International consensus on mitotane treatment in pediatric patients with adrenal cortical tumors: indications, therapy, and management of adverse effects.

OBJECTIVE: Mitotane is an important cornerstone in the treatment of pediatric adrenal cortical tumors (pACC), but experience with the drug in the pediatric age group is still limited and current practice is not guided by robust evidence. Therefore, we have compiled international consensus statements from pACC experts on mitotane indications, therapy, and management of adverse effects. METHODS: A Delphi method with 3 rounds of questionnaires within the pACC expert consortium of the international network groups European Network for the Study of Adrenal Tumors pediatric working group (ENSAT-PACT) and International Consortium of pediatric adrenocortical tumors (ICPACT) was used to create 21 final consensus statements. RESULTS: We divided the statements into 4 groups: environment, indications, therapy, and adverse effects. We reached a clear consensus for mitotane treatment for advanced pACC with stages III and IV and with incomplete resection/tumor spillage. For stage II patients, mitotane is not generally indicated. The timing of initiating mitotane therapy depends on the clinical condition of the patient and the setting of the planned therapy. We recommend a starting dose of 50 mg/kg/d (1500 mg/m²/d) which can be increased up to 4000 mg/m2/d. Blood levels should range between 14 and 20 mg/L. Duration of mitotane treatment depends on the clinical risk profile and tolerability. Mitotane treatment causes adrenal insufficiency in virtually all patients requiring glucocorticoid replacement shortly after beginning. As the spectrum of adverse effects of mitotane is wide-ranging and can be life-threatening, frequent clinical and neurological examinations (every 2-4 weeks), along with evaluation and assessment of laboratory values, are required. CONCLUSIONS: The Delphi method enabled us to propose an expert consensus statement, which may guide clinicians, further adapted by local norms and the individual patient setting. In order to generate evidence, well-constructed studies should be the focus of future efforts.

Multiple TP53 p.R337H haplotypes and implications for tumor susceptibility.

The germline TP53 p.R337H mutation is reported as the most common germline TP53 variant. It exists at a remarkably high frequency in the population of southeast Brazil as founder mutation in two distinct haplotypes with the most frequent co-segregating with the p.E134∗ variant of the XAF1 tumor suppressor and an increased cancer risk. Founder mutations demonstrate linkage disequilibrium with neighboring genetic polymorphic markers that can be used to identify the founder variant in different geographic regions and diverse populations. We report here a shared haplotype among Brazilian, Portuguese, and Spanish families and the existence of three additional distinct TP53 p.R337H alleles. Mitochondrial DNA sequencing and Y-STR profiling of Brazilian carriers of the founder TP53 p.R337H allele reveal an excess of Native American haplogroups in maternal lineages and exclusively European haplogroups in paternal lineages, consistent with communities established through male European settlers with extensive intermarriage with Indigenous women. The identification of founder and independent TP53 p.R337H alleles underlines the importance for considering the haplotype as a functional unit and the additive effects of constitutive polymorphisms and associated variants in modifier genes that can influence the cancer phenotype.

Steroidogenic Factor 1, a Goldilocks Transcription Factor from Adrenocortical Organogenesis to Malignancy.

Steroidogenic factor-1 (SF-1, also termed Ad4BP; NR5A1 in the official nomenclature) is a nuclear receptor transcription factor that plays a crucial role in the regulation of adrenal and gonadal development, function and maintenance. In addition to its classical role in regulating the expression of P450 steroid hydroxylases and other steroidogenic genes, involvement in other key processes such as cell survival/proliferation and cytoskeleton dynamics have also been highlighted for SF-1. SF-1 has a restricted pattern of expression, being expressed along the hypothalamic-pituitary axis and in steroidogenic organs since the time of their establishment. Reduced SF-1 expression affects proper gonadal and adrenal organogenesis and function. On the other hand, SF-1 overexpression is found in adrenocortical carcinoma and represents a prognostic marker for patients' survival. This review is focused on the current knowledge about SF-1 and the crucial importance of its dosage for adrenal gland development and function, from its involvement in adrenal cortex formation to tumorigenesis. Overall, data converge towards SF-1 being a key player in the complex network of transcriptional regulation within the adrenal gland in a dosage-dependent manner.

Impact of Pesticides on Cancer and Congenital Malformation: A Systematic Review.

