A Nutrigenomic View on the Premature-Aging Disease Fanconi Anemia

Fanconi anemia, a rare disorder with an incidence of 1 in 300,000, is caused by mutations in FANC genes, which affect the repair of DNA interstrand crosslinks. The disease is characterized by congenital malformations, bone marrow failure within the first decade of life, and recurrent squamous cell carcinomas of the oral cavity, esophagus, and anogenital regions starting around age 20. In this review, we propose that Fanconi anemia should be considered a premature-aging syndrome. Interestingly, the onset and severity of the life-limiting clinical features of Fanconi anemia can be influenced by lifestyle choices, such as a healthy diet and physical activity. These factors shape the epigenetic status of at-risk cell types and enhance the competence of the immune system through nutritional signaling. Fanconi anemia may serve as a model for understanding the aging process in the general population, addressing research gaps in its clinical presentation and suggesting prevention strategies. Additionally, we will discuss how the balance of genetic and environmental risk factors—affecting both cancer onset and the speed of aging—is interlinked with signal transduction by dietary molecules. The underlying nutrigenomic principles will offer guidance for healthy aging in individuals with Fanconi anemia as well as for the general population

A New Multi-Color FISH Assay for Brush Biopsy-Based Detection of Chromosomal Aneuploidy in Oral (Pre)Cancer in Patients with Fanconi Anemia

Background: Fanconi anemia (FA) is a rare inherited DNA instability disorder with a remarkably elevated risk of oral squamous cell carcinoma. These cancers can be detected with oral brush biopsy-based cytology even at early stages. This study aims to determine the diagnostic accuracy of a new multi-color fluorescent in situ hybridization (FISH) assay consisting of probes for CCND1, TERC, MYC and centromere of chromosome 6, as well as a 9p21 FISH assay consisting of probes for CDKN2A and centromere of chromosome 9 for the detection of oral (pre) malignant lesions in FA. Methods: (I) Cutoffs for the dichotomization of positive or negative multi-color FISH results are determined and (II) retrospectively validated by using archived oral brush biopsy specimens from individuals with Fanconi anemia. In addition, the specimens for cutoff determination were re-hybridized with the 9p21 FISH assay. Results: A cutoff of six or more chromosomal aneuploid cells for a positive FISH result was determined in the cutoff study on 160 biopsy specimens. The validating of this cutoff on 152 specimens showed at best a sensitivity of 87% and a specificity of 82.9%. Conclusion: Multi-color FISH is a sufficient tool to detect chromosomal aneuploidy in oral (pre) malignant lesions of individuals with Fanconi anemia. However, some false positive results may hamper the application as an adjuvant method to oral brush biopsy-based cytology in an oral cancer surveillance program

Oral cancer prediction by noninvasive genetic screening

Oral squamous cell carcinomas (OSCCs) develop in genetically altered epithelium in the mucosal lining, also coined as fields, which are mostly not visible but occasionally present as white oral leukoplakia (OL) lesions. We developed a noninvasive genetic assay using next-generation sequencing (NGS) on brushed cells to detect the presence of genetically altered fields, including those that are not macroscopically visible. The assay demonstrated high accuracy in OL patients when brush samples were compared with biopsies as gold standard. In a cohort of Fanconi anemia patients, detection of mutations in prospectively collected oral brushes predicted oral cancer also when visible abnormalities were absent. We further provide insight in the molecular landscape of OL with frequent changes of TP53, FAT1 and NOTCH1. NGS analysis of noninvasively collected samples offers a highly accurate method to detect genetically altered fields in the oral cavity, and predicts development of OSCC in high-risk individuals. Noninvasive genetic screening can be employed to screen high-risk populations for cancer and precancer, map the extension of OL lesions beyond what is visible, map the oral cavity for precancerous changes even when visible abnormalities are absent, test accuracy of promising imaging modalities, monitor interventions and determine genetic progression as well as the natural history of the disease in the human patient

Limited detection of human polyomaviruses in Fanconi anemia related squamous cell carcinoma.

Fanconi anemia is a rare genome instability disorder with extreme susceptibility to squamous cell carcinoma of the head and neck and anogenital tract. In patients with this inherited disorder, the risk of head and neck cancer is 800-fold higher than in the general population, a finding which might suggest a viral etiology. Here, we analyzed the possible contribution of human polyomaviruses to FA-associated head and neck squamous cell carcinoma (HNSCC) by a pan-polyomavirus immunohistochemistry test which detects the T antigens of all known human polyomaviruses. We observed weak reactivity in 17% of the HNSCC samples suggesting that based on classical criteria, human polyomaviruses are not causally related to squamous cell carcinomas analyzed in this study

Dominant families detected by NGS after OTU clustering in the microbiotas of patients and healthy individuals.

<p>Frequency of families (in %) present in at least one of the 13 analysed samples analysed at a frequency >4%; A., site B of healthy probands H1-H5; B., sites A, B, E and F of FA patient FA_L1; and C., sites A, B, E and F of FA patient FA_T1.</p

Real time PCRs for the detection of <i>M. salivarium</i>, <i>P. aeruginosa</i>, and <i>Candida</i>.

<p>DNA samples, which were derived from healthy probands (H1-H5, site B), FA-patients with leukoplakia (FA_L1 with a leukoplakia at site F and FA_ L2 with leukoplakia at site B) and FA patients with tumour (FA_T1 with a tumour at site B and FA_T2 with tumour at site D) were subjected to real time PCR in duplicates. A. TaqMan qPCR-derived copy numbers of <i>M. salivarium</i> (red), <i>P. aeruginosa</i> (hatched) and <i>Candida</i> (grey). Due to targeting a single copy gene, copy numbers correspond to genome equivalents for <i>M. salivarium</i> and <i>P. aeruginosa</i>. B., scatter plot of <i>M. salivarium</i> load in the different proband groups: Healthy (H1-H5, blue circles), samples of FA-patients with leukoplakia (FA_L1 and FA_L2, green squares) and with tumour (FA_T1 and FA_T2, red triangles). C., scatter plot of <i>M. salivarium</i> load of FA patients FA_T1 (dark red) and FA_T2 (red) at the tumour surface (Tumour) in relation to the contralateral non-tumour surface (Non-T.) and of the FA patients FA_L1 (green) and FA_L2 (dark green) at the site of leukoplakia (Leukopl.) in relation to the contralateral non-leukoplakia site (Non-L.).</p

Total number of reads generated per sample.

<p>Total number of reads generated per sample.</p

Oral cavity and sites of sampling.

<p>A. Anatomic chart of the oral cavity with arrows indicating the sites of sampling (A to F) and a red circle labelling the tumour region of patient FA_T1. Samples C and D were collected from the ventral portion of the tongue (indicated by arrows with hatched centres). B. Photography of the tumour on the FA_T1 patient's left side of the tongue encroaching the centre line. The red circle indicates the sampling site of the tumour region as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0092297#pone-0092297-g001" target="_blank">Figure 1, A</a>.</p