The investigation of the origin of Southern Tunisians using HLA genes.

The south of Tunisia is characterized by marked ethnic diversity, highlighted by the coexistence of native Berbers with Blacks, Jews and Arab-speaking populations. Despite this heterogeneity, genetic anthropology studies investigating the origin of current Southern Tunisians were rarely reported. We examined human leukocyte antigen (HLA) class I (A, B) and class II (DRB1, DQB1) gene profiles of 250 unrelated Southern Tunisians, and compared them with those of Arab-speaking communities, along with Mediterranean and sub-Sahara African populations using genetic distances, neighbor-joining dendrograms, correspondence and haplotype analysis. In total, 137 HLA alleles were detected, which comprised 32 HLA-A, 52 HLA-B, 32 DRB1 and 21 DQB1 alleles. The most frequent alleles were HLA-A*02:01(18.02%), HLA-B*50:01 (9.11%), HLA-DRB1*07:01 (22.06%) and HLA-DQB1*02:01 (17.21%). All pairs of HLA loci show significant linkage disequilibrium. The four loci depict negative Fnd (the normalized deviate of the homozygosity) values indicating an overall trend to balancing selection. Southern Tunisians appear to be closely related to others Tunisian populations including Berbers, North Africans and Iberians. On the contrary, Southern Tunisians were distinct from Palestinian, Lebanese and Jordanian Middle Eastern Arab-speaking population, despite the deep Arab incursions and Arabization that affected Southern Tunisia. In addition, Southern Tunisians were distant from many sub-Saharan communities, evidenced by genetic distance analysis. Collectively, this indicates a limited genetic contribution of Arab invasion and Black caravans on the makeup of Southern Tunisian gene pool.

Clinical significance of T-bet, GATA-3, and Bcl-6 transcription factor expression in bladder carcinoma.

BACKGROUND: The aim of this study was to investigate the clinical significance of three immune cell-related transcription factors, T-bet, GATA-3 and Bcl-6 in bladder cancer in Tunisian patients. METHODS: Expression of T-bet, GATA-3 and Bcl-6 genes was assessed using RT-qPCR in 65 bladder cancers from patients: 32 being diagnosed as low- and medium-grade, 31 as high-grade, 25 as muscle invasive stage and 39 as non-muscle invasive stage. Gene expression was statistically correlated according to the grade, the stage, tobacco consumption, the BCG response and disease severity. RESULTS: T-bet levels in patients with high-grade bladder cancer were significantly elevated compared to patients with low- or medium-grade bladder cancer (p = 0.005). In invasive carcinoma (T2-T4), the T-bet levels were significantly higher than in superficial non-invasive bladder tumors (Tis, Ta, and T1) (p = 0.02). However, T-bet is predictive of the response to BCG. Its expression is high in good responders to BCG (p = 0.02). In contrast, the expression of GATA-3 and Bcl-6 in non-invasive carcinoma (p = 0.008 and p = 0.0003) and in patients with low- and medium-grade cancers (p = 0.001 and p < 0.0001) is significantly higher than in invasive bladder tumors and in patients with high-grade bladder carcinoma, respectively. In addition, heavy smokers, whose tumors express low levels of GATA-3 and Bcl-6, are poor responders to BCG (p = 0.01 and p = 0.03). Finally, better patient survival correlated with GATA-3 (p = 0.04) and Bcl-6 (p = 0.04) but not T-bet expression. CONCLUSIONS: Our results suggest that T-bet expression in bladder tumors could be a positive prognostic indicator of BCG therapy, even if high levels are found in high-grade and stage of the disease. However, GATA-3 and Bcl-6 expression could be considered as predictive factors for good patient survival.

Akt activation correlates with the tumor aggressiveness in Tunisian patients with bladder cancer.

Various studies in western countries found Akt amplification to be a frequent event in human cancers, including bladder, but the correlation with clinicopathological features is controversial. Such studies have not been reported in African populations, including Tunisians. The purpose of this study was to assess expression of the phosphorylated/activated forms of Akt in tumors from Tunisian patients with bladder cancer and to correlate its expression with pathological and clinical parameters of the disease. The study included 72 patients of whom 34 were diagnosed as low- to medium-grade and 35 as high-grade; 30 were muscle stage and 39 non-muscle stage. Primary tumors from these patients, normal adjacent tissues, or bladder cancer cell-lines were analyzed for Ser473 phosphorylated Akt expression by Western blot. Seventy-two percent of primary tumors from patients with bladder cancer had increased levels of p-Akt. The p-Akt levels in patients with high-grade bladder cancer were significantly elevated compared to patients with low- or medium-grade bladder cancer. In invasive carcinoma, the p-Akt level was significantly higher than in superficial non-invasive bladder tumors. Concerning the influence of tobacco on Akt activation, no significant differences of p-Akt expression were found between non-smoker and smoker patients. Altogether, our results suggest that Akt activation can provide useful prognostic information and that tobacco represents a serious risk factor for recurrence in a cohort of Tunisian patients.

