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Supplementary Figure legends

Supplementary Figure S1. Identification of AA signature by ultra-low coverage sequencing reads.

Supplementary Table S1: Clinicopathological and epidemiological patient data and urothelial tumor sample description. Supplementary Table S2: Complete list of annotated SBS identified in all LC-WES analyzed urothelial tumors (HiSeq2500 data only). Supplementary Table S3: GO and KEGG Classification of genes mutated by non-synonymous A:T>T:A mutations. Supplementary Table S4: List of cancer driver genes found recurrently mutated in the EN and Taiwan UTUC sample sets. Supplementary Table S5: GO and KEGG classification of the recurrently mutated cancer driver genes (see Figure 3B).

Supplementary Methods

International audience; Infectious disease outbreaks, such as 'Coronavirus disease 2019' (COVID-19), can constitute major global health threats with far-reaching consequences. As outbreaks develop, the international scientific community must provide high-quality scientific research-ready biological samples to solve the existing clinical and epidemiological questions to better combat the pandemic. Such examples are provided by dedicated biobank facilities, the latter collecting increasingly high volumes of biological samples. However, the more significant concentrations of infectious or potentially infectious biological materials can create a safety risk. The current short report describes the first attempt to identify the published scientific works on biobanking and safety. Three broad thematic areas have been identified: the physical security relevant to staff and sample integrity, the data safety aspects, and the governance parameters relating to the previous two. While the current publications reflect a broad alignment with existing standards and best practices in the biobanking field, they also demonstrate an opportunity for further in-depth work on this field in the post-COVID-19 era.

Recurrent upper tract urothelial carcinomas (UTUCs) arise in the context of nephropathy linked to exposure to the herbal carcinogen aristolochic acid (AA). Here we delineated the molecular programs underlying UTUC tumorigenesis in patients from endemic aristolochic acid nephropathy (AAN) regions in Southern Europe. We applied an integrative multiomics analysis of UTUCs, corresponding unaffected tissues and of patient urines. Quantitative microRNA (miRNA) and messenger ribonucleic acid (mRNA) expression profiling, immunohistochemical analysis by tissue microarrays and exome and transcriptome sequencing were performed in UTUC and nontumor tissues. Urinary miRNAs of cases undergoing surgery were profiled before and after tumor resection. Ribonucleic acid (RNA) and protein levels were analyzed using appropriate statistical tests and trend assessment. Dedicated bioinformatic tools were used for analysis of pathways, mutational signatures and result visualization. The results delineate UTUC-specific miRNA:mRNA networks comprising 89 miRNAs associated with 1, 862 target mRNAs, involving deregulation of cell cycle, deoxyribonucleic acid (DNA) damage response, DNA repair, bladder cancer, oncogenes, tumor suppressors, chromatin structure regulators and developmental signaling pathways. Key UTUC- specific transcripts were confirmed at the protein level. Exome and transcriptome sequencing of UTUCs revealed AA-specific mutational signature SBS22, with 68% to 76% AA-specific, deleterious mutations propagated at the transcript level, a possible basis for neoantigen formation and immunotherapy targeting. We next identified a signature of UTUC- specific miRNAs consistently more abundant in the patients' urine prior to tumor resection, thereby defining biomarkers of tumor presence. The complex gene regulation programs of AAN-associated UTUC tumors involve regulatory miRNAs prospectively applicable to noninvasive urine-based screening of AAN patients for cancer presence and recurrence.

The information on candidate cancer driver alterations available from public databases is often descriptive and of limited mechanistic insight, which poses difficulties for reliable distinction between true driver and passenger events. To address this challenge, we performed in-depth analysis of whole-exome sequencing data from cell lines generated by a barrier bypass-clonal expansion (BBCE) protocol. The employed strategy is based on carcinogen-driven immortalization of primary mouse embryonic fibroblasts and recapitulates early steps of cell transformation. Among the mutated genes were almost 200 COSMIC Cancer Gene Census genes, many of which were recurrently affected in the set of 25 immortalized cell lines. The alterations affected pathways regulating DNA damage response and repair, transcription and chromatin structure, cell cycle and cell death, as well as developmental pathways. The functional impact of the mutations was strongly supported by the manifestation of several known cancer hotspot mutations among the identified alterations. We identified a new set of genes encoding subunits of the BAF chromatin remodeling complex that exhibited Ras-mediated dependence on PRC2 histone methyltransferase activity, a finding that is similar to what has been observed for other BAF subunits in cancer cells. Among the affected BAF complex subunits, we determined Smarcd2 and Smarcc1 as putative driver candidates not yet fully identified by large-scale cancer genome sequencing projects. In addition, Ep400 displayed characteristics of a driver gene in that it showed a mutually exclusive mutation pattern when compared with mutations in the Trrap subunit of the TIP60 complex, both in the cell line panel and in a human tumor data set. We propose that the information generated by deep sequencing of the BBCE cell lines coupled with phenotypic analysis of the mutant cells can yield mechanistic insights into driver events relevant to human cancer development.

Humans are frequently exposed to acrylamide, a probable human carcinogen found in commonplace sources such as most heated starchy foods or tobacco smoke. Prior evidence has shown that acrylamide causes cancer in rodents, yet epidemiological studies conducted to date are limited and, thus far, have yielded inconclusive data on association of human cancers with acrylamide exposure. In this study, we experimentally identify a novel and unique mutational signature imprinted by acrylamide through the effects of its reactive metabolite glycidamide. We next show that the glycidamide mutational signature is found in a full one-third of approximately 1600 tumor genomes corresponding to 19 human tumor types from 14 organs. The highest enrichment of the glycidamide signature was observed in the cancers of the lung (88% of the interrogated tumors), liver (73%), kidney (>70%), bile duct (57%), cervix (50%), and, to a lesser extent, additional cancer types. Overall, our study reveals an unexpectedly extensive contribution of acrylamide-associated mutagenesis to human cancers.

Free circulating DNA (cfDNA) has been known for several decades. These small DNA fragments are released into the circulation from nucleated cells through necrosis, apoptosis and/or active secretion. These genomic fragments are mainly constitutional (nucleated blood cell DNA), but in patients with cancer, a fraction comes from tumor cells. Although poorly known in the field of thoracic oncology, quantitative and qualitative analysis of the cDNA is nevertheless of great interest. Total cfDNA concentration appears to be an independent prognostic factor in lung cancer. Although changes in total cfDNA concentration is not informative to assess the effectiveness of chemotherapy, following-up the fraction of mutated genes such as EGFR during therapy with tyrosine kinase inhibitors appears to be particularly promising for the early detection of disease progression. The use of cfDNA as liquid biopsy is also very promising for the non-invasive somatic molecular profile either at baseline either for sampling at follow-up. Thus, cfDNA is a very promising tool in thoracic oncology and its translation into practice should be developed quickly.

