Clinical Outcomes of Children With Adrenocortical Carcinoma in the Context of Germline TP53 Status
Connor T.A. Brenna, BSc, ** Orli Michaeli, MD,¿ § Jonathan D. Wasserman, MD, PhD, #| and David Malkin, MDtĮ§
Summary: Adrenocortical carcinoma (ACC) is a rare, aggressive malignancy of the adrenal cortex. This study characterizes a single- institution cohort of children treated for ACC, and explores the relationship between clinical outcomes of ACC and germline TP53 mutation status. We performed a retrospective chart review of 23 consecutive pediatric patients with ACC treated at The Hospital for Sick Children, Toronto, Canada, between 1977 and 2017. Clinical, biochemical, radiologic, pathologic, and genetic data were collected for each patient. ACC diagnosis followed a bimodal age dis- tribution of 0 to 6 (n = 17) and 12+ (n =6) years, with a female:male ratio of 3.6:1. Ten of 20 patients tested for germline TP53 status carried a pathogenic (9) or likely pathogenic (1) variant, including all but 1 male patient. Only 3 patients died of ACC-related causes, each 5 months post-diagnosis. When treated with resection and combination chemotherapy, carriers of germline TP53 mutations may respond more favorably than their wild-type counterparts. In addition, the survival of patients reported in our cohort with high-stage ACC was appreciably greater than previously described (100.0% for stage II, 50.0% for stage III, and 42.9% for stage IV), favoring aggressive intervention in these patient populations.
Key Words: adrenocortical carcinoma, TP53, p53, Li-Fraumeni syndrome, pediatric
(J Pediatr Hematol Oncol 2021;43:e635-e641)
A drenocortical carcinoma (ACC) is a rare and aggressive malignancy that forms from the cortex of the adrenal gland. It is classically differentiated from adrenocortical adenoma by the scoring of 9 histopathologic “Weiss” cri- teria, and more recently by molecular and immunohis- tochemical analyses.1-3 A modified version of the Weiss criteria are used to distinguish ACCs from adrenocortical adenomas in the pediatric context.4 The age of ACC onset
Received for publication July 3, 2020; accepted September 28, 2020.
From the Departments of *Medicine; ¿ Pediatrics, University of Toronto; ¡Program in Genetics and Genome Biology, The Hospital for Sick Children; Divisions of §Hematology/Oncology; and | Endocrinology, The Hospital for Sick Children Toronto, ON, Canada.
This original research manuscript has been revised for the Journal of Pediatric Hematology/Oncology in accordance with the journal’s published requirements.
Supported in part by grants from the Canadian Institutes for Health Research (D.M.), a New Frontiers Program Project of the Terry Fox Research Institute (D.M.) with funds from the Terry Fox Founda- tion, and the University of Toronto Mclaughlin Centre (C.T.A.B.). The authors declare no conflict of interest.
Reprints: David Malkin, MD, Division of Hematology/Oncology, The Hospital for Sick Children, 555 University Avenue, Toronto, ON, Canada M5G 1X8 (e-mail: david.malkin@sickkids.ca).
Supplemental Digital Content is available for this article. Direct URL citations appear in the printed text and are provided in the HTML and PDF versions of this article on the journal’s website, www.jpho- online.com.
Copyright @ 2020 Wolters Kluwer Health, Inc. All rights reserved.
