Despite the introduction of new technologies in molecular diagnostics, one should not underestimate the traditional routine methods for studying tumor DNA. Here we present the evidence that short tandem repeat (STR) profiling of tumor DNA relative to DNA from healthy cells might identify chromosomal aberrations affecting therapy outcome. Tumor STR profiles of 87 adult patients with de novo Ph-negative ALL (40 B-ALL, 43 T-ALL, 4 mixed phenotype acute leukemia (MPAL)) treated according to the “RALL-2016” regimen were analyzed. DNA of tumor cells was isolated from patient bone marrow samples taken at diagnosis. Control DNA samples were taken from the buccal swab or the blood of patients in complete remission. Overall survival (OS) analysis was used to assess the independent impact of the LOH as a risk factor. Of the 87 patients, 21 were found with LOH in various STR loci (24%). For B-ALL patients, LOH (except 12p LOH) was an independent risk factor (OS hazard ratio 3.89, log-rank p-value 0.0395). In contrast, for T-ALL patients, the OS hazard ratio was 0.59 (log-rank p-value 0.62). LOH in particular STR loci measured at the onset of the disease could be used as a prognostic factor for poor outcome in B-ALL, but not in T-ALL.

BACKGROUND LiFraumeni syndrome (LFS) is a rare, autosomal dominant, hereditary disorder that is characterized by an increased risk for certain types of cancer, acute lymphoblastic leukemia (ALL), particularly. Germline TP53 mutations are associated with LFS. Genetic counseling and follow-up is essential for patients with LFS and their relatives. Special therapeutic approaches are needed for treatment of oncological disease in these patients. The article presents a series of clinical cases of patients with ALL and SLF, considers general issues of diagnosis and treatment of adult patients with this hereditary genetic syndrome. AIM Describe clinical observations of patients with acute lymphoblastic leukemia (ALL) and LFS and consider general issues of diagnosis and treatment of adult patients with LFS and ALL. MATERIALS AND METHODS TP53 gene mutations were screened using Sanger sequencing in 180 de novo patients with Ph-negative (B- and T-cell) and Ph-positive ALL treated by Russian multicenter protocols (ALL-2009, ALL-2012, ALL-2016) at the National Research Center for Hematology, Moscow, Russia, and at the hematology departments of regional clinics of Russia (multicenter study participants). RESULTS TP53 gene mutations were found in 7.8% (n=14) of de novo ALL patients. In patients, whose biological material was available TP53 gene mutational status was determined in non-tumor cells (bone marrow and peripheral blood during remission, bone marrow samples after allogeneic hematopoietic stem cells transplantation and in tissue of non-hematopoietic origin) for discriminating germline mutations. The analysis included 5 patients (out of 14 with TP53 mutations), whose non-tumor biological material was available for research. Germline status was confirmed in 4 out of 5 B-cell ALL (n=3), T-cell ALL (n=1) investigated patients. CONCLUSION Practical value of the research is the observation that the greater part of TP53 gene mutations in patients with Ph-negative B-cell ALL are germinal and associated with LFS.

Introduction. The Russian multicentre trial on treatment of Acute Lymphoblastic Leukaemia (ALL) in adults (RALL-2016) regulates a centralised detection of minimal residual disease (MRD) on days 70 (end of induction II), 133 (end of consolidation III) and 190 (end of consolidation V).Aim — the assessment of tumour cell clearance and normal B-cell precursor, mature B-cell and plasma cell dynamics.Materials and methods. The study period of December 2016 — August 2019 covered 59 B-ALL patients; the control cohort included four allogeneic blood stem cell donors. The MRD detection was performed with bone marrow samples in flow cytometry.Results. The patient majority have reached MRD-negativity at control timepoints, 60.4, 75.6 and 91.2% on days 70, 133 and 190, respectively. No correlation was observed between MRD values and tumour cell immunophenotype. B-cell precursors were undetected in 54.4, 38.8 and 59.4% patients on days 70, 133 and 190, respectively. Mature B-cells were fewer compared to donors’ on days 133 and 190. The relative plasma cell count did not differ significantly over therapy and between patients and donors.Conclusion. The RALL-2016 protocol facilitates MRD-negativity in the patient majority despite reduced cytostatic intensity.