BackgroundIn Latin America (LA), there is a high incidence rate of breast cancer (BC) in premenopausal women, and the genomic features of these BC remain unknown. Here, we aim to characterize the molecular features of BC in young LA women within the framework of the PRECAMA study, a multicenter population-based case-control study on breast cancer in premenopausal women.MethodsPathological tumor tissues were collected from incident cases from four LA countries. Immunohistochemistry (IHC) was performed centrally for ER, PR, HER2, Ki67, EGFR, CK5/6 and p53 protein markers. Targeted deep sequencing was done on genomic DNA extracted from formalin-fixed paraffin-embedded (FFPE) tumour tissues and their paired blood samples to screen for somatic mutations in eight genes frequently mutated in BC. A subset of samples was analyzed by exome sequencing to identify somatic mutational signatures.ResultsThe majority of cases were positive for ER or PR (168/233; 72%) and there were 21% triple negative (TN) cases, mainly of basal type. Most tumors were positive for Ki67 (189/233; 81%). In 126 sequenced cases,TP53andPIK3CAwere the most frequently mutated genes (32.5% and 21.4%, respectively), followed byAKT1(9.5%).TP53mutations were more frequent in HER2-enriched and TN IHC subtypes, whilePIK3CA/AKT1mutations were more frequent in ER positive tumors, as expected. Interestingly, a higher proportion of G:C>T:A mutations was observed inTP53gene in PRECAMA cases compared to TCGA and METABRIC breast cancer series (27% vs 14%). Exome-wide mutational patterns in 10 TN cases revealed alterations in signaling transduction pathways and major contributions of mutational signatures caused by altered DNA repair pathways.ConclusionsThis pilot results on PRECAMA tumors gives a preview of the molecular features of premenopausal BC in LA. Although, the overall mutation burden was as expected from data in other populations, mutational patterns observed inTP53and exome-wide suggested possible differences in mutagenic processes giving rise to these tumors compared to other populations. Further omics analyses of a larger number of cases in the near future will allow investigating relationships between these molecular features and risk factors.

Plasma circulating cell-free (cf)DNA is of interest in oncology because it has been shown to contain tumour DNA and may thus be used as liquid biopsy. In nonsmall cell lung cancer (NSCLC), cfDNA quantification has been proposed for the monitoring and follow-up of patients. However, available studies are limited and need to be confirmed by studies with larger sample sizes and including patients who receive more homogenous treatments. Our objective was to assess the predictive and prognostic value of plasma cfDNA concentration in a large series of patients with NSCLC and treated with a standard chemotherapy regimen. We included samples from lung cancer patients recruited into the Pharmacogenoscan study. The cfDNA of 218 patients was extracted and quantified by fluorometry before and after two or three cycles of platinum-based chemotherapy. The association between baseline and post-chemotherapy concentrations and treatment response, assessed by RECIST (response evaluation criteria in solid tumours) or patient survival was analysed. Patients with high cfDNA concentrations (highest tertile) at baseline had a significantly worse disease-free and overall survival than those with lower concentrations (lowest and middle tertiles) (median overall survival 10 months (95% CI 10.7–13.9) versus 14.2 months (95% CI 12.6–15.8), respectively; p=0.001). In multivariate analysis, increased baseline concentration of cfDNA was an independent prognostic factor. However, we did not find any association between cfDNA concentration and response to treatment. cfDNA may be a biomarker for the assessment of prognosis in NSCLC. However, total concentration of cfDNA does not appear to predict chemotherapy response. Total cell-free DNA is not associated with chemotherapy response in advanced nonsmall cell lung cancer

Acrylamide, a probable human carcinogen, is ubiquitously present in the human environment, with sources including heated starchy foods, coffee and cigarette smoke. Humans are also exposed to acrylamide occupationally. Acrylamide is genotoxic, inducing gene mutations and chromosomal aberrations in various experimental settings. Covalent haemoglobin adducts were reported in acrylamide-exposed humans and DNA adducts in experimental systems. The carcinogenicity of acrylamide has been attributed to the effects of glycidamide, its reactive and mutagenic metabolite capable of inducing rodent tumors at various anatomical sites. In order to characterize the pre-mutagenic DNA lesions and global mutation spectra induced by acrylamide and glycidamide, we combined DNA-adduct and whole-exome sequencing analyses in an established exposure-clonal immortalization system based on mouse embryonic fibroblasts. Sequencing and computational analysis revealed a unique mutational signature of glycidamide, characterized by predominant T:A>A:T transversions, followed by T:A>C:G and C:G>A:T mutations exhibiting specific trinucleotide contexts and significant transcription strand bias. Computational interrogation of human cancer genome sequencing data indicated that a combination of the glycidamide signature and an experimental benzo[a]pyrene signature are nearly equivalent to the COSMIC tobacco-smoking related signature 4 in lung adenocarcinomas and squamous cell carcinomas. We found a more variable relationship between the glycidamide‐ and benzo[a]pyrene-signatures and COSMIC signature 4 in liver cancer, indicating more complex exposures in the liver. Our study demonstrates that the controlled experimental characterization of specific genetic damage associated with glycidamide exposure facilitates identifying corresponding patterns in cancer genome data, thereby underscoring how mutation signature laboratory experimentation contributes to the elucidation of cancer causation.A 40-word summaryInnovative experimental approaches identify a novel mutational signature of glycidamide, a metabolite of the probable human carcinogen acrylamide. The results may elucidate the cancer risks associated with exposure to acrylamide, commonly found in tobacco smoke, thermally processed foods and beverages.

