Immunogenicity and clinical effectiveness of mRNA vaccine booster against SARS-CoV-2 Omicron in patients with haematological malignancies: A national prospective cohort study.

Information regarding the protective anti-SARS-CoV-2 antibody levels and the effectiveness of the mRNA vaccines against the Omicron variant in patients with haematological malignancies is limited. We prospectively followed two times BNT162b2 vaccinated oncohaematological patients (n = 1010) without prior COVID-19 for PCR-confirmed breakthrough infections during the Alpha/Delta and the Omicron phases of the pandemic. Anti-S1-IgG levels were longitudinally monitored in patients who had received the third (booster) vaccine dose. Patients with anti-S1-IgG levels <50 BAU/mL 1 month after the booster had a higher risk of Omicron infections (RR 1.91; 95% CI 1.39-2.63; p = 0.0001) and severe infections (RR 8.74; 95% CI 3.99-19.1; p < 0.0001). Conversely, the risk of severe COVID-19 was <1% with anti-S1-IgG levels >500 BAU/mL and neutralizing antibody concentrations >50 U/mL. The risks of breakthrough Omicron infections (HR 0.55; 95% CI 0.32-0.96; p = 0.034) and severe COVID-19 (HR 0.27; 95% 0.11-0.7; p = 0.0074) were lower among patients who had received the booster dose. In conclusion, low antibody levels are associated with significantly increased risk of both the breakthrough Omicron infections and severe COVID-19. The third mRNA vaccine dose improved the protection against the Omicron and reduced the risk of severe disease.

Genetic and Epigenetic Signatures in Acute Promyelocytic Leukemia Treatment and Molecular Remission.

Acute myeloid leukemia (AML) is an aggressive, heterogeneous group of malignancies with different clinical behaviors and different responses to therapy. For many types of cancer, finding cancer early makes it easier to treat. Identifying prognostic molecular markers and understanding their biology are the first steps toward developing novel diagnostic tools or therapies for patients with AML. In this study, we defined proteins and genes that can be used in the prognosis of different acute leukemia cases and found possible uses in diagnostics and therapy. We analyzed newly diagnosed acute leukemia cases positive for t (15; 17) (q22; q21) PML-RAR alpha, acute promyelocytic leukemia (APL). The samples of bone marrow cells were collected from patients at the diagnosis stage, as follow-up samples during standard treatment with all-trans retinoic acid, idarubicin, and mitoxantrone, and at the molecular remission. We determined changes in the expression of genes involved in leukemia cell growth, apoptosis, and differentiation. We observed that WT1, CALR, CAV1, and MYC genes' expression in all APL patients with no relapse history was downregulated after treatment and could be potential markers associated with the pathology, thereby revealing the potential value of this approach for a better characterization of the prediction of APL outcomes.

Population-based targeted RNA sequencing reveals novel disease-related gene fusions in pediatric and adult T-ALL.

BACKGROUND: T-cell acute lymphoblastic leukemia (T-ALL) is a genetically diverse disease characterized by a heterogeneous profile of genetic lesions. Recent transcriptome studies identified a number of new T-ALL-related gene fusions. Novel gene fusions could be employed as disease-specific molecular markers and provide a better understanding of T-ALL biology, proving the need for further omics sequencing studies. METHODS: We performed a population-based analysis of 65 (26 pediatric and 39 adults) Lithuanian T-ALL patients. Targeted RNA sequencing (RNA-seq) was used to detect recurrent and novel T-ALL-related fusion transcripts and gene sequence variants. RT-qPCR was used to calculate the relative gene expression of fusion transcripts. RESULTS: We identified disease-related gene alterations in 57/65 (87.7%) T-ALL cases, of which four patients harbored gene fusions that affect ABL-class or JAK-STAT signaling pathways. Five novel gene fusions were detected in 4/65 (6.2%) T-ALL cases: CD99::ABL2 and ZEB1::GNAS in the same case, CTCF::ENKD1, DIAPH1::JAK2, and CDK6::NEK1. Novel fusion transcripts encode chimeric proteins that can potentially affect T-cell proliferation, apoptosis, and help to maintain leukemic cells. Importantly, novel fusion transcripts were not detected in the clinical remission samples attributing these fusions to the leukemic compartment. CONCLUSIONS: We report a similar incidence rate of recurrent gene alterations affecting T-ALL-related molecular signaling pathways in both Lithuanian T-ALL patients and other T-ALL studies. Our data suggest that newly identified gene fusions are specific to leukemic T-cells and provide new molecular insights on T-ALL pathogenesis. Further research is needed to confirm these findings.

