Carboxymethyl Cellulose-Based Hydrogel Film Combined with Umbilical Cord Blood Platelet gel as an Innovative Tool for Chronic Wound Management: A Pilot Clinical Study.

Treatment of chronic leg ulcers remains a major challenge and it is a substantial financial burden on individuals, families, caregivers, and health care system. There is increasing evidence on using of autologous Platelet-rich-plasma in wound repair but limited clinical data are available on the efficacy and safety of the use of umbilical cord blood platelet gel (CBPG). In our pilot study, for the first time, we aimed to evaluated the safety and efficacy of the use of umbilical CBPG combined with a hydrogel dressing in 10 patients with chronic venous ulcers (VU). The protocol consisted of application of umbilical cord blood platelet-rich plasma (PRP) combined with a Carboxymethyl cellulose (CMC)-based hydrogel dressing once a week for 4 weeks. The 80% of patients after 4 weeks of treatment had a significantly decrease in wound size. Moreover, we obtained an improvement in terms of mean Wound Bed Score (WBS), numeric rating scale (NRS) value and the EQ-5D index score. This pilot study showed that the topically therapeutic administration of umbilical CBPG associated with a CMC-based hydrogel dressing has the potential to accelerate the healing of chronic lesions without adverse reaction. However, additional studies with larger sample size and longer follow-up periods are required to confirm our findings.

Defining TCRγδ lymphoproliferative disorders by combined immunophenotypic and molecular evaluation.

Tγδ large granular lymphocyte leukemia (Tγδ LGLL) is a rare lymphoproliferative disease, scantily described in literature. A deep-analysis, in an initial cohort of 9 Tγδ LGLL compared to 23 healthy controls, shows that Tγδ LGLL dominant clonotypes are mainly public and exhibit different V-(D)-J γ/δ usage between patients with symptomatic and indolent Tγδ neoplasm. Moreover, some clonotypes share the same rearranged sequence. Data obtained in an enlarged cohort (n = 36) indicate the importance of a combined evaluation of immunophenotype and STAT mutational profile for the correct management of patients with Tγδ cell expansions. In fact, we observe an association between Vδ2/Vγ9 clonality and indolent course, while Vδ2/Vγ9 negativity correlates with symptomatic disease. Moreover, the 7 patients with STAT3 mutations have neutropenia and a CD56-/Vδ2- phenotype, and the 3 cases with STAT5B mutations display an asymptomatic clinical course and CD56/Vδ2 expression. All these data indicate that biological characterization is needed for Tγδ-cell neoplasm definition.

Tyrosine Kinase Inhibitors Play an Antiviral Action in Patients Affected by Chronic Myeloid Leukemia: A Possible Model Supporting Their Use in the Fight Against SARS-CoV-2.

SARS-CoV-2 is the viral agent responsible for the pandemic that in the first months of 2020 caused about 400,000 deaths. Among compounds proposed to fight the SARS-CoV-2-related disease (COVID-19), tyrosine kinase inhibitors (TKIs), already effective in Philadelphia-positive acute lymphoblastic leukemia (Ph+ ALL) and chronic myeloid leukemia (CML), have been proposed on the basis of their antiviral action already demonstrated against SARS-CoV-1. Very few cases of COVID-19 have been reported in Ph+ ALL and in CML Italian cohorts; authors suggested that this low rate of infections might depend on the use of TKIs, but the biological causes of this phenomenon remain unknown. In this study, the CML model was used to test if TKIs would sustain or not the viral replication and if they could damage patient immunity. Firstly, the infection and replication rate of torquetenovirus (TTV), whose load is inversely proportional to the host immunological control, have been measured in CML patients receiving nilotinib. A very low percentage of subjects were infected at baseline, and TTV did not replicate or at least showed a low replication rate during the follow-up, with a mean load comparable to the measured one in healthy subjects. Then, after gene expression profiling experiments, we found that several "antiviral" genes, such as CD28 and IFN gamma, were upregulated, while genes with "proviral" action, such as ARG-1, CEACAM1, and FUT4, were less expressed during treatment with imatinib, thus demonstrating that TKIs are not detrimental from the immunological point of view. To sum up, our data could offer some biological explanations to the low COVID-19 occurrence in Ph+ ALL and CML patients and sustain the use of TKIs in COVID-19, as already proposed by several international ongoing studies.

Immunoglobulin kappa deleting element rearrangements are candidate targets for minimal residual disease evaluation in mantle cell lymphoma.

