Estrogen-like metabolites and DNA-adducts in urogenital schistosomiasis-associated bladder cancer

Highlights

• An estrogen-DNA adduct may be involved in pathogenesis of the cell carcinoma of the bladder associated with S. haematobium.

• Metabolites can be expected to provide deeper understanding of the carcinogenesis UGS-induced bladder cancer.

• Biomarkers for diagnosis and/or prognosis of this neglected tropical disease-linked cancer.

Abstract

An estrogen-DNA adduct mediated pathway may be involved in the pathogenesis of the squamous cell carcinoma of the bladder associated with infection with the blood fluke Schistosoma haematobium. Extracts from developmental stages of S. haematobium, including eggs, induce tumor-like phenotypes in cultured cells. In addition, estrogen-derived, reactive metabolites occur in this pathogen and in sera of infected persons. Liquid chromatography-mass spectrometry analysis was performed on urine from 40 Angolans diagnosed with urogenital schistosomiasis (UGS), half of who also presented UGS-associated squamous cell carcinoma and/or urothelial cell carcinoma. The analysis revealed numerous estrogen-like metabolites, including seven specifically identified in UGS cases, but not reported in the database of metabolites in urine of healthy humans. These schistosome infection-associated metabolites included catechol estrogen quinones (CEQ) and CEQ-DNA-adducts, two of which had been identified previously in S. haematobium. In addition, novel metabolites derived directly from 8-oxo-7, 8-dihydro-2′-deoxyguanosine (8-oxodG) were identified in urine of all 40 cases of UGS. These metabolites can be expected to provide deeper insights into the carcinogenesis UGS-induced bladder cancer, and as biomarkers for diagnosis and/or prognosis of this neglected tropical disease-linked cancer.

Introduction

Schistosomiasis is one of the major neglected tropical diseases and it is considered the most important helminthic disease of humanity in terms of morbidity and mortality. More than 90% of the cases occur in Africa, of which about two-thirds are caused by Schistosoma haematobium [1], [2], [3], [4], [5]. Indeed, the number of cases of infection with S. haematobium may far exceed that previously predicted so that urogenital schistosomiasis (UGS) may represent the most common infection or even adverse health condition in sub-Saharan Africa [6]. In addition, female genital schistosomiasis increases the risk of transmission of HIV [7], [8], [9], and a recent outbreak in Corsica confirms its re-emergence in Europe [10], [11]. Many cases of UGS result ostensibly in only mild symptoms and disease such as hematuria, dysuria, anemia and inflammation of the genital-urinary tract [6], [12], [13], [14], [15], [16]. However, between 25 and 50% of the UGS cases experience moderate to severe morbidity [6], including renal dysfunction, obstruction of the ureters, and squamous cell carcinoma of the urinary bladder [17], [18], [19], [20]. Bladder cancer is a frequent and dire complication of chronic UGS. Case reports indicate that patients with schistosomiasis may develop bladder cancer earlier than uninfected people. The severity and frequency of the sequelae of UGS and of its complications are related to the intensity and duration of the infection [21], [22], [23]. Moreover, infection with S. haematobium is classified as a Group 1 biological carcinogen by the World Health Organization (WHO)'s International Agency for Research on Cancer (WHO IARC) [24] although the cellular and/or molecular mechanisms linking S. haematobium infection with carcinogenesis have yet to be defined [25], [26]. It has been known for several decades that bladder cancer, especially squamous cell carcinoma (SCC), a distinctly malignant, poorly differentiated neuroendocrine neoplasm [16], was geographically associated with UGS, i.e. areas endemic for schistosomiasis haematobia [20], [24], [27]. In regions with high worm burdens, and a high risk of exposure to S. haematobium infection, SCC is the most frequent histological type, whereas urothelial cell carcinoma (UCC) is predominant in non-endemic locations [22], [23], [24], [26], [28], [29].

An estrogen-DNA adduct mediated pathway in S. haematobium-infection associated bladder cancer has been postulated [27], [30], [31]. We have identified and characterized by liquid chromatography-mass spectrometry (LC-MS/MS) novel estrogen-like metabolites, present in the lysates and secretions of S. haematobium worms and eggs [27], and in sera of UGS cases [30]. Moreover, lysates of S. haematobium induce tumor-like phenotypes; Chinese Hamster Ovary cells exposed in vitro to the lysates exhibit marked proliferation and induce sarcoma formation when inoculated into nude mice. Additionally, the cells display increased duration of S phase, decreased apoptosis, down-regulation of the p27 tumor suppressor, and up-regulation of anti-apoptotic protein Bcl-2 [25], [27], [30], [31]. Eggs of S. haematobium stimulate cellular proliferation, interfere with apoptosis, increase oxidative stress, and induce a genotoxicity on HCV-29 cells, derived from human urothelial cells [27], [30], [31], [32], [33]. In addition, 8-nitroguanine forms via inducible expression of nitric oxide synthase in Oct3/4-positive stem cells in UGS-associated bladder cancer tissue [34], and DNA nitrative and oxidative mutations characterized by 8-nitroguanine and 8-hydroxy-2′-deoxyguanosine (8-oxodG), have been implicated in the promotion of inflammation-mediated carcinogenesis by infection with S. haematobium [35].

Here we undertook analysis of urine from persons with urogenital schistosomiasis (UGS) living in endemic areas in Angola. Liquid chromatography diode array detection electron spray ionization mass spectrometry (LC-ESI-MS) revealed the presence of numerous estrogen metabolites in the urine of the study participants. Seven of these molecules were specifically identified in urine of the UGS cases but, notably, were not described in the recently reported metabolome of urine from healthy humans [36]. These metabolites were potentially reactive with host DNA: the molecules were either catechol estrogen quinone (CEQ) derivatives or CEQ-DNA-adducts. In addition, novel molecules derived from 8-oxodG during UGS were also identified.