Biology subjects

Lamy, P.

Publications and source records attributed to Lamy, P..

5 recordsLinked to original sources

bioRxiv2023

Field cancerization impacts tumor development, T-cell exhaustion and clinical outcomes in bladder cancer

Bladder field cancerization may be associated with disease outcome in patients with bladder cancer. To investigate this, we analyzed biopsies from bladder urothelium and urine samples by genomics and proteomics analyses. Samples were procured from multiple timepoints from 134 patients with early stage bladder cancer and detailed long term follow-up. We measured the field cancerization in normal-appearing bladder biopsies and found that high levels were associated with high tumor mutational burden, high neoantigen load, and high tumor-associated CD8 T-cell exhaustion. Non-synonymous mutations in known bladder cancer driver genes such as KDM6A and TP53 were identified as early disease drivers in normal urothelium. High field cancerization was associated with worse outcome but not with response to BCG. The level of urinary tumor DNA (utDNA) reflected the bladder tumor burden and originated from both tumors and field cancerization. High utDNA levels after BCG were associated with worse clinical outcomes for the patients. Our results indicate that the level of field cancerization may affect clinical outcome, tumor development and immune responses. utDNA measurements have significant prognostic value and reflect the disease status of the bladder.

cancer biology↗

bioRxiv2022

Single nucleus and spatially resolved intra-tumor subtype heterogeneity in bladder cancer

Current transcriptomic classification systems for bladder cancer do not consider the level of intra-tumor subtype heterogeneity. Here we present an investigation of the extent and possible clinical impact of intra-tumor heterogeneity across early and more advanced disease stages of bladder cancer. We performed single nucleus RNA-sequencing of 48 bladder tumors and four of these tumors were additionally analyzed using spatial transcriptomics. Total bulk RNA-sequencing and spatial proteomics data were available from the same tumors for comparison, along with detailed clinical follow-up of the patients. We demonstrate that tumors display varying levels of intra-tumor subtype heterogeneity and show that a higher class 2a weight estimated from bulk RNA-sequencing data is associated with worse outcome in patients with molecular high-risk class 2a tumors. Our results indicate that discrete subtype assignments from bulk RNA-sequencing data may lack biological granularity and continuous class scores could improve clinical risk stratification of patients. HighlightsO_LISingle nucleus RNA-sequencing of tumors from 48 bladder cancer patients. C_LIO_LITumors display varying levels of intra-tumor subtype heterogeneity at single nucleus and bulk tumor level. C_LIO_LIThe level of subtype heterogeneity could be estimated from both single nucleus and bulk RNA-sequencing data with a high concordance between the two. C_LIO_LIHigh class 2a weight estimated from bulk RNA-sequencing data is associated with worse outcome in patients with molecular high-risk class 2a tumors. C_LI

cancer biology↗

bioRxiv2022

Improved Protocol for Single Nucleus RNA-sequencing of Frozen Human Bladder Tumor Biopsies

This paper provides a laboratory workflow for single-nucleus RNA-sequencing (snRNA-seq) including a protocol for gentle nuclei isolation from fresh frozen tumor biopsies, making it possible to analyze biobanked material. To develop this protocol, we used non-frozen and frozen human bladder tumors and cell lines. We tested different lysis buffers (IgePal and Nuclei EZ) and incubation times in combination with different approaches for tissue and cell dissection; sectioning, semi-automated dissociation, manual dissociation with pestles, and semi-automated dissociation combined with manual dissociation with pestles. Our results showed a combination of IgePal lysis buffer, tissue dissection by sectioning and short incubation time was the best conditions for gentle nuclei isolation applicable for snRNA-seq, and we found limited confounding transcriptomic changes based on the isolation procedure. This protocol makes it possible to analyze biobanked material from patients with well described clinical and histopathological information and known clinical outcomes with snRNA-seq.

cancer biology↗

bioRxiv2020

Isoform-specific characterization implicates alternative splicing in APOBEC3B as a mechanism restricting APOBEC-mediated mutagenesis

APOBEC3A (A3A) and APOBEC3B (A3B) enzymes drive APOBEC-mediated mutagenesis, but the understanding of the regulation of their mutagenic activity remains limited. Here, we showed that mutagenic and non-mutagenic A3A and A3B enzymes are produced by canonical and alternatively spliced A3A and A3B isoforms, respectively. Notably, increased expression of the canonical A3B isoform, which encodes the mutagenic A3B enzyme, predicted shorter progression-free survival of bladder cancer patients. Expression of the mutagenic A3B isoform was reduced by exon 5 skipping, generating a non-mutagenic A3B isoform. The exon 5 skipping, which was dependent on the interaction between SF3B1 splicing factor and weak branch point sites in intron 4, could be enhanced by an SF3B1 inhibitor, decreasing the production of the mutagenic A3B enzyme. Thus, our results underscore the role of A3B, especially in bladder cancer, and implicate alternative splicing of A3B as a mechanism and therapeutic target to restrict APOBEC-mediated mutagenesis.

genomics↗

bioRxiv2019

Mutational analysis of field cancerization in bladder cancer

The multifocal and recurrent nature of bladder cancer has been explained by field cancerization of the bladder urothelium. To shed light on field cancerization in the bladder, we investigated the mutational landscape of normal appearing urothelium and paired bladder tumors from four patients. Sequencing of 509 cancer driver genes revealed the presence of 2-16 mutations exclusively localized in normal tissue (average target read depth 634x). Furthermore, 6-13 mutations were shared between tumor and normal samples and 8-75 mutations were exclusively detected in tumor samples. More mutations were observed in normal samples from patients with multifocal disease compared to patients with unifocal disease. Mutations in normal samples had low allele frequencies compared to tumor mutations (p<2.2*10-16). Furthermore, significant differences in the type of nucleotide changes between tumor, normal and shared mutations (p=2.7*10-8) were observed, and mutations in APOBEC context were observed primarily among tumor mutations (p=0.026). No differences in functional impact between normal, shared and tumor mutations were observed (p=0.23). Overall, these findings support the theory of multiple fields in the bladder, and document non-tumor specific driver mutations to be present in normal appearing bladder tissue.

cancer biology↗