An investigation of the genetic diversity of the Kerkennah islands and Mahdia (Tunisia) using biparental markers.

BACKGROUND: Kerkennah is one of the main inhabited islands of Tunisia. The origin of the population of Kerkennah has not been established and no well-defined ethnic groups have been identified nor are genetic studies available. Mahdia, a Tunisian coastal city, has a long history dating back to ancient times. AIM: To discover the genetic diversity of the two studied populations and analyse their relationships with other Mediterranean populations. SUBJECT AND METHODS: Seven human-specific Alu insertion polymorphisms were typed in 99 individuals born in Kerkennah and Mahdia. RESULTS: A neighbour-joining tree and MDS multidimensional scaling analysis showed that these Tunisian populations are scattered amongst North African and Europeans populations, indicating their high genetic diversity and mosaic aspect. The important finding of this study was the proximity of Kerkennah to Moroccans. Hence, the actual gene pool of this insular population may descend from the ancestral population known to be of Moroccan origin. Concerning Mahdia, its closeness to Eurasian populations and some Tunisian groups reflected a high Eurasian genetic component for North African populations and confirmed their heterogeneity. CONCLUSION: The strategic location of the two studied populations and their fortifications have allowed them to play a leading role in the Mediterranean basin.

Gut microbiota, an emergent target to shape the efficiency of cancer therapy.

It is now well-acknowledged that microbiota has a profound influence on both human health and illness. The gut microbiota has recently come to light as a crucial element that influences cancer through a variety of mechanisms. The connections between the microbiome and cancer therapy are further highlighted by a number of preclinical and clinical evidence, suggesting that these complicated interactions may vary by cancer type, treatment, or even by tumor stage. The paradoxical relationship between gut microbiota and cancer therapies is that in some cancers, the gut microbiota may be necessary to maintain therapeutic efficacy, whereas, in other cancers, gut microbiota depletion significantly increases efficacy. Actually, mounting research has shown that the gut microbiota plays a crucial role in regulating the host immune response and boosting the efficacy of anticancer medications like chemotherapy and immunotherapy. Therefore, gut microbiota modulation, which aims to restore gut microbial balance, is a viable technique for cancer prevention and therapy given the expanding understanding of how the gut microbiome regulates treatment response and contributes to carcinogenesis. This review will provide an outline of the gut microbiota's role in health and disease, along with a summary of the most recent research on how it may influence the effectiveness of various anticancer medicines and affect the growth of cancer. This study will next cover the newly developed microbiota-targeting strategies including prebiotics, probiotics, and fecal microbiota transplantation (FMT) to enhance anticancer therapy effectiveness, given its significance.

Internal diversification of mitochondrial haplogroup R0a reveals post-last glacial maximum demographic expansions in South Arabia.

Widespread interest in the first successful Out of Africa dispersal of modern humans ∼60-80 thousand years ago via a southern migration route has overshadowed the study of later periods of South Arabian prehistory. In this work, we show that the post-Last Glacial Maximum period of the past 20,000 years, during which climatic conditions were becoming more hospitable, has been a significant time in the formation of the extant genetic composition and population structure of this region. This conclusion is supported by the internal diversification displayed in the highly resolved phylogenetic tree of 89 whole mitochondrial genomes (71 being newly presented here) for haplogroup R0a-the most frequent and widespread haplogroup in Arabia. Additionally, two geographically specific clades (R0a1a1a and R0a2f1) have been identified in non-Arabic speaking peoples such as the Soqotri and Mahri living in the southern part of the Arabian Peninsula where a past refugium was identified by independent archaeological studies. Estimates of time to the most recent common ancestor of these lineages match the earliest archaeological evidence for seafaring activity in the peninsula in the sixth millennium BC.

[Immunogenetics of psoriasis: update]. / Immunogénétique du psoriasis : actualités.

