Agnieszka Janus

Università Vita-Salute San Raffaele, Milano, Lombardy, Italy

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Publications (18)76.84 Total impact

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    ABSTRACT: The differences in clinical course of chronic lymphocytic leukemia could have an impact on variations in a patient's response to therapy. Our published results revealed that thermal transition (95 ± 5°C) in differential scanning calorimetry profiles appear to be characteristic for the advanced stage of CLL. Moreover, a decrease/loss of this transition in nuclei from leukemic cells exposed to drugs ex vivo could indicate their diverse efficacy. It seems that the lack of changes in thermal profile could predict patient's drug resistance. In this study, we demonstrate the results obtained after drug treatment of leukemic cells by calorimetry, apoptosis-related parameters involved in expression of genes using cDNA microarray and western blot. These data were compared with the patients' clinical parameters before and after RCC therapy (rituximab + cladribine + cyclophosphamide). The complementary analysis of studied cases with opposite clinical response (CR or NR) revealed a strong relationship between clinical data, differences in thermal scans and apoptosis-related gene expression. We quantified expression of eight of apoptosis-related 89 genes, i.e., NOXA, PUMA, APAF1, ESRRBL1, CASP3, BCL2, BCL2A1 and MCL1. Particular differences in NOXA and BCL2 expression were revealed. NOXA expression in cells of patients who achieved a complete response to RCC therapy was 0.44 times higher in comparison to control ones. Interestingly, in the case of patients who did not respond to immunotherapy, NOXA expression was highly downregulated (RQ = 4.39) as compared with untreated cells. These results were confirmed by distinct cell viability, protein expression as well as by differences in calorimetry profiles.
    Cancer biology & therapy 10/2012; 14(1). · 3.29 Impact Factor
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    ABSTRACT: The study was aimed to investigate modifications of apoptotic gene expression profile by microarray technique in 10 patients with chronic lymphocytic leukemia by treatment with rituximab, cladribine and cyclophosphamide (RCC) according to IGHV mutational status. The TaqMan Low Density Array for 96 gene transcripts was used. Those modifications followed two distinctive patterns largely overlapping the IGHV mutational status. In the IGHV-mutated group, the expression of many proapoptotic genes increased after treatment as compared to initial value. Our results suggest that RCC drugs may act through influence on the expression of some apoptosis-involved genes dependently on the IGVH mutational status.
    Leukemia research 05/2012; 36(9):1134-40. · 2.36 Impact Factor
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    ABSTRACT: Mounting evidence indicates that grouping of chronic lymphocytic leukemia (CLL) into distinct subsets with stereotyped BCRs is functionally and prognostically relevant. However, several issues need revisiting, including the criteria for identification of BCR stereotypy and its actual frequency as well as the identification of "CLL-biased" features in BCR Ig stereotypes. To this end, we examined 7596 Ig VH (IGHV-IGHD-IGHJ) sequences from 7424 CLL patients, 3 times the size of the largest published series, with an updated version of our purpose-built clustering algorithm. We document that CLL may be subdivided into 2 distinct categories: one with stereotyped and the other with nonstereotyped BCRs, at an approximate ratio of 1:2, and provide evidence suggesting a different ontogeny for these 2 categories. We also show that subset-defining sequence patterns in CLL differ from those underlying BCR stereotypy in other B-cell malignancies. Notably, 19 major subsets contained from 20 to 213 sequences each, collectively accounting for 943 sequences or one-eighth of the cohort. Hence, this compartmentalized examination of VH sequences may pave the way toward a molecular classification of CLL with implications for targeted therapeutic interventions, applicable to a significant number of patients assigned to the same subset.
    Blood 03/2012; 119(19):4467-75. · 9.78 Impact Factor
  • Blood. 2012 Mar 13. 03/2012;
  • Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K 11/2011; 26(5):1127-31. · 10.16 Impact Factor
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    ABSTRACT: Malignant B lymphocytes from chronic lymphocytic leukemia (CLL) patients maintain the capacity to respond to CD40 ligation, among other microenvironmental stimuli. In this study, we show that (i) leukemic CLL cells stimulated with the soluble form of CD40L in vitro show differential responses in terms of upregulation of surface markers (CD95 and CD80) and induction of chemokines (CCL22 and CCL17) expression/secretion, and that (ii) these changes are mirrored by a distinct activation of intracellular signalling pathways including increase in IKKalpha/beta phosphorylation and upregulation of antiapoptotic proteins (BCL-2 and MCL-1). CLL patients can then be segregated into two distinct functional subsets. We defined the responsive subset of cases CD40L dependent, considering the capacity to respond as a sign of persistent need of this stimulation for the leukemic expansion. Conversely, we named the unresponsive cases CD40L independent, considering them less dependent on this microenvironmental signal, presumably because of a higher autonomous proliferative and survival potential. Importantly, we report that (iii) the two functional subsets show an opposite clinical outcome, with CD40L-independent cases having a shorter time to progression. This indicates that the functional differences observed in vitro may reflect a different leukemic potential in vivo likely responsible for a distinct clinical course.
    Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K 11/2011; 25(11):1794. · 10.16 Impact Factor