Introduction The analysis of somatic mutational signatures in a single tumour can reveal clues on the natural history of this tumour, including past exposures to carcinogens. Characterising the mutational signatures of potential carcinogens used in industry may thus help identify cancers that are due to occupational exposure. Here, we used experimental models to define the mutational signatures of dichloromethane (DCM) and 1,2-dichloropropane (DCP), two solvents suspected to cause cholangiocarcinomas in occupational settings. Material and methods Experimentally induced tumours and matching normal tissues from mice exposed to DCP (by gavage), DCM (by gavage or inhalation) or DCP +DCM (by gavage), and spontaneous tumours from vehicle/sham-exposed mice were analysed by whole-exome sequencing. Somatic mutation calling was performed to define exome-wide mutation patterns induced by DCP and DCM in these assays. Mutation data obtained in biliary tract cancers of workers in the printing industry who have been exposed to DCP and DCM as single agents or as mixtures, as well as with public somatic mutation data from biliary tract cancers were mined for the presence of the experimentally defined DCP and DCM mutational signatures. Results and discussions Liver tumours from DCP and DCM exposed mice had distinct somatic mutations patterns compared to spontaneous liver tumours from vehicle/sham-exposed mice. While mutations in DCP-exposed mice were dominated by C:G>T:A transitions, the most frequent types of mutations in DCM-exposed mice were T:A>A:T and T:A>C:G substitutions. An average of 10.3 somatic mutations was observed per Mb in tumours from DCM-exposed mice, approximately 3-fold higher than in DCP-exposed or sham-treated mice. The mutation patterns found in DCP-exposed mice, but not DCM-exposed mice, presented some similarities with the mutational signature observed in cholangiocarcinomas of Japanese patients with occupational DCP/DCM exposure history. Conclusion These results show that the analysis of tumour genomes from mouse carcinogenicity assays can support the characterisation of mutational signatures of carcinogenic compounds relevant to human exposures.

Over 100 single nucleotide polymorphisms (SNP) are validated in the TP53 tumor suppressor gene. They define haplotypes, which may differ in their activities. Therefore, mutation in cancer may occur at different rates depending upon haplotypes. However, these associations may be masked by differences in mutations types and causes of mutagenesis. We have analyzed the associations between 19 SNPs spanning the TP53 locus and a single specific aflatoxin-induced TP53 mutation (R249S) in 85 in hepatocellular carcinoma cases and 132 controls from Thailand. An association with R249S mutation (P = 0.007) was observed for a combination of two SNPs (rs17882227 and rs8064946) in a linkage disequilibrium block extending from upstream of exon 1 to the first half of intron 1. This domain contains two coding sequences overlapping with TP53 (WRAP53 and Hp53int1) suggesting that sequences in TP53 intron 1 encode transcripts that may modulate R249S mutation rate in HCC.

Aflatoxin B1 (AFB1) is a mutagen and IARC (International Agency for Research on Cancer) Group 1 carcinogen that causes hepatocellular carcinoma (HCC). Here, we present the first whole-genome data on the mutational signatures of AFB1 exposure from a total of >40,000 mutations in four experimental systems: two different human cell lines, in liver tumors in wild-type mice, and in mice that carried a hepatitis B surface antigen transgene—this to model the multiplicative effects of aflatoxin exposure and hepatitis B in causing HCC. AFB1 mutational signatures from all four experimental systems were remarkably similar. We integrated the experimental mutational signatures with data from newly sequenced HCCs from Qidong County, China, a region of well-studied aflatoxin exposure. This indicated that COSMIC mutational signature 24, previously hypothesized to stem from aflatoxin exposure, indeed likely represents AFB1 exposure, possibly combined with other exposures. Among published somatic mutation data, we found evidence of AFB1 exposure in 0.7% of HCCs treated in North America, 1% of HCCs from Japan, but 16% of HCCs from Hong Kong. Thus, aflatoxin exposure apparently remains a substantial public health issue in some areas. This aspect of our study exemplifies the promise of future widespread resequencing of tumor genomes in providing new insights into the contribution of mutagenic exposures to cancer incidence.

In regions with high prevalence of chronic hepatitis B virus (HBV) infection and dietary aflatoxin B(1) (AFB(1)) exposure, hepatocellular carcinomas (HCCs) often contain TP53 mutation at codon 249 (R249S). Furthermore, a C-terminal truncated HBx protein expressed from hepatocyte integrated HBV is associated with HCC development. This study evaluates the association between R249S and HBX status in relation to HCC in West African population. HBX (complete or 3'-truncated) and HBS genes were assessed by PCR in cell-free DNA (CFDNA) from plasma of subjects recruited in a hospital-based case-control study (325 controls, 78 cirrhotic patients and 198 HCC cases) conducted in The Gambia. These samples had been previously analyzed for R249S and HBV serological status. Complete HBX sequence was frequently detected in CFDNA of HCC-R249S positive (77%, 43/56) compared with HCC-R249S-negative cases (44%, 22/50). Conversely, the proportion of 3'-truncated HBX gene was significantly higher in HCC-R249S negative than positive cases (34%, 17/50, compared with 12%, 7/56) (χ(2) = 12.12; P = 0.002; distribution of R249S negative and positive according to HBX status). Occult HBV infection (detected by PCR) was present in 24% of HCC previously considered as negative by HBV serology. Moreover, HBV mutation analysis revealed that double mutation at nucleotides 1762(T)/1764(A) was associated with diagnosis of cirrhosis or HCC {cirrhosis: odds ratio (OR): 9.50 [95% confidence interval (CI) 1.50-60.11]; HCC: OR: 11.29 [95% CI 2.07-61.47]}. These findings suggest that in HCC from The Gambia, complete HBX sequences are often associated with the presence of TP53 R249S mutation.

Infection by hepatitis B virus (HBV) and dietary exposure to aflatoxin B1 (AFB1) are the main risk factors for the development of chronic liver disease and hepatocellular carcinoma (HCC). How these factors cooperate is still largely unknown. AFB1 activation leads to DNA adduction and mutagenesis, with a specific mutation at codon 249 in TP53 (p.R249S). So far, only limited studies have addressed the effects of AFB1 on HBV replication. We have analysed the effects of both risk factors on p53 induction during HBV infection in HepaRG, a cell line with hepatocyte-like morphology that metabolizes AFB1 and supports HBV infection. Exposure to AFB1 up to 5 µM induced a downregulation of HBV replication after 48 h, as measured by a decrease in viral antigens in the culture medium (HBsAg, HBeAg and large envelope protein) and in intracellular levels of HBV transcripts, DNA and HBsAg. Conversely, HBV infection did not significantly modify AFB1-DNA adduct formation or repair as assessed by immunodot-blot assay, and the induction of p53 in response to AFB1 was similar in infected and non-infected HepaRG cells. Overall, our results suggest that AFB1 exposure decreases HBV replication, whereas DNA damage by AFB1 and subsequent p53 induction is not affected by the presence of the virus. Thus, in HepaRG cell line, AFB1 and HBV do not cooperate to increase DNA damage by AFB1. Further studies on the effects of both factors in a context of chronicity are needed to better understand synergistic effects.

