We describe a case of Sweet syndrome, an acute neutrophilic dermatosis that may occur as a paraneoplastic manifestation. A 25-year-old man presented with recurrent abdominal pain and painful erythematous plaques on the trunk and upper limbs, without symptoms of catecholamine excess. Imaging revealed a retroperitoneal mass and extensive hypermetabolic lymphadenopathy on positron emission tomography/computed tomography using fluorine-18 fluorodeoxyglucose (18F-FDG) and gallium-68 DOTA-Tyr3-octreotate (68Ga-DOTATATE), raising concern for metastatic disease. Skin biopsy confirmed Sweet syndrome, while lymph node biopsies demonstrated reactive lymphoid hyperplasia. A computed tomography-guided biopsy of the retroperitoneal mass established the diagnosis of paraganglioma, but was complicated by hypertensive crisis. Subsequent biochemical evaluation revealed markedly elevated plasma normetanephrine levels despite normotension. The patient underwent preoperative alpha-adrenergic blockade followed by surgical resection. Cutaneous lesions resolved rapidly after tumor removal, and plasma normetanephrines normalized, confirming biochemical cure. This case highlights an association between Sweet syndrome and succinate dehydrogenase complex iron-sulfur subunit B (SDHB)-related paraganglioma and underscores the risk of false-positive metastatic findings on functional imaging and reinforces the importance of biochemical exclusion of paraganglioma before biopsy of vascularized retroperitoneal masses.

Background:Carney-Stratakis syndrome (CSS), a rare condition characterized by paragangliomas and/or pheochromocytomas and gastrointestinal stromal tumors (GIST), is caused by germline heterozygous pathogenic variants in the succinate dehydrogenase subunit genes (SDHB, SDHC, SDHD). Methods:Histological, genetic, and functional analyses were conducted in a 59-year-old female with CSS (9 cm left pheochromocytoma, 4.8 cm paraganglioma, and 9.3 cm GIST). Whole-exome sequencing (WES) of germline DNA paired with tumor DNA was performed. Results:WES identified a rare heterozygous germline variant (c.293G>A/p.Arg98His) in the mitochondrial 2-oxoglutarate/malate carrier gene (SLC25A11). This variant, located in a highly conserved residue of the SLC25A11 mitochondrial carrier domain, is predicted to be deleterious in silico (REVEL score = 0.81). WES of pheochromocytoma, paraganglioma, and GIST did not reveal somatic pathogenic variants in genes previously associated with these tumors. A significant reduction in SLC25A11 expression was observed in the tumors of this patient with the SLC25A11 c.293G>A variant (0.69 ± 0.003) compared to tumors from cluster 1 (1.39 ± 0.45; P = 0.0229) and cluster 2 (1.79 ± 0.71; P = .0154). Consistent with the mRNA findings, SLC25A11 protein levels were markedly reduced in the pheochromocytoma and paraganglioma compared to other tumors. Negative staining for 5-hydroxymethylcytosine in all 3 tumors suggests a DNA hypermethylation profile characteristic of cluster 1A, despite normal SDHB expression levels. However, genome-wide copy number variation analysis did not reveal any loss of heterozygosity at the SLC25A11 locus. Conclusion:The loss of SLC25A11 expression in tumors, the absence of somatic drivers, and the hypermethylation status strongly support the role of SLC25A11 in CSS pathogenesis.