BACKGROUND: Psoriasis is a chronic inflammatory skin disease often benign, affecting 2-3% of the total world population. Psoriasis is a multifactorial disease. AIM: To present recent advances in the immunologic mechanisms and susceptibility genes involved in the pathogenesis of psoriasis. METHODS: We presented a literature review of recent genetic and immunological basis of psoriasis to better understand the pathomecanisms of this disease and discuss the contribution of the Tunisian work in this area. RESULTS: Recent works focalized mainly in immunology and genetics. Current progresses in molecular biology have allowed to better characterize the immunogenetic abnormalities in psoriasis. CONCLUSION: Psoriasis is a multifactorial disease model in which environmental factors (psychological, climate, traumatic, infectious, and viral) seem to be triggering factors when associated with a particular immunogenetics predisposition.

New Insight into the human genetic diversity in North African populations by genotyping of SNPs in DRD3, CSMD1 and NRG1 genes.

BACKGROUND: The single nucleotide polymorphisms (SNPs) of the dopamine D3 receptor (DRD3), the CUB and sushi multiple domains 1 (CSMD1) and the neuregulin 1 (NRG1) genes were used to study the genetic diversity and affinity among North African populations and to examine their genetic relationships in worldwide populations. METHODS: The rs3773678, rs3732783 and rs6280 SNPs of the DRD3 gene located on chromosome 3, the rs10108270 SNP of the CSMD1 gene and the rs383632, rs385396 and rs1462906 SNPs of the NRG1 gene located on chromosome 8 were analysed in 366 individuals from seven North African populations (Libya, Kairouan, Mehdia, Sousse, Kesra, Smar and Kerkennah). RESULTS: The low values of FST indicated that only 0.27%-1.65% of the genetic variability was due to the differences between the populations. The Kairouan population has the lowest average heterozygosity among the North African populations. Haplotypes composed of the ancestral alleles ACC and ACAT were more frequent in the Kairouan population than in other North African populations. The PCA and the haplotypic analysis showed that the genetic structure of populations in North Africa was closer to that of Europeans, Admixed Americans, South Asians and East Asians. However, analysis of the rs3732783 and rs6280 SNPs revealed that the CT microhaplotype was specific to the North African population. CONCLUSIONS: The Kairouan population exhibited a relatively low rate of genetic variability. The North African population has undergone significant gene flow but also evolutionary forces that have made it genetically distinct from other populations.

Post-last glacial maximum expansion from Iberia to North Africa revealed by fine characterization of mtDNA H haplogroup in Tunisia.

The first large-scale fine characterization of Tunisian H lineages clarifies that the post-Last glacial maximum expansion originating in Iberia not only led to the resettlement of Europe but also of North Africa. We found that 46% of 81 Tunisian H lineages subscreened for 1,580 bp in mtDNA coding region were affiliated with H1 and H3 subhaplogroups, which are known to have originated in Iberia. Although no signs of local expansion were detected, which would allow a clear dating of their introduction, the younger and less diverse Tunisian H1 and H3 lineages indicate Iberia as the radiating centre. Major contributions from historical migrations to this Iberian genetic imprint in Tunisia were ruled out by the mtDNA gene pool similarity between Berber/Arab/cosmopolitan samples and some "Andalusian" communities, settled by the descendents of the "Moors" who once lived in Iberia for 10 centuries (between 8th and 17th centuries), before being expelled to Tunisia.

[Penetrance of BRCA1 gene mutation and DNA mitochondrial in Tunisian breast cancer occurrence]. / Pénétrance des mutations du gène BRCA1 et de l'ADN mitochondrial dans l'occurrence du cancer du sein en Tunisie.

AIM: The aim of this study is to evaluate the implication of BRCA1 gene and the mitochondrial micro satellite (situated between 303 and 315 positions) mutations in the occurrence of breast cancer in Tunisia. METHODS: Nine Tunisian patients with hereditary breast cancer have been analyzed. For each patient, total genomic DNA was extracted and used as a template for the amplification of 24 exons of the BRCA1 gene and an hyper variable mitochondrial region. The obtained products were purified and automatically sequenced. RESULTS: The results revealed five types of mutations for the micro satellite situated between the 303 and 315 positions and two deleterious BRCA1 mutations for two unrelated patients which present the same mitochondrial mutation (315.insC) suggesting his implication in the modulation of the BRCA1 deleterious mutations penetrance.