Low incidence of ABL-class and JAK-STAT signaling pathway alterations in uniformly treated pediatric and adult B-cell acute lymphoblastic leukemia patients using MRD risk-directed approach - a population-based study.

BACKGROUND: ABL-class and JAK-STAT signaling pathway activating alterations have been associated with both a poor post-induction minimal residual disease (MRD) response and an inferior outcome in B-cell acute lymphoblastic leukemia (B-ALL). However, in most of the studies patients received non-uniform treatment. METHODS: We performed a population-based analysis of 160 (122 pediatric and 38 adult) Lithuanian BCR-ABL1-negative B-ALL patients who had been uniformly treated according to MRD-directed NOPHO ALL-2008 protocol. Targeted RNA sequencing and FISH analysis were performed in cases without canonical B-ALL genomic alterations (high hyperdiploids and low hypodiploids included). RESULTS: We identified ABL-class fusions in 3/160 (1.9%) B-ALL patients, and exclusively in adults (p = 0.003). JAK-STAT pathway fusions were present in 4/160 (2.5%) cases. Of note, P2RY8-CRLF2 fusion was absent in both pediatric and adult B-ALL cases. Patients with ABL-class or JAK-STAT pathway fusions had a poor MRD response and were assigned to the higher risk groups, and had an inferior event-free survival (EFS) / overall survival (OS) compared to patients without these fusions. In a multivariate analysis, positivity for ABL-class and JAK-STAT fusions was a risk factor for worse EFS (p = 0.046) but not for OS (p = 0.278) in adults. CONCLUSIONS: We report a low overall frequency of ABL-class and JAK-STAT fusions and the absence of P2RY8-CRLF2 gene fusion in the Lithuanian BCR-ABL1 negative B-ALL cohort. Future (larger) studies are warranted to confirm an inferior event-free survival of ABL-class/JAK-STAT fusion-positive adult patients in MRD-directed protocols.

Outcomes of relapsed or refractory acute myeloid leukemia patients failing venetoclax-based salvage therapies.

OBJECTIVES AND METHODS: We conducted a retrospective analysis to evaluate the outcomes of 28 heavily pretreated (median 3 (2-6) treatment lines, sixteen (57%) allotransplanted) relapsed/refractory acute myeloid leukemia patients who had failed salvage venetoclax-based therapies. RESULTS: The median age was 59 years (20-80), 20 patients (71%) had ECOG 2-4 status, and 18 patients (64%) were stratified to European Leukemia Network 2017 adverse risk group. The most common mutations were ASXL1 (21%), RUNX1 (18%), FLT3 ITD/TKD (18%), PTPN11 (15%), NRAS/KRAS (15%), and WT1 (15%). Twenty-two patients (79%) received different post-venetoclax salvage therapies with the overall response rate of 23% (complete remission + morphological leukemia-free state). Three of six (50%) patients achieved complete remissions after therapy with venetoclax + actinomycin D ± low-dose cytarabine. The remaining 6 patients did not receive any further salvage treatment mainly due to poor general condition. The median overall survival was 3.9 months for all patients (4.3 for those receiving post-venetoclax salvage vs 1.3 months receiving palliative care alone, P < .001). CONCLUSIONS: Though the remission rate and survival of patients failing venetoclax are poor, a small proportion of these R/R AML patients may still respond to cautious intensification of chemotherapy with venetoclax.

Corrigendum to "HDAC and HMT Inhibitors in Combination with Conventional Therapy: A Novel Treatment Option for Acute Promyelocytic Leukemia".

[This corrects the article DOI: 10.1155/2019/6179573.].

Dose Intensive Rituximab and High-Dose Methylprednisolone in Elderly or Unfit Patients with Relapsed Chronic Lymphocytic Leukemia.