Disclosure: F. Freitas-Castro: None. G.F. Fagundes: None. L.S. Santana: None. A.F. Afonso: None. F.L. Ledesma: None. I.C. Soares: None. B.B. Mendonca: None. A. Latronico: None. M.Q. Almeida: None. Background: Pheochromocytomas and paragangliomas (PPGLs) are associated with germline genetic defects in 30-50% of cases and are categorized into three transcriptional clusters. Cluster 1, subdivided into Clusters 1A and 1B, involves genes associated with cellular pseudohypoxia. Cluster 1A encompasses mutations in genes encoding Krebs cycle enzymes (e.g., succinate dehydrogenase complex subunits SDHA, SDHB, SDHC, and SDHD), leading to the accumulation of oncometabolites such as succinate and pyruvate, which increase the stabilization of hypoxia-inducible factor 2-alpha (HIF-2α). Aim: To investigate novel genetic etiologies in patients with PPGLs. Methods: Whole-exome sequencing (WES) of paired germline and tumor DNA was performed on 50 patients with PPGLs who lacked germline defects in previously known susceptibility genes. A targeted analysis enriched for 3,485 genes involved in cellular responses to hypoxia, mitochondrial function, the Krebs cycle, and tumorigenesis was conducted. Results: A germline c.280A>T (p.Lys94*) variant in the Pyruvate Dehydrogenase Phosphatase Catalytic Subunit 2 (PDP2) gene was identified in a 43-year-old female diagnosed with a 10.5 cm abdominal PGL. This stop-codon variant is absent from population databases and has been classified as a variant of uncertain significance. Tumor WES revealed no oncogenic variants. Additionally, immunohistochemistry for SDHB was positive in the tumor. The patient had no family history of cancer. Among her six siblings, one was tested for the variant, with a negative result. She has no children, and her parents are deceased. PDP2 gene encodes an enzyme responsible for dephosphorylating and reactivating the E1 alpha subunit of the pyruvate dehydrogenase complex (PDC), a critical regulator of mitochondrial metabolism. While the PDC has been implicated in glucose metabolism and tumor aggression in other cancers, such as neuroblastoma, it has not been previously associated with PPGLs. No specific phenotype has been associated with this gene. Conclusion: We report, for the first time, a rare loss-of-function PDP2 variant in a patient with abdominal PGL, which may contribute to HIF-2α stabilization and the pathogenesis of PPGL. Ongoing functional studies aim to further investigate this hypothesis. Support: Sao Paulo Research Foundation (FAPESP) grant 2019/15873-6 (to M.Q.A.), and FAPESP post-doctoral fellowship 2021/11240-9 (to F.F-C.). Presentation: Sunday, July 13, 2025

Objective Germline and somatic drivers are identified in 30% and 40% of pheochromocytomas and paragangliomas (PPGLs), respectively. In this study, we investigated the genetic landscape of PPGLs in a Brazilian cohort. Methods We studied 182 index patients with PPGLs (116 females and 66 males), comprising 118 pheochromocytoma and 70 paraganglioma cases. Our optimized sequencing strategy included SANGER sequencing, targeted next-generation sequencing panel, and whole-exome sequencing. Results Germline and somatic pathogenic or likely pathogenic variants in susceptibility genes were identified in 88 (48.4%) and 18 (10.4%) cases, respectively. SDHB was the most frequently affected gene, identified in 30 patients (16.5%), with a germline SDHB exon 1 deletion present in 46.7% of these cases. The Brazilian cohort exhibited a higher rate of germline diagnoses when compared to the European (31%), American (27%), and Chinese (21%) cohorts (P < .001). Five germline variants in new susceptibility genes were identified: (1) Three CHEK2 likely pathogenic or pathogenic variants (c.475T > C/p.Tyr159His; c.362G > A/p.Cys121Tyr; c.319 + 2T > A); and (2) Two BRCA2 pathogenic variants (c.3680_3681delTG/p.Leu1227fs and c.7806-2A > C). These variants are unreported in the Brazilian genomic variant repository. CHEK2 immunostaining was negative in the three tumors, with one case exhibiting CHEK2 loss of heterozygosity. Moreover, the prevalence of CHEK2 or BRCA2 pathogenic or likely pathogenic variants in our cohort was significantly higher compared to global population databases (P < .0001 and P = .0004, respectively). Conclusion Our cohort of PPGLs demonstrated a high frequency of germline diagnoses. Additionally, our findings suggest CHEK2 and BRCA2 as potential susceptibility genes for PPGLs.