BRCA1/BRCA2 Mutations Shaped by Ancient Consanguinity Practice in Southern Mediterranean Populations

The aim of this study is to investigate the involvement of consanguinity on BRCA1/2 mutation incidence in Southern Mediterranean populations and to confirm their low penetrance by comparison of their recurrence in sporadic and familial breast cancer in a context of ancient consanguinity practice. Our study comprises of two parts: First, a comparison of the consanguinity rates of the South Mediterranean countries in a relationship with the frequency of BRCA1 deleterious mutations in breast cancer families and the recurrence of these mutations. Second, we investigated 23patients with a family history of breast cancer, 51 patients without a family history of breast cancer using next-generation sequencing of BRCA2 and then confirmed by Sanger sequencing for the novel mutation. As results, we clearly show a strong relationship between the frequency of BRCA1 deleterious mutations in breast cancer families and rate of consanguinity, since they are significantly inversely correlated. Four deleterious mutations were found in BRCA2 gene including a novel frame-shift mutationc.9382_9383dup in a patient with familial breast cancer and three other frame-shift mutations c.6591_6592del, c.1310_1313del and c.7654dup in patients with sporadic breast cancer.These results are discussed in a context of selective pressure of ancient consanguinity practice. In conclusion, the study of BRCA1/2 gene in Southern Mediterranean countries revealed low penetrance recurrent mutations in sporadic and familial breast cancer. These mutations have been selected in a context of ancient consanguinity practice along with protective genetic and environmental factors.

Identification of the CCR5-Delta32 HIV resistance allele and new mutations of the CCR5 gene in different Tunisian populations.

Polymorphisms in some chemokine receptor genes are associated with susceptibility to and progression of human immunodeficiency virus-1 (HIV-1) infection. Most mutations detected in the CC-chemokine receptor 5 (CCR5) gene are specific to different populations. In this study, we focused on polymorphisms of the CCR5 coding region in three healthy populations from Tunisia, corresponding to a cosmopolitan population from Tunis, and two isolated Berber populations. In addition to the CCR5-Delta32 deletion, eleven single nucleotide polymorphisms were detected. Some of these point mutations were associated with the same genotype and even the same haplotype. The (L55Q-C101X), I124, V131F, T143N, A159V, I237, T239A and G301R alleles have not been described previously, whereas the CCR5-Delta32, L55Q, A335V and Y339F variants have already been reported in the literature. The distribution and frequency of these variants were different among the three groups studied, a result in agreement with the mosaic genetic structure of the Tunisian population. To determine whether these alleles affect HIV-1 transmission, we compared allele frequencies between healthy and HIV-1 infected individuals from Tunis. The frequency of the CCR5-Delta32 variant was significantly different between the two groups, leading us to conclude that this mutation might confer protection against HIV infection in Tunisian populations.

Methylenetetrahydrofolate reductase and methionine synthase polymorphisms and risk of bladder cancer in a Tunisian population.

Folate insufficiency can induce carcinogenesis by decreasing DNA methylation. It is well known that DNA hypomethylation is a common feature in a number of cancers. Methylenetetrahydrofolate reductase (MTHFR) and methionine synthase (MS) are enzymes that play central roles in the folate metabolic pathway. Two common polymorphisms in the MTHFR gene (C677T and A1298C) and one in the MS gene (A2756G) are associated with decreased enzymatic activity. In this work, we have conducted a case-control study to assess the role of these three polymorphisms in bladder cancer development in North Tunisia. For MS A2756G, gene and genotypic distributions differed significantly between cases and controls. Furthermore, individuals carrying at least one copy of the variant allele presented a 2.33 times increased risk of developing bladder cancer than their control group [P = 0.001, odds ratio (OR) = 2.33; 95% confidence interval (CI) 1.34-4.06]. Statistically significant odds ratios were also found in patients heterozygous for MTHFR A1298C, who have a 1.8-fold higher risk of developing bladder cancer (P = 0.03, OR = 1.86; CI 95% 1.04-3.33). While the isolated polymorphism C677T did not appear to influence bladder cancer susceptibility, results suggest that it might act with an additive contribution determined by variation at MTHFR A1298C. Identical cumulative effect was detected for the MTHFR A1298C and MS 2756 genotypes. Patients harboring at least one mutant allele for each of the three positions analyzed showed a 4.76-fold increased risk of developing bladder cancer in comparison to their reference group (P = 0.02, OR = 4.76; CI 95% 1.26-17.98).

