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Heat shock proteins (HSPs) attract the attention of scientists and clinicians due to their potential role as diagnostic and prognostic factors in a variety of cancers. HSP90 is one of the most important and well-known family members, necessary for maintaining intracellular homeostasis. In the extracellular space, it is responsible for the transmission of alarm signals to the immune system. Numerous reports have indicated that the level of intra - and extracellular HSP90 can correlate either with a poorer prognosis or with a better outcome, depending on the type of cancer. Still, little is known why the level of this chaperone is increased in some tumors and decreased in others, reflecting dual role of protein in cell death processes. Currently, there is no database reporting levels of serum HSP90 in children with acute lymphoblastic leukemia (ALL). As such, using enzyme-linked immunosorbent assay (ELISA) method, we aimed to determine this parameter in a group of 21 patients with newly diagnosed ALL. We found decreased protein serum levels in patients at disease presentation and after induction block of chemotherapy in comparison to healthy controls. Furthermore, we observed a negative correlation between HSP90 serum levels and one of the earliest prognostic factors of the treatment response - peripheral blood lymphoblasts on the 8th day of treatment. Our results indicate that HSP90 serum may play an important role in leukemogenesis and could be used as a marker to predict treatment failure in children with ALL.

IntroductionEpidemiological analysis of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infections in pediatric hematology and oncology (PHO) and hematopoietic cell transplant (HCT) centers in a Polish nationwide study, as well as analysis of the preventive strategies in these centers.MethodsAll of the 18 PHO/HCT centers participated in eight surveys and questionnaires conducted over the first 5 months of the SARS-CoV-2/coronavirus disease 2019 (COVID-19) pandemic in Poland. Epidemiological data were collected at eight regular time points, and the strategy of preventive management was done once after 4 months of the pandemic.ResultsDuring this analyzed period, eight patients were positive for SARS-CoV-2. The estimated incidence of SARS-CoV-2 positivity in Polish PHO/HCT centers was 0.5%. After exclusion of HCT patients (with one patient being infected), the estimated incidence of SARSCoV-2 positivity was between 0.5 and 0.6%. In all but one case, the course of COVID-19 was asymptomatic or mild, and it was moderate in one case. None of them developed SARS or respiratory insufficiency, none of them required treatment in the intensive care unit (ICU), and no patient died due to SARS-CoV-2 infection. As of July 1, parents staying in the hospital together with their children were regularly tested for the virus in 13 centers. Asymptomatic healthcare personnel were regularly tested for the virus in seven centers.ConclusionsThe estimated incidence of SARS-CoV-2 infection among PHO/HCT patients is lower than in Western Europe; however, these patients cannot be regarded as a low-risk group. The low COVID-19 incidence should be interpreted as a result of strictly and continuously implemented detailed preventive measures in the PHO/HCT wards and in hospitals.

This study presents current recommendations of the Polish Association of Pediatric Surgeons (PTChD) regarding diagnostics and treatment of ovarian lesions in girls. They are based on many years of the authors' clinical experience as well as a review of international literature and include practical clinical guidelines. The recommendations were formulated in cooperation with the Polish Association of Pediatric Oncology and Hematology (PTOHD), Polish Pediatric and Adolescent Gynecology Section of the Polish Society of Gynecologists and Obstetricians (PTG) and Polish Pediatric Section of the Polish Society of Radiology (PLTR). Only better understanding of prepubertal ovarian biology and natural history of its pathology may help to introduce efficient and safe diagnostic and therapeutic strategies for girls. The prepared document has been supplemented with treatment algorithms.

Although isolated central nervous system (CNS) relapses are rare, they may become a serious clinical problem in intensively treated patients with high-risk neuroblastoma (NBL). The aim of the study is presentation, assessment of incidence and the clinical course of isolated CNS relapses. Retrospective analysis involved 848 NBL patients treated from 2001 to 2019 at 8 centers of the Polish Pediatric Solid Tumors Study Group (PPSTSG). Group characteristics at diagnosis, treatment and patterns of relapse were analyzed. Observation was completed in December 2020. We analyzed 286 high risk patients, including 16 infants. Isolated CNS relapse, defined as presence of a tumor in brain parenchyma or leptomeningeal involvement, was found in 13 patients (4.5%, 8.4% of all relapses), all of them were stage 4 at diagnosis. Isolated CNS relapses seem to be more common in young patients with stage 4 MYCN amplified NBL, and in this group they may occur early during the first line therapy. The only or the first one symptom may be bleeding into CNS, especially in younger children, even without clear relapse picture on imaging, or the relapse may be clinically asymptomatic and found on routine screening. Although incidence of isolated CNS relapses is not statistically significantly higher in patients after immunotherapy, their occurrence should be carefully monitored, especially in intensively treated infants, with potential disruption of brain-blood barrier.

The wide scope of neuropathology is also connected to different systemic pathology findings. Neuroectodermal-type female genital tract lesions are not frequent. This article presents a short review of such entities in gynecological pathology and reports on two rare cases. The first described lesion is an endometrial glial polyp in a young woman. The second is a mature cystic teratoma accompanied by a peritoneal gliomatosis in an adolescent girl. Both presented entities posed diagnostic difficulties from the clinical and pathological perspective. They demanded careful sampling, immunophenotyping, as well as neuropathological consultation.

Objective: The study was to assess whether tumour expressions of hypoxia-inducible factor (HIF)-1α, glucose transporter (GLUT)-1, carbonic anhydrase (CA) IX and vascular endothelial growth ...