Minimal residual disease (MRD) assessment is of high clinical relevance in patients with mantle cell lymphoma (MCL). In mature B-cell malignancies, the presence of somatic hypermutations (SHM) in Variable-Diversity-Joining Heavy chain (VDJH) rearrangements leads to frequent mismatches between primers, probes, and the target, thus impairing tumor cells quantification. Alternative targets, such as immunoglobulin kappa-deleting-element (IGK-Kde) rearrangements, might be suitable for MRD detection. We aimed at evaluating the applicability of IGK-Kde rearrangements for MRD quantification in MCL patients by real-time quantitative polymerase chain reaction (RQ-PCR)/digital-droplet-PCR (ddPCR). IGK screening was performed on bone marrow samples from two cohorts: the first from Turin (22 patients enrolled in the FIL-MCL0208 trial, NCT02354313) and the second from Rome (15 patients). IGK-Kde rearrangements were found in 76% (28/37) of cases, representing the sole molecular marker in 73% (8/11) of IGH-BCL1/IGH negative cases. MRD RQ-PCR monitoring was possible in 57% (16/28) of cases, showing a 100% concordance with the conventional targets. However, the frequent background amplification affected the sensitivity of the assay, that was lower in MCL compared to acute lymphoblastic leukemia and in line with multiple myeloma published results. ddPCR had a good concordance with RQ-PCR and it might help to identify false positive/negative results. From a clinical perspective, we suggest that IGK-Kde can be a candidate target for MRD monitoring and deserves a validation of its predictive value in prospective MCL series.

Digital Droplet PCR is a Specific and Sensitive Tool for Detecting IDH2 Mutations in Acute Myeloid LeuKemia Patients.

Isocitrate dehydrogenase 1 and 2 (IDH1 and IDH2) interfere with cellular metabolism contributing to oncogenesis. Mutations of IDH2 at R140 and R172 residues are observed in 20% of acute myeloid leukemias (AML), and the availability of the IDH2 inhibitor Enasidenib made IDH2 mutational screening a clinical need. The aim of this study was to set a new quantitative polymerase chain reaction (PCR) technique, the drop-off digital droplet PCR (drop-off ddPCR), as a sensitive and accurate tool for detecting IDH2 mutations. With this technique we tested 60 AML patients. Sanger sequencing identified 8/60 (13.5%) mutated cases, while ddPCR and the amplification refractory mutation system (ARMS) PCR, used as a reference technique, identified mutations in 13/60 (21.6%) cases. When the outcome of IDH2-mutated was compared to that of wild-type patients, no significant difference in terms of quality of response, overall survival, or progression-free survival was observed. Finally, we monitored IDH2 mutations during follow-up in nine cases, finding that IDH2 can be considered a valid marker of minimal residual disease (MRD) in 2/3 of our patients. In conclusion, a rapid screening of IDH2 mutations is now a clinical need well satisfied by ddPCR, but the role of IDH2 as a marker for MRD still remains a matter of debate.

Droplet Digital PCR Quantification of Mantle Cell Lymphoma Follow-up Samples From Four Prospective Trials of the European MCL Network.

Minimal residual disease (MRD) has been increasingly investigated in mantle cell lymphoma (MCL), including for individual therapeutic stratification and pre-emptive treatment in clinical trials. Although patient/allele specific real-time quantitative polymerase chain reaction (qPCR) of IGH or BCL1-IGH clonal markers is the gold-standard method, its reliance on a standard curve for relative quantification limits quantification of low-level positivity within the 1E-4 to 1E-5 range; over half of positive MRD samples after treatment fall below the quantitative range (BQR) of the standard curve. Droplet digital PCR (ddPCR), in contrast, allows absolute quantification, including for samples with no baseline determination of tumor infiltration by multicolor flow cytometry (MFC), avoiding the need for a reference standard curve. Using updated, optimized, ddPCR criteria we compared it with qPCR in 416 MRD samples (and with MFC in 63), with over-representation (61%) of BQR results by qPCR, from a total of 166 patients from four prospective MCL clinical trials. ddPCR, qPCR and MFC gave comparable results in MRD samples with at least 0.01% (1E-4) positivity. ddPCR was preferable to qPCR since it provided more robust quantification at positivity between 1E-4 and 1E-5. Amongst 240 BQR samples with duplicate or triplicate analysis, 39% were positive by ddPCR, 49% negative and only 12% remained positive below quantifiable ddPCR limits. The prognostic relevance of ddPCR is currently under assessment in the context of prospective trials within the European MCL Network.

