Association of the Highly Prevalent TP53 R337H Mutation With Pediatric Choroid Plexus Carcinoma and Osteosarcoma in Southeast Brazil
Ana Luiza Seidinger, BSc1; Maria José Mastellaro, MD2; Fernanda Paschoal Fortes, BSc1; Juliana Godoy Assumpção, PhD1; Izilda Aparecida Cardinalli, PhD3; Mônica Aparecida Ganazza, MSc1; Raul Correa Ribeiro, MD4; Silvia Regina Brandalise, PhD2,5; Simone dos Santos Aguiar, PhD2; and José Andrés Yunes, PhD1
BACKGROUND: The inherited, low-penetrance arginine-to-histidine substitution at codon 337 (R337H) of the tumor protein 53 gene (TP53) is clustered in southeast Brazil (estimated frequency, 0.3%). Although its tumorigenic effect initially appeared to be tissue-specific, recent evidence suggests its association with a broader range of tumors. Therefore, the authors of this report investigated the spectrum of pediatric malignancies associated with the TP53 R337H mutation at a single referral institution in southeast Brazil. METHODS: Genomic DNA samples from 493 chil- dren with malignancies were screened for the R337H mutation. Available tumor samples from carriers were investi- gated for loss of heterozygosity (LOH) and nuclear p53 accumulation. Clinical data were obtained from medical records. RESULTS: Sixty-five of 70 patients (93%) with adrenocortical tumors (ACTs), 9 of 13 patients (69%) with choroid plexus carcinoma (CPC), and 3 of 41 patients (7.3%) with osteosarcoma carried the mutation. The proportion of CPC to choroid plexus papilloma (CPP) was much higher than that reported elsewhere. Osteosarcoma in carriers had a significantly poorer outcome (P = . 02). The mutation was not identified in patients who had acute lymphoblas- tic leukemia (ALL) (n = 187), recurrent ALL (n = 49), acute myeloid leukemia (n = 44), lymphoma (n = 30), non-CPC central nervous system tumors (n = 26), Ewing sarcoma (n = 25), or rhabdomyosarcoma (n = 8). Among the tumors that were available for analysis, LOH with retention of the mutant allele was confirmed in 21 of 21 ACTs, in 2 of 2 CPCs, and in 2 of 3 osteosarcomas that were positive for R337H. CPCs and osteosarcomas that were positive for R337H had marked nuclear accumulation of p53. CONCLUSIONS: The current findings demonstrated compellingly that the TP53 R337H mutation is associated not only with ACT but also with CPC and, to a lesser extent, with osteosarcoma, both of which are core-component tumors of the Li-Fraumeni syndrome. Cancer 2011;117:2228-35. @ 2010 American Cancer Society.
KEYWORDS: TP53, R337H, adrenocortical tumors, choroid plexus carcinoma, osteosarcoma.
The inherited germline arginine-to-histidine substitution at codon 337 (R337H) mutation of the tumor protein 53 gene (TP53) is unusually common in southeast Brazil, occurring in an estimated 0.3% of the population,1 and its fre- quency is higher than that predicted for other TP53 germline mutations.2 Germline mutations in the TP53 gene usually are associated with Li-Fraumeni syndrome (LFS) and with LFS-like syndrome (LFLS),3,4 which predispose carriers to a broad spectrum of early onset tumors,4,5 including pediatric soft tissue and bone sarcomas, central nervous system (CNS) tumors, leukemia, and adrenocortical tumors (ACTs).4,5 The vast majority of TP53 germline mutations that have a known association with LFS and LFLS span sequences that encode the DNA-binding domain of the protein.3,4 Other
Corresponding author: José Andrés Yunes, PhD, Centro Infantil Boldrini, R. Dr. Gabriel Porto, 1270, CEP 13083-210, Campinas, SP, Brazil; Fax: (011) 55-19-32893571; andres@boldrini.org.br
1Molecular Biology Laboratory, Boldrini Children’s Center, Campinas, Sao Paulo, Brazil; 2Department of Oncology, Boldrini Children’s Center, Campinas, Sao Paulo, Brazil; 3Anatomical Pathology Laboratory, Boldrini Children’s Center, Campinas, Sao Paulo, Brazil; 4International Outreach Program, St. Jude Children’s Research Hospital, Memphis, Tennessee; 5Pediatric Hematology and Oncology, State University of Campinas, Campinas, Brazil
We thank Sharon Naron for expert editorial review.