The tumor microenvironment (TME) consists of numerous biologically relevant elements. One of the most important components of the TME is the extracellular matrix (ECM). The compounds of the ECM create a network that provides structural and biochemical support to surrounding cells. The most important substances involved in the regulation of the ECM degradation process are matrix metalloproteinases (MMPs) and their endogenous inhibitors (tissue inhibitors of metalloproteinases, TIMPs). The disruption of the physiological balance between MMP activation and deactivation could lead to progression of various diseases such as cardiovascular disease, cancer, fibrosis arthritis, chronic tissue ulcers, pathologies of the nervous system (such as stroke and Alzheimer’s disease), periodontitis, and atheroma. MMP-TIMP imbalance results in matrix proteolysis associated with various pathological processes such as tumor invasion. The present review discusses the involvement of two MMPs, MMP-2 and MMP-7, in cancer pathogenesis. These two MMPs have been proven in several studies, conducted mostly on adults, to make an important contribution to cancer development and progression. In the current review, several studies that indicate the importance of MMP-TIMP balance determination for the pediatric population are also highlighted. The authors of this review believe that carrying out biochemical and clinical studies focused on metalloproteinases and their inhibitors in tumors in children will be of great relevance for future patient diagnosis, determination of a prognosis, and monitoring of therapy.

Background Cardiovascular diseases (CVD) are one of long-term side effects of the childhood cancers treatment. Salusin β is an indicator of developing atherosclerosis. Aim To assess the prevalence of established risk factors for CVD and the assessment of new indicator for CVD risk - salusin β in long-term acute lymphoblastic leukemia (ALLs) and Wilms tumour (WTs) survivors. Methods 37 ALLs and 11 WTs at least 5 years after the end of oncological treatment undergone physical examination and laboratory tests after an overnight fast. Laboratory tests included lipid profile, serum level of glucose, renal parameters and salusin β. Results The both groups didn’t vary in age, time from the end of the treatment, number of obese persons, BP, lipid profile, serum creatinine and glucose level. ALLs had higher weight and greater waist circumference. Serum cystatin was higher and cystatin-based eGFR lower among WTs. Salusin β was higher in group WTs, but the result was not statistically significant. Conclusions ALLs and WTs differ in types of long-term side effects. ALLs present rather metabolic problems, WTs - lower eGFR. Salusin β seems to implicate development of hypertension rather than metabolic disorders like obesity. Further investigations are necessary to confirm this statement.

Acute lymphoblastic leukemia (ALL) is the leading cause of cancer related death in children despite recent advances showing improved responses to chemotherapy treatment. Gamma-delta (γδ) T cells are a recent topic of growing interest in the field of adoptive immunotherapy. These cells have also been proven to be an optimal predictor of a favorable outcome in numerous malignancies. We evaluated subgroups of γδ T cells in the peripheral blood of 19 children with newly diagnosed ALL at the time of diagnosis and following chemotherapy induction. Due to the fact that serum HSP90 serves as a ligand for γδ T cells, we also checked the correlation between HSP90 and γδ T cells. As a result, we found, that the number of CD8+ γδ T cells in peripheral blood at disease presentation was almost three times higher in the intermediate risk group compared to patients in high risk group. Furthermore, we observed higher percentages of the subset in younger patients at diagnosis and after induction, but not in healthy controls. We also noticed negative correlations between CD8+ γδ T cells and minimal residual disease (MRD) before chemotherapy and after induction. We showed a positive association between activated CD3+ γδ T cell and serum HSP90 at presentation. In conclusion, our data suggests that CD8+ γδ T cells may be an early marker of favorable prognosis in childhood ALL while serum HSP90 may act as an agent activating CD3+ γδ T effector cells.

The aim of the study was to retrospectively compare the effectiveness of the ALL IC-BFM 2002 and ALL IC-BFM 2009 protocols and the distribution of risk groups by the two protocols after minimal residual disease (MRD) measurement as well as its impact on survival. We reviewed the medical records of 3248 patients aged 1–18 years with newly diagnosed ALL who were treated in 14 hemato-oncological centers between 2002 and 2018 in Poland. The overall survival (OS) of 1872 children with ALL treated with the ALL IC 2002 protocol was 84% after 3 years, whereas the OS of 1376 children with ALL treated with the ALL IC 2009 protocol was 87% (P P = 0.006). Our study shows that the ALL IC-BFM 2009 protocol improved the results of children with ALL compared to the ALL IC-BFM 2002 protocol in Poland. This analysis confirms that MRD marrow assessment on day 15 of treatment by FCM-MRD is an important predictive factor.

11q23/MLL rearrangements are frequently detected in pediatric acute myeloid leukemia. The analysis of their clinical significance is difficult because of the multitude of translocation fusion partners and their low frequency. The presence of t(10;11)(p12;q23) translocation was previously identified in pediatric acute myelogenous leukemia (AML). It is considered as the second most common translocation detected in pediatric 11q23/MLL-rearranged (present KMT2A) AML, after t(9;11)(p22;q23). The presence of the above translocation was previously identified as an unfavorable prognostic factor. Since June 2015, the Polish Pediatric Leukemia/Lymphoma Study Group has applied the therapeutic protocol requiring extensive diagnostics of genetic changes in pediatric AML. Until November 2019, molecular genetic studies were performed in 195 children with diagnosed AML to identify carriers of fusion gene transcripts for 28 most common chromosomal translocations in acute leukemia. The fusion gene transcript for translocation t(10;11)(p12;q23) involving MLL gene was detected with unexpectedly high frequency (8.9%) in our research. It was the highest frequency of all detected MLL rearrangements, as well as other detected fusion gene transcripts from chromosomal aberrations characteristic for AML. It seems that chromosomal aberration between chromosomes 10 and 11 can be relatively frequent in some populations. Paying attention to this fact and ensuring proper genetic diagnosis seem to be important for appropriate allocation of patients to risk groups of pediatric AML treatment protocols.

