Identification of new driver and passenger mutations within APOBEC-induced hotspot mutations in bladder cancer

Abstract

Background APOBEC-driven mutagenesis and functional positive selection of mutated genes may synergistically drive the higher frequency of some hotspot driver mutations compared to other mutations within the same gene, as we reported for FGFR3 S249C. Only a few APOBEC-associated driver hotspot mutations have been identified in bladder cancer (BCa). Here, we systematically looked for and characterised APOBEC-associated hotspots in BCa. Methods We analysed 602 published exome-sequenced BCas, for part of which gene expression data were also available. APOBEC-associated hotspots were identified by motif-mapping, mutation signature fitting and APOBEC-mediated mutagenesis comparison. Joint analysis of DNA hairpin stability and gene expression was performed to predict driver or passenger hotspots. Aryl hydrocarbon receptor (AhR) activity was calculated based on its target genes expression. Effects of AhR knockout/inhibition on BCa cell viability were analysed. Results We established a panel of 44 APOBEC-associated hotspot mutations in BCa, which accounted for about half of the hotspot mutations. Fourteen of them overlapped with the hotspots found in other cancer types with high APOBEC activity. They mostly occurred in the DNA lagging-strand templates and the loop of DNA hairpins. APOBEC-associated hotspots presented systematically a higher prevalence than the other mutations within each APOBEC-target gene, independently of their functional impact. A combined analysis of DNA loop stability and gene expression allowed to distinguish known passenger from known driver hotspot mutations in BCa, including loss-of-function mutations affecting tumour suppressor genes, and to predict new candidate drivers, such as AHR Q383H. We further characterised AHR Q383H as an activating driver mutation associated with high AhR activity in luminal tumours. High AhR activity was also found in tumours presenting amplifications of AHR and its co-receptor ARNT. We finally showed that BCa cells presenting those different genetic alterations were sensitive to AhR inhibition. Conclusions Our study identified novel potential drivers within APOBEC-associated hotspot mutations in BCa reinforcing the importance of APOBEC mutagenesis in BCa. It could allow a better understanding of BCa biology and aetiology and have clinical implications such as AhR as a potential therapeutic target. Our results also challenge the dogma that all hotspot mutations are drivers and mostly gain-of-function mutations affecting oncogenes.

Review of Experimental Studies to Improve Radiotherapy Response in Bladder Cancer: Comments and Perspectives

“…At the present time no significant difference has been found in local relapse-free survival between bladder tumour subtypes in MIBC patients treated by TURBT followed by CCRT [44] or RT alone [45]. [36]); (b) cell line subtypes used in experimental studies of RT in BCa in vitro (classified by [46]); (c) cell line subtypes used in preclinical studies of radiotherapy in BCa in vivo (mice xenografts) (classified by [46]).…”

“…Then, Earl et al assigned the subtypes to a series of 40 BCa cell lines [ 47 ]. Our group has made more stringent classification [ 46 ] For example, five cell lines discussed in this review were identified as “basal” by Earl et al, while classified as “non-luminal, non-basal” by Shi et al [ 46 ]. These non-luminal, non-basal cell lines expressed epithelium to mesenchyme transition markers and do not express E-cadherin.…”

“…We identified 29 different BCa cell lines used in the experimental studies of RT. We assigned molecular subtypes to the human BCa using a classification recently proposed by our team [ 46 ] ( Table 1 , additional information regarding the mutational status and origin are available in Table S2 ). We found that from all the BCa cell lines used in RT studies, 26% are of luminal subtype, which is in contrast to 47% of human tumours considered having luminal features ( Figure 1 a,b).…”

“… Comparison of muscle-invasive bladder cancer (MIBC) subtype frequency in patient tumours and human bladder cancer (BCa) cell lines used in radiotherapy (RT) experimental studies in vitro and preclinical studies in vivo. ( a ) MIBC tumour subtypes (classified by [ 36 ]); ( b ) cell line subtypes used in experimental studies of RT in BCa in vitro (classified by [ 46 ]); ( c ) cell line subtypes used in preclinical studies of radiotherapy in BCa in vivo (mice xenografts) (classified by [ 46 ]). …”

KDM6A mutations promote acute cytoplasmic DNA release, DNA damage response and mitosis defects

“…A prevalent nonsense mutation is the Q555* with additional partial deletion of the IDR. This mutation site has been identi ed as an APOBEC3 hotspot (20). Other variants, like the moderately frequent Q333*, lack both IDR and JmjC and could potentially impair TPR8 functionality.…”