CONTEXT:Aldosterone excess chronically induces oxidative stress and cell proliferation. Previously, a single study investigated primary aldosteronism (PA) in patients with papillary thyroid cancer (PTC), albeit without a matched control group. OBJECTIVE:We conducted a propensity score-matched, case-control study to investigate the association between PA and PTC in individuals with arterial hypertension (HT). METHODS:PA was investigated in 137 patients with PTC and HT. The control group included 137 (1:1) age-, sex-, and body mass index-matched individuals with HT. We conducted a secondary analysis in which controls were also matched according to HT stage. RESULTS:The prevalence of PA was 29.20% (95% CI, 21.91%-37.68%) in the PTC group and 20.44% (95% CI, 14.22%-28.35%) in the controls not matched by HT stage (P = .093). Although the PA prevalence was similar in both groups, the frequency of severe HT (stage III or resistant) was significantly lower in the PTC group (23%) compared to the HT controls (73%; P < .001). After matching the controls by HT stage, the prevalence of PA in the PTC group was significantly higher compared to the hypertensive controls (9.56%; 95% CI, 5.39%-16.1%; P < .0001). In the multivariable analysis, PTC was independently associated with PA both in unmatched HT individuals (odds ratio [OR] 4.74; 95% CI, 2.26-10.55; P < .001) and in those matched by HT stage (OR 5.88; 95% CI, 2.79-13.37; P < .001). CONCLUSION:PTC was an independent variable associated with a diagnosis of PA in HT individuals. Therefore, we propose the association between PTC and HT as a new recommendation for PA screening regardless of HT severity.

Context Paragangliomas (PGLs) are rare tumors in adrenal and extra-adrenal locations. Metastasis are found in approximately 5% to 35% of PGLs, and there are no reliable predictors of metastatic disease.Objective This work aimed to develop a prognostic score of metastatic potential in PGLs.Methods A retrospective analysis was conducted of clinical data from a cohort with PGLs and tumor histological assessment. Patients were divided into metastatic PGL (presence of metastasis) and nonmetastatic PGL (absence of metastasis >= 96 months of follow-up) groups. Univariate and multivariable analysis were performed to identify predictors of metastatic potential. A prognostic score was developed based on coefficients of multivariable analysis. Kaplan-Meier curves were generated to estimate disease-specific survival (DSS).Results Out of 263 patients, 35 patients had metastatic PGL and 110 patients had nonmetastatic PGL. In multivariable analysis, 4 features were independently related to metastatic disease and composed the Prognostic Score of Paragangliomas (PSPGL): presence of central or confluent necrosis (33 points), more than 3 mitosis/10 high-power field (HPF) (28 points), extension into adipose tissue (20 points), and extra-adrenal location (19 points). A PSPGL of 24 or greater showed similar sensitivity with higher specificity than the Pheochromocytoma of the Adrenal Gland Scaled Score (PASS) and Grading System for Adrenal Pheochromocytoma and Paraganglioma (GAPP). PSPGL less than or equal to 20 was associated with a risk of metastasis of approximately 10%, whereas a PSPGL of 40 or greater was associated with approximately 80%. The presence of metastasis and Ki-67 of 3% or greater were related to lower DSS.Conclusion The PSPGL, composed of 4 easy-to-assess parameters, demonstrated good performance in predicting metastatic potential and good ability in estimating metastasis risk.