A simple, rapid, reliable and sensitive method based on liquid chromatography with fluorescence detection (LC-FL) for the quantification of doxorubicin (DOX) in human plasma and urine samples was developed. The assay was carried out after the solid-phase extraction procedure (SPE) with hydrophilic-lipophilic balance (HLB) cartridges, and with daunorubicin hydrochloride (DAU) used as the internal standard. Chromatographic separation was performed on a Discovery HS C18 column in isocratic elution mode, and the detection of the analytes set at excitation and emission wavelengths of 487 and 555 nm, respectively. The developed LC-FL method has been validated for accuracy, precision, selectivity, linearity, recovery and stability. The limits of detection and quantification for DOX were 0.5 and 1 ng/mL in both biological fluids, respectively. Linearity was confirmed in the range of 1–1000 ng/mL and 0.001–25 μg/mL in plasma and urine samples, respectively, with a correlation coefficient greater than 0.9994. The proposed LC-FL method is selective, precise and accurate, and has been successfully applied for drug monitoring in pediatric cancer patients treated with DOX as a component of OEPA (Oncovin (Vincristine)-Etoposide-Prednisone-Adriamycin) and IOA (Ifosfamide-Oncovin-Adriamycin) chemotherapeutic schemes. Moreover, real exposure of hospital personnel to the anthracycline drugs in plasma and urine was evaluated in clinical practice.

Rosai-Dorfman disease (RDD), also known as sinus histiocytosis with massive lymphadenopathy, is a rare, benign clinical entity of unknown cause. RDD is characterised by the overproduction and accumulation of histiocytes, primarily in the lymph nodes, although it may affect every organ and system. It predominantly affects children and young adults. Typically, patients are in good general condition, with massive cervical lymphadenopathy and fever. In about 40% of cases extranodal localisation of RDD is diagnosed. In laboratory tests the most common abnormalities are increased erythrocyte sedimentation rate (ESR), leukocytosis with neutrophilia, normocytic anaemia, and hypergammaglobulinaemia. Histopathological examination remains the mainstay of diagnosis - lymph nodes have massive sinusoidal dilation, containing histiocytes positive for S-100 and CD68, and negative for CD1a. Most patients do not require treatment as spontaneous remissions are observed. We present a brief review of the literature and the case of a six-year-old boy with cervical lymphadenopathy diagnosed with RDD. So far, the patient has not required systemic treatment and has been kept under observation.

Introduction The authors aimed to answer the following questions: 1) What level of knowledge of type 1 diabetes do the parents of children and young adults with this disease have? 2) Will this level of knowledge increase after 1 year of observation? 3) Does improving the knowledge of young adults and their parents result in better metabolic control of the patients? Material and methods This study included 227 patients between the ages of 5 and 20 years with type 1 diabetes. The research was conducted from March 2009 to June 2011. The following two time points were examined: the beginning of the study (test 1a) and one year later (test 1b). The knowledge levels of the patients and parents were obtained using a survey and a knowledge test. Results Comparison of the results from the two study time points showed that the respondents had a significantly higher level of knowledge after 1 year (p = 0.001). The comparison of glycated hemoglobin levels between the two time points in patients with type 1 diabetes revealed that the levels were significantly higher at test 1b compared to test 1a (p = 0.0005). Conclusions The parents of children and young adults with type 1 diabetes demonstrate a satisfactory level of theoretical knowledge of therapeutic conduct and self-monitoring principles. The test 1b results demonstrated a higher level of theoretical knowledge in all respondents and poorer metabolic control. Poorer metabolic control in some patients suggests that metabolic control in type 1 diabetes depends on factors other than education. Further research is necessary to determine these additional factors.

Sinusoidal obstruction syndrome (SOS), also termed veno-occlusive disease (VOD) of the liver, is a well-known complication of haematopoietic stem cell transplantation (HSCT) both in children and adults. In the medical literature there are occasional reports of SOS in patients receiving conventional chemotherapy. In children with solid tumours this entity occurs during treatment of nephroblastoma, rhabdomyosarcoma, and medulloblastoma. In the late 1990s SOS was quite often observed as the complication of oral 6-thioguanine (6-TG) in patients suffering from acute lymphoblastic leukaemia (ALL), who received 6-TG throughout maintenance. In current protocols, the syndrome has become uncommon because treatment with 6-TG is limited to two weeks of oral therapy. Here, we report a case of a nine-year-old boy with ALL, who developed sinusoidal obstruction syndrome shortly after completing the reinduction block of chemotherapy (cyclophosphamide, cytarabine, thioguanine) according to the ALL Intercontinental Berlin-Frankfurt-Münster 2009 (ALL IC BFM 2009) treatment protocol.

Iron participates in oxygen transport, energetic, metabolic, and immunologic processes. There are 2 main causes of iron overload: hereditary hemochromatosis which is a primary cause, is a metabolic disorder caused by mutations of genes that control iron metabolism and secondary hemochromatosis caused by multitransfusions, chronic hemolysis, and intake of iron rich food. The most common type of hereditary hemochromatosis is caused by HFE gene mutation. In this study, we analyzed iron metabolism in 100 healthy Polish children in relation to their HFE gene status. The wild-type HFE gene was predominant being observed in 60 children (60%). Twenty-five children (25%), presented with heterozygotic H63D mutation, and 15 children (15%), presented with other mutations (heterozygotic C282Y and S65C mutation, compound heterozygotes C282Y/S65C, C282Y/H63D, H63D homozygote). The mean concentration of iron, the level of ferritin, and transferrin saturation were statistically higher in the group of HFE variants compared with the wild-type group. H63D carriers presented with higher mean concentration of iron, ferritin levels, and transferrin saturation compared with the wild-type group. Male HFE carriers presented with higher iron concentration, transferrin saturation, and ferritin levels than females. This preliminary investigation demonstrates allelic impact on potential disease progression from childhood.