Pesticide exposure has deleterious effects on human health and development; however, no review has been conducted on human exposure to pesticides and the risk of congenital malformations and cancer in the same cohort. We systematically reviewed the evidence for this relationship following the Preferred Reporting Items for Systematic Reviews and Meta-Analysis guidelines. Four databases, namely, PubMed, Scopus, Cochrane Library, and BVS, were searched for studies deposited till July 2020 that examined the influence of pesticide exposure on congenital malformations and cancer outcomes in the same cohort. Seven studies were systematically included in this review. Among these, four were case-control studies, two were cross-sectional studies, and one was a longitudinal cohort study. The sources of contamination were food, water, or exposure during agricultural work. A link between the occurrence of cancer, congenital malformations, and exposure to pesticides was observed in most studies.

Comprehensive Characterization of the Regulatory Landscape of Adrenocortical Carcinoma: Novel Transcription Factors and Targets Associated with Prognosis.

We reconstructed a transcriptional regulatory network for adrenocortical carcinoma (ACC) using transcriptomic and clinical data from The Cancer Genome Atlas (TCGA)-ACC cohort. We investigated the association of transcriptional regulatory units (regulons) with overall survival, molecular phenotypes, and immune signatures. We annotated the ACC regulons with cancer hallmarks and assessed single sample regulon activities in the European Network for the Study of Adrenal Tumors (ENSAT) cohort. We found 369 regulons associated with overall survival and subdivided them into four clusters: RC1 and RC2, associated with good prognosis, and RC3 and RC4, associated with worse outcomes. The RC1 and RC3 regulons were highly correlated with the 'Steroid Phenotype,' while the RC2 and RC4 regulons were highly correlated with a molecular proliferation signature. We selected two regulons, NR5A1 (steroidogenic factor 1, SF-1) and CENPA (Centromeric Protein A), that were consistently associated with overall survival for further downstream analyses. The CENPA regulon was the primary regulator of MKI-67 (a marker of proliferation KI-67), while the NR5A1 regulon is a well-described transcription factor (TF) in ACC tumorigenesis. We also found that the ZBTB4 (Zinc finger and BTB domain-containing protein 4) regulon, which is negatively associated with CENPA in our transcriptional regulatory network, is also a druggable anti-tumorigenic TF. We anticipate that the ACC regulons may be used as a reference for further investigations concerning the complex molecular interactions in ACC tumors.

Prolactin as an adrenocorticotropic hormone: Prolactin signalling is a conserved key regulator of sexually dimorphic adrenal gland function in health and disease.

A large number of previous reports described an effect of the pituitary hormone prolactin (PRL) on steroid hormone production by the adrenal cortex. However, those studies remained anecdotal and were never converted into a conceptual and mechanistic framework, let alone being translated into clinical care. In the light of our recently published landmark study where we described PRL signalling as a pivotal regulator of the sexually dimorphic adrenal phenotype in mouse and of adrenal androgen production in humans, we present here the overarching hypothesis that PRL signalling increases the activity of Steroidogenic Factor-1 (SF-1/NR5A1), a transcription factor that has an essential role in adrenal gland development and function, to regulate adrenal cortex growth and hormonal production in physiological and pathological conditions. PRL can then be considered as a bona fide adrenocorticotropic hormone synergizing with ACTH in the endocrine control of adrenal cortex function.

Psychological Impact of TP53-Variant-Carrier Newborns and Counselling on Mothers: A Pediatric Surveillance Cohort.

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

Environmental Contaminants Modulate Breast Cancer Development and Outcome in TP53 p.R337H Carriers and Noncarriers.

Two major concerns associated with cancer development in Paraná state, South Brazil, are environmental pollution and the germline TP53 p.R337H variant found in 0.27−0.30% of the population. We assessed breast cancer (BC) risk in rural (C1 and C2) and industrialized (C3) subregions, previously classified by geochemistry, agricultural productivity, and population density. C2 presents lower organochloride levels in rivers and lower agricultural outputs than C1, and lower levels of chlorine anions in rivers and lower industrial activities than C3. TP53 p.R337H status was assessed in 4658 women aged >30 years from C1, C2, and C3, subsequent to a genetic screening (Group 1, longitudinal study). BC risk in this group was 4.58 times higher among TP53 p.R337H carriers. BC prevalence and risk were significantly lower in C2 compared to that in C3. Mortality rate and risk associated with BC in women aged >30 years (n = 8181 deceased women; Group 2) were also lower in C2 than those in C3 and C1. These results suggest that environmental factors modulate BC risk and outcome in carriers and noncarriers.

Newborn Screening for the Detection of the TP53 R337H Variant and Surveillance for Early Diagnosis of Pediatric Adrenocortical Tumors: Lessons Learned and Way Forward.