follows a striking bimodal distribution with major peaks in the first and fifth decades of life.5-7 The incidence of ACC in children is estimated to be in the range of 0.2 to 0.4 cases per million individuals per year.8-10 In children, ACCs often present with clinical signs and symptoms of adrenal hor- mone excess.11 Virilization is common with or without manifestations of glucocorticoid excess.11-15 A long delay between initial symptom onset and diagnosis is thought to be partly responsible for the poor prognosis of ACC.14,16,17
Known predictors of a favorable prognosis in pediatric ACC include a younger age at diagnosis, lower primary tumor stage, lower mitotic rate, and the absence of endo- crine signs or atypical mitoses.13,18-24 In addition, recent findings suggest that several common somatic mutations and methylation markers reliably predict prognosis in ACC tumors.25-27 A widely accepted staging system for ACCs classifies these tumors as stage I-IV based on size, resect- ability, and the presence of lymph node involvement or distant metastases.17,28 Current clinical approaches stratify patients by ACC stage to determine treatment with surgical intervention or systemic therapy (mitotane plus chemo- therapeutic agents such cisplatin, etoposide, and doxor- ubicin), or a combination of the two.28-32 Pediatric ACC has strong genetic associations, and is commonly seen in the context of Li-Fraumeni syndrome (LFS).33
Germline TP53 mutations have been reported in 50% to 80% of children presenting with ACC.6 TP53 encodes the p53 nuclear phosphoprotein, which is central to the regu- lation of cell cycle, DNA repair, apoptosis, cellular metab- olism, and senescence.34,35 Germline TP53 mutations are associated with LFS, an autosomal dominant disorder that is clinically characterized by a wide spectrum of early-onset cancers in both children and adults.36-40 A wide range of TP53 mutations have been reported in LFS, most com- monly in the core DNA-binding domain of the p53 protein.35,37 Somatic TP53 mutations are observed in roughly half of all human cancers, and these neoplasms tend to be biologically more aggressive than their TP53 wild-type counterparts.41,42 Interestingly, one of the most common hotspot mutations in the TP53 gene (R175H, arginine> histidine) has strong associations with LFS, whereas the R175L variant (arginine > leucine) has been linked to a more specific failure to suppress ACC formation.43 In addition, the R337H (arginine > histidine) variant has been implicated in a uniquely high population incidence of pediatric ACC observed in Brazil.44,45
The aim of this study was to provide a broad clinical characterization of children treated for ACCs at a large, single, tertiary care institution, and to explore the relation- ship between clinical outcomes of ACC and germline TP53 mutation status. We suggest that germline TP53 mutation status may help reconcile the difference between clinical outcomes of ACC treatment in our cohort and those
reported recently in a multi-institutional clinical trial.28 Our findings provide a new perspective to the value of germline TP53 testing in guiding prognosis and treatment for pedia- tric patients presenting with ACC.
PATIENTS AND METHODS
We retrospectively reviewed patients treated for ACC at The Hospital for Sick Children (SickKids, Toronto, ON, Canada) between 1977 and 2017. Fifteen patients from this cohort have been previously described.46,47 All patients 0 to 18 years of age with a pathologic diagnosis of ACC were included. There were no exclusion criteria. Clinical, bio- chemical, radiologic, pathologic, and genetic data were collected for each patient (summarized in Supplementary Table S1, Supplemental Digital Content 1, http://links.lww. com/JPHO/A411). Germline TP53 mutation status was ascertained for each patient using Sanger sequencing and multiplex ligation-dependent probe amplification to ensure that any small copy number alterations were captured. A list of unique TP53 variants observed in this patient cohort can be found in Supplementary Table S2, Supplemental Digital Content 2 (http://links.lww.com/JPHO/A412). Statistical analysis was limited to the Student t test, with significance set at P=0.05. This study was approved by the institutional Research Ethics Board.
RESULTS
Twenty-three patients were included in the study with a mean age at diagnosis of 5.50 years. These included 18 females (mean age at diagnosis = 5.51 y) and 5 males (mean age at diagnosis =5.25 y) (female:male =3.6:1). Age dis- tribution was bimodal with 17 patients diagnosed before 6 years of age, and the remaining 6 patients all diagnosed after the age of 12. The mean tumor size and weight at presentation were 7.82 cm (length of longest edge) and 211.6 g, respectively. A descriptive analysis of all 23 patients is shown in Table 1. Twenty patients underwent germline TP53 sequencing and 10/20 (50%) were found to harbor a pathogenic (9) or likely pathogenic (1) TP53 mutation. Only 1 patient, within the TP53 variant group, had previously been diagnosed with cancer (rhabdomyosarcoma). One patient in our cohort had a previous diagnosis of LFS before his ACC, and 2 had a substantial family history consistent with LFS in accordance with Chompret criteria.48 Six patients developed distant metastatic disease, and sites of metastases included lung (5) and abdomen (liver, peri- toneum) (4). The 20 patients for whom TP53 gene testing results were available are described in Table 2. No sig- nificant difference was seen in age, sex, stage, or family history between patients with wild-type TP53 and germline TP53 alterations.