DOI: 10.1002/cncr.25826, Received: October 5, 2010; Revised: November 8, 2010; Accepted: November 9, 2010, Published online December 29, 2010 in Wiley Online Library (wileyonlinelibrary.com)
TP53 mutations, including the R337H mutation, also have been implicated in tumorigenesis,6,7 although the spectrum of tumors associated with these mutations is not well established.
The TP53 R337H mutation was discovered origi- nally among children with ACT in southern Brazil, where the incidence of ACT is high.6 Although it was believed initially that the mutation selectively promoted pediatric ACT,6,8,9 our recent study suggested its involvement in the pathogenesis of breast cancer.1º Moreover, Achatz et al11 identified R337H carriers among individuals from LFS/LFLS families. It is noteworthy that the cancer his- tory of most R337H-positive families does not meet the criteria for LFS/LFLS,6 suggesting that the R337H muta- tion may be associated with a unique spectrum of tumors. Therefore, we investigated the frequency of the TP53 R337H mutation in different pediatric tumors at a pediat- ric cancer center serving southeast Brazil.
MATERIALS AND METHODS
Treating Institution and Patient Population
The Boldrini Children’s Center (Boldrini), a referral cen- ter for childhood hematologic and oncologic diseases, is located in Campinas, Brazil, in the southeastern state of Sao Paulo. Approximately 95% of its patients are referred from the states of Sao Paulo (86.3%) and Minas Gerais (8.5%). It is known that Sao Paulo and the neighboring state of Parana are within the TP53 R337H mutation cluster. During the period 2002 through 2009, an average of 268 children with newly diagnosed malignancies were admitted to Boldrini annually, including an annual aver- age of 93 patients with leukemia (lymphoblastic and mye- loid), 34 patients with lymphoma, 38 patients with brain tumors (2.6 choroid plexus tumor), 15 patients with Wilms tumor, 10 patients with neuroblastoma, 10 patients with soft tissue sarcoma, 10 patients with osteo- sarcoma, 5 patients with ACT, and 53 patients with other malignancies. Since 2002, all tumor diagnoses at Boldrini have been reviewed histologically for accuracy by 1 of the authors (I.A.C.). Demographic data were obtained from patients’ medical records. This study was approved by the Ethical Research Committee of the Faculty of Medical Sciences at the State University of Campinas. That com- mittee waived the signature of informed consent, because this research was conducted using retrospective samples from the tumor bank. The genetic testing of patients’ rela- tives was performed after the families had been informed about the test and had signed the informed consent.
Tumors and Samples
Peripheral blood mononuclear cells (MNCs) and/or tu- mor samples (bone marrow MNCs, tumor sections, and biopsy specimens) were used for genomic DNA testing.
Adrenocortical tumors
Since 2006, all children at Boldrini with newly diag- nosed ACT have been tested for the TP53 R337H muta- tion. Testing also has been offered to patients with ACT on active follow-up, and available samples from patients with ACT who have died or lost to follow-up have been evaluated retrospectively. Genomic DNA samples from 70 of 102 patients who had ACT diagnosed at Boldrini between 1982 and 2009 were available for testing.
Central nervous system tumors
Genomic DNA samples from 25 patients with CNS tumors were tested for the TP53 R337H mutation. These patients were selected only on the basis of availability of frozen tumor samples. After finding that only children with choroid plexus carcinoma (CPC) carried the R337H mutation, we tested samples from an additional 14 patients with choroid plexus tumors (CPT), 9 of whom had CPC and 5 of whom had choroid plexus papilloma (CPP).