Context: It is well known that measurable residual disease (MRD) has a significant prognostic impact in patients with acute leukemia. However, the effect of the immunophenotype of MRD cells has not been studied yet. Objective: The aim of the study was to assess the significance of CD38 and CD58 expression for relapse-free survival (RFS) in B-ALL patients. Design: This study included data from 2016 to 2020 for patients enrolled in the Russian multicenter study RALL-2016 (NCT03462095). MRD was assessed in the bone marrow at the end of induction (day +70) in patients who achieved a complete remission. Patients and Methods: The RALL-2016 study included 207 adult patients. The MRD was assessed in 150 patients (80 B-ALL patients and 70 T-ALL patients). Analysis of MRD was performed by 6- or 10-color flow cytometry. The sensitivity was at least 0.01%. CD19, CD34, CD10, CD20, CD38, CD45, and CD58 were analyzed to detect MRD. The main immunophenotypic features of B-ALL blasts are low CD38 expression and high CD58 expression. CD38 expression in MRD cells was considered low if it was lower than that on normal B-cell progenitors. CD58 expression was considered high if it was equal to or higher than that on plasma cells. The RFS was estimated with the Kaplan–Meier method, and comparisons were assessed using the log-rank test. Results: Among B-ALL patients, 48 (60%) patients were MRD-negative, and 32 (40%) patients remained MRD-positive at the end of induction. The duration of RFS was lower in MRD-positive patients (median 29.5 months vs median undefined, p=0.004). Among MRD cells, high CD38 expression was found in 8 patients, and 7 (87.5%) of these relapsed or died. RFS was shorter in CD38-high-MRD patients than in CD38-low-MRD (median 5.2 months vs median undefined, p<0.001). RFS in CD58-high-MRD was also shorter (medians 9.7 vs 34.7 months, p<0.001). Conclusion: MRD at the end of induction was confirmed as an important prognostic factor in the RALL-2016 study. However, immunophenotyping residual blasts could improve the prognostic impact of MRD. High expression of CD58 and CD38 on MRD cells dramatically decreases the duration of RFS in B-ALL patients. It is well known that measurable residual disease (MRD) has a significant prognostic impact in patients with acute leukemia. However, the effect of the immunophenotype of MRD cells has not been studied yet. The aim of the study was to assess the significance of CD38 and CD58 expression for relapse-free survival (RFS) in B-ALL patients. This study included data from 2016 to 2020 for patients enrolled in the Russian multicenter study RALL-2016 (NCT03462095). MRD was assessed in the bone marrow at the end of induction (day +70) in patients who achieved a complete remission. The RALL-2016 study included 207 adult patients. The MRD was assessed in 150 patients (80 B-ALL patients and 70 T-ALL patients). Analysis of MRD was performed by 6- or 10-color flow cytometry. The sensitivity was at least 0.01%. CD19, CD34, CD10, CD20, CD38, CD45, and CD58 were analyzed to detect MRD. The main immunophenotypic features of B-ALL blasts are low CD38 expression and high CD58 expression. CD38 expression in MRD cells was considered low if it was lower than that on normal B-cell progenitors. CD58 expression was considered high if it was equal to or higher than that on plasma cells. The RFS was estimated with the Kaplan–Meier method, and comparisons were assessed using the log-rank test. Among B-ALL patients, 48 (60%) patients were MRD-negative, and 32 (40%) patients remained MRD-positive at the end of induction. The duration of RFS was lower in MRD-positive patients (median 29.5 months vs median undefined, p=0.004). Among MRD cells, high CD38 expression was found in 8 patients, and 7 (87.5%) of these relapsed or died. RFS was shorter in CD38-high-MRD patients than in CD38-low-MRD (median 5.2 months vs median undefined, p<0.001). RFS in CD58-high-MRD was also shorter (medians 9.7 vs 34.7 months, p<0.001). MRD at the end of induction was confirmed as an important prognostic factor in the RALL-2016 study. However, immunophenotyping residual blasts could improve the prognostic impact of MRD. High expression of CD58 and CD38 on MRD cells dramatically decreases the duration of RFS in B-ALL patients.

Context: TPMTandNUDT15polymorphisms are involved in the toxicity and therapeutic efficacy of thiopurine drugs (6-mercaptopurine (6-MP)). The frequencies of polymorphisms of these genes are different across diverse populations. The role ofTPMTandNUDT15genes in adult patients (pts) with Ph-negative acute lymphoblastic leukemia (ALL) in Russia is still unclear. Objective:Evaluation of frequency and significance ofTPMTandNUDT15polymorphisms in the cohort of adult Ph-negative ALL pts treated by the RALL-2016 protocol. Design and Patients:Since April 2017 till July 2020 56 adult Ph-negative ALL patients treated by RALL-2016 protocol were included in the research ofTPMTandNUDT15polymorphisms. Median age was 32 у (range, 18-54), m/f: 39 (70%) / 17 (30%). B-ALL/LBL were diagnosed in 26 (46,4%) pts, T-ALL- in 26 (46,4%) and MPAL- in 4 (7,2%). Genomic DNA was extracted from peripheral blood samples of given pts. Genetic polymorphisms inNUDT15(*2, *3)and TPMT(*2, *3A, *3B, *3C)genes were detected using the allele-specific RT-PCR. The dose of 6-MP was calculated only for 54 pts from 56 (one of these pts