Biology subjects

Sant, P.

Publications and source records attributed to Sant, P..

4 recordsLinked to original sources

bioRxiv2024

Comparison of spatial transcriptomics technologies used for tumor cryosections

BackgroundSpatial transcriptomics (ST) technologies are revolutionizing our understanding of intra-tumor heterogeneity and the tumor microenvironment by revealing single-cell molecular profiles within their spatial tissue context. The rapid evolution of ST methods, each with unique features, presents a challenge in selecting the most appropriate technology for specific research objectives. Here, we compare four imaging-based ST methods - RNAscope HiPlex, Molecular Cartography, MERFISH/Merscope, and Xenium - together with sequencing-based ST (Visium). These technologies were used to study cryosections of medulloblastoma with extensive nodularity (MBEN), a tumor chosen for its distinct microanatomical features. ResultsOur analysis reveals that automated imaging-based ST methods are well suited to delineating the intricate MBEN microanatomy, capturing cell-type-specific transcriptome profiles. We devise approaches to compare the sensitivity and specificity of the different methods together with their unique attributes to guide method selection based on the research aim. Furthermore, we demonstrate how reimaging of slides after the ST analysis can markedly improve cell segmentation accuracy and integrate additional transcript and protein readouts to expand the analytical possibilities and depth of insights. ConclusionsThis study highlights key distinctions between various ST technologies and provides a set of parameters for evaluating their performance. Our findings aid in the informed choice of ST methods and delineate approaches for enhancing the resolution and breadth of spatial transcriptomic analyses, thereby contributing to advancing ST applications in solid tumor research.

genomics↗

bioRxiv2023

HIPSD&R-seq enables scalable genomic copy number and transcriptome profiling

Single-cell DNA-sequencing (scDNA-seq) enables decoding somatic cancer variation. Existing methods are hampered by low throughput or cannot be combined with transcriptome sequencing in the same cell. We propose HIPSD&R-seq (HIgh-throughPut Single-cell Dna and Rna-seq), a scalable yet simple assay to profile low-coverage DNA and RNA in thousands of cells in parallel. Our approach builds on an accessible modification of the 10X Genomics platform for scATAC and multiome profiling. In applications to human cell models and primary tissue, we demonstrate the feasibility to detect rare clones and we combine the assay with combinatorial indexing to profile over 16,000 cells.

cancer biology↗

bioRxiv2023

Enhancing sensitivity and versatility of Tn5 based single cell omics

The analysis of chromatin features in single cells centers around Tn5 transposase and exploits its activity to simultaneously fragment target DNA and integrate adapter sequences of choice. This reaction provides a direct readout in the assay for transposase-accessible chromatin in single cells (scATAC-seq) to map open chromatin regions. However, a current limitation is the sparse coverage of open chromatin loci that are detected in a given single cell by droplet-based methods. Thus, enhancing Tn5 activity to improve genomic coverage of scATAC-seq or facilitating multi-omics readouts of chromatin features via Tn5 together with the transcriptome is of great interest. Here, we address these issues by optimizing scATAC-seq for an increased number of integrations per cell. In addition, we provide a protocol that combines mapping of histone modification with scRNA-seq from the same cell by targeting Tn5 to antibody-bound chromatin epitopes. Our experimental workflows improve the results obtained from the downstream data analysis and serve to better resolve epigenetic heterogeneity and transcription regulation in single cells.

molecular biology↗

bioRxiv2023

Spatial and temporal transcriptomics of SHH-medulloblastoma with chromothripsis identifies multiple genetic clones that resist to treatment and lead to relapse

Paediatric medulloblastomas with chromothripsis are characterised by high genomic instability and are among the tumours with the worst prognosis. However, the determinants of their aggressiveness and the molecular makeup of chromothriptic medulloblastoma are not well understood. Here, we applied spatial transcriptomics to profile a cohort of 13 chromothriptic and non-chromothriptic medulloblastomas from the same molecular subgroup. Our data reveal a higher extent of spatial intra-tumour heterogeneity in chromothriptic medulloblastomas, which is associated with increased proliferation and stemness, but lower immune infiltration and differentiation. Spatial mapping of genetic subclones of the same tumour identify a regionally distinct architecture and clone-specific phenotypic features, with distinct degrees of differentiation, proliferation and immune infiltration between clones. We conducted temporal profiling of 11 patient-derived xenografts from chromothriptic medulloblastomas, covering the transition from the minimal residual disease stage to treatment-resistant regrown tumours. In chromothriptic medulloblastoma, an ecosystem of cells from multiple genetic clones resisting treatment and leading to relapse highlighted the importance of multi-clone interplay. Finally, we identified a potential role for tumour microtubes in treatment resistance in chromothriptic medulloblastoma, suggesting cell network communication as a putative target. HighlightsO_ST_ABSBiological insightsC_ST_ABSO_LIMedulloblastomas with chromothripsis are characterised by higher spatial intra-tumour heterogeneity, proliferation and stemness, but lower immune infiltration and differentiation, as compared with non-chromothriptic medulloblastomas C_LIO_LISpatially aware genetic clone assignment identifies phenotypic features such as degree of differentiation, proliferation and immune infiltration enriched in specific clones C_LIO_LICells from distinct genetic clones resist to treatment and give rise to relapse in patient-derived xenografts of medulloblastoma with chromothripsis C_LIO_LITumour microtubes point to putative molecular processes implicated in treatment resistance in chromothriptic medulloblastoma C_LI Technological advancesO_LIValidation of the spatial representation of patient-derived xenograft models, providing novel insights into the faithfulness of such models to study cancer C_LIO_LISpatial mapping of clones based on spatial transcriptomics and copy-number variant profiling C_LI

cancer biology↗