Other tumors
Tumor samples other than ACT and CNS tumors that were tested for the R337H mutation included diag- nostic bone marrow MNCs from 187 patients with acute lymphoblastic leukemia (ALL) who were enrolled consec- utively on the Brazilian ALL protocol GBTLI-99,12 49 patients with relapsed ALL, and 44 patients with newly diagnosed acute myeloid leukemia; diagnostic lymph node biopsy samples from 30 patients who were diagnosed consecutively with lymphoma (Hodgkin lym- phoma, = 20; non-Hodgkin lymphoma, n = 10); periph- eral blood MNCs from 41 patients who were diagnosed consecutively with osteosarcoma; and tumor biopsy speci- mens from 25 patients with Ewing sarcoma and 8 patients with rhabdomyosarcoma.
DNA Analysis
Genomic DNA was isolated from peripheral blood MNCs (n = 120) or from samples of tumor or leukemia (n = 399) by using standard phenol-chloroform extrac- tion.13 In some patients, both tumor tissue and MNCs were available. TP53 exon 10 was amplified by using pri- mers and polymerase chain reaction (PCR) conditions
described previously .? The PCR product was digested with HhaI (Fermentas Inc., Glen Burnie, Md). This restriction enzyme yields 2 fragments (293 base pairs [bp] and 154 bp) of the wild-type amplicon but only 1 frag- ment when the R337H mutation is present (the mutation eliminates the restriction site within the amplicon) (Fig. 1A,B). Exon 10 of mutation-positive samples was sequenced by using the BigDye Terminator Cycle Sequencing Ready Reaction Kit (Applied Biosystems, Foster City, Calif) in an ABI PRISM 3700 automated sequencer (Applied Biosystems) to confirm the guanine- to-adenine transition at the second base of codon 337 (Fig. 1C).
Immunohistochemistry
After dewaxing and rehydration, 5-um-thick CPC and os- teosarcoma tumor sections were treated with 3% H2O2 to reduce endogenous peroxidase activity and then under- went wet heat-mediated antigen retrieval with TRIS-eth- ylene diamine tetraacetic acid, pH 8.9. The sections were incubated for 1 hour at 37℃ with mouse monoclonal anti-p53 antibody (clone DO-7; Dako A/S, Glostrup, Denmark) diluted 1:300. This antibody is specific for an N-terminal epitope and reacts with both wild-type and mutant human p53 proteins. Colon adenocarcinoma sec- tions were stained in parallel as positive controls. For the analysis of INI1 (SWI/SNF-related, matrix associated, actin-dependent regulator of chromatin, subfamily b, member 1 or SMARCB1), sections were incubated as described above with the mouse monoclonal anti-INI1 antibody (clone 3E10; ABD Serotec, Raleigh, NC) diluted 1:100. The results were observed by using the standard avidin-biotin complex method with the Dako LSAB+ System horseradish peroxidase kit.
Statistical Analysis
Age at tumor diagnosis was compared between patients with TP53 R337H mutation-positive and mutation-nega- tive tumors using the Mann-Whitney test. Nominal varia- bles were compared by using the chi-square test (the Fisher test was used when the expected number in a cell was <10). The criterion for statistical significance was a = . 05. Statis- tical analyses were conducted with GraphPad Prism soft- ware (GraphPad Software Inc., San Diego, Calif).
RESULTS
Adrenocortical Tumors
Like in other pediatric oncology units6,8,9 within the TP53 R337H cluster region, most of our patients with
M
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E
| Tumor Type | No. of Nontumor Samples Tested | No. of Tumor or Leukemia Samples Tested | No. of Patients Positive/ Totalª | Prevalence of R337H, % |
|---|---|---|---|---|
| AML | 0 | 44 | 0/44 | 0 |
| ALL (nonrelapsed)b | 0 | 187 | 0/187 | 0 |
| ALL (Relapsed)" | 0 | 49 | 0/49 | 0 |
| CNS tumord | 2 | 24 | 3/25 | 12 |
| Choroid plexus carcinomae | 7 | 8 | 9/13 | 69 |
| Choroid plexus papillomae | 2 | 3 | 0/5 | 0 |
| Ewing sarcoma | 0 | 25 | 0/25 | 0 |
| Rhabdomyosarcoma | 0 | 8 | 0/8 | 0 |
| Osteosarcoma | 41 | 3 | 3/41 | 7 |
| Lymphoma‘ | 0 | 30 | 0/30 | 0 |
| Adrenocortical tumor | 70 | 21 | 65/70 | 93 |
AML indicates acute myeloid leukemia; ALL, acute lymphocytic leukemia; CNS, central nervous system.