Background: Chronic hepatitis B virus (HBV) infection and dietary aflatoxin B1 (AFB1) exposure are etiological factors for hepatocellular carcinoma (HCC) in countries with hot, humid climates. HCC often harbors a TP53 (tumor protein p53) mutation at codon 249 (R249S). In chronic carriers, 1762T/1764A mutations in the HBV X gene are associated with increased HCC risk. Both mutations have been detected in circulating cell-free DNA (CFDNA) from asymptomatic HBV carriers. Objective: We evaluated seasonal variation in R249S and HBV in relation to AFB1 exposure. Methods: R249S was quantitated by mass spectrometry in CFDNA in a cross-sectional survey of 473 asymptomatic subjects (237 HBV carriers and 236 noncarriers) recruited in three villages in the Gambia over a 10-month period. 1762T/1764A HBV mutations were detected by quantitative polymerase chain reaction. In addition, the HBV S gene was sequenced in 99 subjects positive for HBV surface antigen (HBsAg). Results: We observed a seasonal variation of serum R249S levels. Positivity for R249S and average concentration were significantly higher in HBsAg-positive subjects surveyed during April–July (61%; 5,690 ± 11,300 R249S copies/mL serum) than in those surveyed October–March [32% and 480 ± 1,030 copies/mL serum (odds ratio = 3.59; 95% confidence interval: 2.05, 6.30; p < 0.001)]. Positivity for HBV e antigen (HBeAg) (a marker of HBV replication) and viral DNA load also varied seasonally, with 15–30% of subjects surveyed between April and June HBeAg positive, compared with < 10% surveyed during other months. We detected 1762T/1764A mutations in 8% of carriers, half of whom were positive for R249S. We found HBV genotype E in 95 of 99 HBsAg-positive subjects. Conclusion: R249S is detectable in CFDNA of asymptomatic subjects. Evidence of temporal and quantitative variations suggests an interaction among AFB1 exposure, HBV positivity, and replication on TP53 mutation formation or persistence.

Cancers of the upper aerodigestive tract [(UADT): oral cavity, pharynx, larynx and oesophagus] have high incidence rates in some parts of South America. Alterations in the TP53 gene are common in these cancers. In our study, we have estimated the prevalence and patterns of TP53 mutations (exons 4-10) in 236 UADT tumours from South America in relation to lifestyle risk factors, such as tobacco smoking and alcohol drinking. Moreover, we have conducted a pilot study of EGFR mutations (exons 18-21) in 45 tumours from the same population. TP53 mutation prevalence was high: 59% of tumours were found to carry mutant TP53. We found an association between TP53 mutations and tobacco smoking and alcohol drinking. The mutation rate increased from 38% in never-smokers to 66% in current smokers (P-value for trend 5 0.09). G:C>T:A transversions were found only in smokers (15%). Alcohol drinkers carried more G:C>A:T transitions (P 5 0.08). Non-exposed individuals were more probable to carry G:C>A:T transitions at CpG sites (P 5 0.01 for neversmokers and P < 0.001 for never-drinkers). EGFR mutations were found in 4% of cases. Inactivation of TP53 by mutations is a crucial molecular event in the UADT carcinogenesis and it is closely related to exposure to lifestyle risk factors. EGFR mutations do not appear to be a common event in UADT carcinogenesis in this population. © The Author 2009. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oxfordjournals.org.

Loss of imprinting (LOI) of the insulin-like growth factor II (IGFII) gene is a frequent phenomenon in colorectal tumor tissues. Previous reports indicated that subjects with colorectal neoplasias show LOI of IGFII in circulating lymphocytes. Furthermore, LOI of IGFII is strongly related to the methylation of a differentially methylated region (DMR) in intron 2 of IGFII, suggesting that the methylation status could serve as a biomarker for early detection. Thus, hypermethylation of this DMR, even at a systemic level, e.g., in lymphocyte DNA, could be used for screening for colon cancer. To validate this, we performed a case-control study of 97 colon cancer cases and 190 age-matched and gender-matched controls, nested within the prospective Northern Sweden Health and Disease Study cohort. Methylation levels of the IGFII-DMR in lymphocyte DNA were measured at two specific CpG sites of the IGFII-DMR using a mass-spectrometric method called short oligonucleotide mass analysis, the measurements of which showed high reproducibility between replicate measurements for the two CpG sites combined and showed almost perfect validity when performed on variable mixtures of methylated and unmethylated standards. Mean fractions of CpG methylation, for the two CpG sites combined, were identical for cases and controls (0.47 and 0.46, respectively; Pdifference = 0.75), and logistic regression analyses showed no relationship between colon cancer risk and quartile levels of CpG methylation. The results from this study population do not support the hypothesis that colon cancer can be predicted from the different degrees of methylation of DMR in the IGFII gene from lymphocyte DNA. [Cancer Res 2009;69(13):5400–5]

Hepatocellular carcinoma (HCC) has a high mortality in East Asia and Sub-Saharan Africa, two regions where the main etiologic factors are chronic infections with hepatitis B virus and dietary exposure to aflatoxin. A single base substitution at the third nucleotide of codon 249 of TP53 (R249S) is common in HCC in these regions and has been associated with aflatoxin-DNA adducts. To determine whether R249S may be detected in plasma DNA before HCC diagnosis, we conducted a case-control study nested in a cohort of adult chronic hepatitis B virus carriers from Qidong County, People's Republic of China. Of the 234 plasma specimens that yielded adequate DNA, only 2 (0.9%) were positive for R249S by restriction fragment length polymorphisms, and both of them were controls. Of the 249 subjects tested for aflatoxin-albumin adducts, 168 (67%) were positive, with equal distribution between cases and controls. Aflatoxin-albumin adduct levels were low in the study, suggesting an overall low ongoing exposure to aflatoxin in this cohort. The R249S mutation was detected in 11 of 18 (61%) available tumor tissues. To assess whether low levels of mutant DNA were detectable in pre-diagnosis plasma, 14 plasma specimens from these patients were analyzed by short oligonucleotide mass analysis. Nine of them (64%) were found to be positive. Overall, these results suggest that HCC containing R249S can occur in the absence of significant recent exposure to aflatoxins. The use of short oligonucleotide mass analysis in the context of low ongoing aflatoxin exposure may allow the detection of R249S in plasma several months ahead of clinical diagnosis. (Cancer Epidemiol Biomarkers Prev 2009;18(5):1638–43)