Background and Objectives: BTK and BCL2 inhibitors have changed the treatment paradigms of high-risk and elderly patients with chronic lymphocytic leukemia (CLL), but their long-term efficacy and toxicity are still unknown and the costs are considerable. Our previous data showed that Rituximab (Rtx) and high-dose methylprednisolone (HDMP) can be an effective and safe treatment option for relapsed high-risk CLL patients. Materials and Methods: We explored the efficacy and safety of a higher Rtx dose in combination with a shorter (3-day) schedule of HDMP in relapsed elderly or unfit CLL patients. Results: Twenty-five patients were included in the phase-two, single-arm trial. The median progression free survival (PFS) was 11 months (range 10-12). Median OS was 68 (range 47-89) months. Adverse events (AE) were mainly grade I-II° (77%) and no deaths occurred during the treatment period. Conclusions: 3-day HDMP and Rtx was associated with clinically meaningful improvement in most patients. The median PFS in 3-day and 5-day HDMP studies was similar and the toxicity of the 3-day HDMP schedule proved to be lower. The HDMP and Rtx combination can still be applied in some relapsed high-risk and elderly or unfit CLL patients if new targeted therapies are contraindicated or unavailable. (ClinicalTrials.gov identifier: NCT01576588).

HDAC and HMT Inhibitors in Combination with Conventional Therapy: A Novel Treatment Option for Acute Promyelocytic Leukemia.

Acute promyelocytic leukemia (APL) is characterized by PML-RARA translocation, which causes the blockage of promyelocyte differentiation. Conventional treatment with Retinoic acid and chemotherapeutics is quite satisfactory. However, there are still patients who relapse or develop resistance to conventional treatment. To propose new possibilities for acute leukemia treatment, we studied the potential of histone deacetylase (HDAC) inhibitor and histone methyl transferase (HMT) inhibitor to enhance conventional therapy in vitro and ex vivo. NB4 and HL60 cell lines were used as an in vitro model; APL patient bone marrow mononuclear cells were used as an ex vivo model. Cell samples were treated with Belinostat (HDAC inhibitor) and 3-Deazaneplanocin A (HMT inhibitor) in combination with conventional treatment (Retinoic acid and Idarubicin). We demonstrated that the combined treatment used in the study had slightly higher effect on cell proliferation inhibition than conventional treatment. Also, enhanced treatment showed stronger effect on induction of apoptosis and on suppression of metabolism. Moreover, the treatment accelerated granulocytic cell differentiation and caused chromatin remodelling (increased H3K14 and H4 acetylation levels). In vitro and ex vivo models showed similar response to the treatment with different combinations of 3-Deazaneplanocin A, Belinostat, Retinoic acid, and Idarubicin. In conclusion, we suggest that 3-Deazaneplanocin A and Belinostat enhanced conventional acute promyelocytic leukemia treatment and could be considered for further investigations for clinical use.

Significance of BRAF V600E Mutation and Cytomorphological Features for the Optimization of Papillary Thyroid Cancer Diagnostics in Cytologically Indeterminate Thyroid Nodules.

BACKGROUND: Ultrasound guided fine needle aspiration biopsy with cytologic analysis is an initial step in diagnostic of thyroid nodules. Unfortunately, up to 30% of biopsies are indeterminate and diagnostic surgery is required. The aim of this study was to estimate the diagnostic value of BRAF V600E mutation status combined with cytomorphological features for diagnosis of papillary thyroid cancer (PTC) in cytologically indeterminate thyroid nodules. METHODS: A prospective study analyzed patients who had ultrasound suspicious thyroid nodules, underwent fine needle aspiration and cytological examination, and were classified according to the Bethesda system. Patients from indeterminate diagnostic categories were examined for BRAF V600E mutation and 22 cytomorphological features, and underwent thyroid surgery. A binary logistic regression model was used to evaluate the diagnostic utility. RESULTS: A total of 219 patients met study criteria. After histological examination, 77 (35.2%) patients were diagnosed with PTC and 142 (64.8%) with benign nodular thyroid disease. According to logistic regression model, significant features for PTC diagnosis were: liquid colloid consistency, papillary structures, eosinophilic colloid bodies, and BRAF V600E mutation. Risk groups classified by this model have sensitivity of 80.5% (95% CI: 69.9 to 88.7), specificity of 99.3% (95% CI: 96.1 to 100), positive predictive value of 98.4% (95% CI: 89.8 to 99.8), negative predictive value of 90.4% (95% CI: 85.7 to 93.7), and accuracy of 92.7% (95% CI: 88.4 to 95.8) for PTC diagnosis. CONCLUSIONS: Evaluation of BRAF V600E mutation status combined with cytomorphological features for diagnosis of PTC in cytologically indeterminate thyroid nodules can significantly improve diagnostic accuracy and reduce the number of diagnostic operations (calculator available at www.ptc-calc.we2host.lt).