aBoth nontumor (peripheral blood mononuclear cells) and tumor tissues from some patients were analyzed; therefore, the total number of patients is less than the totals in the second and third columns.
b Biphenotypic leukemia (n=1), B-cell ALL (n=159), and T-cell ALL (n=27).
CB-cell ALL (n=37) and T-cell ALL (n=12).
d The original cohort of 25 patients with astrocytoma (n=7), ependymoma (n=4), choroid plexus carcinoma (n=4), medulloblastoma (n=02), and others (n=8) was used for initial screening.
” The total number analyzed, including patients who were added after the initial screening (choroid plexus carcinoma, n=13; choroid plexus papilloma, n=5). ‘Hodgkin lymphoma (n=20) and non-Hodgkin lymphoma (n=10).
ACT (65 of 70; 93%) carried this mutation (Table 1). All 21 of the available tumor samples from carriers had loss of heterozygosity (LOH) with retention of the mutated al- lele. Genomic TP53 sequence analysis of the 5 children with ACT who did not carry the TP53 R337H mutation revealed the wild-type sequence in exons 5 through 8. We observed no statistically significant difference in survival between carriers and noncarriers of the mutation (P = .579; Fisher exact test).
Central Nervous System Tumors
Three of the 25 CNS tumors that were tested initially on the basis of availability had the TP53 R337H mutation. Remarkably, all 3 were CPC. Therefore, we investigated additional CPTs. During the study period, 21 of 309 (6.8%) CNS tumors had been diagnosed as CPT. All 21 CPTs that were included in this analysis were immuno- positive for INI1 (data not shown). We tested 14 of these tumors for which samples were available. Six of the 9 CPC samples that were tested, but none of the 5 CPP samples that were tested, were positive for the TP53 R337H mutation. Hence, the mutation was detected in a total of 9 of 13 (69%) CPC samples that were tested (Ta- ble 1). Next, we assessed whether the TP53 R337H muta- tion played a role in CPC tumorigenesis. Marked nuclear accumulation of p53 (>80% of cells) was noted in all 8 tested CPC samples from carriers of the mutation (Fig.
1D, Table 2). Moreover, LOH with retention of the mu- tant allele was noted in 2 of 2 tested tumors. Although no statistically significant difference could be detected, CPC appeared to present at an earlier age in patients who car- ried the TP53 R337H mutation than in noncarriers (me- dian age at onset: 9 months vs 38 months; P = . 06; Mann-Whitney test). This difference was not observed in ACT or osteosarcoma (P = . 873 and P = . 136, respec- tively; Mann-Whitney test). CPC also appeared to have a worse outcome in carriers (5 patients with CPC died, 1 patient relapsed, and 3 were in remission) than in noncar- riers (2 patients died, and 2 were in remission), although no statistically significant difference could be detected (P = 1; Fisher exact test) in these small subsets.
