Expression of the miR-9-5p, miR-125b-5p and its target gene NFKB1 and TRAF6 in childhood-onset systemic lupus erythematosus (cSLE).

Childhood- onset systemic lupus erythematosus (cSLE) is a multisystem inflammatory disease that can lead to severe clinical conditions resulting in early comorbidities. Several genetic, environmental, and immunological factors are known to influence the onset of the disease. MiRNAs have been already considered as potential actors involved in the development and activity of the SLE. Thus, understanding the behavior of these regulators can contribute to clarify the inflammatory process affecting SLE patients. Among miRNAs, miR-125b-5p and miR-9-5p targeting NFKB1 and TRAF6 genes can be involved in the etio-pathogenesis of the disease by modulating inflammation. In this study we evaluated miR-9-5p and miR-125b-5p expression and its target genes NFKB1 and TRAF6 in peripheral blood samples (PBMC) from the 35 cSLE patients and 35 healthy controls. MiRNAs and gene target expression have been evaluated by using RT-PCR with specific TaqMan® probes. Both miR-9-5p [Fold Change (FC) = -2.21; p = 0.002] and miR-125b-5p (FC= -3.30; p < 0.0001) and NFKB1 (FC = -1.84; p < 0.001) were downregulated in cSLE patients, while TRAF6 was upregulated (FC = 1.80; p = 0.006) in cSLE patients when compared to controls. A significant correlation was found between miR-125b-5p and its target gene NFKB1 [Spearman (r) = 0.47; p = 0.023]. Our results showed miR-125b-5p and miR-9-5p differential expression in cSLE patients, possibly contributing to better understanding the role of these regulators in cSLE development and disease pathogenesis.

Identification of ITPR1 as a Hub Gene of Group 3 Medulloblastoma and Coregulated Genes with Potential Prognostic Values.

The Group 3 Medulloblastoma (Grp3-MB) is an aggressive molecular subtype with a high incidence of metastasis and deaths. In this study, were used an RNA sequencing data (RNA-Seq) from a Brazilian cohort of MBs to identify hub genes associated with the metastatic risk. Data validation were performed by using multiple large datasets from MBs (GSE85217, GSE37418, and EGAS00001001953). DESeq2 package in R software was used to identify the differentially expressed genes (DEGs) in our RNA-Seq data. The DEGs data were accessed to construct the modules/graphs of co-expression and to identify hub genes through Cytoscape platform. The coregulated genes were enriched by the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway, and the protein-protein interaction (PPI) network was visualized by Cytoscape. The Kaplan-Meier plotter and ROC curves were used to validate the diagnostic and prognostic values of specific biomarkers identified through this model. We identified that inositol 1,4,5-trisphosphate receptor type 1 (ITPR1) as a downregulated hub gene, with a high diagnostic accuracy to Grp3-MBs and associated with tumor metastasis. In addition, we identified genes significantly correlated with ITPR1 that were associated with metastasis in Grp3-MB (ATP1A2, MTTL7A, and RGL1) and worst overall survival in MBs (ANTXR1 and RGL1). Our findings suggest that the ITPR1 hub gene is potentially involved in the metastatic process for Grp3-MB. Our data also provide evidence of targets that may serve as prognostic predictors and/or regulators for the metastatic process that maybe explored for further research of individualized therapy to Grp3-MBs.

Frequency of the TP53 p.R337H mutation in a Brazilian cohort of pediatric patients with solid tumors.

TP53 p.R337H germline mutation is highly prevalent in the Southern region of Brazil. We sought to investigate TP53 p.R337H mutation in pediatric tumor samples from a population settled in a geographic area of high prevalence for this variant. Mutation assessment and genetic counseling for carriers/relatives were provided. 6/57 tumor samples were heterozygous for TP53 p.R337H. As expected, a high frequency was observed within adrenocortical tumors (3/3) and choroid plexus carcinomas (2/2). Interestingly, the TP53 R337H mutation was found in one case of pediatric rhabdomyosarcoma with Li-Fraumeni pedigree. Our finding expands the spectrum of childhood cancer associated with this germline mutation.

A new complex rearrangement in infant ALL: t(X;11;17)(p11.2;q23;q12).