Minimal residual disease (MRD) in non-Hodgkin lymphomas: Interlaboratory reproducibility on marrow samples with very low levels of disease within the FIL (Fondazione Italiana Linfomi) MRD Network.

In 2009, the four laboratories of the Fondazione Italiana Linfomi (FIL) minimal residual disease (MRD) Network started a collaborative effort to harmonize and standardize their methodologies at the national level, performing quality control (QC) rounds for follicular lymphoma (FL) and mantle cell lymphoma (MCL) MRD assessment. In 16 QC rounds between 2010 and 2017, the four laboratories received 208 bone marrow (BM) samples (126 FL; 82 MCL); 187 were analyzed, according to the EuroMRD Consortium guidelines, by both nested (NEST) polymerase chain reaction (PCR) and real-time quantitative (RQ) PCR for BCL2/IGH MBR or IGHV rearrangements. Here, we aimed at analyzing the samples that challenged the interlaboratory reproducibility and data interpretation. Overall, 156/187 BM samples (83%) were concordantly classified as NEST+/RQ+ or NEST-/RQ- by all the four laboratories. The remaining 31 samples (17%) resulted alternatively positive and negative in the interlaboratory evaluations, independently of the method and the type of rearrangement, and were defined "borderline" (brd) samples: 12 proved NEST brd/RQ brd, 7 NEST-/RQ brd, 10 NEST brd/RQ positive not quantifiable (PNQ), and 2 NEST brd/RQ-. Results did not change even increasing the number of replicates/sample. In 6/31 brd samples, droplet digital PCR (ddPCR) was tested and showed no interlaboratory discordance. Despite the high interlaboratory reproducibility in the MRD analysis obtained and maintained by the QC round strategy, samples with the lowest MRD levels can still represent a challenge: 17% (31/187) of our samples showed discordant results in interlaboratory assessments, with 6.4% (12/187) remained brd even applying the two methods. Thus, although representing a minority, brd samples are still problematic, especially when a clinically oriented interpretation of MRD results is required. Alternative, novel methods such as ddPCR and next-generation sequencing have the potential to overcome the current limitations.

The Minimal Residual Disease in Non-Hodgkin's Lymphomas: From the Laboratory to the Clinical Practice.

Minimal residual disease (MRD) in non-Hodgkin's lymphomas (NHLs) still represents matter of interest and debate: indeed, the new available treatments offer higher rates of complete responses and MRD negativity than in the past, with a positive impact on the long-term survival. Furthermore, the introduction of more sensitive and accurate molecular techniques, such as digital PCR (ddPCR) and the next generation sequencing techniques (NGS), increased the possibility of identifying molecular targets to be followed after therapy (such as rearrangement of immunoglobulins, fusion genes, or mutations). This review focused on how molecular biology can help to detect MRD in different types of NHLs and how MRD can change the clinical practice in 2019. In follicular lymphoma (FL), contamination of the grafts and molecular disease persistence after transplantation represent a negative prognostic factors. The combination of Rituximab or Obinutuzumab with Bendamustine seems to be the most effective way to clear MRD in FL patients receiving chemo-immunotherapy (further studies are in progress), and also 90Yttrium-Ibritumomab-Tiuxetan offers a deep clearance of molecular disease. Finally, molecular MRD can further stratify PET-negative cases, with subjects both PET- and MRD-negative presenting the best outcome. In aggressive lymphomas, MRD has a relevant prognostic power and can represent the platform for immunotherapy (such as CAR-T). In diffuse large B-cell lymphoma (DLBCL), the assessment of MRD in the plasma (where cell-free DNA and exosomes circulate) seems to be more predictive than the bone marrow analysis or peripheral blood mononuclear cells. Finally, NGS technologies could be more useful than the classical "patient allele-specific PCR" because they can identify any possible clone emerging during the treatment or follow-up, even if different from that identified at diagnosis, thus predicting relapse. After all, the present available molecular approaches can move MRD from the bench side to the clinical practice.

The WNT Pathway Is Relevant for the BCR-ABL1-Independent Resistance in Chronic Myeloid Leukemia.