Osteosarcoma
Three of 41 patients with osteosarcoma (7%) were identi- fied as carriers of the TP53 R337H mutation (Table 1). Their tumor characteristics and outcomes are summarized in Table 3. Two of these patients had osteoblastic osteo- sarcoma, and the other patient had a small cell osteosar- coma. Two of the 3 tumors had chondroblastic differentiation. LOH with retention of the mutant allele and strong nuclear accumulation of p53 were detected in 2 of the 3 tumors (Fig. 1E). In the third tumor, both the mutated and wild-type alleles were detected in normal and tumor tissues. All 3 of these patients had extremely
| Patient | DX | Sex | Age at DX, mo | TP53 R337H Status | TP53 IHCª | Treatment | Outcome | ||
|---|---|---|---|---|---|---|---|---|---|
| Normal Tissue | Tumor Tissue | Intensity | Localization | ||||||
| 1 | CPC | Boy | 13 | NA | Positive homozygous | ++++ | Nuclear | S, C | Death; PD |
| 2 | CPC | Girl | 12 | Negative | NA | NA | NA | S, C | Alive; 6-y DFS |
| 3 | CPC | Boy | 27 | Positive heterozygous | Positive homozygous | ++++ | Nuclear | S, C, R | Death; PD |
| 4 | CPC | Girl | 84 | NA | Negative | ++++ | Nuclear | S, C, R | Alive; 5-y DFS |
| 5 | CPC | Girl | 5 | NA | Positive homozygous | ++++ | Nuclear | S, C | Death unrelated to |
| disease | |||||||||
| 6 | CPC | Girl | 27 | NA | Negative | ++++ | Nuclear | S | Death; S complications |
| 7 | CPC | Boy | 49 | NA | Negative | þ | Nuclear | C | Death; sepsis during C |
| 8 | CPC | Boy | 9 | Positive heterozygous | NA | ++++ | Nuclear | S | Death; bleeding during S |
| 9 | CPC | Girl | 8 | Positive heterozygous | NA | ++++ | Nuclear | S, C, R | Alive; 3-mo DFS |
| 10 | CPC | Girl | 17 | Positive heterozygous | NA | ++++ | Nuclear | S | Alive; relapsed |
| 11 | CPC | Boy | 36 | Positive heterozygous | Positive homozygous | ++++ | Nuclear | S, C | Alive; receiving C |
| 12 | CPC | Girl | 8 | Positive heterozygous | NA | NA | NA | S, C | Alive; receiving C |
| 13 | CPC | Boy | 3 | NA | Positive homozygous | +++ | Nuclear | S | Death; PD |
| 14 | CPP | Girl | 24 | NA | Negative | 0 | — | S | Alive |
| 15 | CPP | Girl | 1 | Negative | Negative | 0 | — | S | Alive; 4-y DFS |
| 16 | CPP | Boy | 10 | NA | NA | 0 | — | S | Alive; 3-y DFS |
| 17 | CPP | Girl | 52 | NA | NA | NA | NA | S | Lost to follow- up |
| 18 | CPP | Girl | 29 | NA | NA | +++ | Nuclear | S | Lost to follow- up |
| 19 | CPP | Boy | 88 | NA | Negative | 0 | — | S | Alive; 2-y DFS |
| 20 | CPP | Boy | 11 | NA | Negative | 0 | — | S | Alive; 2-y DFS |
| 21 | CPP | Boy | 9 | Negative | Negative | ++++ | Nuclear | S | Death; bleeding during S |
TP53 R337H indicates the arginine-to-histidine substitution at codon 337 in the tumor protein 53 gene; IHC, immunohistochemistry; CPC, choroid plexus carci- noma; NA not applicable; S, surgery; C, chemotherapy; PD, progressive disease; DFS, disease-free survival; R, radiotherapy; CPP, choroid plexus papilloma. ªImmunohistochemistry was scored as 0 (no cells positive), + (up to 25% of cells positive), ++ (26% to 50% of cells positive), +++ (51% to 75% of cells positive), or ++++ (>75% of cells positive).
aggressive tumors with pulmonary metastasis at diagnosis, and all 3 died of progressive disease after a median of 15 months, whereas only 9 of the other 38 patients with os- teosarcoma who were not carriers of the TP53 R337H mutation had died at the time of this report (P = . 021; Fisher exact test).
Other Tumors
The TP53 R337H mutation was not detected among patients with ALL (n = 187), relapsed ALL (n = 49), acute myeloid leukemia (n = 44), Hodgkin or non-Hodg- kin lymphoma (n = 30), Ewing sarcoma (n = 25), or rhabdomyosarcoma (n = 8) (Table 1).
Geographic Distribution of the TP53 R337H Mutation
Altogether, 22 carriers of the mutation (ACT, n = 20; CPC, n = 2) resided in the state of Minas Gerais, suggest- ing that the TP53 R337H cluster region includes not only the states of Sao Paulo and Parana but also the southern portion of the state of Minas Gerais (Fig. 2).