We present a case of an infant who developed pro-B acute lymphoblastic leukemia with a rare and complex MLL-translocation. Cytogenetic analysis of bone marrow cells at diagnosis showed a 46,XY,t(X;11)(p11.2;q23)[13]/46,XY[7] karyotype. Fluorescence in situ hybridization analysis using a break apart specific probes showed a split in the MLL gene. Long distance inverse-PCR and next generation sequencing analysis depicted a complex rearrangement t(X;11;17)(p11.2;q23;q12) involving MLL, MLLT6 and the genomic region Xp11.23, 41 bases upstream of the WDR45 gene. WDR45 encodes a beta-propeller protein essential for autophagocytosis. MLL rearrangements with involvement of Xp have not been previously described.

Fcγ receptor gene polymorphisms in childhood immune thrombocytopenic purpura.

Immune thrombocytopenic purpura (ITP) is a common hematological disorder in the childhood, and it is one of the most common forms of autoimmune disease in pediatric patients. The ITP basis is a primary dysfunction of the immune system. This study aimed to analyze the genetic polymorphisms of the Fcγ receptors IIA and IIIA. The genetic polymorphisms of the Fc receptors γIIA (131H/R) and γRIIIA (158V/F) were analyzed by polymerase chain reaction-restriction fragment length polymorphism technique. Odds ratio and 95% confidence interval were calculated by χ(2) test. Homozygous polymorphic genotype for the FcγRIIIA was significantly more frequent among patients compared with controls (odds ratio = 0.27; 95% confidence interval, 0.09-0.80; P = 0.03). There was no statistical difference between the ITP group and the controls in the analysis of combinations of alleles of the high-affinity Fc receptor, but the ITP individuals with this combination had a lower duration of disease (P = 0.01). Genetic polymorphisms in immune system genes can be important for ITP pathogenesis and disease outcome.

Protein expression of matrix metalloproteinase (MMP-1, -2, -3, -9 and -14) in Ewing family tumors and medulloblastomas of pediatric patients.

The matrix metalloproteinases (MMP) are endopeptidases performing proteolytic functions in the extracellular matrix and their overexpression has been suggested to be a characteristic of malignant tumors. Molecular changes such as the presence of chimeric protein Ewing's sarcoma protein-friend leukemia virus integration 1 (EWS-FLI1) in the Ewing family of tumors (EFT) and the oncogenes C-ERBB-2, N-MYC, C-MYC in medulloblastoma (MB) promote the overexpression of MMP. In the present study, protein expression of MMP-1, -2, -3, -9 and -14 was qualitatively evaluated in 17 EFT and MB samples of children and adolescent by western blotting and optical densitometry, and the level of gene expression of some MMPs was determined by real-time quantitative polymerase chain reaction. Five MB samples (45.4%) presented expression of the five MMPs and six samples (54.6%) presented expression of at least one of them. Four EFT samples (66.6%) presented expression of MMP-2, -9 and -14, and two samples (33.4%) presented expression of at least one of these MMPs, whereas the presence of MMP-1 and -3 was not observed. Gene analysis showed that MMP-2 had a high expression in MB, while the expression of MMP-9 and MMP-14 was higher in EFT. It has been established that the expression of the MMPs might be related to a complex pathway of gene regulation.

Vascular endothelial growth factor (VEGF) and endothelial nitric oxide synthase (NOS3) polymorphisms are associated with high relapse risk in childhood acute lymphoblastic leukemia (ALL).

BACKGROUND: Angiogenesis has been shown as an important process in hematological malignancies. It consists in endothelial proliferation, migration, and tube formation following pro-angiogenic factors releasing, specially the vascular endothelial growth factor (VEGF), which angiogenic effect seems to be dependent on nitric oxide (NO). We examined the association among functional polymorphisms in these two angiogenesis related genes: VEGF (-2578C>A, -1154G>A, and -634G>C) and NOS3 (-786T>C, intron 4 b>a, and Glu298Asp) with prognosis of childhood acute lymphoblastic leukemia (ALL). METHODS: The genotypes were determined and haplotypes estimated in 105 ALL patients that were divided in 2 groups: high risk (HR) and low risk of relapse (LR) patients. In addition, event-free survival curves according to genotypes were assessed. RESULTS: The group HR compared to the LR showed a higher frequency of the alleles -2578C and -634C and the haplotype CGC for VEGF (0.72 vs. 0.51, p<0.008; 0.47 vs. 0.26, p<0.008; and 42.1 vs. 14.5, p<0.006; respectively) and a lower frequency of the haplotype CbGlu (0.4 vs. 8.8,p<0.006), for NOS3. CONCLUSION: Polymorphisms of VEGF and NOS3 genes are associated with high risk of relapse, therefore may have a prognostic impact in childhood ALL.

mRNA expression of matrix metalloproteinases (MMPs) 2 and 9 and tissue inhibitor of matrix metalloproteinases (TIMPs) 1 and 2 in childhood acute lymphoblastic leukemia: potential role of TIMP1 as an adverse prognostic factor.