The utility of the Bethesda category and its association with BRAF mutation in the prediction of papillary thyroid cancer stage.

PURPOSE: This study aims to determine the utility of the Bethesda category and its association with BRAF mutation in prediction of the papillary thyroid cancer (PTC) stage. METHODS: A prospective study analyzed patients who had ultrasound-suspicious thyroid nodules, underwent FNA and cytological examination, and were classified according to the Bethesda system. Patients from Undetermined Significance Or Follicular Lesion Of Undetermined Significance (AUS/FLUS), Follicular Neoplasm or Suspicious for a Follicular Neoplasm (FN/SFN), Suspicious for Malignant Cells (SMC), and Positive for Malignant Cells (PMC) groups were examined for the BRAF mutation and had a thyroid surgery. Demographical and histological features and stage of the disease were evaluated for PTC patients in accordance with the Bethesda category and its association with BRAF mutation. RESULTS: Three hundred eight of all patients underwent operation. One hundred forty-three (46.4%) of them were diagnosed with PTC. In 14 (9.8%) PTC cases, FNA biopsies were classified as AUS/FLUS, 23 (16.1%) as FN/SFN, 41 (28.7%) as SMC, and 65 (45.5%) as PMC. I-II stages of PTC were diagnosed for 88 (61.5%) patients and III-IVA for 55 (38.5%). Patients from the SMC and PMC groups had larger tumors, higher incidence of lymph node metastases, classical PTC type, B-type Raf (BRAF) positive, and III-IVA stage cancer, than patients from the AUS/FLUS and FN/SFN groups. When comparing 27 (18.9%) BRAF-negative patients from the AUS/FLUS and FN/SFN groups with 116 (81.1%) BRAF-negative patients from the SMC and PMC groups and all BRAF-positive patients, the prediction of more aggressive histological features and stage was slightly improved. CONCLUSIONS: Higher Bethesda categories are associated with higher stages of PTC. Association of the Bethesda category with BRAF mutation can slightly improve the value of stage prediction.

Defining the significance of IGF2BP1 overexpression in t(12;21)(p13;q22)-positive leukemia REH cells.

The IGF2 mRNA binding protein 1 (IGF2BP1) belongs to a family of regulatory RNA-binding proteins and controls stability, transport or translation of its target transcripts. Re-expression of IGF2BP1 is frequently found in different tumors and has been associated with aggressive disease phenotypes. IGF2BP1 has also been identified to be exclusively specific for t(12;21)(p13;q22)-positive acute lymphoblastic leukemia (ALL) but biological significance of IGF2BP1 overexpression has not been investigated to date. We have recently reported that ETV6/RUNX1 transcript is a target of RNA-binding protein IGF2BP1 in t(12;21)(p13;q22)-positive ALL suggesting a direct role of IGF2BP1 in ETV6/RUNX1-mediated leukemogenesis. To address this question we have employed stable clones of REH cells - a model cell line of t(12;21)(p13;q22)-positive ALL - with downregulated IGF2BP1 expression. Here we show that downregulation of IGF2BP1 impairs proliferation by attenuating cell cycle progression and increasing the rate of spontaneous cell death. We also provide evidence that downregulation of IGF2BP1 induce reduction of STAT3 mRNA levels and augments sensitivity to STAT3 selective inhibitor S3I-201. These data imply that IGF2BP1 indirectly potentiates ETV6/RUNX1-RAC1-STAT3 signaling axis by sustaining appropriate ETV6/RUNX1 and STAT3 transcript levels in REH cells. Further studies are warranted to specify the role of IGF2BP1 in t(12;21)(p13;q22)-positive ALL.