Aim of the study : Examination of copy number changes in a group of intracranial germ cell tumors (GCTs) with particular focus on putative aberrations of the main genes coding SHh pathway proteins. Material and methods : The study was performed on DNA isolated from fresh-frozen tumor tissue samples from eight GCTs, including six intracranial GCTs. The intracranial group consisted of three germinomas, two mature teratomas and one mixed germ cell tumor. Comparative genomic profiling analysis was carried out using microarray-CGH method (Cytosure ISCA UPD 4×180k, OGT). The results were analyzed with Feature Extraction (Agilent Technologies) and Nexus Copy Number (BioDiscovery) softwares. Results and conclusions : Chromosomal aberrations were found in two intracranial germinomas. These tumors were characterized by complex genomic profiles encompassing chromosomes 7, 8, 9, 10, 11, 12, 16, 17 and 19. Common findings were gain at 12p13.33p11.1 of 35 Mbp and gain at 17q11.1q25.3 of 55 Mbp. In one tumor, also SHh (7q36.3), SMO (7q32.1) and GLI3 (7p14.1) copy gains occurred together with 9q21.11q34.3 loss, including PTCH1 , all being elements of SHh signaling pathway. Moreover, both tumors showed various copy gain of genes being ligands, regulators, receptors or target genes of SHh ( MTSS1; PRKACA and FKBP8 ) as well as gain of genes of SHh coopting WNT pathway ( WNT3 , WNT5B , WNT9B in both tumors; WNT16 , WNT2 in pineal lesion). Further studies on larger group are needed to characterize SHh-related gene alterations in intracranial GCTs and for searching genotype-phenotype relations.

The unbalanced secretion of biogenic amines (BAs) is considered to be a relevant biochemical biomarker in the screening for neuroendocrine tumors, such as: neuroblastoma and pheochromocytoma. However, there is still a need to improve the bioanalytical procedures for BA determination in biological samples due to their instability (photo- and thermosensitivity, easy oxidation) and low concentration in the body fluids. In this study, the primary analytical challenge was to optimize the method of extraction of seven compounds from among BAs and their precursors from urine samples. Several methods based on liquid-liquid extraction (LLE) or solid phase extraction (SPE) techniques were tested. By optimization of the extraction and data analysis using chemometric tool, the dispersive liquid-liquid microextraction (DLLME) has been chosen due to its low solvents consumption, high efficiency of isolation, preconcentration and suitable clean-up of biological matrix. Further, α-cyclodextrin-modified micellar electrokinetic chromatography (MEKC) with ultraviolet detection (UV) has been applied for quantification of the analyzed biologically active compounds with limits of detection (LOD) and limits of quantification (LOQ) at 0.15 and 0.5μgmL-1, respectively. Finally, the optimized and validated DLLME-MEKC-UV method has been employed for the analysis of real urine samples, obtained from 6 children with neuroendocrine tumors and 6 healthy children. It was stated that concentrations of BA could serve to differentiate between the patients and healthy children. This pilot study indicates that the elaborated fast and sensitive DLLME-MEKC-UV method for determination of panel of biomarkers could be successfully applied in everyday clinical practice to help to confirm the clinical diagnosis of neuroendocrine tumors in children.

The analysis of biogenic amines (BAs) and their metabolites is helpful for the diagnosis of central nervous system disorders and other neuroendocrine and cancer disturbances. In the study, a developed micellar electrokinetic chromatography method, coupled with diode array detection (MEKC-DAD), was validated to monitor levels of adrenaline (A), noradrenaline (NA), dopamine (DA), L-Tryptophan (L-Tryp) and L-Tyrosine (L-Tyr) in real human urine samples. These neurotransmitters were isolated from urine samples using solid-phase microextraction (SPME) and methanol containing 1-ethyl-3-methylimidazolium tetrafluoroborate ionic liquid as the desorption phase. The method was linear for DA, A and L-Tyr in the range of 0.5–20 μg/mL and for NA and L-Tryp in the range of 0.25–20 μg/mL. The good linearity for BAs was confirmed by the correlation coefficient (R2) from 0.9989 for A to 0.9997 for NA and L-Tryp, respectively. The validation assays for accuracy, precision, limit of detection, limit of quantification, absolute recovery, and stability of the analytes were consistent with the requirements recommended by the FDA and ICH guidelines. Next, the validated SPME-MEKC method was successfully used for the quantification of A, NA, DA, L-Tryp and L-Tyr in real human urine samples collected from pediatric patients suffering from neuroblastoma, ganglioneuroblastoma, Wilms’ tumor, rhabdoid tumor and lipoblastomatosis, as well as from healthy volunteers. Finally, the levels of BAs in cancer patients were evaluated as to whether they can be used as biomarkers of various health disturbances.

The objective of this nation-wide study was to evaluate the epidemiology and profile of bacterial (BI), viral (VI), and invasive fungal disease (IFD) in patients treated for non-Hodgkin lymphoma (NHL) and Hodgkin lymphoma (HL) between the years 2013-2015. In the analyzed period of time, within the studied group of 328 children diagnosed and treated for lymphomas, at least one infectious complication (IC) was diagnosed i.e. 39.3% children. In these patients there were 350 episodes of IC, therein 80.6% episodes of BI, 11.1% episodes of VI, and 8.3% episodes of IFD. In both groups, NHL and HL patients, a stable level of bacterial infections, with an increase in resistance rates, and increased levels of viral and fungal infections were observed. Profile of BI does not depend on lymphoma type, with predominance of Gram-negative bacteria and higher prevalence of MDR pathogens. The overall survival of lymphoma patients with IC was comparable for different types of infections.

