data analysis, and their applications in disciplines such as cancer research, microbiology, neuroscience, reproductive biology, and immunology. It highlights the critical role of single-cell sequencing tools in characterizing the dynamic nature of individual cells.
    Keywords:  high-throughput; single-cell RNA-sequencing; spatial transcriptomics; technology development; transcriptome
    DOI:  https://doi.org/10.3389/abp.2025.13922
  7. Sci Adv. 2025 Feb 21. 11(8): eads7405
      Increased infiltration of CD3+ and CD8+ T cells into ovarian cancer (OC) is linked to better prognosis, but the specific antigens involved are unclear. Recent reports suggest that HLA class I can present peptides from noncoding genomic regions, known as noncanonical or cryptic peptides, but their immunogenicity is underexplored. To address this, we used immunopeptidomic analysis and RNA sequencing on five metastatic OC samples, which identified 311 cryptic peptides (40 to 83 per patient). Despite comprising less than 1% of total peptides, cryptic peptides from noncoding transcripts emerged as the predominant antigen class when compared to the other major classes of known tumor-specific and tumor-associated antigens in OC samples. Notably, nearly 70% of the prioritized cryptic peptides elicited T cell activation, as evidenced by increased 4-1BB and IFN-γ expression in autologous CD8+ T cells. This study reveals noncoding cryptic peptides as an important class of immunogenic antigens in OC.
    DOI:  https://doi.org/10.1126/sciadv.ads7405
  8. Int J Gynecol Cancer. 2025 Jan 23. pii: S1048-891X(25)00179-3. [Epub ahead of print] 101656
      Circulating tumor DNA (ctDNA) is a promising non-invasive tool that has been demonstrated to be a clinically useful biomarker in several tumor types for risk stratification, prognosis, and early detection of recurrence. However, there are limited data on the clinical utility of ctDNA in endometrial cancer (EC) compared with other solid tumors. The evolution of EC management through the integration of molecular characterization into the treatment algorithm has intensified the need to develop more effective predictive biomarkers to optimize treatment and reduce clinical toxicities. Given its non-invasive nature and its ability to represent and complement tumor multiclonal spatial and temporal heterogeneity, ctDNA could act as a valid surrogate for tissue sampling. In addition to plasma ctDNA detection being associated with clinicopathologic features of tumor aggressiveness at pre-operative assessment, an association with reduced disease-free survival and overall survival has been observed in patients with detectable ctDNA. Moreover, the half-life of ctDNA is significantly shorter than CA125, and plasma levels are reported to be completely cleared from the blood within 1 week from surgical debulking. Therefore, ctDNA may serve as a dynamic biomarker for occult microscopic residual disease when assessed within the first 4 to 8 weeks after eradicative surgery. Few studies have reported high sensitivity of ctDNA in detecting disease recurrence at longitudinal follow-up, although there are limited data comparing ctDNA and traditional serum biomarkers (CA125 and HE4) in identifying recurrence. In the perspective of personalized oncology, ctDNA may potentially help improve adjuvant therapeutic management by escalating/de-escalating treatment based on ctDNA detection after surgery, during maintenance, or in the recurrent/metastatic setting, in addition to acting as a sensitive biomarker for early detection of recurrence. Several challenges hinder the use of ctDNA in EC, including the lack of standardized protocols, the low mutational burden, tumor heterogeneity, and background normal DNA, which limit assay sensitivity and specificity. In addition, the high cost of ctDNA analysis, particularly, next-generation sequencing, restricts its accessibility. Future trials should focus on cost-effective approaches to ensure sustainability and efficient resource allocation.
    Keywords:  Circulating Tumor DNA; Endometrial Cancer; Liquid Biopsy
    DOI:  https://doi.org/10.1016/j.ijgc.2025.101656
  9. bioRxiv. 2025 Jan 27. pii: 2025.01.24.634791. [Epub ahead of print]
       Motivation: Reconstructing clonal lineage trees ("tumor phylogenetics") has become a core tool of cancer genomics. Earlier approaches based on bulk DNA sequencing (DNA-seq) have largely given way to single-cell DNA-seq (scDNA-seq), which offers far greater resolution for clonal substructure. Available data has lagged behind computational theory, though. While single-cell RNA-seq (scRNA-seq) has become widely available, scDNA-seq is still sufficiently costly and technically challenging to preclude routine use on large cohorts. This forces difficult tradeoffs between the limited genome coverage of scRNA-seq, limited availability of scDNA-seq, and limited clonal resolution of bulk DNA-seq. These limitations are especially problematic for studying structural variations and focal copy number variations that are crucial to cancer progression but difficult to observe in RNA-seq.
    Results: We develop a method, TUSV-int, combining advantages of these various genomic technologies by integrating bulk DNA-seq and scRNA-seq data into a single deconvolution and phylogenetic inference computation while allowing for single nucleotide variant (SNV), copy number alteration (CNA) and structural variant (SV) data. We accomplish this by using integer linear programming (ILP) to deconvolve heterogeneous variant types and resolve them into a clonal lineage tree. We demonstrate improved deconvolution performance over comparative methods lacking scRNA-seq data or using more limited variant types. We further demonstrate the power of the method to better resolve clonal structure and mutational histories through application to a previously published DNA-seq/scRNA-seq breast cancer data set.
    Availability: The source code for TUSV-int is available at https://github.com/CMUSchwartzLab/TUSV-INT.git.