The value of the repeated examination of BRAF V600E mutation status in diagnostics of papillary thyroid cancer.

INTRODUCTION: Nodular thyroid disease is one of the most frequently diagnosed pathologies of the adult population in iodine-deficient regions. Approximately 30% of thyroid aspirates are classified as nondiagnostic/unsatisfactory or indeterminate. However, patients with indeterminate cytology still undergo surgery. The object of this study was to determine the diagnostic value of re-examining the BRAF V600E mutation in papillary thyroid carcinoma patients. MATERIAL AND METHODS: All patients underwent ultrasound guided fine-needle aspiration of a thyroid nodule. They were assigned to one of the four groups (indeterminate or positive for malignant cells) of the Bethesda System for Reporting Thyroid Cytopathology. Genetic investigation of the BRAF V600E mutation was performed for all of the fine-needle aspiration cytology specimens. All of the patients underwent surgery. Subsequently, histological investigation of the removed tissues was performed. Additional analysis of the BRAF V600E mutation from the histology specimen was then performed for the initially BRAF-negative cases. RESULTS: Two hundred and fourteen patients were involved in the study. One hundred and six (49.53%) patients were diagnosed with thyroid cancer. Of these 106 patients, 95 (89.62%) patients were diagnosed with papillary thyroid cancer. The BRAF V600E mutation was positive in 62 (65.26%) and negative in 33 (34.74%) histologically confirmed papillary thyroid cancer cases. After the genetic investigation, a total of 74 (77.89%) papillary thyroid cancer cases were positive for the BRAF V600E mutation and 21 (22.11%) were negative. CONCLUSIONS: Repeated examination of the BRAF V600E mutation status in the fine-needle aspiration may potentially increase the sensitivity of papillary thyroid cancer diagnostics.

ETV6/RUNX1 transcript is a target of RNA-binding protein IGF2BP1 in t(12;21)(p13;q22)-positive acute lymphoblastic leukemia.

The oncofetal RNA-binding protein IGF2BP1 (IGF2 mRNA binding protein 1) is overexpressed in a subset of cancers and promotes cell cycle, migration and aggressive phenotype by regulating post-transcriptionally a number of key mRNAs (e. g, ACTB, CD44, CTNNB1, KRAS, MAPK4, MYC, PTEN and others). IGF2BP1 is also overexpressed in t(12;21)(p13;q22)-positive acute lymphoblastic leukemia (ALL), but the biological significance of this phenomenon has not been addressed so far. We have identified leukemia fusion gene ETV6/RUNX1 mRNA to be highly enriched in immunoprecipitated fraction of endogenous IGF2BP1 from a model cell line REH and t(12;21)(p13;q22)-positive ALL samples. Furthermore, downregulation of IGF2BP1 by two-fold has resulted in a corresponding decrease of ETV6/RUNX1 mRNA validating this transcript as a target of IGF2BP1 protein in t(12;21)(p13;q22)-positive ALL. These data infer that IGF2BP1 is a potent regulator of ETV6/RUNX1 mRNA stability and potentially link this evolutionary-highly conserved protein to cell transformation events in ETV6/RUNX1-mediated leukemogenesis of t(12;21)(p13;q22)-positive ALL.

Therapeutic Plasma Exchange in Multiple Sclerosis Patients with Abolished Interferon-beta Bioavailability.