Abstract Disclosure: F. Freitas-Castro: None. L.S. Santana: None. G.F. Fagundes: None. A.F. Afonso: None. E.C. Lobato: None. F.L. Ledesma: None. I.C. Soares: None. B.B. Mendonca: None. M.C. Fragoso: None. A. Latronico: None. C.A. Stratakis: None. M.Q. Almeida: None. Background: Carney-Stratakis syndrome (CSS, OMIM #606864), also reported as “paraganglioma and gastrointestinal stromal tumor syndrome” or Carney-Stratakis dyad, is an autosomal dominant rare syndrome with incomplete penetrance, characterized by the association of paragangliomas (PGL) and/or pheochromocytomas (PHEO) and gastrointestinal stromal tumors (GIST). CSS is mainly caused by germline heterozygous pathogenic variants (PVs) in the succinate dehydrogenase subunit genes (SDHB, SDHC, SDHD), with SDHB and SDHD being the most frequently affected. Aim: To investigate a novel genetic etiology for CSS not associated with germline SDHx defects. Methods: Genetic investigation using SANGER sequencing and multiplex ligation-dependent probe amplification (MLPA) rule out germline defects for SDHx in a 59-year-old woman diagnosed with a 9 cm left PHEO, 4.8 cm PGL and 9.3 cm GIST. Thus, we performed whole exome sequencing of germline DNA (paired with tumor DNA from PHEO, PGL, and GIST) and applied a targeted analysis with the enrichment of 3,485 potential neuroendocrine tumors susceptibility genes associated with cellular response to hypoxia, mitochondrion, Krebs cycle, and tumorigenesis (MAPK, mTOR, ERK, and Wnt signaling). Additionally, we performed RT-qPCR to determine SLC25A11 expression levels in PHEO and PGL samples, stratified into three groups: tumors from the case harboring the SLC25A11 variant, Cluster 1 (n= 4), and Cluster 2 (n= 8). Tumor DNA methylation status was assessed using immunostaining for 5-hydroxymethylcytosine (5hmC). Results: Exome sequencing revealed a new heterozygous germline variant (c.293G>A / p.Arg98His) in the mitochondrial 2-oxoglutarate/malate carrier gene (SLC25A11). This variant, located in a highly conserved residue of the SLC25A11 mitochondrial carrier domain, is predicted to be deleterious in silico (REVEL score= 0.81). Exome sequencing of the PHEO and PGL did not reveal somatic PVs in genes previously associated with PHEO and PGL. Moreover, somatic c-KIT and PDGFRA PVs were not identified in GIST. Notably, a significant downregulation of SLC25A11 expression was observed in tumor samples harboring the SLC25A11 p.Arg98His variant (0.57 ± 0.13) compared with tumors from cluster 1 (1.39 ± 0.45; p = 0.025) and cluster 2 (1.79 ± 0.71; p < 0.001). Interestingly, immunostaining for 5hmC was negative in all tumors (PHEO, PGL and GIST), indicating tumor hypermethylation. Conclusion: A rare germline deleterious variant in SLC25A11 was identified in a patient with CSS. Moreover, the absence of somatic drivers, hypermethylation status and loss of SLC25A11 expression in the CSS tumors support the involvement of this gene with CSS. Support: Sao Paulo Research Foundation (FAPESP) grant 2019/15873-6 (to M.Q.A.), and FAPESP post-doctoral fellowship 2021/11240-9 (to F.F-C.). Presentation: 6/1/2024

Context Invasive and somatostatin receptor ligand (SRL)-resistant pituitary tumors represent a challenge in the clinical practice of endocrinologists. Efforts have been made to elucidate reliable makers for both. Survivin and eukaryotic translation initiation factor-binding protein 1 (4EBP1) are upregulated in several cancers and involved in apoptosis and cell proliferation. Objective We explored the role of these markers in somatotropinomas. Methods Immunostains for survivin and 4EBP1, and also for somatostatin receptor type 2 (SSTR2), Ki-67, and cytokeratin 18, were analyzed in tissue microarrays containing 52 somatotropinoma samples. Tumor invasiveness was evaluated in all samples while drug resistance was evaluated in 34 patients who received SRL treatment. All these parameters were correlated with first-generation SRL (fg-SRL) responsiveness and tumor invasiveness. Results Low survivin expression (P = 0.04), hyperintense signal on T2 weighted image (T2WI) (P = 0.01), younger age (P = 0.01), sparsely granular adenomas (SGA) (P = 0.04), high postoperative growth hormone (GH) and insulin-like growth factor-1 (IGF-1) levels (P = 0.049 and P < 0.001, respectively), and large postoperative tumor size (P = 0.02) were associated with resistance to fg-SRL. Low survivin and SSTR2 expression and high 4EBP1 expression were associated with SGA (P = 0.04, P = 0.01, and P = 0.001, respectively). Younger age (P = 0.03), large tumor pre- and postoperative (P = 0.04 and P = 0.006, respectively), low SSTR2 expression (P = 0.03), and high baseline GH and IGF-1 (P = 0.01 and P = 0.02, respectively) were associated with tumor invasiveness. However, survivin, 4EBP1, Ki-67, and granulation patterns were not associated with tumor invasion. Conclusion This study suggests that low survivin expression is predictive of resistance to fg-SRL in somatotropinomas, but not of tumor invasiveness.