The incidence of pediatric adrenocortical tumors (ACT) is high in southern Brazil due to the founder TP53 R337H variant. Neonatal screening/surveillance (NSS) for this variant resulted in early ACT detection and improved outcomes. The medical records of children with ACT who did not participate in newborn screening (non-NSS) were reviewed (2012-2018). We compared known prognostic factors between the NSS and non-NSS cohorts and estimated surveillance and treatment costs. Of the 16 non-NSS children with ACT carrying the R337H variant, the disease stages I, II, III, and IV were observed in five, five, one, and five children, respectively. The tumor weight ranged from 22 to 608 g. The 11 NSS children with ACT all had disease stage I and were alive. The median tumor weight, age of diagnosis, and interval between symptoms and diagnosis were 21 g, 1.9 years, and two weeks, respectively, for the NSS cohort and 210 g, 5.2 years, and 15 weeks, respectively, for the non-NSS cohort. The estimated surveillance/screening cost per year of life saved is US$623/patient. NSS is critical for improving the outcome of pediatric ACT in this region. Hence, we strongly advocate for the inclusion of R337H in the state-mandated universal screening and surveillance.

Adrenocortical Carcinoma Steroid Profiles: In Silico Pan-Cancer Analysis of TCGA Data Uncovers Immunotherapy Targets for Potential Improved Outcomes.

Despite progress in understanding the biology of adrenocortical carcinoma (ACC), treatment options have not dramatically changed in the last three decades, nor have we learned how to avoid some of its long-term side effects. Our goal was to improve the understanding of immune pathways that may include druggable targets to enhance immune responses of patients with ACC, focusing on immune evasion and the activation of immune cells against ACC. Our strategy was aimed at improving insight regarding gene expression without steroid interference. Using approaches based on high and low steroid phenotypes (HSP and LSP, respectively), we characterized immune pathways using The Cancer Genome Atlas (TCGA) ACC cohort data. Although previous studies have suggested that patients with ACC receive minimal benefit from immunotherapy, high expression of immune modulators was noted in patients with LSP, suggesting the activation of these biomarkers may be an important adjuvant therapy target after clearance of excess glucocorticoids. In addition, patients with LSP ACC had higher immune cell infiltration than patients with HSP ACC and other cancer subtypes. Our findings can be summarized as follows (1): we confirmed and improved the definition of two immune response pathways to ACC (HSP and LSP) based on in silico transcriptome analysis (2), we demonstrated the steroid profile should be considered, otherwise analyses of ACC immune characteristics can generate confounding results (3), among the overexpressed immunotherapy targets, we demonstrated that LSP was rich in PDCD1LG2 (PD-L2) and both HSP and LSP overexpressed CD276 (B7-H3), which was associated with resistance to anti-PD1 therapy and may have accounted for the modest results of previous clinical trials, and (4) identification of patients with LSP or HSP ACC can be used to help determine whether immunotherapy should be used. In conclusion, we highlighted the differences between LSP and HSP, drawing attention to potential therapeutic targets (CD276, PDCD1, and PDCD1LG2). Treatments to reduce immune evasion, as well as the use of other natural and pharmacological immune activators, should include prior pharmacological inhibition of steroidogenesis. Attempts to combine these with tumor cell proliferation inhibitors, if they do not affect cells of the immune system, may produce interesting results.

The Common Germline TP53-R337H Mutation Is Hypomorphic and Confers Incomplete Penetrance and Late Tumor Onset in a Mouse Model.

The TP53-R337H founder mutation exists at a high frequency throughout southern Brazil and represents one of the most common germline TP53 mutations reported to date. It was identified in pediatric adrenocortical tumors in families with a low incidence of cancer. The R337H mutation has since been found in association with early-onset breast cancers and Li-Fraumeni syndrome (LFS). To study this variability in tumor susceptibility, we generated a knockin mutant p53 mouse model (R334H). Endogenous murine p53-R334H protein was naturally expressed at high levels in multiple tissues and was functionally compromised in a tissue- and stress-specific manner. Mutant p53-R334H mice developed tumors with long latency and incomplete penetrance, consistent with many human carriers being at a low but elevated risk for cancer. These findings suggest the involvement of additional cooperating genetic alterations when TP53-R337H occurs in the context of LFS, which has important implications for genetic counseling and long-term clinical follow-up. SIGNIFICANCE: A p53-R334H knockin mouse serves as an important model for studying the most common inherited germline TP53 mutation (R337H) that is associated with variable tumor susceptibility.