BACKGROUND: Neutralizing antibodies (NAb) to interferon-beta (IFN-ß) are associated with reduced bioactivity and efficacy of IFN-ß in multiple sclerosis (MS). The myxovirus resistance protein A (MxA) gene expression is one of the most appropriate markers of biological activity of exogenous IFN-ß. We hypothesized that therapeutic plasma exchange (TPE) can restore the ability of IFN-ß to induce the MxA mRNA expression and that maintenance plasmapheresis can sustain the bioavailability of IFN-ß. MATERIAL AND METHODS: Eligible patients underwent 4 primary separate plasma exchange sessions. After the induction TPE sessions, they were transferred to maintenance plasmapheresis. Bioactivity of IFN-ß was expressed as in vivo MxA mRNA induction in whole blood using RT-qPCR. RESULTS: Six patients with low IFN-ß bioavailability detected by the MxA mRNA response were included. Four patients became biological responders after induction plasmapheresis. In 2 patients an increase of MxA mRNA expression was found, but the values persisted below the cut-off and the patients remained as "poor biological responders". The effect of maintenance plasmapheresis was transient: MxA mRNA expression values reverted to the baseline levels after 1-2 months. CONCLUSIONS: Therapeutic plasma exchange is able to restore the bioavailability of IFN-ß in the majority of studied patients, but the effect of TPE on the IFN-ß bioavailability was transient.

Cure rates of childhood acute lymphoblastic leukemia in Lithuania and the benefit of joining international treatment protocol.

BACKGROUND: Childhood acute lymphoblastic leukemia (ALL) represents the largest group of pediatric malignancies with long-term survival rates of more than 80% achieved in developed countries. Epidemiological data and survival rates of childhood ALL in Lithuania were lacking. Therefore, the aim of this study was to analyze the population-based long-term treatment results of childhood ALL in Lithuania during 1992-2012. MATERIALS AND METHODS: Data of all 459 children with T-lineage and B-cell precursor ALL treated in Lithuania from 1992 to 2012 were collected and analyzed. Results were compared among four time-periods: 1992-1996 (N=132), 1997-2002 (N=136), 2003-2008 (N=109) and 2009-2012 (N=82). RESULTS: The incidence of childhood ALL in Lithuania was 3.2-3.6 cases per 100000 children per year during the study period. Five-year probability of event-free survival increased from 50%± 4% in 1992-1996 to 71%± 4% in 2003-2008 (P<0.001). Five-year cumulative incidence of relapses reduced from 27%± 4.5% in 1992-1996 to 14%± 3.6% in 2003-2008 (P=0.042). After introduction of high-dose methotrexate of 5 g/m(2), cumulative incidence of CNS-involving relapses reduced from 17%± 3.9% in 1992-1996 to 1%± 1.0% in 2003-2008 (P<0.001). Trend for further improvement in survival was seen in 2009-2012 when Lithuania joined international the Nordic Society of Pediatric Hematology and Oncology (NOPHO) ALL-2008 treatment protocol. CONCLUSIONS: Cure rates of childhood ALL in Lithuania are improving steadily and are now approaching those reported by the largest international study groups. The reasons for such a positive effect are both better financial support for treatment of children with cancer in Lithuania and international collaboration with joining international treatment protocol for childhood ALL.

Dose-dense high-dose methylprednisolone and rituximab in the treatment of relapsed or refractory high-risk chronic lymphocytic leukemia.

This study evaluated the efficacy and safety of dose-dense high-dose methylprednisolone (HDMP) plus rituximab (Rtx) in patients with high-risk CLL. Twenty-nine patients with relapsed or progressive CLL with adverse cytogenetics (17p deletion, TP53 mutation, 11q deletion, and/or trisomy 12) and/or progression within 12 months of fludarabine treatment were included. HDMP (1 g/m(2)) was administered daily for 5 days of each treatment course. Rtx was administered on days 1 (375 mg/m(2)) and 5 (500 mg/m(2)) of the first treatment course, on days 1 (500 mg/m(2)) and 5 (500 mg/m(2)) of the second course, and on day 1 (500 mg/m(2)) of courses 3-6. The cycles were repeated every 21 days. The overall response rate (ORR) was 62%, and 28% of patients had stable disease. In 13 patients with 17p deletion/TP53 mutation, ORR was 69%. After 22 months, the median progression-free and overall survivals were 12 and 31 months, respectively. The most frequent toxicity was hyperglycemia, and three deaths occurred in the study. Dose-dense treatment with HDMP and Rtx is an effective therapy with a favorable safety profile in patients with high-risk CLL, including those with 17p deletion/TP53 mutation.