Carney-Stratakis syndrome (CSS) is an autosomal dominant rare syndrome, with incomplete penetrance, characterized by the association of paragangliomas and/or pheochromocytomas and gastrointestinal stromal tumors (GISTs). CSS is caused by germline heterozygous loss-of-function pathogenic variants (PVs) in the succinate dehydrogenase subunit genes (SDHB, SDHC, SDHD), with SDHB and SDHD being the most frequent. To date, only 2 germline SDHC PVs (c.43 C > T; c.405 + 1G > A) have been described in 3 patients with CSS. Three patients with CSS and very distinct clinical presentations are reported here: 1 caused by a germline SDHC large deletion and the others with metastatic GIST and negative genetic investigation for SDHx defects. Two cases (1 and 2) presented with pheochromocytoma (case 1 also with abdominal paraganglioma) and metastatic GIST. Although these 2 cases fulfilled the diagnostic criteria for CSS, the genetic investigation for SDHx PVs by next-generation sequencing and multiplex ligation-dependent probe amplification was negative. Case 3 had a large abdominal paraganglioma and a small low-grade GIST not associated with recurrence or metastasis. This case harbored a germline SDHC exon 3 deletion, not previously reported. In conclusion, CSS is a rare and morbid disease with distinct clinical presentations and genetic heterogeneity, which can contribute to underdiagnosis.

Pediatric adrenocortical tumors (PACTs) represent rare causes of malignancies. However, the south/southeast regions of Brazil are known to have a high incidence of PACTs because of the founder effect associated with a germline pathogenic variant of tumor suppressor gene TP53. We aimed to retrospectively analyze the types of variables among hormone production, radiological imaging, tumor staging, histological and genetic features that were associated with the occurrence of malignancy in 95 patients (71% females) with PACTs from a unique center. The worst prognosis was associated with those aged > 3 years (p < 0.05), high serum levels of 11-desoxicortisol (p < 0.001), tumor weight ≥ 200 g (p < 0.001), tumor size ≥ 5 cm (p < 0.05), Weiss score ≥ 5 (p < 0.05), Wieneke index ≥ 3 (p < 0.001) and Ki67 ≥ 15% (p < 0.05). Furthermore, patients with MacFarlane stage IV had an overall survival rate almost two times shorter than patients with other stages (p < 0.001). Additionally, the subtractions of BUB1B-PINK1 (<6.95) expression (p < 0.05) and IGF-IR overexpression (p = 0.0001) were associated with malignant behavior. These results helped identify patients who are likely to have an aggressive course; further multicenter prospective studies are required to confirm our results. In conclusion, PACTs with these patterns of prognostic factors could be treated using an adjuvant approach that may improve the overall survival in such patients.