This study evaluates the mRNA expression profile of genes TIMP1, TIMP2, MMP2 and MMP9 in diagnostic bone marrow samples from 134 consecutive ALL children by real-time quantitative PCR. A significant association was observed between higher expression levels of MMP9 and low risk group and absence of extramedullary infiltration and higher expression levels of TIMP2 and MMP2 with T-ALL. TIMP1 gene expression values higher than the median were associated with a significantly lower 5-year event free-survival in univariable (P=0.04) and multivariable analysis (P=0.01). Our data address new information in the complex interaction of the migration/adhesion genes and childhood ALL.

Role of the CYP2D6, EPHX1, MPO, and NQO1 genes in the susceptibility to acute lymphoblastic leukemia in Brazilian children.

Polymorphic variations of several genes associated with dietary effects and exposure to environmental carcinogens may influence susceptibility to leukemia development. The objective of the present study was to evaluate the effect of the polymorphisms of debrisoquine hydroxylase (CYP2D6), epoxide hydrolase (EPHX1), myeloperoxidase (MPO), and quinone-oxoreductase (NQO1), which have been implicated in xenobiotic metabolism, on the risk of childhood acute lymphoblastic leukemia (ALL). We evaluated the frequency of polymorphisms in the CYP2D6 (*3 and *4), EPHX1 (*2 and *3), MPO (*2), and NQO1 (*2) genes in 206 patients with childhood ALL and in 364 healthy individuals matched for age and gender from a Brazilian population separated by ethnicity (European ancestry and African ancestry), using the PCR-RFLP method. The CYP2D6 polymorphism variants were associated with an increased risk of ALL. The EPHX1, NQO1, and MPO variant genotypes were significantly associated with a reduced risk of childhood ALL. A significantly stronger protective effect is observed when the EPHX1, NQO1, and MPO variant genotypes are combined suggesting that, CYP2D6 polymorphisms may play a role in the susceptibility to pediatric ALL, whereas the EPHX1, NQO1, and MPO polymorphisms might have a protective function against leukemogenesis.

Acute lymphoblastic leukemia following severe congenital neutropenia or de novo ALL?

Acute lymphoblastic leukemia (ALL) presenting with neutropenia alone is very rare. We describe a newborn with an early life-threatening infection, severe neutropenia and bone marrow findings compatible with severe congenital neutropenia (SCN). She was treated with granulocyte colony-stimulating factor (G-CSF) with complete neutrophil recovery. Three months later she developed a pro-B ALL. We identified a rare loss of 5'-MLL present at the diagnosis of SCN and ALL by FISH analysis using two different MLL (11q23) probes. Molecular analyses for SCN causing mutations (ELA-2, HAX-1 and G6PC3) and for somatic mutations of the CSF3R gene were negative. The early presence of 5'-MLL loss in bone marrow samples may favor the diagnosis of de novo ALL. Nevertheless, the genetic background for SCN is heterogeneous and a non-described mutation for SCN followed by a secondary ALL cannot be excluded. Further genetic investigation may be useful to gain insight into this rare condition in children.

Polymorphisms in genes encoding drugs and xenobiotic metabolizing enzymes in a Brazilian population.