    DOI:  https://doi.org/10.1101/2025.01.24.634791
  10. bioRxiv. 2025 Feb 03. pii: 2025.02.03.636285. [Epub ahead of print]
      Centromere location is specified by CENP-A, a centromere-specific histone that epigenetically propagates centromere identity. How CENP-A is epigenetically maintained at one location in rapidly evolving centromeric DNA is unknown. Using single cell-derived clones of human cell lines, we demonstrate heterogeneity in CENP-A position within cell populations at neocentromeres and a native centromere. CENP-A heterogeneity is accompanied by heterogenous DNA methylation and H3K9me3 patterns, with DNA methylation shifting according to CENP-A position. We further reveal temporary precise CENP-A maintenance but evolution of CENP-A, DNA methylation and H3K9me3 position over prolonged proliferation, with neocentromeres exhibiting DNA methylation instability, H3K9me3 gain, boundary loss and increased epigenetic heterogeneity. Lastly, prolonged CENP-A and HJURP overexpression leads to neocentromere expansion, gradual CENP-A depletion, neocentromere destabilization and re-localization. This study reveals the evolving epigenetic plasticity of human centromeres and neocentromeres and highlights the importance of repressive chromatin boundaries in maintaining centromere stability across cellular proliferation.
    DOI:  https://doi.org/10.1101/2025.02.03.636285
  11. Nature. 2025 Feb 19.
      Neoantigen vaccines are under investigation for various cancers, including epidermal growth factor receptor (EGFR)-driven lung cancers1,2. We tracked the phylogenetic history of an EGFR mutant lung cancer treated with erlotinib, osimertinib, radiotherapy and a personalized neopeptide vaccine (NPV) targeting ten somatic mutations, including EGFR exon 19 deletion (ex19del). The ex19del mutation was clonal, but is likely to have appeared after a whole-genome doubling (WGD) event. Following osimertinib and NPV treatment, loss of the ex19del mutation was identified in a progressing small-cell-transformed liver metastasis. Circulating tumour DNA analyses tracking 467 somatic variants revealed the presence of this EGFR wild-type clone before vaccination and its expansion during osimertinib/NPV therapy. Despite systemic T cell reactivity to the vaccine-targeted ex19del neoantigen, the NPV failed to halt disease progression. The liver metastasis lost vaccine-targeted neoantigens through chromosomal instability and exhibited a hostile microenvironment, characterized by limited immune infiltration, low CXCL9 and elevated M2 macrophage levels. Neoantigens arising post-WGD were more likely to be absent in the progressing liver metastasis than those occurring pre-WGD, suggesting that prioritizing pre-WGD neoantigens may improve vaccine design. Data from the TRACERx 421 cohort3 provide evidence that pre-WGD mutations better represent clonal variants, and owing to their presence at multiple copy numbers, are less likely to be lost in metastatic transition. These data highlight the power of phylogenetic disease tracking and functional T cell profiling to understand mechanisms of immune escape during combination therapies.
    DOI:  https://doi.org/10.1038/s41586-025-08586-y
  12. bioRxiv. 2025 Feb 03. pii: 2025.02.01.636056. [Epub ahead of print]
      Many tumors evolve through cellular mutation and selection, where subpopulations of cells (subclones) with shared ancestry compete for dominance. Introduction of next generation sequencing enables subclone identification using small somatic. However, there are several advantages to marking subclones with structural variants: they have greater functional impact, play a crucial role in late-stage tumors, and provide a more complete view of genomic instability driving tumor evolution. Here, we present SVCFit, a scalable method to estimate the cellular prevalence of somatic deletions, duplications and inversions. We demonstrate that cellular prevalence estimation can be improved by incorporating distinct read patterns for each structural variant type. Additionally, this improvement is achieved without prior knowledge of tumor purity, which is often inaccurate. Using a combination of simulated data and patient-derived metastatic samples with known mixture proportions, we show that our algorithm achieves significantly greater accuracy than state-of-the-art in estimating the structural variants cellular prevalence (p<0.05). The speed of SVCFit estimation from cost-effective bulk whole-genome sequencing (WGS) makes it well-suited for analyzing large cohorts of sequenced tumor samples, enhancing the accessibility of SV-based clonal reconstruction.
    DOI:  https://doi.org/10.1101/2025.02.01.636056
  13. Oncoimmunology. 2025 Dec;14(1): 2466308
      Immune checkpoint blockade (ICB) is the standard of care for recurrent/metastatic head and neck squamous cell carcinoma (HNSCC), yet efficacy remains low. The combined positive score (CPS) for PD-L1 is the only biomarker approved to predict response to ICB and has limited performance. Tertiary Lymphoid Structures (TLS) have shown promising potential for predicting response to ICB. However, their exact composition, size, and spatial biology in HNSCC remain understudied. To elucidate the impact of TLS spatial biology in response to ICB, we utilized pre-ICB tumor tissue sections from 9 responders (complete response, partial response, or stable disease) and 11 non-responders (progressive disease) classified via RECISTv1.1. A custom multi-immunofluorescence (mIF) staining assay was applied to characterize tumor cells (pan-cytokeratin), T cells (CD4, CD8), B cells (CD19, CD20), myeloid cells (CD16, CD56, CD163), dendritic cells (LAMP3), fibroblasts (α Smooth Muscle Actin), proliferative status (Ki67) and immunoregulatory molecules (PD1). A machine learning model was employed to measure the effect of spatial metrics on achieving a response to ICB. A higher density of B cells (CD20+) was found in responders compared to non-responders to ICB (p = 0.022). The presence of TLS within 100 µm of the tumor was associated with improved overall (p = 0.04) and progression-free survival (p = 0.03). A multivariate machine learning model identified TLS density as a leading predictor of response to ICB with 80% accuracy. Immune cell densities and TLS spatial location play a critical role in the response to ICB in HNSCC and may potentially outperform CPS as a predictor of response.
    Keywords:  Head and neck cancer; immune checkpoint blockade; spatial biology; tertiary lymphoid structures
    DOI:  https://doi.org/10.1080/2162402X.2025.2466308