stopped the therapy and another received the 1 st phase of induction by RALL-2016 without 6-MP). According to the RALL-2016 protocol 6-MP dose correction imply: if the level leukocytes<2,0*109/l, thrombocytes <100*109/l pts got 50% from the required dose. The therapy of 6-MP was stopped, if the level leukocytes <1,0*109/l, thrombocytes <50*109/l. On the 2 induction therapy by the protocol, the doses of 6-MP were calculated only for 47 pts with CR. The group of pts who didn't have remission on the 36 day by the protocol took the drug without corrections. Results:From 54 pts CR rate was 87 % (n=47) and resistance - 13% (n=7). One patient died in CR. The frequency ofTPMTandNUDT15polymorphisms in study group was 17,8 % (n=10): 6 (23%) of 26 pts with B-ALL, 3 (11,5 %) of 26 pts -T-ALL and 1 (25%) of 4 pts - MPAL. In our cohort these polymorphisms were found significantly more frequently in pts with B-ALL (p<0.001). These polymorphisms were detected in 8 (80%) men and 2 (20%) women (p=0.432). All detected polymorphisms were presented as heterozygous variants:NUDT15*3was in 2 (20%) pts,TPMT*2-1(10%),TPMT*3A-6 (60%),TPMT*3C-1 (10%). The doses of 6-MP in 5 pts withTPMTandNUDT15and 42 pts withWTwere 54% (27-89%) vs 71% (25-100%), respectively. Statistics analysis didn't show correlation between the 6-MP toxicity (difference of the fact dose of 6-MP therapy) and the polymorphisms in our cohort (fig.1). The frequency of resistance cases didn't different in cohort with/withoutTPMTandNUDT15polymorphisms in 3 (37, 5%) from 8 pts and 4 (10, 5%) from 42 pts, respectively. Only 1 patient with heterozygousTPMT*2died in CR due to toxicity and infection. Two-years overall survival (OS) in pts with the genetic polymorphisms was worse 75 % than in pts without them - 83,6% (p=0,67) (fig.2). Conclusions:In our study the frequency ofTPMTandNUDT15polymorphisms in adult pts with Ph-negative ALL was 17,8%. The statistical analysis showed that polymorphisms of these genes were detected more frequently in pts with В-ALL. Only one patient withTPMT*2had significant toxicity on the therapy of 6-MP and died in CR. However, we don't know whether this is due to deficient 6-MP metabolism. We didn't find a correlation between the 6-MP toxicity and the polymorphisms in our pts. There is a tendency that OS within 2-years in pts withoutTPMTandNUDT15polymorphisms is better than in the group of pts with them. But our cohorts are small and require further study. Keywords:acute lymphoblastic leukemia, toxicity, 6-mercaptopurine,TPMT,NUDT15 Disclosures No relevant conflicts of interest to declare.

Context Testing for NRAS, KRAS, JAK2, CRLF2 and TP53 mutations in adult ALL patients and studying of molecular pathways are critical for understanding the biology of ALL. Objective Assessment of mutations in NRAS, KRAS, JAK2, CRLF2, TP53 genes and analysis of signaling pathways in de novo adult ALL patients. Design NRAS, KRAS, JAK2, CRLF2, TP53 mutations were studied B-ALL (Ph+/Ph−). TP53 gene mutations were investigated in B-ALL (Ph+/Ph−) and T-ALL patients.TP53 gene was also evaluated in non-tumor cells of 5 patients to exclude germline mutations. Gene expression profiling was performed for 5 B-ALL patients. Setting Sanger sequencing of hot-spot regions NRAS, KRAS, JAK2, CRLF2 and TP53 (exons 2-10) was conducted. PanCancer Pathways Panel by NanoString was used for gene expression analysis. KEGG pathway analysis for overexpressed genes (of the 13 canonical pathways) was carried out using the DAVID tool. Patients or other participants The study included 95 B-ALL patients, m:f (39:45), median age 31 years; 64 T-ALL, m:f (44:20), median age 27 years, 27 Ph+ ALL, m:f (10:17), median age 33 years. 109 patients treated by protocol ALL-2016 (NCT03462095), 50 patients – by ALL-2009 (NCT01193933), 23 – by ALL-2012 (for Ph«+»ALL) and 4 – by other protocols. Median follow-up 20.8 months. Results NRAS (9.7%), KRAS (12.9%), JAK2 (3.16%) mutations was detected only in B-ALL patients. CRLF2 mutations mutation was not detected. NRAS, KRAS, JAK2 mutations didn't impact disease outcome in B-ALL patients treated by ALL-2009, ALL-2016 protocols. TP53 mutations more often were detected in B-ALL patients (11.2%) compared with T (3.1%) and Ph+ (7.4%). They were associated with poorer OS (TP53-WT – 66.9% vs 48% TP53 mut, p=0.094) and a lower incidence of CR (TP53-WT – 90.6% vs 70.4% TP53 mut, p = 0.033). In 4 out of 5 patients, TP53 mutations were detected in non-tumor cells what considers leukemia as a manifestation of Li-Fraumeni syndrome. The most significant pathways affected were pathways in cancer (p = 6.7E-33) and cell cycle (p = 3.3E-31). Conclusions TP53 mutations are associated with unfavorable prognosis. Most TP53 mutations are of hereditary origin. NRAS, KRAS, JAK2 mutations did not affect the disease outcome. Pathway analysis provides a useful tool for understanding the ALL biology, and we plan to continue it and recruit more patients. The reported study was funded by RFBR, project number 19-31-27001. Testing for NRAS, KRAS, JAK2, CRLF2 and TP53 mutations in adult ALL patients and studying of molecular pathways are critical for understanding the biology of ALL. Assessment of mutations in NRAS, KRAS, JAK2, CRLF2, TP53 genes and analysis of signaling pathways in de novo adult ALL patients. NRAS, KRAS, JAK2, CRLF2, TP53 mutations were studied B-ALL (Ph+/Ph−). TP53 gene mutations were investigated in B-ALL (Ph+/Ph−) and T-ALL patients.TP53 gene was also evaluated in non-tumor cells of 5 patients to exclude germline mutations. Gene expression profiling was performed for 5 B-ALL patients. Sanger sequencing of hot-spot regions NRAS, KRAS, JAK2, CRLF2 and TP53 (exons 2-10) was conducted. PanCancer Pathways Panel by NanoString was used for gene expression analysis. KEGG pathway analysis for overexpressed genes (of the 13 canonical pathways) was carried out using the DAVID tool. The study included 95 B-ALL patients, m:f (39:45), median age 31 years; 64 T-ALL, m:f (44:20), median age 27 years, 27 Ph+ ALL, m:f (10:17), median age 33 years. 