Polymorphic variations of several genes associated with drugs and xenobiotic metabolism have been linked to the factors that predispose to the carcinogenesis process. As considerable interindividual and interethnic variation in metabolizing enzyme activity has been associated with polymorphic alleles, we evaluated the frequency of the polymorphisms of CYP2D6, EPHX1 and NQO1 genes in 361 Brazilian individuals separated by ethnicity (European and African ancestry), using the polymerase chain reaction-restriction fragment length (PCR-RFLP) method. The allele frequencies of the variants *3 and *4 for the gene CYP2D6 were 0.04 and 0.14 for white subjects and 0.03 and 0.10 for black individuals, respectively. For the both variants of the gene EPHX1, we found higher allele frequencies among white individuals compared with mulatto subjects (0.62 vs 0.54 and 0.18 vs 0.14, respectively); however, these differences were not statistically significant (p = 0.39 and 0.56, respectively). For the NQO1 gene we observed a higher frequency of the homozygous genotype among black individuals (7.9%) compared with white subjects (6.3%) (p = 0.003). The genotype frequencies were within the Hardy-Weinberg equilibrium. We concluded that the allele frequencies of CYP2D6, EPHX1 and NQO1 gene polymorphisms in this Brazilian population showed ethnic variability when compared with those observed in other populations.

Polymorphisms of xenobiotic metabolizing enzymes and DNA repair genes and outcome in childhood acute lymphoblastic leukemia.

The interindividual variation in the activity of xenobiotic metabolizing enzymes and DNA repair genes could modify an individual's risk of recurrent malignancy and response to therapy. We investigated whether ALL outcome was related to polymorphisms in genes CYP2D6, MPO, EPHX1, NQO1, TS, XPD and XRCC1 in 95 consecutive ALL children by PCR or PCR-FRLP techniques. Polymorphisms in genes NQO1 and TS were associated with a significantly slow response to induction chemotherapy and NQO1 was also associated with a lower five-year event-free survival. This study suggests that polymorphisms of NQO1 and TS could be important for patient response to induction therapy and for treatment outcome.

MLL leukemia-associated rearrangements in peripheral blood lymphocytes from healthy individuals

Chromosomal translocations are characteristic of hematopoietic neoplasias and can lead to unregulated oncogene expression or the fusion of genes to yield novel functions. In recent years, different lymphoma/leukemia-associated rearrangements have been detected in healthy individuals. In this study, we used inverse PCR to screen peripheral lymphocytes from 100 healthy individuals for the presence of MLL (Mixed Lineage Leukemia) translocations. Forty-nine percent of the probands showed MLL rearrangements. Sequence analysis showed that these rearrangements were specific for MLL translocations that corresponded to t(4;11)(q21;q23) (66 percent) and t(9;11) (20 percent). However, RT-PCR failed to detect any expression of t(4;11)(q21;q23) in our population. We suggest that 11q23 rearrangements in peripheral lymphocytes from normal individuals may result from exposure to endogenous or exogenous DNA-damaging agents. In practical terms, the high susceptibility of the MLL gene to chemically-induced damage suggests that monitoring the aberrations associated with this gene in peripheral lymphocytes may be a sensitive assay for assessing genomic instability in individuals exposed to genotoxic stress.

Fungal infection by Paracoccidioides brasiliensis mimicking bone tumor.

Paracoccidioides brasiliensis infection causes a systemic mycosis originally described in Latin America but with current reports of worldwide distribution. The clinical presentation of paracoccidiodomycosis as an isolated long-bone lesion in children is quite unusual. This article describes a 10-year-old male with a lytic femoral bone lesion caused by P. brasiliensis infection that was first suspected of being of neoplasic etiology. The text also emphasizes the importance of including endemic fungal infections in the differential diagnosis of bone lesions.

Quantitative PCR analysis of the expression profile of genes related to multiple drug resistance in tumors of the central nervous system.

OBJECTIVES: To evaluate and compare the profile of expression of genes related to drug resistance in brain tumors and to analyze the impact of the increased expression of these genes on overall survival. METHODS: Eighty microdissected brain tumor samples from 79 patients were analyzed by RQ-PCR for the genes MDR1, MRP1, MRP3, LRP and BCRP. Protein expression was assessed by immunohistochemistry for MRP1 and LRP genes. Pediatric cases (0 to 20 years): 46 (17F:29M, median age 7.3 +/- 5.9 years); adult tumors: 33 (17F:16M, median age 46.6 +/- 14.5 years). Histological diagnoses: 21 astrocytomas I and II, 28 astrocytomas III and glioblastomas, 17 medulloblastomas, 8 ependymomas, and 6 oligodendrogliomas. RESULTS: glial tumors expressed higher MDR1 (P = 0.003) and BCRP (P = 0.03) levels than embryonic tumors. Low-grade astrocytomas expressed high MDR1 (P = 0.001), MRP3 (P = 0.01) and LRP (P = 0.02) levels. The MRP1 gene was preferentially expressed by medulloblastomas (P = 0.04) and ependymomas (P = 0.04); ependymomas also presented an increase of LRP (P = 0.02). The mRNA levels of MRP1 and LRP genes were associated to protein expression. The profile of gene expression of primary pilocytic astrocytomas of the hypothalamus and of the other locations was similar. Increased expression of resistance genes, separately or jointly, was not correlated with shorter overall survival in patients with medulloblastomas/PNET and malignant gliomas. CONCLUSIONS: Drug resistance genes do not explain the higher sensitivity of gliomas of the hypothalamus/pituitary/optic pathways to chemotherapy. The increased expression of resistance genes had no impact on the overall survival of patients with medulloblastomas/PNET and high grade gliomas. High MDR1, MRP3 and LRP levels may be implicated in the primary resistance of pilocytic astrocytomas to chemotherapy.