Notwithstanding the introduction of Tyrosine Kinase Inhibitors (TKIs) revolutionized the outcome of Chronic Myeloid Leukemia (CML), one third of patients still suspends treatment for failure response. Recent research demonstrated that several BCR/ABL1-independent mechanisms can sustain resistance, but the relationship between these mechanisms and the outcome has not yet been fully understood. This study was designed to evaluate in a "real-life" setting if a change of expression of several genes involved in the WNT/BETA-CATENIN, JAK-STAT, and POLYCOMB pathways might condition the outcome of CML patients receiving TKIs. Thus, the expression of 255 genes, related to the aforementioned pathways, was measured by quantitative PCR after 6 months of therapy and compared with levels observed at diagnosis in 11 CML patients, in order to find possible correlations with quality of response to treatment and event-free-survival (EFS). These results were then re-analyzed by the principal component method (PCA) for tempting to better cluster resistant cases. After 12 months of therapy, 6 patients achieved an optimal response and 5 were "resistant;" after application of both statistical methods, it was evident that in all pathways a significant overall up-regulation occurred, and that WNT was the pathway mostly responsible for the TKIs resistance. Indeed, 100% of patients with a "low" up-regulation of this pathway achieved an optimal response vs. 33% of those who showed a "high" gene over-expression (p = 0.016). Analogously, the 24-months EFS resulted significantly influenced by the degree of up-regulation of the WNT signaling: all patients with a "low" up-regulation were event-free vs. 33% of those who presented a "high" gene expression (p = 0.05). In particular, the PCA analysis confirmed the role of WNT pathway and showed that the most significantly up-regulated genes with negative prognostic value were DKK, WNT6, WISP1, and FZD8. In conclusion, our results sustain the need of a wide and multitasking approach in order to understand the resistance mechanisms in CML.

The assessment of minimal residual disease versus that of somatic mutations for predicting the outcome of acute myeloid leukemia patients.

BACKGROUND: In addition to morphological and cytogenetic features, acute myeloid leukemias are characterized by mutations that can be used for target-therapy; also the minimal/measurable residual disease (MRD) could be an important prognostic factor. The purpose of this retrospective study was to investigate if somatic mutations could represent an additional prognostic value in respect of MRD alone. METHOD: At baseline, 98 patients were tested for NPM1, FLT3, and for WT1 expression; 31 for ASXL1, TET2, IDH1, IDH2, N-RAS, WT1, c-KIT, RUNX1, and DNMT3A. The same genes have been also tested after induction and consolidation. RESULTS: Overall, 60.2% of our patients resulted mutated: 24.5% carried mutations of FLT3-ITD, 38.7% of NPM1, 48.4% of c-KIT, 25.8% of N-RAS and 19.3% of IDH2. The probability of achieving a complete response (CR) was higher for younger patients, with low ELN risk score, NPM1-mutated, with low WT1 levels, and without FLT3. The presence of additional mutations represented a poor predictive factor: only 19% of these cases achieved CR in comparison to 43% of subjects without any of it. Concerning survival, it was conditioned by a lower ELN risk score, younger age, reduction > 1 log of the NPM1 mutational burden, disappearance of FLT3 mutations and lower WT1 expression. Regarding the role of the additional mutations, they impaired the outcome of 20% of the already MRD-negative patients. Concerning the possibility of predicting relapse, we observed an increase of the NPM1 mutational burden at the time-point immediately preceding the relapse (about 2 months earlier) in 50% of subjects. Similarly concerning WT1, an increase of its expression anticipated disease recurrence in 64% of cases. CONCLUSIONS: We demonstrated that additional somatic mutations are able to impair outcome of the already MRD-negative subjects. About MRD, we suggest a prognostic role also for the WT1 expression. Finally, we considered as relevant the assessment of NPM1 quantity clearance instead of the presence/absence of mutations alone. Still remains in doubt the utility in terms of long-term prognosis of a baseline more complex mutational screening; we could hypothesize that it would be useful for those patients where other markers are not available or who reached the MRD negativity.

The Polycomb BMI1 Protein Is Co-expressed With CD26+ in Leukemic Stem Cells of Chronic Myeloid Leukemia.