Abstract Evaluate expression and amplification of EGFR in cholangiocarcinoma (CCA) and correlate with the different histological types. 74 patients with CCA from 1992 to 2017 were evaluated. Cases were classified in large duct subtype (DL), cholangiolocarcinoma (CLC), intermediate cell carcinoma (ICC) and papillary (LP).The immunohistochemistry (IHQ) was conducted in 71 cases and the amplification of EGFR was using the fluorescence in situ hybridization (FISH) in 48 cases. From the 74 patients, most lesions affected the perihilar topography (54%, 40/74), extrahepatic portion (27%, 20/74) and the least frequent was the intrahepatic (19%, 14/74). Periductal infiltrative macroscopic growth patterns 60.9% (45/74) and the mass forming 33.7% (25/74) were the predominant, intraductal pattern 5.4% (4/74) lower frequency. The DL subtype was the most frequent (66.2%, 49/74), followed by the CLC (21.7%, 16/74). The LP (8.1%, 6/74) and the ICC (4.0%, 3/74) had a lower frequency. In the IHQ, EGFR showed positivity in 80.2% (57/71), presenting moderate intensity 2+ in 55.0% (39/71) of the cases and strong intensity 3+ in 25.3% (18/71), 14 were detected as negative 19.8%. The FISH, of the 48 cases, 10.5% (5/48) were amplified by the gain in the number of copies of the EGFR gene and 89.5% (43/48) were considered negative. The amplified cases were distributed in 12.5% (4/32) of the DL subtype and 12.5% (1/8) of the CLC subtype. The IHQ expression of EGFR in the tumor is high in all histological subtypes of CCA. EGFR amplification occurred in a small portion of the DL and CLC subtypes.

Purpose Breast cancer (BC) is considered a heterogeneous disease composed of distinct subtypes with diverse clinical outcomes. Luminal subtype tumors have the best prognosis, and patients benefit from endocrine therapy. However, resistance to endocrine therapies in BC is an obstacle to successful treatment, and novel biomarkers are needed to understand and overcome this mechanism. The RET, BCAR1, and BCAR3 genes may be associated with BC progression and endocrine resistance. Methods Aiming to evaluate the expression profile and prognostic value of RET, BCAR1, and BCAR3, we performed immunohistochemistry on tissue microarrays (TMAs) containing a cohort of 361 Luminal subtype BC. Results Low expression levels of these three proteins were predominantly observed. BCAR1 expression was correlated with nuclear grade ( p = 0.057), and BCAR3 expression was correlated with lymph node status ( p = 0.011) and response to hormonal therapy ( p = 0.021). Further, low expression of either BCAR1 or BCAR3 was significantly associated with poor prognosis ( p = 0.005; p = 0.042). Pairwise analysis showed that patients with tumors with low BCAR1/low BCAR3 expression had a poorer overall survival ( p = 0.013), and the low BCAR3 expression had the worst prognosis with RET high expression stratifying these patients into two different groups. Regarding the response to hormonal therapy, non-responder patients presented lower expression of RET in comparison to the responder group ( p = 0.035). Additionally, the low BCAR1 expression patients had poorer outcomes than BCAR1 high ( p = 0.015). Conclusion Our findings suggest RET, BCAR1, and BCAR3 as potential candidate markers for endocrine therapy resistance in Luminal BC.

Adrenocortical carcinoma (ACC) is a rare malignancy that is associated with a dismal prognosis. Pan-genomic studies have demonstrated the involvement of ATRX and ZNRF3 genes in adrenocortical tumorigenesis. Our aims were to evaluate the protein expression of ATRX and ZNRF3 in a cohort of 82 adults with ACC and to establish their prognostic value. Two pathologists analyzed immuno-stained slides of a tissue microarray. The low protein expression of ATRX and ZNRF3 was associated with a decrease in overall survival (OS) (p = 0.045, p = 0.012, respectively). The Cox regression for ATRX protein expression of >1.5 showed a hazard ratio (HR) for OS of 0.521 (95% CI 0.273–0.997; p = 0.049) when compared with ≤1.5; for ZNRF3 expression >2, the HR for OS was 0.441 (95% CI, 0.229–0.852; p = 0.015) when compared with ≤2. High ATRX and ZNRF3 protein expressions were associated with optimistic recurrence-free survival (RFS) (p = 0.027 and p = 0.005, respectively). The Cox regression of RFS showed an HR of 0.332 (95%CI, 0.111–0.932) for ATRX expression >2.7 (p = 0.037), and an HR of 0.333 (95%CI, 0.140–0.790) for ZNRF3 expression >2 (p = 0.013). In conclusion, low protein expression of ATRX and ZNRF3 are negative prognostic markers of ACC; however, different cohorts should be evaluated to validate these findings.