109 patients treated by protocol ALL-2016 (NCT03462095), 50 patients – by ALL-2009 (NCT01193933), 23 – by ALL-2012 (for Ph«+»ALL) and 4 – by other protocols. Median follow-up 20.8 months. NRAS (9.7%), KRAS (12.9%), JAK2 (3.16%) mutations was detected only in B-ALL patients. CRLF2 mutations mutation was not detected. NRAS, KRAS, JAK2 mutations didn't impact disease outcome in B-ALL patients treated by ALL-2009, ALL-2016 protocols. TP53 mutations more often were detected in B-ALL patients (11.2%) compared with T (3.1%) and Ph+ (7.4%). They were associated with poorer OS (TP53-WT – 66.9% vs 48% TP53 mut, p=0.094) and a lower incidence of CR (TP53-WT – 90.6% vs 70.4% TP53 mut, p = 0.033). In 4 out of 5 patients, TP53 mutations were detected in non-tumor cells what considers leukemia as a manifestation of Li-Fraumeni syndrome. The most significant pathways affected were pathways in cancer (p = 6.7E-33) and cell cycle (p = 3.3E-31). TP53 mutations are associated with unfavorable prognosis. Most TP53 mutations are of hereditary origin. NRAS, KRAS, JAK2 mutations did not affect the disease outcome. Pathway analysis provides a useful tool for understanding the ALL biology, and we plan to continue it and recruit more patients. The reported study was funded by RFBR, project number 19-31-27001.

ISSUE The study of activating mutations (NRAS,KRAS,FLT3,JAK2,CRLF2genes) of RAS/RAF/MEK/ERK and JAK/STAT signaling pathways in B-cell acute lymphoblastic leukemia (B-ALL) in adult patients which are included in Russian multicenter clinical trials. MATERIALS AND METHODS Within the multicenter study there were 119 adult patients included withde novoB-ALL. The study was considered as prospective and retrospective. The group withBCR-ABL1-negative B-ALL consisted of up to 93 patients (45 male and 48 female, at the age of 17 to 59, the median age 31), they were treated according to the protocols ALL-2009, ALL-2016. The median follow-up lasted for 19 months (1119). The group withBCR-ABL1-positive B-ALL with up to 26 patients (10 male and 16 female, at the age of 23 to 78, the median age 34 years) was included in the study as well. The treatment was carried out according to the protocols ALL-2009 and ALL-2012 in combination with tyrosine kinase inhibitors. The median follow-up lasted for 23 months (4120). The molecular analysis of activating mutations inNRAS,KRASgenes (RAS/RAF/MEK/ERK signaling pathway) andJAK2,CRLF2genes (JAK/STAT signaling cascade) was performed via Sanger sequencing. The internal tandem duplications (ITDs) inFLT3gene were studied by fragment analysis. The evaluation of CRLF2 expression was fulfilled via flow cytometry. RESULTS Activating mutations inNRAS,KRAS,FLT3genes were found in 22 (23.6%) patients withBCR-ABL1-negative B-ALL. In total, 23 mutations were revealed in theNRAS(n=9),KRAS(n=12), andFLT3(n=2) genes, according to statistics that was significantly more frequent than withBCR-ABL1-positive B-ALL, these genes mutations were not identified in patients (p=0.007). The frequency of mutations detection inKRASandNRASgenes in patients withBCR-ABL1-negative B-ALL was comparable as 12.9% (12 of 93) to 9.7% (9 of 93), respectively (p=0.488). One patient was simultaneously revealed 2 mutations in theKRASgene (in codons 13 and 61).FLT3-ITD mutations were detected in 3.5% (2 of 57) cases ofBCR-ABL1-negative B-ALL. In patients withBCR-ABL1-positive B-ALLFLT3-ITD mutations were not assessed. Violations in the JAK/STAT signaling cascade were detected in 4 (4.3%) patients withBCR-ABL1-negative B-ALL. They were represented by the missense mutations ofJAK2gene (n=3) and the overexpression of CRLF2 (n=2); in one patient were detected the overexpression of CRLF2 and a mutation inJAK2gene simultaneously. No mutations were found inCRLF2gene. In patients withBCR-ABL1-positive B-ALL noJAK2mutations were detected. As long as analyzing demographic and clinical laboratory parameters between groups of patients with and without mutations, there were no statistically significant differences obtained. In the analyzed groups of patients, long-term therapy results did not differentiate according to the mutations presence inNRAS,KRAS,FLT3,JAK2genes. Also, substantive differences were not shown in the rate of the negative status achievement of the minimum residual disease between patients with and without activating mutations in the control points of the protocol (on the 70th, 133rd and 190th days). CONCLUSION NRAS,KRAS,FLT3,JAK2activating mutations do not affect the long-term results of the therapy and the rate of the negative status achievement of the minimum residual disease in patients withBCR-ABL1-negative B-ALL treated by the Russian multicenter clinical trials.