A majority of patients (18/23, 78.3%) presented symptomatically, although 1 patient was identified in the context of presymptomatic LFS surveillance,49,50 and information regarding the presenting symptoms was not available for 5 patients. Presentation as the result of endo- crine manifestations outnumbered presentations due to mass effect. Of those patients who underwent genetic testing and presented symptomatically, most presented with more than 1 symptom. The most common presenting symptoms were virilization (30% of patients), change in weight (30%), hypertension (25%), and abdominal pain (15%) (Table 2). No significant differences in presenting symptoms were
| Characteristic | No. Patients (n = 23) |
|---|---|
| Sex | |
| Male | 5 |
| Female | 18 |
| Stage | |
| I | 9 |
| II | 4 |
| III | 2 |
| IV | 7 |
| Not Specified | 1 |
| Age at diagnosis (y) | |
| 0-6 | 17 |
| 6-12 | 0 |
| 12+ | 6 |
| Family history | |
| Cancer before the age of 50 in first-degree siblings or parents | 3 |
| None | 16 |
| Not specified | 4 |
| Treatment | |
| Resection only | 12 |
| Chemotherapy and resection | 10 |
| Radiation | 0 |
found between children with and without known germline TP53 mutations.
Hormone levels measured at the time of diagnosis were available for 17 of 23 patients (Fig. 1). Dihydroepiano- drostene (DHEAS) was elevated in 14 of 15 patients tested, androstenedione and testosterone in 12 of 15, and 17-hydroxyprogesterone in 7 of 11. Urinary free cortisol was elevated in 5 of 11 patients. In addition, 7 of 11 patients had an increased lactic dehydrogenase level.
Of the 22 patients whose ACC stage (as per criteria from Sandrini and colleagues, based on disease stage and tumor size)51,52 was known or could be determined from pathologic features, 13 were lower stage (I/II) and 9 were higher stage (III/IV) (Table 3). All except 1 patient in this study underwent complete tumor resection. Of the 8 patients with higher stage (III/IV) disease who underwent germline TP53 sequencing, 2 of 3 with wild-type TP53 died of ACC and 1 of 5 with mutated TP53 died of ACC. All 3 patients who succumbed to ACC were adolescents. The sole patient who died of ACC with both higher-stage disease and mutant germline TP53 succumbed to rapidly progressive and unresectable disease, having declined chemotherapy after 1 cycle.
The most common pathologic findings were vascular invasion (50%), nuclear atypia (45%), breach of the adrenal gland’s fibrous capsule (30%), and high mitotic rate (25%; defined as > 10 mitoses per 50 HPF) (Table 2). Surprisingly, stage II ACCs were comparable in size and weight to higher stage tumors. Nuclear atypia correlated with presence of germline TP53 gene variants; however, no statistically sig- nificant relationship was observed between pathologic findings and ACC stage at diagnosis.
No patients in this cohort were treated curatively with radiation. Eight patients with stage III/IV disease were treated as per the Children’s Oncology Group ARAR0332 protocol, which uses a combination of mitotane, cisplatin, etoposide, and doxorubicin therapy together with surgical resection and retroperitoneal lymph node dissection for local disease control. In addition, 1 stage I patient was
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| TABLE 2. Descriptive Analysis of TP53 Wild Type and Germline Predisposition Mutation Patients Included in the Current Study | |||
|---|---|---|---|
| Characteristic | No. Patients | ||
| Wild Type (n = 10) | Confirmed TP53 Predisposition Syndrome Mutation (n = 10) | Total (n = 20) | |
| Sex | |||