Background Hepatitis B (HB) infection is common in Mali. However, there is little information on molecular and biochemical characteristics of HB carriers. Methods A group of 1466 adult volunteers was recruited in the district of Bamako. Confirmed HB carriers were tested for HB viral load by quantitative PCR and HBV was genotyped by sequencing of HBS. Fibrosis and hepatitis activity were measured using the Fibrotest-Actitest. A mutation of TP53 at codon 249 (R249S), specific for exposure to aflatoxin, was detected in cell-free DNA extracted from plasma. Results Overall, 276 subjects were HBsAg-positive (18.8%). Among 152 subjects tested for HBV load, 49 (32.2%) had over 104 copies/mL and 16 (10.5%) had levels below the limit of detection. The E genotype was found in 91.1% of carriers. Fibrotest scores ≥ F2 were observed in 52 subjects (35.4%). Actitest scores ≥ A2 were detected in 15 subjects (10.2%) and were correlated with Fibrotest scores (p = 0.0006). Among 105 subjects tested, 60% had detectable levels of R249S copies (>40 copies/mL plasma). Conclusion Chronic HB carriage in adults in Bamako district is well over epidemic threshold. About 1/3 of carriers have moderate to severe liver fibrosis and 60% have detectable aflatoxin-related TP53 R249S mutation. These results support introduction of anti-HB therapies to reduce the progression towards severe liver disease.

Background: Dietary exposure to cytotoxic and carcinogenic aristolochic acid (AA) causes severe nephropathy typically associated with urologic cancers. Monitoring of AA exposure uses biomarkers such as aristolactam-DNA adducts, detected by mass spectrometry in the kidney cortex, or the somatic A>T transversion pattern characteristic of exposure to AA, as revealed by previous DNA-sequencing studies using fresh-frozen tumors. Methods: Here, we report a low-coverage whole-exome sequencing method (LC-WES) optimized for multisample detection of the AA mutational signature, and demonstrate its utility in 17 formalin-fixed paraffin-embedded urothelial tumors obtained from 15 patients with endemic nephropathy, an environmental form of AA nephropathy. Results: LC-WES identified the AA signature, alongside signatures of age and APOBEC enzyme activity, in 15 samples sequenced at the average per-base coverage of approximately 10×. Analysis at 3 to 9× coverage revealed the signature in 91% of the positive samples. The exome-wide distribution of the predominant A>T transversions exhibited a stochastic pattern, whereas 83 cancer driver genes were enriched for recurrent nonsynonymous A>T mutations. In two patients, pairs of tumors from different parts of the urinary tract, including the bladder, harbored overlapping mutation patterns, suggesting tumor dissemination via cell seeding. Conclusions: LC-WES analysis of archived tumor tissues is a reliable method applicable to investigations of both the exposure to AA and its biologic effects in human carcinomas. Impact: By detecting cancers associated with AA exposure in high-risk populations, LC-WES can support future molecular epidemiology studies and provide evidence-base for relevant preventive measures. Cancer Epidemiol Biomarkers Prev; 24(12); 1873–81. ©2015 AACR.

Aristolochic acid (AA) is a potent nephrotoxin and carcinogen (IARC Group 1) associated with urothelial, hepatobiliary, and renal carcinomas. Exposure to AA from dietary intake of traditional herbal medicines containing Aristolochia species poses a global health hazard, yet also an opportunity for disease prevention. Molecular epidemiology studies employing sensitive and specific biomarkers for screening of populations at risk are thus warranted. Genome-scale studies established a characteristic mutational signature of AA. Exploiting its unique features (predominant A>T transversions), alongside the knowledge of key affected cancer genes, we devised a cost-effective targeted resequencing (TRseq) method to detect AA etiology in upper tract urothelial carcinoma (UTUC) samples from South Korean patients. Aristolochia species (e.g., A. contorta, Bunge) are included in the Korean traditional pharmacopoeia although the local occurrence of AA-associated cancers remains unexplored. DNA from archived, paired kidney and UTUC samples of 16 Korean patients was used for adduct and TRseq analyses. Thirty-one cancer genes, previously found recurrently mutated in AA-induced UTUCs from Asia and Europe, were selectively captured using SeqCap EZ enrichment kit (Nimblegen Roche) and sequenced using Illumina MiSeq. Sets of UTUCs from Croatia (harboring AA signature) and from US patients (unlikely to be exposed) were included as respective positive and negative controls. Somatic variants were called against the patient-matched non-tumor DNA, using two distinct variant calling strategies. Three out of 16 kidney cortex samples tested positive for aristolactam-dA (AL-dA) adducts, indicative of past exposure to AA. One UTUC sample with A>T-mutated TP53 harbored 32 A>T mutations across the 31-gene panel, and another 8 cases, 3 of which were adduct-positive, harbored lower counts of A>T mutations (up to 3 per gene panel per sample). The allelic frequencies of the considered A>T mutations ranged between 10-83%. The positive control samples exhibited a strong A>T signal while the US samples were all A>T-negative, as expected. We propose that the TRseq approach can detect AA-driven mutagenesis in UTUC tumors, providing a sensitive complement to the AL-dA adduct analysis, at low per-sample cost. Importantly, the presence of adducts and AA signature mutations in South Korean UTUC cases highlights a previously uncharacterized population at risk. The future use of TRseq, either alone or in combination with adduct analysis, can address exposure to AA in extended UTUC case series, thereby assisting in the design of preventive measures against this global public health hazard. Funding: IARC; SW is Recipient of research support from the Drs. Martin & Dorothy Spatz Charitable Foundation; Henry and Marsha Laufer grant to KGD, VSS, APG. Citation Format: Shuhan Wang, Daeeun Choi, Jaesung Lim, Kathleen G. Dickman, Magali Olivier, Stephanie Villar, Viktoriya S. Sidorenko, Maude Ardin, Byeong H. Yun, Robert J. Turesky, Minho Shong, Jiri Zavadil, Arthur P. Grollman. New molecular evidence associating exposure to aristolochic acid with urothelial cancers in South Korean patients: Implications for global public health risk linked to carcinogenic herbal medicines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4252. doi:10.1158/1538-7445.AM2017-4252