Multiple drug resistance protein (MDR-1), multidrug resistance-related protein (MRP) and lung resistance protein (LRP) gene expression in childhood acute lymphoblastic leukemia.

CONTEXT: Despite the advances in the cure rate for acute lymphoblastic leukemia, approximately 25% of affected children suffer relapses. Expression of genes for the multiple drug resistance protein (MDR-1), multidrug resistance-related protein (MRP), and lung resistance protein (LRP) may confer the phenotype of resistance to the treatment of neoplasias. OBJECTIVE: To analyze the expression of the MDR-1, MRP and LRP genes in children with a diagnosis of acute lymphoblastic leukemia via the semiquantitative reverse transcription polymerase chain reaction (RT-PCR), and to determine the correlation between expression and event-free survival and clinical and laboratory variables. DESIGN: A retrospective clinical study. SETTING: Laboratory of Pediatric Oncology, Department of Pediatrics, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Brazil. METHODS: Bone marrow aspirates from 30 children with a diagnosis of acute lymphoblastic leukemia were assessed for the expression of messenger RNA for the MDR-1, MRP and LRP genes by semi-quantitative RT-PCR. RESULTS: In the three groups studied, only the increased expression of LRP was related to worsened event-free survival (p = 0.005). The presence of the common acute lymphoblastic leukemia antigen (CALLA) was correlated with increased LRP expression (p = 0.009) and increased risk of relapse or death (p = 0.05). The relative risk of relapse or death was six times higher among children with high LRP expression upon diagnosis (p = 0.05), as confirmed by multivariate analysis of the three genes studied (p = 0.035). DISCUSSION: Cell resistance to drugs is a determinant of the response to chemotherapy and its detection via RT-PCR may be of clinical importance. CONCLUSIONS: Evaluation of the expression of genes for resistance to antineoplastic drugs in childhood acute lymphoblastic leukemia upon diagnosis, and particularly the expression of the LRP gene, may be of clinical relevance, and should be the object of prospective studies.

Multiple drug resistance protein (MDR-1), multidrug resistance-related protein (MRP) and lung resistance protein (LRP) gene expression in childhood acute lymphoblastic leukemia

CONTEXTO: Apesar dos avanços nos índices de cura da leucemia linfoblástica aguda (LLA) aproximadamente 25% das crianças sofrem recaídas da doença. A expressão dos genes de resistência múltipla a drogas (MDR-1), genes relacionados à proteína de resistência múltipla a drogas (MRP) e genes da proteína de resistência pulmonar (LRP) podem conferir o fenótipo de resistência ao tratamento das neoplasias. OBJETIVO: Analisar a expressão dos genes de resistência MDR-1, MRP e LRP em crianças diagnosticadas com LLA por meio da técnica da reação em cadeia da polimerase da transcriptase reversa (RT-PCR) semiquantitativa, associando estas expressões à sobrevida livre de eventos (SLE) e a variáveis clínico-laboratoriais. TIPO DE ESTUDO: Estudo clínico retrospectivo. LOCAL: Laboratório de Oncologia Pediátrica do Departamento de Puericultura e Pediatria da Faculdade de Medicina de Ribeirão Preto - Universidade de São Paulo, Brasil. MÉTODOS: Amostras de medula óssea de 30 crianças com o diagnóstico de leucemia linfoblástica aguda foram avaliadas quanto à expressão do RNA-mensageiro para os genes MDR-1, MRP e LRP, pela reação em cadeia da RT-PCR semiquantitativa. RESULTADOS: Dos três genes estudados, somente a expressão aumentada de LRP esteve relacionada a uma pior SLE (p = 0.005). A presença do antígeno para leucemia linfoblástica aguda comum (CALLA) se correlacionou à expressão aumentada de LRP (p = 0.009) e a risco aumentado de ocorrência de recaída ou óbito (p = 0.05). O risco relativo de ocorrência de recaída ou óbito é seis vezes maior em crianças com alta expressão de LRP ao diagnóstico (p = 0.05), o que se confirma na análise multivariada dos três genes estudados (p = 0.035). DISCUSSAO: A resistência celular a drogas é um determinante de resposta ao tratamento oncológico e sua avaliação por RT-PCR pode ser de importância. CONCLUSÕES: A avaliação da expressão dos genes de resistência a drogas antineoplásicas na leucemia linfoblástica aguda da criança ao diagnóstico, particularmente do gene LRP, pode ser de relevância clínica e deve ser objeto de estudos prospectivos.