| Male | 1 | 4 | 5 |
| Female | 9 | 6 | 15 |
| Stage | |||
| I | 4 | 3 | 7 |
| II | 3 | 1 | 4 |
| III | 0 | 2 | 2 |
| IV | 3 | 3 | 6 |
| Not specified | 0 | 1 | 1 |
| Family history | |||
| Cancer before the age of 50 in first-degree siblings or parents | 0 | 2 | 2 |
| None | 9 | 7 | 16 |
| Not specified | 1 | 1 | 2 |
| Outcomes | |||
| Deceased (due to ACC) | 2 | 1 | 3 |
| Event-free survival (%) | 80 | 90 | 85 |
| Average time from diagnosis to death by ACC, when applicable (y) | 0.42 | 0.42 | 0.42 |
| Deceased (all-cause) | 2 | 3 | 5 |
| Average time from diagnosis to death (all-cause), when applicable (y) | 0.42 | 9.19 | 4.81 |
| Alive | 8 | 7 | 15 |
| Average time from diagnosis to last follow-up (y) | 5.78 | 9.89 | 7.84 |
| Presenting symptoms, n (%) | |||
| Virilization | 2 (20.0) | 4 (40.0) | 6 (30.0) |
| Change in weight | 4 (40.0) | 2 (20.0) | 6 (30.0) |
| Hypertension | 3 (30.0) | 2 (20.0) | 5 (25.0) |
| Abdominal pain | 2 (20.0) | 1 (10.0) | 3 (15.0) |
| Hirsutism | 1 (10.0) | 1 (10.0) | 2 (10.0) |
| Pathologic features | |||
| Average tumor size (longest edge, cm) | 8.15 | 7.95 | 8.05 |
| Average tumor weight (g) | 232.6 | 164.4 | 198.5 |
| Vascular invasion, n (%) | 5 (50.0) | 4 (40.0) | 9 (45.0) |
| Nuclear atypia, n (%) | 1 (10.0) | 6 (60.0) | 7 (35.0) |
| High mitotic activity (rate > 10 per 50 HPF), n (%) | 3 (30.0) | 4 (40.0) | 7 (35.0) |
| Capsular breach, n (%) | 3 (30.0) | 2 (20.0) | 5 (25.0) |
Descriptive analysis of those patients included in this current study for whom genetic testing information was available. It contains demographic information, presenting symptoms, and tumor pathology for these patients. ACC indicates adrenocortical carcinoma.
treated with mitotane and 5-fluorouracil, and 1 stage IV patient received mitotane alone. Twelve patients underwent resection without adjuvant therapy, and 1 died before undergoing either resection or chemotherapy. Impact of the evolution of treatment from the 1980s to the more recent ARAR0332 protocol is difficult to assess due to the small numbers of patients ascertained before the late 1990s.
Six patients experienced disease recurrence, and 5 of these 6 harbored germline TP53 mutations. Only 2 of these 6 patients died from disease, and both of these 2 harbored germline TP53 mutations. Seventeen patients (73.9%) are alive after a mean follow-up of 7.84 years (range, 1 mo to 20.08 y). There was no statistically significant difference in overall event-free survival between the group harboring TP53 germline mutations and the group without (Table 2, Supplementary Fig. 1, Supplemental Digital Content 3, http://links.lww.com/JPHO/A413). Four of the 23 patients in our cohort (17.3%) died as the result of ACC, although 1 of these 4 did not undergo germline TP53 testing and so was excluded from this subgroup analysis (Supplementary Fig. 2, Supplemental Digital Content 4, http://links.lww. com/JPHO/A414). Two other patients, both with germline TP53 mutations, died indirectly as the result of subsequent
malignancy (1 due to osteosarcoma, and 1 related to mye- lodysplastic syndrome).
Of the 6 deceased patients in this cohort, 3 harbored germline TP53 mutations, 2 were wild type, and 1 did not undergo genetic testing. For the 5 patients whose mutation status was identified, the mean survival time from diagnosis to death from any cause was longer for patients harboring a germline TP53 mutation (110.3 mo, n=3) than those who were confirmed to be TP53 wild type by genetic testing (5.0 mo, n=2), although this finding was not statistically significant.
