The TP53 R337H mutation is associated with increased incidence of pediatric adrenocortical tumor (ACT). The different environmental conditions where R337H carriers live have not been systematically analyzed. Here, the R337H frequencies, ACT incidences, and R337H penetrance for ACT were calculated using the 2006 cohort with 4165 R337H carriers living in Paraná state (PR) subregions. The effectiveness of a second surveillance for R337H probands selected from 42,438 tested newborns in PR (2016 cohort) was tested to detect early stage I tumor among educated families without periodical exams. Estimation of R337H frequencies and ACT incidence in Santa Catarina state (SC) used data from 50,115 tested newborns without surveillance, ACT cases from a SC hospital, and a public cancer registry. R337H carrier frequencies in the population were 0.245% (SC) and 0.306% (PR), and 87% and 95% in ACTs, respectively. The ACT incidence was calculated as ~6.4/million children younger than 10 years per year in PR (95% CI: 5.28; 7.65) and 4.15/million in SC (CI 95%: 2.95; 5.67). The ACT penetrance in PR for probands followed from birth to 12 years was 3.9%. R337H carriers living in an agricultural subregion (C1) had a lower risk of developing pediatric ACT than those living in industrial and large urban subregion (relative risk = 2.4). One small ACT (21g) without recurrence (1/112) was detected by the parents in the 2016 cohort. ACT incidence follows R337H frequency in each population, but remarkably environmental factors modify these rates.

Adrenocortical carcinoma (ACC) is a rare disease among children. Our goal was to identify prognostic biomarkers in 48 primary ACCs from children (2.83 ± 2.3 y; mean age ± SD) by evaluating the tumor stage and outcome for an age of diagnosis before or after 3 years, and association with ACC cluster of differentiation 8 positive (CD8⁺) cytotoxic T lymphocytes (CD8⁺-CTL) and Ki-67 immunohistochemical expression (IHC). Programmed death 1(PD-1)/Programmed death-ligand 1 (PD-L1) immunohistochemistry (IHC) in ACC was analyzed in a second, partially overlapping cohort (N = 19) with a similar mean age. All patients and control children were carriers of the germline TP53 R337H mutation. Survival without recurrence for less than 3 years and death unrelated to disease were excluded. Higher counts of CD8⁺-CTL were associated with patients diagnosed with ACC at a younger age and stage I, whereas a higher percentage of the Ki-67 labeling index (LI) and Weiss scores did not differentiate disease free survival (DFS) in children younger than 3 years old. No PD-1 staining was observed, whereas weakly PD-L1-positive immune cells were found in 4/19 (21%) of the ACC samples studied. A high CD8⁺-CTL count in ACC of surviving children is compelling evidence of an immune response against the disease. A better understanding of the options for enhancement of targets for CD8⁺ T cell recognition may provide insights for future pre-clinical studies.

In South Brazil, the incidence of pediatric adrenocortical carcinoma (ACC) is higher than in other regions and countries worldwide. The ACC treatment includes therapy with mitotane, the only adrenolytic drug approved by the FDA. The mitotane metabolism occurs via two main reactions: the β-hydroxylation, which yields the final product o,p’-DDA, and the α-hydroxylation, which will give the final product o,p’-DDE. It is speculated that o,p’-DDE may be an active metabolite since it has a cytotoxic effect on adrenocortical carcinoma cells (H295R). No further studies have been conducted to confirm this hypothesis; however, it was found that mitotane and its metabolites are present at significantly different concentrations in the plasma of the patients. Our study aimed to assess the in vitro effects of o,p’-DDE and o,p’-DDD in cell death pathways, oxidative parameters, and interaction with adrenal CYP’s involved in the steroidogenic process in the H295R cell line. It was found that o,p’-DDE had a different effect than the o,p’-DDD on apoptosis, inhibiting this cell death pathway, but it promotes cell necrosis at higher concentrations. In contrast to o,p’-DDD, the o,p’-DDE did not have effects on the different oxidative parameters evaluated, but exhibited stimulatory interactions with steroidogenic CYP’s, at intermediate concentrations. Therefore, we demonstrated important cell effects of o,p’-DDE; its plasma levels during mitotane therapy should be monitored as an important therapeutic parameter.