DISCUSSION
Here, we present for the first time compelling evidence that the TP53 R337H germline mutation is strongly asso- ciated not only with ACT but also with CPC in this geo- graphic cluster region. Nine of 13 patients (69%) with CPC who were tested at our institution were positive for the mutation. Approximately 7% of pediatric brain tumors evaluated since 2002 have been CPT, for a relative frequency 2 to 3 times that reported in other pediatric brain tumor series.14,15 This observation cannot be attrib- uted to referral bias, because our institution has no specific program or unusual expertise in managing this malig- nancy. Moreover, the ratio of CPP to CPC, which typi- cally is reported as 5:1 in pediatric series,14 was only 0.6:1 at our center, indicating a surprisingly high relative fre- quency of CPC, which we estimate to be 5 to 10 times that reported in other series.1 14
The incidence of CPC at other institutions within the TP53 R337H cluster region is not yet known. A high relative frequency of pediatric CPC has been noted at Clinics Hospital in Curitiba, Brazil, which is within the cluster region.16 However, it remains to be determined whether these tumors are associated with the TP53
| Characteristic | Patient 1 | Patient 2 | Patient 3 |
|---|---|---|---|
| Osteosarcoma type | Small-cell osteosarcoma; focal chondroblastic differentiation | Osteoblastic osteosarcoma; extensive chondroblastic differentiation | Osteoblastic osteosarcoma |
| Primary site | Medial distal femur | Proximal humerus | Medial distal femur |
| Tumor size, cm | 13×12×8 | 8.7×8.5 | 10×9×9 |
| Soft tissue invasion | Yes | Yes | Yes |
| Articular cartilage invasion | Yes | Yes | Yes |
| Neural invasion | No | NA | No |
| Angiolymphatic invasion | No | NA | No |
| Pulmonary metastasis | Yes | Yes | Yes |
| Tumor necrosisª | 85% | NA | 93% |
| p53 Immunohistochemistryb | +++ | +++ | þ |
| Tumor LOH | Yes | Yes | No |
| Outcome | Death; PD | Death; PD | Death; PD |
NA indicates not available; LOH, loss of heterozygosity; PD, progressive disease.
a After chemotherapy.
b Immunohistochemistry was scored as + (up to 25% of cells positive), ++ (26% to 50% of cells positive), +++ (51% to 75% of cells positive), or ++++ (≥75% of cells positive).
Minas Gerais
São Paulo
Paraná
1 CASE
2 CASES
4 CASES
6 CASES
200 KM
CENTRO INFANTIL BOLDRINI
R337H mutation. Additional studies of pediatric and adult choroid plexus tumors in conjunction with regional population-based registry data should estab- lish the incidence of these tumors within the cluster region.
ACT and choroid plexus tumor reportedly have occurred simultaneously in a single patient or in related children in families that carry germline TP53 muta- tions.17,18 We did not observe both malignancies in any patient in our series. It is possible that few children who
have CPC associated with the TP53 R337H survive long enough to develop ACTs.
Loss of the wild-type allele and retention of the mutated allele is considered a hallmark of tumorigenesis mediated by tumor suppressor genes.19 Virtually all pediat- ric patients with ACTs who are carriers of the TP53 R337H have LOH with retention of the mutated allele.” Although only 2 patients with CPC in our series were tested for LOH, both tumors had LOH with retention of the mutated allele. Moreover, 8 of 8 tested cases of CPC among carriers of the TP53 R337H mutation had strong nuclear localization of p53, suggesting the involvement of this mutation in tumorigenesis. Although the p53 protein expressed by the mutated gene is relatively functional and is not associated typically with LFS or LFLS,20 it may exert a tumor-promoting effect in specific tissues, such as the cho- roid plexus and the adrenal cortex. This premise is consist- ent with the recent observation by Tabori et al21 that children with CPC who had no history of LFS or LFLS carried TP53 polymorphisms that reduced p53 activity.