Exposure to aristolochic acid (AA, IARC Group 1 carcinogen) present in some traditional herbal medicines leads to aristolochic acid nephropathy (AAN), often complicated by development of multiple urothelial carcinomas of sequential onset. We used genome-scale mutational signature analysis of multiple urinary tract tumors of AAN cases from a unique patient group to determine the relationships of the patients’ late-onset second cancers to the AA exposure as well as to the first cancers. Aristolactam-DNA adduct-positive AAN patients (n=4) who developed cancer within 8 years following the initial exposure to AA were chosen for analysis of their first cancers (upper tract urothelial carcinomas, UTUC) and second cancers of delayed onset (1-9 years after first-cancer diagnosis, involving the bladder or ureteral meatus). All patients had received a kidney transplant before developing the second cancers and had a functional renal graft prior to prophylactic nephroureterectomy. Genomic DNAs were isolated from FFPE sections of the renal cortex and from the upper and lower tract tumors of each patient using laser capture micro-dissection or macro-dissection of the tumor areas. Low-coverage (~15x) exome 100-bp paired-end sequencing was performed using Illumina HiSeq2500. Customized variant calling identified somatic variants absent in non-tumor tissues. Non-negative matrix factorization was applied to extract mutational signatures in the tumor tissues. In all cases, we established the mutational signature of AA (the COSMIC signature 22) in the first UTUC as well as second cancers involving the bladder or lower ureter (meatus). Additionally, the first and second cancers harbored considerable overlaps in exposure-specific (A>T) somatic mutations. This finding provides evidence that the delayed onset of bladder urothelial carcinomas in AAN patients is likely due to distal seeding of cancer cells originating from the primary UTUC tumors. Our first-of-its-kind study addresses the risk as well as mechanistic factors leading to the second, late-onset bladder urothelial carcinomas following kidney transplantation and primary UTUC development. Our results underline the importance of long-term bladder follow-up in high-risk populations with established or suspected AA exposure. Funding: IARC; NYU Genome Technology Center is partially supported by the NIH/NCI (P30CA016087) grant. Citation Format: Xavier Castells, Maude Ardin, Sandrine Rorive, Nilufer Broeders, Yan Song, Stephanie Villar, Christine Carreira, Pierre-Paul Bringuier, Adriana Heguy, Thierry Quackels, Thierry Roumeguere, Joelle Nortier, Jiri Zavadil. Tracking the genetic relationship between first and late-onset second urothelial cancers by mutational signature analysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5738. doi:10.1158/1538-7445.AM2017-5738

Purpose: Tumor somatic mutation analysis is part of the standard management of metastatic lung cancer. However, physicians often have to deal with small biopsies and consequently with challenging mutation testing. Circulating free DNA (cfDNA) is a promising tool for accessing the tumor genome as a liquid biopsy. Here, we evaluated next-generation sequencing (NGS) on cfDNA samples obtained from a consecutive series of patients for the screening of a range of clinically relevant mutations. Experimental Design: A total of 107 plasma samples were collected from the BioCAST/IFCT-1002 lung cancer study (never-smokers cohort). Matched tumor DNA (tDNA) was obtained for 68 cases. Multiplex PCR-based assays were designed to target specific coding regions in EGFR, KRAS, BRAF, ERBB2, and PI3KCA genes, and amplicon sequencing was performed at deep coverage on the cfDNA/tDNA pairs using the NGS IonTorrent Personal Genome Machine Platform. Results: CfDNA concentration in plasma was significantly associated with both stage and number of metastatic sites. In tDNA, 50 mutations (36 EGFR, 5 ERBB2, 4 KRAS, 3 BRAF, and 2 PIK3CA) were identified, of which 26 were detected in cfDNA. Sensitivity of the test was 58% (95% confidence interval, 43%–71%) and the estimated specificity was 87% (62%–96%). Conclusion: These data demonstrate the feasibility and potential utility of mutation screening in cfDNA using IonTorrent NGS for the detection of a range of tumor biomarkers in patients with metastatic lung cancer. Clin Cancer Res; 20(17); 4613–24. ©2014 AACR.

Background Esophageal squamous cell carcinoma (ESCC) shows geographic variations in incidence, with high incidences (>50/105 person-years) in central Asia, including North Eastern Iran (Golestan) and Northern India (Kashmir). In contrast to Western countries, smoking does not appear to be a significant risk factor for ESCC in central Asia. In lung adenocarcinoma, activating mutations in the gene encoding epidermal growth factor receptor (EGFR) are frequent in tumors of never smokers of Asian origin, predicting therapeutic sensitivity to Egfr-targeting drugs. Methods In this study 152 cases of histologically confirmed ESCC from Iran (Tehran and Golestan Province) and North India (Kashmir Valley) have been analyzed for EGFR mutation by direct sequencing of exons 18–21. Egfr protein expression was evaluated by immunohistochemistry in 34 samples from Tehran and HER2 mutations were analyzed in 54 cases from Kashmir. Results A total of 14 (9.2%) EGFR variations were detected, including seven variations in exons. Among those, four (2.6%) were already documented in lung cancers, two were reported as polymorphisms and one was a potentially new activating mutation. All but one variation in introns were previously identified as polymorphisms. Over-expression of Egfr was detected in 22/34 (65%) of tested cases whereas no HER2 mutation was found in 54 cases from Kashmir. Conclusion Overall, EGFR mutations appear to be a rare event in ESCC in high incidence areas of central Asia, although a very small proportion of cases may harbor mutations predicting sensitivity to anti-Egfr drugs.