The Polycomb gene BMI1 expression exerts a negative predictive impact on several hematological malignancies, such as acute and chronic myeloid leukemia (CML), myelofibrosis, and follicular lymphoma. As already demonstrated in CML, BMI1 is responsible for the resistance to the tyrosine kinase inhibitors (TKIs) in a BCR-ABL1-independent way. Even if, it is unknown where BMI1 in CML is expressed (in progenitors or more mature cells). We decided, therefore, to evaluate if and where the BMI1 protein is located, focusing mainly on the CD34+/CD38-/CD26+ CML progenitors. To begin we measured, by flow cytometry, the proportion of CD34+/CD26+ cells in 31 bone marrow samples from 20 CML patients, at diagnosis and during treatment with imatinib. After that the bone marrow blood smears were stained with antibodies anti-CD26, BCR-ABL1, and BMI1. These smears were observed by a confocal laser microscope and a 3D reconstruction was then performed. At diagnosis, CD34+/CD26+ cells median value/µL was 0.48; this number increased from diagnosis to the third month of therapy and then reduced during treatment with imatinib. The number and behavior of the CD26+ progenitors were independent from the BCR-ABL1 expression, but they summed up what previously observed about the BMI1 expression modulation. In this work we demonstrate for the first time that in CML the BMI1 protein is co-expressed with BCR-ABL1 only in the cytoplasm of the CD26+ precursors; on the contrary, in other hematological malignancies where BMI1 is commonly expressed (follicular lymphoma, essential thrombocytemia, acute myeloid leukemia), it was not co-localized with CD26 or, obviously, with BCR-ABL1. Once translated into the clinical context, if BMI1 is a marker of stemness, our results would suggest the combination of the BMI1 inhibitors with TKIs as an interesting object of research, and, probably, as a promising way to overcome resistance in CML patients.

The hOCT1 and ABCB1 polymorphisms do not influence the pharmacodynamics of nilotinib in chronic myeloid leukemia.

First-line nilotinib in chronic myeloid leukemia is more effective than imatinib to achieve early and deep molecular responses, despite poor tolerability or failure observed in one-third of patients. The toxicity and efficacy of tyrosine kinase inhibitors might depend on the activity of transmembrane transporters. However, the impact of transporters genes polymorphisms in nilotinib setting is still debated. We investigated the possible correlation between single nucleotide polymorphisms of hOCT1 (rs683369 [c.480C>G]) and ABCB1 (rs1128503 [c.1236C>T], rs2032582 [c.2677G>T/A], rs1045642 [c.3435C>T]) and nilotinib efficacy and toxicity in a cohort of 78 patients affected by chronic myeloid leukemia in the context of current clinical practice. The early molecular response was achieved by 81% of patients while 64% of them attained deep molecular response (median time, 26 months). The 36-month event-free survival was 86%, whereas 58% of patients experienced toxicities. Interestingly, hOCT1 and ABCB1 polymorphisms alone or in combination did not influence event-free survival or the adverse events rate. Therefore, in contrast to data obtained in patients treated with imatinib, hOCT1 and ABCB1 polymorphisms do not impact on nilotinib efficacy or toxicity. This could be relevant in the choice of the first-line therapy: patients with polymorphisms that negatively condition imatinib efficacy might thus receive nilotinib as first-line therapy.

Myelodysplastic syndromes: advantages of a combined cytogenetic and molecular diagnostic workup.

In this study we present a new diagnostic workup for the myelodysplastic syndromes (MDS) including FISH, aCGH, and somatic mutation assays in addition to the conventional cytogenetics (CC). We analyzed 61 patients by CC, FISH for chromosome 5, 7, 8 and PDGFR rearrangements, aCGH, and PCR for ASXL1, EZH2, TP53, TET2, RUNX1, DNMT3A, SF3B1 somatic mutations. Moreover, we quantified WT1 and RPS14 gene expression levels, in order to find their possible adjunctive value and their possible clinical impact. CC analysis showed 32% of patients with at least one aberration. FISH analysis detected chromosomal aberrations in 24% of patients and recovered 5 cases (13.5%) at normal karyotype (two 5q- syndromes, one del(7) case, two cases with PDGFR rearrangement). The aGCH detected 10 "new" unbalanced cases in respect of the CC, including one with alteration of the ETV6 gene. After mutational analysis, 33 patients (54%) presented at least one mutation and represented the only marker of clonality in 36% of all patients. The statistical analysis confirmed the prognostic role of CC either on overall or on progression-free-survival. In addition, deletions detected by aCGH and WT1 over-expression negatively conditioned survival. In conclusion, our work showed that 1) the addition of FISH (at least for chr. 5 and 7) can improve the definition of the risk score; 2) mutational analysis, especially for the TP53 and SF3B1, could better define the type of MDS and represent a "clinical warning"; 3) the aCGH use could be probably applied to selected cases (with suboptimal response or failure).