DISCUSSION
The impact of germline TP53 mutations on patient outcome has not been previously described in the context of childhood ACC. Survival of patients comprising both groups in this study was, overall, relatively high. The results of this institutional cohort study suggest that, while overall survival among children with and without germline TP53 mutations is comparable, germline TP53 status may influ- ence prognosis and management, although the evidence supporting this suggestion is limited by a small sample size
| Wild-Type or Unknown Germline TP53 (n = 9) | TP53 Predisposition Syndrome Mutation (n=8) | ||||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| 1 | 2 | m | 4 | 5 | 0 | 1 | 8 | a | 2 | = | d | 13 | 14 | 15 | 9 | = | |
| DHEAS | N/A | N/A | |||||||||||||||
| Androstenedione | N/A | N/A | |||||||||||||||
| Testosterone | N/A | N/A | |||||||||||||||
| 17OHP | N/A | N/A | N/A | N/A | N/A | N/A | |||||||||||
| Urinary Free Cortisol | N/A | N/A | N/A | N/A | N/A | N/A | N/A | N/A | N/A | N/A | |||||||
| ACTH | N/A | N/A | N/A | N/A | N/A | N/A | N/A | N/A | N/A | N/A | |||||||
| LDH | N/A | N/A | N/A | N/A | N/A | N/A | |||||||||||
| Undetectable | |
| Within Reference Range | |
| 1-2x Upper Limit Reference Range | |
| 2-4x Upper Limit Reference Range | |
| >4x Upper Limit Reference Range |
commensurate with the rarity of ACC. Notably, ACC patients in our cohort with both high-stage (III/IV) disease and germline TP53 mutations appeared to respond favor- ably to treatment. Together, these findings suggest the possibility that a germline TP53 mutation may confer a survival advantage in the context of pediatric ACC through a biologic mechanism that remains yet unclear, although greater statistical power is required to achieve statistical significance for this association. All 3 patients in our cohort who died as a direct consequence of ACC survived ~5 months from diagnosis, regardless of germline TP53 mutation status.
The female-to-male ratio, mean age of diagnosis, and other tumor features of the 23 pediatric patients in our study cohort were consistent with previously published accounts.22,46 Specifically, our female-to-male ratio of 3.6:1 is close to previously published reports in the range of 1.6- 3.5:1.18,53,54 All except 1 of the male patients in our cohort, however, carried a cancer-predisposing germline mutation, leading us to suggest the possibility that there may be 2 distinct mechanisms of pediatric ACC pathogenesis (with approximately equal incidence, in our cohort): one which
affects wild-type TP53 patients in a sex-specific (female- predominant) manner, and another which occurs by a TP53 mutation cancer predisposition syndrome with equal inci- dence between male and female populations. This idea is supported by our findings that the pathologic feature of nuclear atypia is almost exclusive to ACCs with germline TP53 mutations, although our sample size was not large enough to achieve statistical significance for this finding. However, the suggestion that there may be a distinct, germline TP53 mutation-specific mode of pathogenesis is further supported by a recent study of single nucleotide polymorphisms in pediatric ACC which reported that tumors from children with germline TP53 mutations are unique in their genomic alterations.55 It is important to note, however, that other studies of TP53 in the context pediatric ACC have not reported the same extremes of sex-specificity in TP53 mutation and wild-type groups that we see in our cohort.6,56
One patient in our cohort was identified as having a likely pathogenic germline TP53 mutation, and we consid- ered this patient to belong to the germline TP53 mutation group of our study (which otherwise contained only patients
| Pathologic Features | Stage I (n = 7) | Stage II (n =4) | Stage III (n =2) | Stage IV (n =6) | Total (n=19) |
|---|---|---|---|---|---|
| Average tumor size (longest edge, cm) | 4.49 | 9.63 | 9.00 | 10.1 | 7.82 |
| Average tumor weight (g) | 44.3 | 292.4 | NA* | 280.9 | 211.6 |
| Vascular invasion | 4 | 1 | 1 | 4 | 10 |
| Nuclear atypia | 3 | 2 | 1 | 3 | 9 |
| Capsular breach | 3 | 0 | 0 | 2 | 5 |
| High mitotic activity (rate > 10 per 50 HPF) | 2 | 1 | 0 | 3 | 6 |
Patterns of pathologic features in the ACCs resected from patients in this study, divided by tumor stage. This table includes symptoms of virilization, change in weight, hypertension, and abdominal pain. This table includes only patients for whom genetic testing information was available, with the exception of 1 patient whose tumor stage was unavailable.
*Tumor weight was unavailable for either of the 2 patients with stage III ACC in this study.