Villar, Stephanie Ortiz-Cuaran, Sandra Abedi-Ardekani, Behnoush Gouas, Doriane Nogueira da Costa, Andre Plymoth, Amelie Khuhaprema, Thiravud Kalalak, Anant Sangrajrang, Suleeporn Friesen, Marlin D Groopman, John D Hainaut, Pierre eng Research Support, Non-U.S. Gov't 2012/06/08 06:00 PLoS One. 2012;7(6):e37707. doi: 10.1371/journal.pone.0037707. Epub 2012 Jun 4.; International audience; Primary Liver Cancer (PLC) is the leading cause of death by cancer among males in Thailand and the 3(rd) among females. Most cases are hepatocellular carcinoma (HCC) but cholangiocarcinomas represent between 4 and 80% of liver cancers depending upon geographic area. Most HCC are associated with chronic infection by Hepatitis B Virus while a G ―> T mutation at codon 249 of the TP53 gene, R249S, specific for exposure to aflatoxin, is detected in tumors for up to 30% of cases. We have used Short Oligonucleotide Mass Analysis (SOMA) to quantify free circulating R249S-mutated DNA in plasma using blood specimens collected in a hospital case:control study. Plasma R249S-mutated DNA was detectable at low concentrations (>/= 67 copies/mL) in 53 to 64% of patients with primary liver cancer or chronic liver disease and in 19% of controls. 44% of patients with HCC and no evidence of cirrhosis had plasma concentrations of R249S-mutated DNA >/= 150 copies/mL, compared to 21% in patients with both HCC and cirrhosis, 22% in patients with cholangiocarcinoma, 12% in patients with non-cancer chronic liver disease and 3% of subjects in the reference group. Thus, plasma concentrations of R249S-mutated DNA >/= 150 copies/mL tended to be more common in patients with HCC developing without pre-existing cirrhosis (p = 0.027). Overall, these results support the preferential occurrence of R249S-mutated DNA in HCC developing in the absence of cirrhosis in a context of HBV chronic infection.

The inactivation of the p53 tumor suppressor pathway, which often occurs through mutations in TP53 (encoding tumor protein 53) is a common step in human cancer. However, in melanoma-A highly chemotherapy-resistant disease-TP53 mutations are rare, raising the possibility that this cancer uses alternative ways to overcome p53-mediated tumor suppression. Here we show that Mdm4 p53 binding protein homolog (MDM4), a negative regulator of p53, is upregulated in a substantial proportion (∼65%) of stage I-IV human melanomas and that melanocyte-specific Mdm4 overexpression enhanced tumorigenesis in a mouse model of melanoma induced by the oncogene Nras. MDM4 promotes the survival of human metastatic melanoma by antagonizing p53 proapoptotic function. Notably, inhibition of the MDM4-p53 interaction restored p53 function in melanoma cells, resulting in increased sensitivity to cytotoxic chemotherapy and to inhibitors of the BRAF (V600E) oncogene. Our results identify MDM4 as a key determinant of impaired p53 function in human melanoma and designate MDM4 as a promising target for antimelanoma combination therapy. © 2012 Nature America, Inc. All rights reserved.

High incidence of HCC is mostly due to the combination of two major risk factors, chronic infection with hepatitis B (HBV) and/or C (HCV) viruses and exposure to the mycotoxin aflatoxin B1, which induces a particular mutation at codon 249 in TP53 (R249S). Eight genotypes of HBV are diversely found in high and low incidence areas. Regardless of documented strong associations between TP53 R249S mutation and HBV genotypes B, C, A or E, there is no report of such association for genotype D despite of the presence of aflatoxin in areas with high prevalence of HBV genotype D. In Iran, 3% of the population is chronically infected with HBV, predominantly genotype D. Twenty-one histologically confirmed HCC cases from Iran were analyzed for TP53 R249S and HBV double mutations 1762T/1764A, hallmarks of more pathogenic forms of HBV. We did not detect any of these mutations. In addition, we report the only case identified so far carrying both R249S mutation and chronic HBV genotype D, a patient from The Gambia in West Africa. This paper suggests that association between HBV genotype D and aflatoxin-induced TP53 mutation is uncommon, explaining the relatively lower incidence of HCC in areas where genotype D is highly prevalent.

Hepatocellular carcinoma (HCC) may develop according to two major pathways, one involving HBV infection and TP53 mutation and the other characterized by HCV infection and CTNNB1 mutation. We have investigated HBV/HCV infections and TP53/CTNNB1 mutations in 26 HCC patients from Thailand. HBV DNA (genotype B or C) was detected in 19 (73%) of the cases, including 5 occult infections and 3 coinfections with HCV. TP53 and CTNNB1 mutations were not mutually exclusive, and most of TP53 mutations were R249S, suggesting a significant impact of aflatoxin-induced mutagenesis in HCC development.

Renal-cell carcinomas (RCC) are frequent in central and eastern Europe and the reasons remain unclear. Molecular mechanisms, except for VHL, have not been much investigated. We analysed 361 RCCs (334 clear-cell carcinomas) from a multi-centre case-control study for mutations in TP53 (exons 5-9 in the whole series and exons 4 and 10 in a pilot subset of 60 tumours) and a pilot 50 tumours for mutations in EGFR (exons 18-21) or KRAS (codon 12) in relation to VHL status. TP53 mutations were detected in 4% of clear-cell cases, independently of VHL mutations. In non-clear-cell carcinomas, they were detected in 11% of VHL-wild-type tumours and in 0% of tumours with VHL functional mutations. No mutations were found in EGFR or KRAS. We conclude that mutations in TP53, KRAS, or EGFR are not major contributors to the RCC development even in the absence of VHL inactivation. The prevalence of TP53 mutations in relation to VHL status may differ between clear-cell and other renal carcinomas. © 2010 Elsevier Ireland Ltd.

Triple negative breast cancer (TNBC), defined by the lack of expression of the estrogen receptor (ER), progesterone receptor (PR) and human epidermal receptor 2 (HER2), is an aggressive form of breast cancer (BC) that is more prevalent in certain populations, in particular in low and middle-income regions. The detailed molecular features of TNBC in these regions remain unexplored as samples are mostly accessible as formalin-fixed paraffin embedded (FFPE) archived tissues, a challenging material for advanced genomic and transcriptomic studies. We performed an integrative genomic analysis on FFPE samples from TNBC patients to identify molecular programs associated with this disease in an understudied population. We implemented whole exome sequencing of 12 tumor tissues and blood pairs, complemented with miRNA and mRNA profiling of the tumor tissues, using FFPE archived pathological samples from Mexican women. Sequencing analyses found TP53 and RB1 genes as the most frequently mutated. Transcriptional programs were characterized by the overexpression of growth-promoting signals (EGFR, PDGFR, VEGF, PIK3CA, FOXM1), the repression of cell cycle control pathways (TP53, RB1), the deregulation of DNA-repair pathways, and alterations in epigenetic modifiers through miRNA:mRNA network de-regulation. The molecular programs identified were typical of those described in basal-like tumors in other populations. This work demonstrates the utility of archival clinical samples for advanced integrated genomics analyses. It thus opens up opportunities for investigating molecular features of tumors from regions where only FFPE tissues are available, allowing the development of treatment strategies and of retrospective studies on the exploration of the geographic diversity of BC. Citation Format: Felipe Vaca-Paniagua, Rosa María Alvarez-Gomez, Carlos Perez-Plasencia, Hector Aquiles Maldonado-Martínez, Veronica Fragoso-Ontiveros, Federico Lasa-Gonsebatt, Luis Alonso Herrera, David Cantú, Enrique Bargallo-Rocha, Alejandro Mohar, Geoffroy Durand, Nathalie Forey, Catherine Voegele, Maxime Vallee, Florence Le Calvez-Kelm, James McKay, Maude Ardin, Stephanie Villar, Jiri Zavadil, Magali Olivier. Revealing the molecular portrait of triple negative breast tumors from an understudied population through omics analysis of formalin-fixed and paraffin-embedded tissues. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4748. doi:10.1158/1538-7445.AM2015-4748