Counselling and genetic testing (CGT) after neonatal screening may increase depression and anxiety (DA) levels during cancer surveillance. This study assessed the DA scores in mothers of newborns from Paraná state, Southern Brazil, carrying the TP53 p.R337H variant. To understand and adjust DA conditions during term of pregnancy, we initially detected sociodemographic covariates [marital status (MS), number of children (NC), and/or education level (EL): MS-NC-EL] on an independent group of pregnant women (not subjected to genetic testing). The Hospital Anxiety and Depression Scale (HADS) was used to assess risk factors in pregnant (cross-sectional analysis) and unrelated mothers (at 2-month intervals, longitudinal study) of TP53 p.R337H-tested newborns (three sessions of HADS analysis) using Wilcoxon (Mann–Whitney) and Kruskal–Wallis nonparametric tests. Lower anxiety levels were observed in mothers of noncarriers (without MS-NC-EL = 6.91 ± 1.19; with MS-NC-EL = 6.82 ± 0.93) than in mothers of p.R337H carriers in the first session (without MS-NC-EL = 6.82 = 8.49 ± 0.6025, with MS-NC-EL = 6.82 = 9.21 ± 0.66). The anxiety levels significantly decreased 4 months after CGT (third session) in mothers of p.R337H carriers. We did not find a significant change in depression scores. Mothers with mental health instability requiring medications need periodical psychological support during and after CGT.

Adrenal cortex carcinoma (ACC) in children is a rare tumor that is probably of multifactorial origin and is mainly associated with genetic and environmental alterations. In the south and part of the southeast of Brazil, as well as in the Paraguayan region bordering the Brazilian State of Paraná, ACC prevalence is higher than in any other country, which is associated with the high prevalence of the TP53 p.R337H variant in Paraná (0.30%), Santa Catarina (0.249%), cities around Campinas-SP (0.21%), and the Paraguayan cities on the border with Paraná (0.05%). Recent research suggests that the co-segregation of XAF1-E134* and TP53-R337H mutations leads to a more aggressive cancer phenotype than TP53-R337H alone. Breast cancer may be mildly influenced by co-segregation with XAF1 p.E134*, and this variant can also confer risk for sarcoma. Objectives: The objectives of this study were to (1) estimate the prevalence of the germline TP53 p.R337H and XAF1 p.E134* variants in Paraguay (excluding cities on the border with Paraná State, Brazil) and (2) estimate whether the ethnic origin of TP53 p.R337H carriers in Paraguay is similar to that of ethnic groups in Paraná (possible Portuguese/Spanish origin). Materials and methods: DNA tests for the identification of TP53 p.R337H were carried out from 2016 to 2019 at the Bio-Materials Laboratory of Facultad Politecnica, UNA, and at the Research Center in Biotechnology and Informatics (CEBIOTEC), Asunción, Paraguay. Polymerase chain reaction followed by restriction enzyme digestion (PCR-RFLP) was used to identify TP53 p.R337H, and real-time PCR (RT-PCR) was employed for XAF1 p.E134*. Peripheral blood samples from 40,000 Paraguayan newborns (NBs) were used for the TP53 p.R337H tests. The XAF1 p.E134* tests (RT-PCR) were performed on samples from 2000 Paraguayan newborns at the Pelé Pequeno Principe Research Institute, Curitiba, PR, Brazil. Results: The TP53 p.R337H variant was not found in any of the 14 Paraguayan departments investigated. A total of 12 of the 2000 Paraguayan NBs were positive for one XAF1 p.E134* allele. Conclusions: The hypothesis of Spanish immigrants carrying p.R337H to Paraguay was disproved. TP53 p.R337H neonatal testing in Paraguay is not recommended, except when there are families with Brazilian ancestry presenting cancer cases. Additional epidemiological studies are required to determine the likelihood of the identified prevalence of the XAF1 p.E134* allele (1/153) in NBs from Paraguay without TP53 p.R337H to present cancer risk. This study complements the first national initiative for the DNA screening of newborns aimed at mapping the TP53 p.R337H and XAF1 p.E134* variants in Paraguay (based on the regions of residence of the newborns).