ACC indicates adrenocortical carcinoma; NA, not available.
with pathogenic mutations) as per the ACMG Guidelines which, on the basis of several criteria, define likely patho- genic mutations as those with a high degree of certainty to support the claim that they are disease causing.57 Age of diagnosis, tumor size, tumor weight, and cancer stage did not differ significantly between the germline TP53 mutation group and germline TP53 wild-type group. Our cohort included 13 stage I/II patients, 9 stage III/IV patients, and 1 patient whose disease stage was unknown. Of all the deceased patients in our cohort (6), 4 were classified as stage IV ACC and 1 as stage III ACC. The sixth was classified as stage I ACC; this patient died as a consequence of lung metastases from an osteosarcoma, 7 years following ACC treatment. These findings are concordant with previously published trends of incidence and survival for each ACC stage in pediatric patient populations, except that high-stage ACC patients in our cohort displayed markedly greater overall survival (100.0% for stage II, 50.0% for stage III, and 42.9% for stage IV) than has been previously reported.18,51,58,59 We hypothesize that this may be due to favorable surgical results and adherence to systemic che- motherapy regimens in a large tertiary center, as well as a cohort of predominately younger pediatric patients. In particular, we note that the youngest patients diagnosed with ACC have an impressive survival rate, even at an advanced stage, supporting an argument that aggressive treatment in this population is not futile. Furthermore, only 1 of 5 patients with stage III/IV disease who harbored a germline TP53 mutation died of disease, compared with 2 of 3 patients with stage III/IV disease harboring wild-type germline TP53. This patient was the only one in the higher- stage ACC group that did not complete a standard che- motherapy cycle, again suggesting that therapy may be more successful in the context of germline TP53 mutation, warranting further study and raising the possibility that aggressive treatment of these patients should be a focus of advocacy.
In contrast to our findings of pediatric ACC survival, previous studies of pediatric ACC have reported a 5-year survival for stage II patients of 52% and a 2-year survival for stage III and IV patients below 25%, although the treatment of patients in both study cohorts preceded current standards of care; data from studies of adult ACC patients report survival of stage IV disease at 13% and a median survival time of 0.89 years.18,20,30,60 In our cohort, 20 of 23 (87%) children did not die as a direct consequence of ACC (mean follow-up of 7.84y), although it is challenging to draw certain conclusions from this event-rate given limitations in sample size. Notably, however, only 1 of the 3 children who died due to ACC had a germline TP53 mutation, and unlike the other 2, this patient discontinued chemotherapy treat- ment after receiving only a single cycle.
DHEAS was demonstrated to be a remarkably sensi- tive marker of ACC (14/15, 93%), elevated in all but one of the patients in our study tested at the time of ACC diag- nosis, although DHEAS concentration was not found to be a reliable correlate of tumor size.61,62 This is consistent with previous reports suggesting that high concentrations of DHEAS are a feature of tumors originating from the androgen-producing cells in the zona reticularis of the adrenal cortex.17,59 Of the 5 deceased patients in our study for whom biochemical information was available, 4 had DHEAS testing at the time of diagnosis and these levels were below the cohort average (1 within reference range, and 3 elevated to 2 to 4 times the upper limit of the reference
range, compared with the remainder of the cohort in which 11 of 13 patients were elevated >4 times the normal refer- ence range and 2 of 13 were elevated to 2 to 4 times the normal reference range). We propose that this may reflect tumor dedifferentiation, and comparatively reduced responsiveness of dedifferentiated disease to therapy. These findings may imply that pediatric ACC patients with higher DHEAS elevations have a better survival outlook, although low statistical significance from a small sample size again limits firm conclusions. Androstenedione and testosterone levels correlated positively in most patients tested for both hormones, because the former is a metabolic precursor of the latter. Stage IV ACCs were associated with the greatest proportion of presenting symptoms, although no single presenting symptom correlated specifically with any stage of ACC.
Our data regarding average survival time and event- free survival of patients with germline TP53 mutations compared with patients who are germline TP53 wild-type contrasts with a previously published abstract from the American Society of Clinical Oncology which suggested that TP53 mutation status was not prognostically significant in the context of pediatric ACC.28 Our findings suggest a possibility that TP53 germline mutation status may be a clinically valuable feature both for defining prognosis and informing ACC treatment options, in addition to the utility it provides in advising families of pediatric patients with ACC about their own risks of carrying cancer-predisposing mutations and the risk of subsequent cancers in the patient. In addition, our data indicate that the long-term survival of children with high-stage ACC receiving current treatments such as the ARAR0332 protocol is substantially greater than has been previously described, favoring the application of aggressive intervention for these patient populations. Owing to the rarity of ACC, a further exploration of these findings in a larger, multicenter study is warranted.
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