Triple negative breast cancer (TNBC), defined by the lack of expression of the estrogen receptor, progesterone receptor and human epidermal receptor 2, is an aggressive form of breast cancer that is more prevalent in certain populations, in particular in low- and middle-income regions. The detailed molecular features of TNBC in these regions remain unexplored as samples are mostly accessible as formalin-fixed paraffin embedded (FFPE) archived tissues, a challenging material for advanced genomic and transcriptomic studies. Using dedicated reagents and analysis pipelines, we performed whole exome sequencing and miRNA and mRNA profiling of 12 FFPE tumor tissues collected from pathological archives in Mexico. Sequencing analyses of the tumor tissues and their blood pairs identified TP53 and RB1 genes as the most frequently mutated genes, with a somatic mutation load of 1.7 mutations/exome Mb on average. Transcriptional analyses revealed an overexpression of growth-promoting signals (EGFR, PDGFR, VEGF, PIK3CA, FOXM1), a repression of cell cycle control pathways (TP53, RB1), a deregulation of DNA-repair pathways, and alterations in epigenetic modifiers through miRNA:mRNA network de-regulation. The molecular programs identified were typical of those described in basal-like tumors in other populations. This work demonstrates the feasibility of using archived clinical samples for advanced integrated genomics analyses. It thus opens up opportunities for investigating molecular features of tumors from regions where only FFPE tissues are available, allowing retrospective studies on the search for treatment strategies or on the exploration of the geographic diversity of breast cancer.

A mutation in codon 249 of the TP53 gene (249Ser), related to aflatoxin B1 exposure, has previously been associated with hepatocellular carcinoma risk. Using a novel internal standard plasmid, plasma concentrations of 249Ser-mutated DNA were quantified by electrospray ionization mass spectrometry in 89 hepatocellular carcinoma cases, 42 cirrhotic patients, and 131 nonliver diseased control subjects, all from highly aflatoxin-exposed regions of The Gambia. The hepatocellular carcinoma cases had higher median plasma concentrations of 249Ser (2,800 copies/mL; interquartile range: 500-11,000) compared with either cirrhotic (500 copies/mL; interquartile range: 500-2,600) or control subjects (500 copies/mL; interquartile range: 500-2,000; P < 0.05). About half (52%) of the hepatocellular carcinoma cases had >2,500 copies of 249Ser/mL plasma, corresponding to the prevalence of this mutation in liver tumors in The Gambia. In comparison, only 15% of control group and 26% of cirrhotic participants exceeded this level (P < 0.05). Further subset analysis revealed a statistically significant, quantitative relation between diagnosis of hepatocellular carcinoma and levels of 249Ser detected at 2,501 to 10,000 copies/mL plasma (odds ratio, 3.8; 95% confidence interval, 1.3-10.9) and at >10,000 copies/mL plasma (odds ratio, 62; 95% confidence interval, 4.7-820) when compared with control subjects and after adjusting for age, gender, recruitment site, hepatitis B and C serologic status, and total DNA concentration. Levels of >10,000 copies of 249Ser/mL plasma were also significantly associated with the diagnosis of hepatocellular carcinoma (odds ratio, 15; 95% confidence interval, 1.6-140) when compared with cirrhotic patients. Potential applications for the quantification of 249Ser DNA in plasma include estimation of long-term, cumulative aflatoxin exposure and selection of appropriate high-risk individuals for targeted intervention. (Cancer Epidemiol Biomarkers Prev 2005;14(12):2956–62)

11046 Background: Plasma circulating free DNA (cfDNA) has raised interest in oncology because it has been shown to contain tumor DNA and may thus be used as liquid biopsy. In non-small cell lung ca...

International audience; Background: The natural history of chronic HBV infection in sub-Saharan Africa is unknown. Data are required to inform WHO guidelines that are currently based on studies in Europe and Asia.Methods: Between 1974 and 2008, serosurveys were repeated in two Gambian villages, and an open cohort of treatment-naive chronic HBV carriers was recruited. Participants were followed to estimate the rates of hepatitis B e (HBeAg) and surface antigen (HBsAg) clearance and incidence of hepatocellular carcinoma (HCC). In 2012-2013, a comprehensive liver assessment was conducted to estimate the prevalence of severe liver disease.Results: 405 chronic carriers (95% genotype E), recruited at a median age of 10.8 years, were followed for a median length of 28.4 years. Annually, 7.4% (95% CI 6.3% to 8.8%) cleared HBeAg and 1.0% (0.8% to 1.2%) cleared HBsAg. The incidence of HCC was 55.5/100 000 carrier-years (95% CI 24.9 to 123.5). In the 2012-2013 survey (n=301), 5.5% (95% CI 3.4% to 9.0%) had significant liver fibrosis. HBV genotype A (versus E), chronic aflatoxin B1 exposure and an HBsAg-positive mother, a proxy for mother-to-infant transmission, were risk factors for liver fibrosis. A small proportion (16.0%) of chronic carriers were infected via mother-to-infant transmission; however, this population represented a large proportion (63.0%) of the cases requiring antiviral therapy.Conclusions: The incidence of HCC among chronic HBV carriers in West Africa was higher than that in Europe but lower than rates in East Asia. High risk of severe liver disease among the few who are infected by their mothers underlines the importance of interrupting perinatal transmission in sub-Saharan Africa.