Bispecifi c monoclonal antibody blinatumomab is targeting CD19, being applied for treatment of acute lymphoblastic leukemia (ALL).Several additional targets could be used for combined chemo-free treatment with thyrosine kinase inhibitors: BCR-ABL, FLT3 and IKZF1 deletions are amongst them.In the following study, we aimed to assess toxicity and clinical eff ectiveness of the combination of blinatumomab and several thyrosine kinase inhibitors.

CONTEXT:A substantial number of patients with acute lymphoblastic leukemia (ALL) die from progressive disease after relapse that is frequently associated with clonal evolution. Clinical exome sequencing (CES) is a useful tool for mutation investigation, particularly in leukemic subclones during relapse. OBJECTIVE:The objective of this pilot study was to determine the specific genetic abnormalities associated with primary tumor comparing with normal hematopoietic tissue and to define clonal evolution patterns at relapse. DESIGN:CES (TruSightTM One Sequencing Panel) was performed on samples obtained from the patient with B-cell Ph-negative ALL at diagnosis, remission and relapse. SETTING:DNA was extracted from bone marrow samples according to modified salting out method. Libraries were pooled together and underwent two rounds of hybridization and capture, and additional quantification and size assessment. Between 21 and 26 million reads for each sample were obtained with a MiSeq (Illumina, USA), with a coverage of at least 20x for 97% of all sequences. More than 99% of reads passed quality and length control and were used in the analysis. De-multiplexing of indexed reads and the generation of FASTQ files were performed by the MiSeq system on-instrument software. The sequencing data, composed of 150 bp paired-end reads, were processed for MiSeq Reporter (Illumina), which is endowed with Burrows-Wheeler Aligner (BWA) for alignment on a reference sequence of hg19 and Genome Analysis Toolkit (GATK) for variant calling. Then, the Variant Call format (VCF) file was processed for analyzing on VariantStudio (Illumina) and wANNOVAR (Yang H. et al, 2015), which contain annotation databases, such as the ClinVar database, OMIM, the dbSNP database), the Polymorphism Phenotyping v2 database (Polyphen-2), and the Sorting Intolerant from Tolerant database (SIFT) and some others. PATIENTS OR OTHER PARTICIPANTS :A 27-year-old woman, who was admitted to the National Research Center for Hematology (Moscow, Russia) on March, 2017, presented with a three-month history of paleness and ossalgia. Common B-ALL (BII) ALL with complex karyotype and extramedullary lesions in ovaries, kidneys, intra-abdominal and peripheral lymph nodes has been diagnosed. Patient achieved complete remission (CR) after first induction therapy according to ALL-2016 protocol (ClinicalTrials.gov NCT03462095). After 16 months of CR during maintenance therapy (July, 2019) was diagnosed a relapse, which was refractory to multiple chemotherapy approaches and the patient died from the disease (January, 2020). RESULTS:Primary dataset analysis included more than 9000 gene variations for each of the three samples. Only clonal variations (at diagnosis) that are pathogenic either by the type of mutations (insertions (frameshift) and splice site mutations) or by prediction databases (PolyPhen-2, SIFT) were selected. As a result, about 70 variations were identified for each of the samples. Table 1 shows all clonal somatic mutations by which tumor tissue differs from normal hematopoietic tissue. At the time of diagnosis, a clone with monosomy of chromosome 3 was identified, that disappeared at CR and reappeared as a minor clone at relapse. Taking in consideration that the loss of one allele was caused by monosomy, we searched for pathogenic variations in genes localized on the chromosome 3 in order to identify genes inactivated according to Knudson's "Two-Hit" Theory of cancer causation (Knudson A., 1971). Out of 500 genes variations on chromosome 3, mutations in 5 genes according to prediction databases (PolyPhen-2, SIFT) were defined as pathogenic. All mutations were germinal. Moreover, four out of five mutations were homozygous. Table 2 shows the dynamics of pathogenic variations in genes localized on the chromosome 3 at different stages of the disease. The missense mutation ofCOL7A1gene (c.2791C> T), which was found in a small percentage (8%), was localized on the disappeared copy of chromosome 3, and its detection in this case was possible due to the presence of normal cells. CONCLUSIONS:There is an evident change in tumor clones at relapse: a major clone diagnosed inde novoB-cell ALL patient became minor at relapse. The data obtained from this analysis suggest that at the time of diagnosis, several clones are present in the tumor, and that most of the diagnosed pathogenic variations are not associated with oncogenesis and are germinal. Disclosures No relevant conflicts of interest to declare.