Currently, many new possible biomarkers and mechanisms are being searched and tested to analyse pathobiology of pediatric tumours for the development of new treatments. One such candidate molecular factor is BARD1 (BRCA1 Associated RING Domain 1)—a tumour-suppressing gene involved in cell cycle control and genome stability, engaged in several types of adult-type tumours. The data on BARD1 significance in childhood cancer is limited. This study determines the expression level of BARD1 and its isoform beta (β) in three different histogenetic groups of pediatric cancer – neuroblastic tumours, and for the first time in chosen germ cell tumours (GCT), and rhabdomyosarcoma (RMS), using the qPCR method. We found higher expression of beta isoform in tumour compared to healthy tissue with no such changes concerning BARD1 full-length. Additionally, differences in expression of BARD1 β between histological types of neuroblastic tumours were observed, with higher levels in ganglioneuroblastoma and ganglioneuroma. Furthermore, a higher expression of BARD1 β characterized yolk sac tumours (GCT type) and RMS when comparing with non-neoplastic tissue. These tumours also showed a high expression of the TERT (Telomerase Reverse Transcriptase) gene. In two RMS cases we found deep decrease of BARD1 β in post-chemotherapy samples. This work supports the oncogenicity of the beta isoform in pediatric tumours, as well as demonstrates the differences in its expression depending on the histological type of neoplasm, and the level of maturation in neuroblastic tumours.

Mastocytosis is a disease caused by the accumulation of mast cells (MC) in the skin and/or in other tissues. Both the cutaneous form of the disease (CM) predominating in children and the systemic form (SM) typical for adults are associated with the occurrence of MC mediator-related symptoms. The release of mediators can be induced by physical stimuli and/or specific triggering factors. The routine vaccination program performed in the majority of children in infancy can be considered as an additional factor provoking exacerbation of CM. Conscious of the important role of MC in the innate immunity, we have analysed retrospective data concerning the levels of immunoglobulins, an adaptive factor, in a group of 74 infants and toddlers with CM. The values corresponding to transient hypogammaglobulinaemia of infants (THI) were found in 8 (10.81%) of cases. Classification of the antibody deficiency was done according to the working definitions for clinical diagnosis of primary immunodeficiency of the European Society of Immunodeficiencies (ESID) Registry - version May 11, 2015. Following the retrospective data, the final diagnosis of THI cannot be made due to the young age of the study group. The percentage may significantly exceed the published incidence of THI, i.e. about 0.11%. The results of our study may indicate, importantly, a higher incidence of THI in childhood-onset mastocytosis than in the general paediatric population and strengthen indications for vaccinations. In conclusion, we suggest that THI may be considered as a new aspect of paediatric mastocytosis that requires further investigation.

The aim of the work was to develop a simple, sensitive and accurate liquid chromatography with fluorescence detection (LC-FL) method for the determination of epirubicin in human urine and plasma. Solid phase extraction with HLB cartridges and mixture of dichloromethane:2-propanol:methanol (2:1:1, v/v/v) as the eluent, was used to prepare the samples. The chromatographic analysis was carried out on a Synergi Hydro-RP column with a mobile phase consisting of 40 mM phosphate buffer (pH 4.1) and acetonitrile (69:31, v/v). Epirubicin was monitored at 497 nm and 557 nm for excitation and emission wavelengths, respectively. Validation data confirmed that the limit of detection and limit of quantification was 0.25 ng/mL and 0.5 ng/mL in both matrices. Next, the optimized LC-FL method was applied to determine the level of epirubicin in real samples taken from a 19-year-old patient with metastatic alveolar rhabdomyosarcoma (RMA) to create a drug profile. Plasma and urine samples were collected for 24 h after the end of a 6-hour infusion of epirubicin. The obtained results confirmed that the optimized and validated LC-FL method can be successfully used in drug monitoring therapy, pharmacokinetic and clinical studies. Moreover, the current work is also drawing attention to the relatively high level of epirubicin in the patient urine, which requires compliance with the safety rules in contact with this biological fluid by both medical staff and others, e.g. family members.

Regulatory T-cells (Tregs) are a very important subtype of lymphocytes when it comes to self-control in the human immunological system. Tregs are decisive not only in the protection against destruction of own tissues by autoimmune immunocompetent cells but also in the immunological answer to developing cancers. On the other hand, Tregs could be responsible for the progression of acute and chronic leukemias. In our study, we review publications available in the PUMED database concerning acute leukemia, with a particular emphasis on child’s leukemias. The percentage of regulatory T-lymphocytes in peripheral blood and bone marrow was elevated compared to those in healthy individuals and correlated with progressive disease. Regulatory T-cells taken from children diagnosed with leukemia showed a higher suppressive capability, which was confirmed by detecting elevated levels of secreted IL-10 and TGF-beta. The possibility of pharmacological intervention in the self-control of the immunological system is now under extensive investigation in many human cancers. Presumably, Treg cells could be a vital part of targeted therapies. Routine Treg determination could be used to assess the severity of disease and prognosis in children with acute lymphoblastic leukemia. This proposition results from the fact that in some studies, higher percentage of Treg cells in peripheral blood was demonstrated. However, observations confirming these facts are scarce; thus, extrapolating them to the population of children with hematological malignancies needs to be verified in additional studies.

This study presents a unique series of malignant supratentorial gliomas in children previously cured from non-CNS primary cancer. On neuroimaging these tumors were not specific, so the patients were suspected of cerebral recurrence of their primary neoplasm: leukemia in four children and sarcoma in one child. Histologically, the group contained four glioblastomas and one anaplastic astrocytoma. Three patients underwent neurosurgical resection, while the other two underwent stereotactic diagnostic biopsy only. Despite combined oncological treatment, four children died during 20 months, and only one glioblastoma patient continued to live for another twelve years. Microscopically, the neoplasms consisted of small cells with some morphologic features of astrocytic lineage, having scanty or prominent processes. Microvascular proliferation and focal or diffuse necrosis were encountered in four cases. The GFAP reactivity in neoplastic cells was low or nil, together with the expression of Olig2, vimentin, and nestin. In two cases a subpopulation of synaptophysin-positive cells was present. Molecular immunohistochemical profiling revealed the expression of phosphorylated forms of PI3Kp110 and AKT, in parallel to a strong PTEN and p53 positivity. The tumors were of IDH1R132H-wild type and immunoreactive for ATRX, HER3, and EGFR. Secondary malignant gliomas in pediatric cancer survivors pose a diagnostic challenge. The present study shows that these tumors are of IDH wild type, PI3K/AKT-activated, having no PTEN and EGFR mutations. Therefore, the biopsy of brain tumors in such patients is crucial both for accurate diagnosis and material preservation for molecular typing.