Second Primary Malignancy after Acute Promyelocytic Leukemia: A Population-Based Study.

Acute promyelocytic leukemia (APL), is now highly curable with treatment approaches that include all-trans retinoic acid (ATRA). The high incidence of APL in the Hispanics suggests an association with genetic variants in this population. Information on second primary malignancies (SPMs) in patients with APL is limited. The Surveillance, Epidemiology, and End Results (SEER) database was used to interrogate whether the rate of SPMs in patients with APL was associated with ethnicity and/or ATRA treatment. Between 2000 and 2016, 116 cases of SPM were diagnosed among 4019 patients with APL. The mean age at diagnosis of primary APL was 53.9 years (±15.7 years), and the mean age at diagnosis of SPMs was 59.0 years (±14.5 years). Comparisons with 3774 APL survivors who did not develop SPMs revealed that age ≥40 years at diagnosis of APL (p < 0.001) and non-Hispanic white ethnicity (p = 0.025) were associated with SPMs in APL survivors. Salivary gland, liver, and soft tissue malignancies were significantly more common in patients with primary APL than in individuals with non-APL malignancies. A risk analysis comparing patients who had APL with patients who had non-APL AML suggests that SPMs after APL is associated with ATRA treatment. Therefore, patient follow-up after APL should focus on early diagnosis of SPMs.

Frequency of the TP53 R337H variant in sporadic breast cancer and its impact on genomic instability.

The R337H is a TP53 germline pathogenic variant that has been associated with several types of cancers, including breast cancer. Our main objective was to determine the frequency of the R337H variant in sporadic breast cancer patients from Paraná state, South Brazil, its association with prognosis and its impact in genomic instability. The genotyping of 805 breast cancer tissues revealed a genotypic and allelic frequency of the R337H variant of 2.36% and 1.18%, respectively. In these R337H+ cases a lower mean age at diagnosis was observed when compared to the R337H-cases. Array-CGH analysis showed that R337H+ patients presented a higher number of copy number alterations (CNAs), compared to the R337H-. These CNAs affected genes and miRNAs that regulate critical cancer signaling pathways; a number of these genes were associated with survival after querying the KMplot database. Furthermore, homozygous (R337H+/R337H+) fibroblasts presented increased levels of copy number variants when compared to heterozygous or R337H- cells. In conclusion, the R337H variant may contribute to 2.36% of the breast cancer cases without family cancer history in Paraná. Among other mechanisms, R337H increases the level of genomic instability, as evidenced by a higher number of CNAs in the R337H+ cases compared to the R337H-.

XAF1 as a modifier of p53 function and cancer susceptibility.

Cancer risk is highly variable in carriers of the common TP53-R337H founder allele, possibly due to the influence of modifier genes. Whole-genome sequencing identified a variant in the tumor suppressor XAF1 (E134*/Glu134Ter/rs146752602) in a subset of R337H carriers. Haplotype-defining variants were verified in 203 patients with cancer, 582 relatives, and 42,438 newborns. The compound mutant haplotype was enriched in patients with cancer, conferring risk for sarcoma (P = 0.003) and subsequent malignancies (P = 0.006). Functional analyses demonstrated that wild-type XAF1 enhances transactivation of wild-type and hypomorphic TP53 variants, whereas XAF1-E134* is markedly attenuated in this activity. We propose that cosegregation of XAF1-E134* and TP53-R337H mutations leads to a more aggressive cancer phenotype than TP53-R337H alone, with implications for genetic counseling and clinical management of hypomorphic TP53 mutant carriers.

Cancer-testis Antigen FATE1 Expression in Adrenocortical Tumors Is Associated with A Pervasive Autoimmune Response and Is A Marker of Malignancy in Adult, but Not Children, ACC.

The SF-1 transcription factor target gene FATE1 encodes a cancer-testis antigen that has an important role in regulating apoptosis and response to chemotherapy in adrenocortical carcinoma (ACC) cells. Autoantibodies directed against FATE1 were previously detected in patients with hepatocellular carcinoma. In this study, we investigated the prevalence of circulating anti-FATE1 antibodies in pediatric and adult patients with adrenocortical tumors using three different methods (immunofluorescence, ELISA and Western blot). Our results show that a pervasive anti-FATE1 immune response is present in those patients. Furthermore, FATE1 expression is a robust prognostic indicator in adult patients with ACC and is associated with increased steroidogenic and decreased immune response gene expression. These data can open perspectives for novel strategies in ACC immunotherapy.