Adrenocortical tumors (ACT) in the adult and pediatric population are generally considered distinct entities due to differences in molecular events related to tumorigenesis, clinical presentation, and outcome. Furthermore, pathological criteria used for diagnosis and prognostication of ACT in adults are usually inadequate for predicting the biological behavior of ACT in children. Here, we analyzed 146 adult and 44 pediatric (< 15y/o) ACT with long-term clinical follow-up and furthered current evidence on the clinical and pathological differences between pediatric and adult tumors. Predilection for female over male gender was observed in both cohorts, but more so in adults (84% vs. 61%, p = 0.003). Cushing syndrome was more frequent in adults (p < 0.001), whereas virilization, either isolated (p < 0.001) or combined to Cushing (p = 0.047), was more common in children. The Ki67 labelling index (LI) of pediatric adenomas and carcinomas was much higher than their corresponding tumors in adults (p < 0.001). Despite these differences, pathological analyses including the evaluation of Ki67 greatly improved patient prognostication in both age cohorts. Indeed, increased Weiss scores and Ki67 indexes correlated with poor overall- and disease-free survival in adult patients with carcinoma. Among the proliferative indexes tested, Ki67 LI ≥ 10% showed the highest hazard ratio (HR) for recurrence and the Ki67 LI ≥ 3% showed the highest HR for survival. In pediatric tumors, the Wieneke score (p < 0.001) and the Ki67 LI (p < 0.001) showed high accuracy for predicting biological behavior, and increased scores/indexes correlated with worse overall and disease-free survival. In this age cohort, Ki67 LI < 10% was able to rule out malignant behavior, whereas Ki67 LI ≥ 15% may be used to predict the patients with higher risks of recurrence and/or poor outcome.

Adrenocortical carcinoma (ACC) is a rare malignant neoplasia that is usually associated with a dismal prognosis. ACC overall survival (OS) depends on the particular biology of the tumor. Pan-genomic studies have demonstrated the involvement of ATRX and ZNRF3 genes in adrenocortical tumorigenesis. Aim: To evaluate ATRX and ZNRF3 protein expression in a large cohort of adults with ACC followed at a single tertiary referral center to establish the prognostic value of these genes. Methods: Two pathologists analyzed immunohistochemically-stained slides for ATRX and ZNRF3 (blinded assessment) proteins using tissue microarrays comprising tissue samples from 82 ACC (kappa 0.854; P<0.001). Cohort: female 76.8%; median age 38.2 (15.3–85.4) years; 67.5% of patients presented with hypercortisolism; ENSAT stage 1, 11.12%; 2, 44.44%; 3 and 4: (44.44%). The median follow-up time was 39.58 (1.4–406.8) months; 39 patients died during this period. Low protein expression was defined as follows: ATRX, when < 25% of tumor cells were immunoreacted; ZNRF3, when a score < 3 (extension + intensity) was established. Results: Low ATRX protein expression was positively associated with the age at diagnosis (P< 0.001), and it was negatively correlated with tumor size (P=0.020) and with Weiss score (P=0.033). Low ZNRF3 protein expression correlated negatively with tumor weight (P=0.026), with tumor size (P= 0.005), and with Weiss score (P=0.002). Regarding OS, the low protein expression of ATRX and ZNRF3 was associated with a decrease of OS (P=0.045 and P=0.012, respectively). The Cox model for ATRX protein expression showed a HR of 0.521 (95% CI 0.273–0.997; P=0.049); for ZNRF3 expression, HR = 0.441 (95% CI 0.229–0.852; P=0.015). The combined protein expression of both genes was also associated with a decrease in OS in this cohort (P=0.02). The high ATRX and ZNRF3 protein expressions were positively associated with optimistic recurrence-free survival (P=0.027, P=0.005, respectively). Conclusion: The knowledge of tumor biology is an important step to deliver personalized medicine, affording the opportunity to design better clinical approaches and targeted therapies. This study brought pan-genomic studies closer to clinical practice with an easily accessible tool for stratifying patients with ACC. We demonstrated that low levels of protein expression of ATRX and ZNRF3 are negative prognostic markers of ACC.