GTF2I Mutations Are Common in Thymic Epithelial Tumors But Not in Hematological Malignancies.

BACKGROUND: Mutation of general transcription factor IIi (GTF2I) (chromosome 7 c.74146970T>A) is common in thymic epithelial tumors and is a candidate driver aberration for cancer growth. To our knowledge, this mutation has not been described in other diseases. We evaluated the presence of GTF2I mutation in hematological malignancies. MATERIALS AND METHODS: We sequenced samples from 31 patients with acute leukemia, 29 with chronic leukemia and 12 with myelodysplastic syndrome. The genomic fragment of exon 15 containing the hotspot of mutation was amplified using polymerase chain reaction (PCR) and sequenced. RESULTS: We did not identify any GTF2I mutation in patients with hematological malignancies. CONCLUSION: Even though our sample size was limited, our data and reports from the literature suggest that GTF2I mutation is not present or is uncommon in these diseases.

The Combination of Rituximab and Bendamustine as First-Line Treatment Is Highly Effective in the Eradicating Minimal Residual Disease in Follicular Lymphoma: An Italian Retrospective Study.

R-Bendamustine is an effective treatment for follicular lymphoma (FL). Previous large trials demonstrated the prognostic role of the molecular minimal residual disease (MRD) during the most frequently adopted chemotherapeutic regimens, but there are not yet conclusive data about the effect of combination of rituximab (R) and bendamustine in terms of MRD clearance. Thus, the aim of this retrospective study was to assess if and in what extent the combination of rituximab and bendamustine would exert a significant reduction of the molecular disease in 48 previously untreated FL patients. The molecular marker at baseline was found in the 62.5% of cases; no significant differences were observed between patients with or without the molecular marker in respect of the main clinical features. Moreover, the quantization of the baseline molecular tumor burden showed a great variability: the median value was 1.4 × 10-2 copies, ranging from 3 × 10-5 to 4 × 104. The initial molecular tumor burden did not correlate with clinical features and did not impact on the subsequent quality of response. After treatment, 93% of cases became MRD-negative; the median reduction of the BCL2/JH load was 4 logs. The 2-years PFS was 85%; it was significantly longer for patients in complete than for those in partial response (91 vs. 57%; p = 0.002), and for cases with lower FLIPI-2 score (88 vs. 60%; p = 0.004). On the contrary, PFS did not differ between patients with or without the molecular marker at baseline; a molecular tumor burden 15 times higher was observed in the relapsed subgroup in comparison to the relapse-free one, but this difference did not change the PFS length. The 2-years OS was 93.6%; the only variable that significantly impacted on it was the FLIPI-2 score; the presence of the molecular marker at baseline or its behavior after treatment did not impact on survival. This study, even if retrospective and conducted on a small series of patients, would represent a proof of concept that R-bendamustine is able to so efficaciously eradicate MRD that it could be able to by-pass the prognostic significance of MRD already demonstrated for other chemotherapeutic regimens in FL.

Association of the hOCT1/ABCB1 genotype with efficacy and tolerability of imatinib in patients affected by chronic myeloid leukemia.

PURPOSE: The present study was aimed at investigating whether imatinib pharmacogenetics is related to its pharmacodynamics in patients affected by chronic myeloid leukemia. METHODS: Through a procedure based on a sequence of classical statistics methods, we investigated the possible relationships between treatment efficacy/tolerability and combinations of time-independent variables as gender and genetic covariates in the form of single nucleotide polymorphisms (SNPs) or combinations thereof. Moreover, since the drug tolerability has a strong incidence on the discontinuation of the therapy, we investigated whether the time of manifestation of the most frequent toxic effects can be related to time-independent patients' characteristics or not. RESULTS: We found that a combination of two polymorphisms, namely hOCT1 c.480C>G (rs683369) and ABCB1 c.3435C>T (rs1045642), seems to play the role of predictor for imatinib in both efficacy and toxicity. Furthermore, the time of manifestation of edema toxicity is found to be associated to a combination of gender and ABCB1 c.3435C>T, whereas the time of manifestation of cramp toxicity appears related to gender. CONCLUSIONS: The novelty of this study is dual: the achievement of results that potentially have a significant clinical interest and the demonstration that the adoption of composed covariates may represent a unique tool to study different aspects of the treatment with imatinib.