Background. Nijmegen breakage syndrome is a rare hereditary autosomal recessive disorder characterized by microcephaly, combined primary immunodeficiency, sensitivity to radioactive radiation and liability to tumours of various nature (in particular, those developing in the lymphatic tissue). This syndrome is part of a group of diseases characterized by chromosomal instability. This disease develops as a result of mutations in the NBS1 gene, which is responsible for repairing DNA double-stranded breaks.Aim. To describe a clinical case of the diagnosis and treatment of T-cell acute lymphoblastic leukemia in a patient with Nijmegen syndrome, which was first diagnosed in adulthood.General findings. A clinical case of the diagnosis and treatment of Nijmegen syndrome in a young man with de novo T-cell acute lymphoblastic leukemia is presented. The difficulty of early diagnosis of hereditary genetic syndromes is demonstrated. The genetic character of such conditions is revealed over time, when children and young adults begin to develop long-term complications, in particular tumours of various origins. Early detection of hereditary genetic syndromes in children is of great importance.

Introduction. No recommendations are currently available on the use of positron emission tomography / computer tomography (PET/CT) for evaluating the response to chemotherapy in patients with acute lymphoblastic leukosis / lymphoblastic lymphoma (ALL/LBL). Aim . The aim of this research was to study the ability of tumour cells to accumulate radiopharmaceuticals during PET/CT in patients with ALL/LBL, as well as to evaluate the prognostic value of PET/CT results performed after completion of consolidation therapy with/without autologous hematopoietic blood stem cell transplantation (auto-HSCT) in patients with Ph-negative ALL/LLL who underwent therapy according to the protocols of a Russian research group ALL-2009/ALL-2016. Materials and methods. PET/CT was performed in 3 patients with various variants of a newly diagnosed ALL before the onset of therapy and after the completion of induction therapy. In 10 patients with Ph-negative ALL/LLL, a PET study was performed after consolidation had been completed according to the ALL-2009/ALL-2016 protocol. Results . The results of PET/CT in 3 patients with different variants of newly detected ALL/LBL were analysed. All patients showed a metabolic activity of 18F-FDG in all morphologically and immunohistochemically (immunophenotypically) confirmed lesions. An analysis of the PET/CT results in 10 patients with Ph-negative ALL/LBL after completion of consolidation therapy with/without auto-HSCT showed that all patients had achieved a PET-negative disease remission. With a median follow-up of 20.5 months (from 15 to 44 months), only one out of 10 patients demonstrated isolated neurorecurrence 10 months after achieving remission. The remaining 9 patients, under a median relapse-free survival rate of 19 months (from 14 to 43 months), demonstrated complete clinical and hematological remission. Conclusion . Specific medullary and extramedullary lesions in ALL/LBL are capable of accumulating 18F-FDG in PET, which allows the method under study to be used for evaluating the completeness of remission in extramedullary lesions. The prognostic feasibility of PET/CT under the involvement of the central nervous system remains to be studied.

Introduction.Modern therapy for relapses and resistant forms of T-cell acute lymphoblastic leukaemia/lymphoma (T-ALL/ LBL) shows poor efficacy. The use of nelarabine can improve the results of therapy in patients with refractory/relapsed T-ALL/LBL.Aim.To evaluate the efficacy and toxicity of nelarabine treatment combined with etoposide and cyclophosphamide in adult patients with refractory/relapsed T-ALL/LBL.Materials and methods. During the 2012–2018 period, 10 patients with refractory/relapsed T-ALL aged from 19 to 41 underwent nelarabine treatment. The patients received from 1 to 3 chemotherapy courses including nelarabine 650 mg/m2 (days 1 to 5), etoposide 100 mg/m2 and cyclophosphamide 440 mg/m2 (days 8 to 12). All the patients having achieved complete remission (СR) underwent transplantation of allogeneic haematopoietic stem cells (allo-HSCT). The development of toxic sequelae (myelosuppression, neurotoxicity, incidence of infectious complications) was considered after each chemotherapy course.Results. Out of 10 patients who received 1–2 chemotherapy courses, 6 (60 %) achieved CR. These 6 patients subsequently underwent allo-HSCT, which was followed by early relapse in 3 (50 %) of 6 patients and the death of 1 patient in persisting CR caused by infectious complications. Only 2 of 6 patients have been monitored for 1.5 years after the allo-HSCT. The five-year overall survival rate in relapsed patients came to 18 %. In terms of toxic sequelae, myelosuppression and infectious complications were observed in all patients. Neurotoxicity was noted in 3 (30 %) out of 10 patients, with two of them experiencing it after each course and one patient — only following the third course of nelarabine treatment.Conclusion. The use of nelarabine for the treatment of refractory/relapsed T-ALL/LBL provides the opportunity to achieve CR in 60 % of cases, as well as to perform allo-HSCT. However, long-term results are not very optimistic, thus further research is required.Conflict of interest:the authors declare no conflict of interest.Financial disclosure:the study had no sponsorship.