CD4+CD25highCD127low/−FoxP3+ regulatory T cells (Tregs) are currently under extensive investigation in childhood acute lymphoblastic leukemia (ALL) and in other human cancers. Usually, Treg cells maintain the immune cell homeostasis. This small subset of T cells has been, in fact, considered to be involved in the pathogenesis of autoimmune diseases and progression of acute and chronic leukemias. However, whether Treg dysregulation in CLL and ALL plays a key role or it rather represents a simple epiphenomenon is still a matter of debate. Treg cells have been proposed as a prognostic indicator of the clinical course of the disease and might also be used for targeted immune therapy. Our study revealed statistically higher percentage of Treg cells in the bone marrow than in peripheral blood in the group of 42 children with acute lymphoblastic leukemia. By analyzing Treg subpopulations, it was shown that only memory Tregs in contact with leukemic antigens showed statistically significant differences. We noticed a low negative correlation between Treg cells in the bone marrow and the percentage of blasts (R=−0.36) as well as a moderate correlation between Treg cells in the bone marrow and Hb level (R=+0.41) in peripheral blood before therapy. The number of peripheral blood blasts on day 8th correlates negatively (R=−0.36) with Tregs. Furthermore, statistical analysis revealed low negative correlation between the number of Tregs in the bone marrow and the minimal residual disease measured on day 15th, the percentage of blasts in the bone marrow and leukocytosis after 15 days of chemotherapy. These results indicate the influence of Tregs on the final therapeutic effect.

The main aim of the presented study was to develop a reliable and non-time-consuming method for the simultaneous separation of biogenic amines (BAs) like noradrenalin, adrenalin, dopamine and their main metabolites - homovanillic acid (HVA), vanillylmandelic acid (VMA), 3,4-dihydroxyphenylacetic acid (DOPAC) - in urine samples. To achieve this, the validated α-cyclodextrin (α-CD)-modified micellar electrokinetic chromatography method with DAD was proposed. The optimized separation parameters were as follows: background electrolyte was composed of 10mM sodium tetraborate decahydrate, 30mM SDS, 15% (v/v) methanol and 25mM α-CD, adjusted to pH 9.36 with 1N NaOH; uncoated fused silica capillary (75μm i.d.×60.2cm length); λ=200nm; injection time 5s, applied voltage 25kV; temperature 25 (±0.1)°C. Next, the developed MEKC method was practically applied to evaluate the levels of selected acidic metabolites of catecholamines like HVA, VMA and DOPAC in urine samples collected from patients diagnosed with neuroblastoma (NB), melanotic neuroectodermal tumor of infancy (MNTI).

Infections in children treated for hematological malignancies pose a direct threat to life and are one of the most common causes of treatment failure in this group of patients. Unequivocal diagnosis at the early stages of infection together with an appropriate and timely treatment may be often difficult due to poor manifestation and nonspecific clinical symptoms of the infection progress. Inflammatory markers make a useful diagnostic tool for this purpose. They significantly help to diagnose, monitor, stratify and predict the outcome in severe infections. This article describes selected biomarkers, both those commonly used in clinical practice, such as erythrocyte sedimentation rate, C-reactive protein, procalcitonin as well as less common like IL-6, IL-8 and moreover one promising novel marker – pentraxin 3. The authors emphasize their diagnostic value, clinical usefulness and significance in the treatment efficacy monitoring.

Langerhans cell histiocytosis (LCH) is a rare disease of unknown etiology and varied picture and clinical course. The intensity and the outcome of LCH treatment depend on the amount and type of the affected organs, their dysfunction and age of the patient. In this paper we present the diagnostic difficulties in a 19-month-old girl with multisystem LCH, manifesting mainly by persistent fever, deep anemia, hepatosplenomegaly and lymphadenopathy, coexisting with EBV infection and urinary tract infection. On diagnosis, LCH was suspected once the bone lesions were detected on CT scans, which, however, were not confirmed in the bone scintigraphy. Histopathological diagnosis of LCH was based on the histopathological examination of the cervical lymph node, in which CD1a expression was detected. Myelogram morphologically was assessed as normal; however, bone marrow smears were respectively found to express CD1a. The patient received aggressive and long-lasting treatment according to the protocol LCH in 2009, yielding a complete clinical remission of LCH.

Aim The aim of the study was to analyse the reason of elevated hemoglobin concentration in childhood. Background Elevated hemoglobin concentration is a rare abnormality during childhood. There are disorders of hereditary or acquired hyperproliferations of red cells or pseudo high hemoglobin conditions. Materials and methods We present 3 asymptomatic children in whom high hemoglobin concentration was diagnosed by a family doctor. Results Given that the iron concentration was elevated, genetic testing for HFE mutation was carried out. Conclusion Patients presented with HFE mutation might have an increased hemoglobin production.

Iron overload resulting from the mutation of genes involved in iron metabolism or excess dietary intake has been reported to negatively influence human physical performance. The aim of this study was to test the hypothesis that adolescents bearing a hemochromatosis gene(HFE)mutation in contrast to adults with the same mutation will not experience iron accumulation and their aerobic capacity will be similar to that of age-matched controls. Thirteen boys participated in the study. Seven of them are carriers of H63D mutation in theHFEgene and six were wild type. Fitness levels were assessed using the cardiopulmonary exercise test. In addition, iron status and inflammatory markers were determined. We observed that cardiovascular fitness was significantly lower in the group bearing theHFEmutation compared to the control group. Moreover, theHFEmutation group achieved lower maximal power output compared to the control group. There were no differences in blood ferritin concentrations between the two groups which indicates similar amounts of stored iron. Obtained data do not confirm our hypothesis. On the contrary, it was demonstrated thatHFEmutation is associated with a lower level of aerobic capacity, even in the absence of iron accumulation.