The TP53 R337H mutation was identified in 7% of our patients with osteosarcoma. This frequency is much higher than that reported for the general population of this region (0.3%) and is higher than that reported for nonselected North American patients with osteosarcoma who had any constitutional TP53 mutation (3%).22 Investigators at the Clinics Hospital in Sao Paulo also observed an association between osteosarcoma and the TP53 R337H mutation in this region of Brazil. Those investigators reported that, of 47 patients with osteosar- coma, 8 had tumors with positive immunostaining for nu- clear p53, and 4 of those 8 tumors harbored the TP53 R337H mutation.23 However, at our institution, the fre- quency of the mutation was much lower among patients with osteosarcoma than among patients with ACT (93%) and CPC (69%). Moreover, unlike patients with ACT and CPC, patients with osteosarcoma had no evidence of increased frequency, and 1 of 3 patients with osteosar- coma who had the TP53 R337H mutation retained the wild-type TP53 allele. Taken together, these results sug- gest that the TP53 R337H mutation plays a different role in the development and progression of osteosarcoma than in those of ACTs and choroid plexus tumors.
The TP53 R337H mutation was not detected in any of our patients with newly diagnosed ALL (n = 187), relapsed ALL (n = 49), or acute myeloid leukemia (n = 44), consistent with the findings of others.24 A recent study identified only 2 patients with leukemia (2.7%) among 75 patients who had TP53 mutations, and both of
those patients had other primary cancers that were typical of LFS.25 Although an association has been reported between acquired TP53 mutations and relapsed ALL,26 our results provided no indication that the TP53 R337H mutation plays a role in ALL relapse in this region. Nor did we identify the TP53 R337H mutation among 30 patients with of lymphoma (Hodgkin lymphoma, n = 20; non-Hodgkin lymphoma, n = 10), 25 patients with Ewing sarcoma, or 8 patients with rhabdomyosarcoma.
The prognostic implications of the TP53 R337H mutation in children with malignancies remain elusive. There is no evidence that the mutation is associated with prognosis in patients with ACTs. Children with com- pletely resected small tumors have an excellent prognosis, and patients with residual or metastatic disease have a very poor outcome regardless of the presence or absence of the germline mutation.27 In patients with pediatric CPC, prognostic analysis is limited by the small number of patients. A recent study21 suggested that TP53 mutations are associated with a poor prognosis in children with CPC; however, all of the mutations in that study’s popu- lation affected the p53 DNA-binding domain. Our 3 patients with osteosarcoma who carried the TP53 R337H mutation had disseminated disease at diagnosis and had dismal outcomes. To fully address the role of R337H in the incidence and prognosis of these tumors, we recom- mend TP53 R337H screening for every pediatric patient with CPC or osteosarcoma in southern Brazil.
Currently, there is no evidence of de novo R337H mutations. In this study, 43 families (41 from patients with ACT and 2 from patients with CPC) were tested for the presence of R337H, and the results demonstrated that it was inherited in all cases. The familial cancer histories of our patients who carried the TP53 R337H mutation were available only from medical records and were not obtained by a genetic counselor. For that reason, we lacked sufficient information to adequately investigate the association between R337H and LFS/LFLS among our patients. However, very few of our carriers’ medical records information was compatible with LFS or related syndromes, consistent with reports by others6,8,9; thus, there had been no reason to suspect a germline TP53 mutation. This finding highlights the need for more in- clusive clinical criteria, such as those described by Gonza- lez et al,25 for the identification of potential carriers of low-penetrance germline TP53 mutations. Although the R337H mutation appears to be unique to Brazil, it is likely that it and other low-penetrance TP53 mutations play a much broader role in human cancers.
CONFLICT OF INTEREST DISCLOSURES
This study was supported by a grant from Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (PROCAD Pro- ject 247/2007). The authors acknowledge support by the St. Jude International Outreach Program through a Center-of-Excel- lence grant from the State of Tennessee and by the American Lebanese Syrian Associated Charities (ALSAC). Sharon Naron, who provided editorial review, is supported by ALSAC.
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