PCR detection of clonal IgH and TCR gene rearrangements at the end of induction as a non-remission criterion in children with ALL: comparison with standard morphologic analysis and risk group classification.

BACKGROUND: The initial response to induction therapy is currently considered one of the most important prognostic factors in acute lymphoblastic leukemias (ALL). A series of methods for the detection of submicroscopic levels of residual disease in patients with ALL mainly based on PCR and immunophenotyping has been developed, demonstrating that the presence of high levels of residual disease at the end of induction therapy is an important, independent prognostic factor. We determined the usefulness of PCR detection of minimal residual disease using consensus primers as a non-remission criterion. PROCEDURE: Bone marrow samples obtained from 49 children with ALL were analyzed at diagnosis and at the end of induction therapy for the detection of clonal IgH, TCRdelta, and TCRgamma rearrangements by PCR. The results were compared with those obtained by standard morphologic analysis and risk group classification. RESULTS: Patients who had clonality detected at the end of induction showed a significantly higher recurrence rate and lower event-free survival than those without detected clonality (24.9% vs. 89.7%) (P < 0.0001). Multivariate analysis revealed that detection of clonality at the end of induction was the most important, independent prognostic factor when associated with age, number of white blood cells, and immunophenotyping. CONCLUSIONS: PCR detection of clonality using consensus primers is a relatively simple technique that is able to identify patients with a high chance of recurrence, and shows a higher sensitivity and a better prognostic value than standard morphologic analysis and risk group classification, defining a new remission criterion. However, further multicentric prospective studies using this technique employing a larger number of cases are necessary to confirm these findings.

Carcinogênese cutânea no sudeste do Brasil: análise do gene TP53 em tumores cutâneos não melanocíticos / Skin carcinogenesis in southeastern Brazil: analysis of TP53 gene in nonmelanoma skin tumors

Mutações no gene TP53 são comuns em muitos tipos de tumores humanos e são associados à imunorreatividade positiva para a proteína p53. Embora tal positividade seja utilizada como um marcador da mutação, evidências recentes sugerem que essa suposição nem sempre é verdadeira. O objetivo era investigar a relação entre a presença de mutação do gene TP53 e a imunocoloração da proteína p53 em tumores da pele. Foi analisada a expressão da proteína p53, com o anticorpo monoclonal Pab 1801, em 92 lesões cutâneas em indíviduos procedentes da Região Sudeste do Brasil, área com alto nível de exposição solar. Além disso, mutações no gene TP53 foram investigadas em 32 amostras imunopositivas para a proteína p53 empregando as técnicas de polimorfismo de conformação do filamento único (SSCP) e se seqüenciamento de DNA. a imunocoloração nuclear foi detectada em 5/30 (16,6 por cento) casos de ceratose solar, em 11/31 (35,5 por cento) carcinomas espinocelulares e em 16/31 (51,6 por cento) carcinomas basocelulares, todos provenientes de áreas do corpo expostas ao sol. Enmtre 32 amostras imunopositivas para a p53, apneas 3 carcinomas basocelulares revelaram alterações no gene, em códons que constituem hotspots para mutações em câncer. Os resultados mostraram baixa correlação entre imunorreatividade positiva e mutação, o que pode ser atribuído a alterações em outros segmentos do gene ou refletir a sensibilidade do sistema de detecção utilizado no estudo