Background:Positron emission tomography/computed tomography (PET/CT) is emerging as an essential diagnostic tool in characterizing suspicious neoplastic lesions and staging malignant diseases. Most studies regarding the possible role of FDG‐PET/CT in the management of acute lymphoblastic leukemia (ALL) patients are very limited. But one can suggest the high role of PET/CT in the assessment of residual lesions after chemotherapy in patients with lymphoblastic lymphoma (LBL) and ALL with extramedullary involvements.Aims:The aim of the study was to estimate the ability to accumulate 18F‐FDG radiopharmaceuticals by tumor cells in PET/CT in patients with ALL/LBL and assess the prognostic value of the PET/CT after the consolidation therapy (with or without autologous stem cell transplantation (autoSCT)) in patients with Ph‐negative ALL/LBL who received therapy according to the protocols of the Russian research group ALL‐2009/ALL‐2016.Methods:PET/CT was performed in 3 patients with different types of newly diagnosed ALL before the start of the therapy and after the end of induction. Characteristic of patients were: the first‐ male, 55 age, with BII type Ph‐negative ALL, with t(6;11) with generalized bone involvement and ossalgia; the second ‐ female, 33 age, with BII type Ph‐positive ALL, p190type of transcript without extramedullary involvement; and the third‐ female, 28 age, with TIII type ALL, with TCR A/D rearrangement, with massive mediastinal lesion, bilateral renal and ovarium involvement.PET/CT was applied in 10 patients with Ph‐negative ALL/LBL after the end of consolidation by ALL‐2009/ALL‐2016 protocols. The characterization of patients (Table): 6/4 (60%/40%) M/F, 9/1 (90%/10%) B/T type Ph‐negative ALL, 8/1/1 (80%/10%/10%) with mediastinum/lymph nodes/renal and ovarium involvements, 3/7 (30%/70%) with or without bone marrow involvement, nobody with CNS involvement, 5/5 (50%/50%) with or without autoSCT, 3/7 (30%/70%) with or without radiotherapy in consolidation.Results:The results of PET/CT in three patients with different variants of newly diagnosed ALL/LBL were analyzed and it was shown that all patients had metabolic activity of 18F‐FDG in all morphologically and immunohistochemically (immunophenotypically) confirmed lesions (SUV 3,9‐8,7).An analysis of the results of PET/CT after the consolidation therapy with/without autoSCT in patients with Ph‐negative ALL was performed, and it has shown that all patients had PET‐negative remission of the disease and MRD negative status in bone marrow by flow cytometry. With a median follow‐up of 20.5 months (from 15 to 44 months), only one patient out of 10 patients had isolated CNS relapse after 10 months of remission. In another 9 patients, with a median CR duration of 19 months (from 14 to 43 months), complete clinical and hematological remission is maintained.Summary/Conclusion:The results demonstrated that all specific lesions in ALL/LBL patients, medullary and extramedullary, were capable to accumule 18F‐FDG in PET. This fact provides opportunities for further studies of this method in assessing the completeness of remission in patients with extramedullary lesions. The value of PET/CT in ALL/LBL patients with central nervous system involvement remained unexplored. If the suggestions arising from this report are confirmed, FDG‐PET/CT may be included in future clinical practice for disease staging, evaluating the response to the chemotherapy in identified lesions and in the restaging for recurrence of extramedullary disease.image

The aim of the study was to investigate the role of IKZF1 deletions in adult patients with Ph-negative (Ph–) and Ph-positive (Ph+) B-cell acute lymphoblastic leukemia (ALL) who participated in the Russian Acute Lymphoblastic Leukemia (RALL) multicenter study. Our study included 67 patients with newly diagnosed B-cell ALL (49 Ph– and 18 Ph+ cases). Patients with Ph– B-cell ALL were treated according to RALL-2009 and RALL-2016 protocols and were followed up for a median of 18.1 months (range 1.5–93.4 months). Patients with Ph+ B-cell ALL were treated according to RALL- 2009 and RALL-2012 protocols with addition of tyrosine kinase inhibitors and were followed up for a median of 21.2 months (range 3.53–91.77 months). Intragenic deletions of IKZF1 were detected using breakpoint-specific fluorescent multiplex polymerase chain reaction. They were more frequently found in patients with Ph+ ALL (n = 10, 56%) than in patients with Ph– ALL (n = 9, 18%; p = 0.0074). No statistically significant association between IKZF1 deletions and age, sex, initial WCC of over 30 × 109/L, LDH above 750 U/mL, splenomegaly or neuroleukemia was observed. Notably, an expression of both myeloid antigens (MyAg) CD13 and СD33 was detected in almost half (n = 4,44%) of the Ph– ALL patients with IKZF1 deletions compared to only 1 patient (2.5%) without IKZF1 deletions (p = 0.0027). The presence of IKZF1 mutations was associated with persistence of minimal residual disease at 2 and 4 months of treatment, with higher leukemic cell counts; however, there seemed to be no observable differences in the long-term results of therapy regardless of whether or not IKZF1 mutations were present. Thus IKZF1 mutations in our study did not seem to be prognostically valuable for either Ph+ B-cell ALL or Ph– B-cell ALL, although they were shown to be associated with a delayed tumor clearance in patients with Ph– B-cell ALL.