Data for Y-chromosome haplotypes defined by 17 STRs (AmpFLSTR Yfiler) in two Tunisian Berber communities.

The 17 Y-chromosomal short tandem repeats (STRs) included in the AmpFLSTR Yfiler PCR Amplification Kit (AB Applied Biosystems) (DYS19, DYS389I, DYS389II, DYS390, DYS391, DYS392, DYS393, DYS385, DYS437, DYS438, DYS439, DYS448, DYS456, DYS458, DYS635 and GATA H4) were typed in two Berber communities, a small village (Takrouna) and a town (Sejenane), from North Tunisia. As expected, diversity was higher in the town, even when compared with a pool of three small Berber communities, probably due to the combination of different founder effects and genetic drifts operating in the small villages.

HLA Class I and Class II Alleles and Haplotypes Confirm the Berber Origin of the Present Day Tunisian Population.

In view of its distinct geographical location and relatively small area, Tunisia witnessed the presence of many civilizations and ethnic groups throughout history, thereby questioning the origin of present-day Tunisian population. We investigated HLA class I and class II gene profiles in Tunisians, and compared this profile with those of Mediterranean and Sub-Sahara African populations. A total of 376 unrelated Tunisian individuals of both genders were genotyped for HLA class I (A, B) and class II (DRB1, DQB1), using reverse dot-blot hybridization (PCR-SSO) method. Statistical analysis was performed using Arlequin software. Phylogenetic trees were constructed by DISPAN software, and correspondence analysis was carried out by VISTA software. One hundred fifty-three HLA alleles were identified in the studied sample, which comprised 41, 50, 40 and 22 alleles at HLA-A,-B,-DRB1 and -DQB1 loci, respectively. The most frequent alleles were HLA-A*02:01 (16.76%), HLA-B*44:02/03 (17.82%), HLA-DRB1*07:01 (19.02%), and HLA-DQB1*03:01 (17.95%). Four-locus haplotype analysis identified HLA-A*02:01-B*50:01-DRB1*07:01-DQB1*02:02 (2.2%) as the common haplotype in Tunisians. Compared to other nearby populations, Tunisians appear to be genetically related to Western Mediterranean population, in particular North Africans and Berbers. In conclusion, HLA genotype results indicate that Tunisians are related to present-day North Africans, Berbers and to Iberians, but not to Eastern Arabs (Palestinians, Jordanians and Lebanese). This suggests that the genetic contribution of Arab invasion of 7th-11th century A.D. had little impact of the North African gene pool.

Obesity and renal cancer: Role of adipokines in the tumor-immune system conflict.

Epidemiological studies link obesity, as measured by increased body mass index (BMI) to the incidence of renal cell carcinoma (RCC) as well as to the cancer-related mortality of RCC patients. RCC is the third cancer most robustly associated with increased BMI. Understanding the role of the adipose tissue in renal carcinogenesis is therefore of major importance for the development of novel paradigms of RCC prevention and treatment. Here, we discuss the current knowledge on the impact of obesity on the development and progression of RCC as well as the role of adipose tissue-derived hormones (adipokines) in the conflict between growing tumors and the immune system.

Thymidylate synthase polymorphism in sporadic colorectal and gastric cancer in Tunisian population: a predictive role in 5-fluorouracil based chemotherapy treatment.

In our study, we investigate the possible association of thymidylate synthase polymorphism, 28 bp tandem repeat in 5'-UTR (transcription enhancer element) with susceptibility of colorectal and gastric cancer in Tunisian population. Because thymidylate synthase provides an effective prediction of chemotherapy treatment based on 5-fluorouracil, our interest in this study was focused on finding an eventual interaction between thymidylate synthase polymorphism and treatment of sporadic colorectal and gastric cancer. Whole blood was collected into EDTA tube, after centrifugation for 15 min, the buffy coat was isolated, and genotyping of TS 5'-UTR polymorphism was carried by polymerase chain reaction method using appropriate primers. Determination of the different genotypes was done directly on the stained agarose gel. Our finding showed that the 5'tandem repeat polymorphism of the thymidylate synthase gene is associated with risk of colorectal cancer; thus, LL (3R/3R) genotype is significantly high in patients with colorectal cancer compared to controls (P = 0.002; OR 2.7; 95 % CI 1.4-5.2). In addition, we found a positive association between SL (2R/3R) genotype in the thymidylate synthase 5'-UTR and gastric cancer risk (P = 0.015; OR 4.46; 95 % CI 1.08-19.64). Furthermore, we found a correlation of thymidylate synthase polymorphism with the fluorouracil-based therapy regimes and also with preoperatory radiation in patients with colorectal cancer. Thymidylate synthase is associated with risk of colorectal cancer but not with gastric cancer; however, heterozygous SL (2R/3R) polymorphism is associated with risk of gastric cancer; moreover, the 5' tandem repeat polymorphism of thymidylate synthase gene was an independent predictor of the clinical treatment.