Neuroblastic tumors (NB) are the most common extracranial pediatric neural crest-derived tumors, with a dismal outcome in a substantial group of patients. The study objective was to evaluate the patho-clinical correlations and prognostic impact of the proliferation index (PI) measured with two markers, Ki67 and topoisomerase II alpha (Topo2A), in a NB series. A retrospective analysis of 118 NB from 103 consecutive patients was performed. Analyzed data included tumor stage, histology, mitosis/karyorrhexis index (MKI), MYCN status, and overall survival. Patients' median follow-up period was 50 months. Ki67 and Topo2A PI were assessed immunohistochemically on representative tissue slides in hot spots. PI for Ki67 was in the range 0-72% (median 18%) and for Topo2A was in the range 0-58% (median 20%), being strongly interrelated (r = 0.83). Median PIs with both markers were lower in children older than 18 months (> 18 m) than in the younger patients, with p = 0.0002 and p = 0.005 respectively. Higher Ki67 and Topo2A correlated with metastatic stage, higher MKI, and inversely with increasing tumor differentiation. The cut-off values of PI Ki67 > 30% and Topo2A > 20% were associated with fatal outcome of the disease. In the subgroup of patients > 18 m already at cut-offs Ki67 > 10% and Topo2A > 15% a fatal outcome was predicted by Kaplan-Meyer analysis. Cox regression analysis identified cumulative PI (joint Ki67 and Topo2A index) as an independent prognostic factor. The conclusion is that the proliferation index measured with the examined markers provides substantial prognostic information in NB, especially in infants. PI assessment should become an element of the standard pathological checkup list of NB tumors.

The lack of registration of ultrasound contrast agents for use in patients below the age of 18 is a significant limitation of their usage. Despite this, examinations with the use of contrast agents are conducted in numerous centers, mainly as part of the diagnostic process of vesicoureteral reflux. Examinations after an intravenous administration of contrast agents are conducted rarely. The reason for this is not only the lack of registration, but also the lack of studies on their safety profile in paediatric patients or no guidelines concerning the dosage. It seems that imaging with the use of such agents could help solve certain clinical problems when other diagnostic methods fail. The paper presents selected cases of pediatric patients treated in oncological departments, in whom the examination with the use of ultrasound contrast agents had a considerable influence on the diagnostic and therapeutic process.

A total number of 817 children with acute lymphoblastic leukemia (ALL) and 181 with acute myeloblastic leukemia (AML) were assessed for individualized tumor response testing (ITRT) profile as a prognostic factor in long-term follow-up. For each patient, ITRT, initial response to therapy and long-term outcome were assessed. In initial ALL, an impact on long-term response was shown in ITRT for 13 drugs, while in initial AML only for cytarabine. For patients with ALL, a combined five-drug ITRT profile for prednisolone, l-asparaginase, vincristine, cytarabine and daunorubicin or doxorubicin had predictive value for probability of disease-free survival (pDFS) in univariate analysis, whereas in multivariate analysis, bone marrow response by day 33 was the only prognostic factor. For patients with AML, no factor had prognostic value for pDFS in univariate analysis, while ITRT to cytarabine almost reached significance. In conclusion, ITRT can possibly be regarded as a risk factor in childhood acute leukemias.

Background. Hematopoietic stem cell transplantation (HSCT) is an established procedure for many acquired and congenital disorders of the hematopoietic system, including malignancies, bone marrow failure syndromes, disorders of the immune system, and metabolic disorders. Objective. Analysis of results of hematopoietic stem cell transplantations performed over a period of 12 years in a single pediatric center. Patients and methods. All transplants performed between 2003 and 2015 in the Department of Pediatric Hematology and Oncology in Bydgoszcz were included in this analysis. The results of therapy with stem cell transplantation were analyzed in three time periods: 2004-2007, 2008-2011 and 2012-2015. Results. A total number of transplants was 318, including 132 auto-HSCT and 186 allo-HSCT. Among allogeneic transplants, 68 were done from matched-sibling donor and 118 from alternative donor. The mean survival for all patients estimated by Kaplan-Meier method was 8.1 years. Probability of overall survival (pOS) after all transplants was 0.64±0.03. pOS after allo-HSCT was 0.62±0.04, and 0.67±0.05 after auto-HSCT. Overall survival for patients transplanted in the second (2008-2011) and third (2012-2015) time period was comparable both for auto- and allo-HSCT. However, it was significantly higher than for patients transplanted in the first period of time (2004-2007) for all patients, and for those undergoing auto-HSCT. In allo-HSCT patients, in spite of increase of over 20% in pOS (43% vs 66% vs 64% in respective time periods), the difference was not statistically significant. Conclusion. Presented results of HSCT obtained in our center are comparable with those from other international registries and centers. Wstęp. Transplantacja komórek krwiotwórczych (HSCT) jest ważną metodą terapeutyczną w wielu wrodzonych i nabytych chorobach, w tym nowotworowych, zespołach niewydolności szpiku oraz zaburzeniach immunologicznych i metabolicznych. Celem pracy jest analiza wyników HSCT w pojedynczym ośrodku pediatrycznym w okresie 12 lat. Pacjenci i metodyka. Analizie poddano wyniki przeszczepień wykonanych w latach 2003-2015 w Klinice Pediatrii, Hematologii i Onkologii w Bydgoszczy. Transplantacje analizowano w trzech przedziałach czasowych: 2004-2007, 2008-2011 oraz 2012-2015. Wyniki. Wykonano 318 HSCT, w tym 186 alloge-nicznych (68 zgodnych rodzinnych i 118 od dawców alternatywnych) oraz 132 autologiczne. Średnie przeżycie po HSCT, wyznaczone metodą Kaplana-Meiera wyniosło 8,1 lat. Całkowite prawdopodobieństwo przeżycia (pOS) wyniosło 0,64±0,03; pOS po allo-HSCT wynosi 0,62±0,04, a po auto-HSCT 0,67±0,05. Nie wykazano znamiennych różnic w pOS zarówno po allo-HSCT, jak i po auto-HSCT pomiędzy drugim (2008-2011) i trzecim (2012-2015) analizowanym okresie. Jednakże, pOS było wyższe w dru-gim i trzecim okresie w stosunku do okresu pierwszego (2004-2007), zarówno dla wszystkich pacjentów, jak i u pacjentów po auto-HSCT. W grupie pacjentów allo-HSCT, uzyskano wzrost pOS o ponad 20% (43% vs 66% vs 64% w kolejnych przedziałach czasu; ns). Wnioski. Wyniki HSCT uzyskiwane aktualnie w na-szym ośrodku są porównywalne z wynikami podawanymi w międzynarodowych rejestrach.