Introduction. MRD-tailored therapy based on pediatric-inspired intensification is a back-bone of the majority of the European study groups in adult ALL. Taking in consideration the major pitfalls of the first Russian acute lymphoblastic leukemia study group trial RALL-2009 (NCT01193933) - high CR death rate, early CNS relapses in T-ALL, selection bias in auto-HSCT vs chemotherapy comparison, absence of MRD monitoring - a new RALL-2016 protocol (NCT03462095) was introduced based on the same principles as the first one - non-intensive but non-interruptive approach with low numbers of allo-HSCT, but with further deintensification of consolidation phase, centralized MRD-monitoring and randomization for autologous HSCT with non-myeloablative conditioning (CEAM). AIM. To analyze the 2,5 years efficacy and to determine significance of MRD status after induction in the new Russian ongoing prospective multicenter study RALL-2016. Materials and patients. RALL-2016 was based on the previous RALL-2009 protocol , but one day high-dose MTX and high-dose ARA-C blocks were eliminated and substituted by 2 months of non-interruptive therapy, L-asparaginase was scheduled for 1 year of treatment instead of 2,5 y, 15 intrathecal injections were increased up to 21 mostly while consolidation phase, CR T-ALL patients were brought to randomization after the informed consent: auto-HSCT vs no auto-HSCT, - with the similar further maintenance. All primary bone samples are collected and tested for cytogenetics and molecular markers, all included patients are monitored by flow cytometry by aberrant immunophenotype in a centralized lab. Results and discussion. From Dec 2016 till Jul 2019 148 Ph-negative ALL pts from 10 centers were included: median age 33 y (18-54) (BCP-ALL-80 (54%) pts, T-ALL- 64 (44%), biphenotypic- 4 (2%)). CR was achieved in 84% pts. The induction death before CR was 8% (n=12), refractory ALL was registered in 12 pts (8%). Death in CR occurred in 4%. After CR achievement 52 T-ALL patients were randomized either to chemotherapy (n=25) or to autoHSCT(n= 27). 15 of 27 T-ALL pts were transplanted at a median time of 6 months from CR (1 of 27 received alloHSCT - Neimegen Syndrom, 2 of 27 died in CR before HSCT, one pt refused the autotransplant ). OS and DFS at 2-years constituted 70,7% and 80%. 2-y OS was 65,8% for BCP-ALL, 80% for T-ALL and 66,7% for MPAL (p=0,5). 2-y DFS was 78,7% for BCP-ALL, 83,4% for T-ALL and 100% for MPAL (p=0,88). AlloHSCT in 1st CR have received only 3 (2%) pts. We have registered the differences in OS in pts who were treated in Federal Center (51 pts) or in Regional centers (97pts): 82% vs 64,6%, respectively (p=0,02). But there were no differences in DFS: 87,7% vs 77,3%, respectively (p=0,66) (Pic1). We have detected very high death rate in induction and in CR in the regional Centers despite the fact that the main pts characteristics were similar (median age, hyperleukocytosis, high risk group). MRD persistence after induction (70th day of protocol) became a significant factor of poor prognosis: 2-yeasr OS and DFS in MRD-negative (59 pts) and MRD-positive (33pts) were 91,8% vs 56,4% (p=0,017) and 88,7% vs 64,3% (p=0,16), respectively (Pic 2). Median of relapse was 7 month. Conclusion. The new RALL-2016 study pitifully continues to demonstrate high induction and CR death rate in regional centers despite of de-intensification of chemotherapy. We've observed significant differences in OS in Federal Center vs Regionals Centers, but not in DFS. MRD monitoring by FCM in ALL patients revealed that the persistence of MRD after induction (day+70) was an independent factor of poor prognosis and high relapse rate suggesting the introduction of new treatment approaches within a very short time after induction (maximum 3 months) in MRD positive patients. Figure Disclosures No relevant conflicts of interest to declare.