A common polymorphism in the retinoic acid pathway modifies adrenocortical carcinoma age-dependent incidence.

BACKGROUND: Genome-wide association studies (GWASs) have enriched the fields of genomics and drug development. Adrenocortical carcinoma (ACC) is a rare cancer with a bimodal age distribution and inadequate treatment options. Paediatric ACC is frequently associated with TP53 mutations, with particularly high incidence in Southern Brazil due to the TP53 p.R337H (R337H) germline mutation. The heterogeneous risk among carriers suggests other genetic modifiers could exist. METHODS: We analysed clinical, genotype and gene expression data derived from paediatric ACC, R337H carriers, and adult ACC patients. We restricted our analyses to single nucleotide polymorphisms (SNPs) previously identified in GWASs to associate with disease or human traits. RESULTS: A SNP, rs971074, in the alcohol dehydrogenase 7 gene significantly and reproducibly associated with allelic differences in ACC age-of-onset in both cohorts. Patients homozygous for the minor allele were diagnosed up to 16 years earlier. This SNP resides in a gene involved in the retinoic acid (RA) pathway and patients with differing levels of RA pathway gene expression in their tumours associate with differential ACC progression. CONCLUSIONS: These results identify a novel genetic component to ACC development that resides in the retinoic acid pathway, thereby informing strategies to develop management, preventive and therapeutic treatments for ACC.

Germline Variants in Phosphodiesterase Genes and Genetic Predisposition to Pediatric Adrenocortical Tumors.

Phosphodiesterases (PDEs) form a superfamily of enzymes that catalyze the hydrolysis of cyclic nucleotides adenosine 3'5'-cyclic monophosphate (cAMP) and guanosine 3'5'-cyclic monophosphate (cGMP) to their inactive 5' monophosphates. cAMP plays a critical role as a second messenger in endocrine tissues, and activation of cAMP signaling has been reported in endocrine tumors. Germline variants in PDEs have been associated with benign cortisol-secreting adrenocortical adenomas and testicular germ cell cancer but not adrenocortical carcinoma. We performed whole genome sequencing (WGS) and whole exome sequencing (WES) of paired blood and tumor samples from 37 pediatric adrenocortical tumors (ACTs). Germline inactivating variants in PDEs were observed in 9 of 37 (24%) patients. Tumor DNA analysis revealed loss of heterozygosity, with maintenance of the mutated allele in all cases. Our results suggest that germline variants in PDEs and other regulators of the cAMP-signaling pathway may contribute to pediatric adrenocortical tumorigenesis, perhaps by cooperating with germline hypomorphic mutant TP53 alleles and uniparental disomy of chromosome 11p15 (Beckwith-Wiedemann syndrome).

Penetrance of the TP53 R337H Mutation and Pediatric Adrenocortical Carcinoma Incidence Associated with Environmental Influences in a 12-Year Observational Cohort in Southern Brazil.

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

The Prognostic Role of CD8+ T Lymphocytes in Childhood Adrenocortical Carcinomas Compared to Ki-67, PD-1, PD-L1, and the Weiss Score.

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

The Impact of Flt3 Gene Mutations in Acute Promyelocytic Leukemia: A Meta-Analysis.

The association of FLT3 mutations with white blood cell (WBC) counts at diagnosis and early death was studied in patients with acute promyelocytic leukemia (APL). Publications indexed in databases of biomedical literature were analyzed. Potential publication bias was evaluated by analyzing the standard error in funnel plots using the estimated relative risk (RR). Mixed-effect models were used to obtain the consolidated RR. All analyses were conducted using the R statistical software package. We used 24 publications in the final meta-analysis. Of 1005 males and 1376 females included in these 24 publications, 645 had FLT3-ITD (internal tandem duplication) mutations. Information on FLT3-D835 mutations was available in 10 publications for 175 patients. Concurrent occurrence of the two mutations was rare. WBC count at diagnosis was ≥10 × 109/L in 351 patients. For patients with the FLT3-ITD mutation, RR was 0.59 for overall survival (OS) and 1.62 for death during induction. For those with FLT3-D835 mutations, the RR was 0.50 for OS and 1.77 for death during induction. RR for WBC count ≥10 × 109/L was 3.29 and 1.48 for patients with FLT3-ITD and FLT3-D835, respectively. APL patients with FLT3-ITD or FLT3-D835 are more likely to present with elevated WBC counts and poorer prognosis than those without these mutations.