Identification of characteristic IGF2BP expression patterns in distinct B-ALL entities.

Insulin-like growth factor 2 mRNA-binding proteins IGF2BP1, IGF2BP2, and IGF2BP3 have been shown to have diagnostic and prognostic utility in a number of epithelial and soft tissue tumors. Still, little is known about the expression of these molecules in different types of leukemia and our study aims to fill this gap. By using an RT-qPCR approach, we have systemically analyzed the expression of three IGF2BP coding genes in normal hematopoietic tissues and distinct acute lymphoblastic leukemia (ALL) entities. We show that low/negative IGF2BP1 and IGF2BP3 and high IGF2BP2 levels are characteristic to healthy donor bone marrow and peripheral blood whereas different B-ALL entities displayed characteristic perturbations of IGF2BP expression patterns. Namely, we have identified significant associations of overexpressed IGF2BP1 with ETV6/RUNX1-positive (r(2)=0.7891, y=0.8105x-0.4471, p<0.0001), underexpressed IGF2BP2 with E2A/PBX1-positive (p<0.01), and overexpressed IGF2BP2 and IGF2BP3 with MLL/AF4-positive (r(2)=0.6571, y=0.1507x-0.2722, p<0.0001, and r(2)=0.7022, y=0.6482x-0.7660, p<0.0001, respectively) leukemia. Secondly, based on transcript expression dynamics during follow-up, we conclude that overexpression of only IGF2BP1 is inherent characteristic of ETV6/RUNX1-positive leukemic blasts in contrast to IGF2BP3 which remained stably expressed throughout the monitoring period and upon the achievement of molecular remission. Finally, our data suggest that IGF2BP3 might be a marker of disease aggressiveness in BCR/ABL1-positive ALL as consistently increasing levels of this transcript during follow-up predicted eventual leukemia relapse by three months. Altogether, our results highlight the potential utility of IGF2BP profiling in precursor B lymphoid neoplasms as the functions of IGF2BPs in normal and malignant hematopoiesis are further delineated.

Recent advances in quantitative chimerism analysis.

Quantitative chimerism analysis is a diagnostic tool used to monitor engraftment kinetics after allogeneic stem cell transplantation. It reflects the proportion of recipient and donor genotypes and is based on the identification of genetic markers characteristic to a given transplant pair. Currently, PCR amplification of short tandem repeats and single-nucleotide polymorphism-specific quantitative real-time PCR are the most widely used techniques for this purpose. In this review, we will address advances as well as technology-specific imperfections, of both techniques that have emerged over the recent years. We will discuss new principles that may simplify assay design, and improve its robustness and reliability. A better chimerism assay could then guide clinical interventions and may, eventually, improve the outcome of allogeneic stem cell transplantation.

Single nucleotide polymorphism-based system improves the applicability of quantitative PCR for chimerism monitoring.

Recently, several studies demonstrated the feasibility of a real-time quantitative PCR (qPCR) approach for chimerism monitoring. qPCR offers a fast, sensitive, and elegant quantification of genotypes. However, before it becomes an established method for routine chimerism monitoring, a qPCR marker set for every transplant pair should be available. This requirement poses a major challenge since the genetic markers for qPCR--short insertions/deletions (Indels) and single nucleotide polymorphisms (SNPs)--published to-date do not guarantee applicability for every transplant pair. The aim of our study was to design and validate a new SNP allele-specific system to supplement an already existing Indel primer panel and improve applicability of the qPCR approach for chimerism status monitoring. Here, we present an approach for an economical in-house design of SNP allele-specific qPCR primers/probe sets with a locus-individualized reference system that allows for the accurate quantification of the respective informative locus using a simple DeltaDeltaCt method. We designed primers/probe sets specific for seven biallelic SNP loci and validated them in a population of 30 transplant pairs. Repeatability varied depending on the amount of quantifiable genotype. The combination of our SNP-qPCR system and Indel primers increased recipient genotype identification from 86.6% to 96.6% when tested in a population of our transplant pairs. These results demonstrate the feasibility of our SNP-based qPCR approach to improve the applicability of a qPCR for chimerism monitoring.