Assessment of Aurora A kinase expression in breast cancer: a tool for early diagnosis?

Aurora A kinase is overexpressed in many cancers but the status of this protein in the breast cancer often varies. We investigate the expression and localization of Aurora A protein in relation with tumor emergence and progression in breast cancer. Aurora A kinase status was evaluated in 107 patients using immunohistochemistry. The experimental findings showed that high expression of the Aurora A protein was correlated with elevated nuclear grade, low expression of progesterone receptor and positive nodal status. The experimental results showed also that the localization of this kinase shifts from cytoplasm in non malignant adjacent tissue to both cytoplasmic and nuclear compartments in tumoral tissue, suggesting an oncogenic role of the nuclear accumulation. We have, furthermore, detected the overexpression of this protein in non malignant adjacent tissue. The expression of the Aurora A kinase in non malignant tissue may represent an earlier diagnosis tool for breast cancer.

Association analysis of LCE3C-LCE3B deletion in Tunisian psoriatic population.

An association between a common deletion comprising the late cornified envelope LCE3B and LCE3C genes (LCE3C_LCE3B-del) and psoriasis has been reported in Caucasian and Asian populations. To investigate whether this deletion plays a role in the genetic of psoriasis in Tunisian population, we determined the LCE3C_LCE3B-del genotype in 180 Ps patients and 208 healthy controls from different regions of Tunisia. The LCE3B and LCE3C gene variant was determined in the patients through PCR amplification and the SPSS software package. The frequency of the LCE3C_LCE3B-del was similar between patients and healthy controls. Subanalyses by family history revealed that the frequency of LCE3C_LCE3B-del was significantly higher in patients with a positive family history than in control individuals, as well as in individuals with a positive family history versus those without in the case cohort. However, no significant difference was observed between psoriatic patients with no family history and controls. We also evaluated the relationship between LCE3C_LCE3B-del and PSORS1. No significant epistatic effect was observed suggesting that there was no significant epistasis of the two loci in the Tunisian population. Our findings indicate that the LCE3C_LCE3B-del might play a role in familial psoriasis in the Tunisian population.

A meta-analysis of the relationship between FGFR3 and TP53 mutations in bladder cancer.

TP53 and FGFR3 mutations are the most common mutations in bladder cancers. FGFR3 mutations are most frequent in low-grade low-stage tumours, whereas TP53 mutations are most frequent in high-grade high-stage tumours. Several studies have reported FGFR3 and TP53 mutations to be mutually exclusive events, whereas others have reported them to be independent. We carried out a meta-analysis of published findings for FGFR3 and TP53 mutations in bladder cancer (535 tumours, 6 publications) and additional unpublished data for 382 tumours. TP53 and FGFR3 mutations were not independent events for all tumours considered together (OR = 0.25 [0.18-0.37], p = 0.0001) or for pT1 tumours alone (OR = 0.47 [0.28-0.79], p = 0.0009). However, if the analysis was restricted to pTa tumours or to muscle-invasive tumours alone, FGFR3 and TP53 mutations were independent events (OR = 0.56 [0.23-1.36] (p = 0.12) and OR = 0.99 [0.37-2.7] (p = 0.35), respectively). After stratification of the tumours by stage and grade, no dependence was detected in the five tumour groups considered (pTaG1 and pTaG2 together, pTaG3, pT1G2, pT1G3, pT2-4). These differences in findings can be attributed to the putative existence of two different pathways of tumour progression in bladder cancer: the CIS pathway, in which FGFR3 mutations are rare, and the Ta pathway, in which FGFR3 mutations are frequent. TP53 mutations occur at the earliest stage of the CIS pathway, whereas they occur would much later in the Ta pathway, at the T1G3 or muscle-invasive stage.