Background We sought to verify the hypothesis that children and young adults with cancer who have completed treatment differ according to the type and degree of renal damage. Procedure This study included 144 children and young adults (73 female) who had completed treatment for leukemias and lymphomas (group L, n=45), Wilms tumor (group W, n=52) and other solid tumors (group S, n=47). The following parameters were evaluated: serum concentrations of creatinine, cystatin C, β2-microglobulin, neutrophil gelatinase-associated lipocalin and urine excretion of albumin, and urinalysis with sediment. Glomerular filtration rate (eGFR) was estimated using the classic Schwartz (eGFRSch), Schwartz redux (eGFRSchred), and Filler (eGFRFiller) formulas and with the new Schwartz equation for patients with chronic kidney disease (eGFRSchCKD). Results Group S had the lowest eGFRSchCKD and eGFRFiller, the highest serum cystatin C and the highest albumin excretion compared with groups L and W. Groups S and W had lower eGFRSch and eGFRSchred and higher serum β2-microglobulin and neutrophil gelatinase-associated lipocalin compared with group L. Group W had lower eGFRSchCKD than group L. Conclusions Children and young adults with cancer who have completed treatment differ in the type and degree of renal damage they sustain.

Introduction Hemochromatosis is a disease of iron overload. There are primary – inherited diseases leading to increased intestinal absorption and secondary caused by multi transfusions or iron pills and iron rich food intake. Hereditary hemochromatosis is a metabolic disorder caused by mutations of genes that control iron metabolism. Mutations in HFE gene localized on the short arm of chromosome 6 are responsible for autosomal recessive hemochromatosis type 1. The disease symptoms are observed in adult males. The aim of the study was to analyse iron metabolism in 41 children. In chosen cases HFE mutations were determined. Results HFE mutations were documented in 15 cases. Conclusions Iron metabolism tests are indispensable in the diagnosis of its elevated status. In chosen cases molecular tests are essential.

Introduction: On June 11, 2009, the World Health Organization (WHO) announced a pandemic influenza caused by pandemic (H1N1) 2009 virus. The aim of the current study was to analyze the clinical course of the pandemic (H1N1) 2009 influenza in children with neoplastic diseases or severe aplastic anemia. As a final point, an attempt to identify risk factors predictive for the severity of the clinical course of the disease was also undertaken. Material&Methods: Thirty-seven children undergoing antineoplastic treatment were enrolled in the study. The analysis comprised an assessment of the number of episodes of AH1N1 pandemic (H1N1) 2009 influenza in the group of patients and an evaluation of clinical signs and symptoms of the infection. We also analyzed the relation between the presence of lymphocytopenia and its duration prior to infection and until? eventual contraction of the pandemic (H1N1) 2009 virus, as well as severity of clinical course of the disease. Results: Pandemic (H1N1) 2009 influenza was diagnosed in 20 of 37 children. In the analyzed group no death related to pandemic (H1N1) 2009 influenza was reported. Conclusions: The clinical manifestations of pandemic (H1N1) 2009 influenza in immunoincompetent children are analogous to symptoms observed in the general population. A severe clinical course of the disease was observed among patients with hematological disorders, especially those presenting with preceding lymphocytopenia.

Introduction The majority of hyperglycaemic incidents in oncohaematological patients treated with glucocorticosteroids remain undiagnosed. The aim of our study was to work out a detailed protocol for the control of carbohydrate metabolism and to evaluate whether such a protocol can help in diagnosis of carbohydrate metabolism disturbances in oncohaematological paediatric patients. Material and methods A one hundred and twenty-eight children treated for proliferative diseases of the haematopoietic system and severe aplastic anaemia with therapeutic protocols including glucocorticosteroids were divided into two groups. Group I consisted of 70 children, whose blood glucose was evaluated on random occasions (retrospective analysis). Group II consisted of 58 children included in the programme of intensive carbohydrate metabolism control (prospective analysis). We compared the incidence of hyperglycaemia in both groups as well as the number of hyperglycaemic incidents per individual therapeutic protocol. Results A significantly higher incidence of transient hyperglycaemia was noted in oncohaematological patients in the programme of early carbohydrate metabolism disturbances diagnosis than in the other group (22.4% vs. 5.7% respectively; p = 0.008), especially in patients treated with the ALL IC-BFM 2002 protocol for the high risk group (arm A and B), the ALL-REZ BFM 2002 protocol, and in a heterogenous group of children (protocols ALCL 99, Euro-LB02, Interfant-06, WPSAA) (p = 0.042, 0.021 and 0.002, respectively). Conclusions The improvement of transient hyperglycaemia detection may constitute the first step towards the reduction of unfavourable consequences of hyperglycaemia. Prospective studies are required to demonstrate the influence of normal carbohydrate metabolism on the frequency of infectious complications in this group.