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    ABSTRACT: Monoclonal B-cell lymphocytosis (MBL) is classified as chronic lymphocytic leukemia (CLL)-like, atypical CLL, and CD5(-) MBL. The number of B cells per microliter divides CLL-like MBL into MBL associated with lymphocytosis (usually detected in a clinical setting) and low-count MBL detected in the general population (usually identified during population screening). After a median follow-up of 34 months we reevaluated 76 low-count MBLs with 5-color flow cytometry: 90% of CLL-like MBL but only 44.4% atypical CLL and 66.7% CD5(-) MBL persisted over time. Population-screening CLL-like MBL had no relevant cell count change, and none developed an overt leukemia. In 50% of the cases FISH showed CLL-related chromosomal abnormalities, including monoallelic or biallelic 13q deletions (43.8%), trisomy 12 (1 case), and 17p deletions (2 cases). The analysis of the T-cell receptor β (TRBV) chains repertoire showed the presence of monoclonal T-cell clones, especially among CD4(high)CD8(low), CD8(high)CD4(low) T cells. TRBV2 and TRBV8 were the most frequently expressed genes. This study indicates that (1) the risk of progression into CLL for low-count population-screening CLL-like MBL is exceedingly rare and definitely lower than that of clinical MBL and (2) chromosomal abnormalities occur early in the natural history and are possibly associated with the appearance of the typical phenotype.
    Blood 08/2011; 118(25):6618-25. · 9.78 Impact Factor
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    ABSTRACT: Malignant B lymphocytes from chronic lymphocytic leukemia (CLL) patients maintain the capacity to respond to CD40 ligation, among other microenvironmental stimuli. In this study, we show that (i) leukemic CLL cells stimulated with the soluble form of CD40L in vitro show differential responses in terms of upregulation of surface markers (CD95 and CD80) and induction of chemokines (CCL22 and CCL17) expression/secretion, and that (ii) these changes are mirrored by a distinct activation of intracellular signalling pathways including increase in IKKalpha/beta phosphorylation and upregulation of antiapoptotic proteins (BCL-2 and MCL-1). CLL patients can then be segregated into two distinct functional subsets. We defined the responsive subset of cases CD40L dependent, considering the capacity to respond as a sign of persistent need of this stimulation for the leukemic expansion. Conversely, we named the unresponsive cases CD40L independent, considering them less dependent on this microenvironmental signal, presumably because of a higher autonomous proliferative and survival potential. Importantly, we report that (iii) the two functional subsets show an opposite clinical outcome, with CD40L-independent cases having a shorter time to progression. This indicates that the functional differences observed in vitro may reflect a different leukemic potential in vivo likely responsible for a distinct clinical course.
    Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K 06/2011; 25(11):1760-7. · 10.16 Impact Factor
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    Leukemia 01/2011; · 10.16 Impact Factor
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    ABSTRACT: The mammalian target of rapamycin (mTOR) kinase is a key regulator of cell growth and proliferation. Overexpression of the mTOR signaling pathway has been described in several tumor cells, including the majority of acute myeloid leukemia (AML) cases. The anti-tumor efficacy of mTOR inhibitors was shown in several preclinical and clinical studies. In AML, however, the potential antineoplastic effect of mTOR inhibitors has received little attention thus far. In this in-vitro study of the human AML cell line, HL-60, we aimed to assess the antileukemic activity of rapamycin (RAPA), an mTOR inhibitor, alone and in combination with cytarabine (Ara-C). The study showed that RAPA in concentrations of 1-10 nmol/l arrested the cell cycle progression of Hl-60 cells in the G1 phase, without evident cytotoxic effect. This effect was associated with significant inhibition of cyclin E expression. At concentrations higher than 10 nmol/l, RAPA exerted a significant proapoptotic effect, with the collapse of mitochondrial potential and caspase-3 activation. The most prominent proapoptotic effect was observed for a combination of 1 nmol/l of RAPA and 50 nmol/l of Ara-C, especially when Ara-C was added at a 24-h interval after RAPA. In conclusion, these data indicate that RAPA might be effective in the treatment of acute leukemia patients, especially in combination with Ara-C, the drug routinely used in AML treatment. On the basis of these results, attempts to combine classical induction chemotherapy with an inhibitor of the mTOR kinase in AML treatment could be warranted.
    Anti-cancer drugs 10/2009; 20(8):693-701. · 2.23 Impact Factor
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    ABSTRACT: The aim of this study was to determine the feasibility, efficacy and toxicity of the combined therapy consisting of cladribine (2-CdA), mitoxantrone and cyclophosphamide (CMC regimen) in patients with refractory or relapsed non-Hodgkin's lymphoma (NHL). Thirty six patients, 14 with mantle cell lymphoma (MCL), 10 with diffuse large B-cell lymphoma (DLBCL), 5 with follicular lymphoma (FL), 3 with small lymphocytic lymphoma (SLL), and 4 with T-cell lymphoma were enrolled to the study. The CMC protocol consisted of 2-CdA at a dose of 0.12 mg/kg in a 2-hour infusion on days 1 through 3, mitoxantrone 10 mg/m(2) i.v. on day 1 and cyclophosphamide 650 mg/m(2) i.v. on day 1. The CMC courses were repeated at intervals of 4 weeks. Thirty three patients were available for evaluation of response. Overall response rate (OR) was 58% (95% CI, 41--75%). Seven patients (21%; 95% CI, 7--35%) achieved a complete response (CR) and 12 patients (36%; 95% CI, 20--52%) achieved a partial response (PR). Seven of 19 patients with CR/PR are still in remission with a median follow-up of 3 months (range, 2-17 months). The median failure-free survival (FFS) was 5 months (range, 2-17 months). The median overall survival (OS) for the entire group was 9 months (range, 0.1-7 months). There was a significant difference in OS between responders and nonresponders after CMC therapy (log rank test, P = 0.015). When different disease status before CMC treatment was considered, a trend toward longer survival of recurrent patients was observed (log rank test, P = 0.08). Grade 3-4 neutropenia developed in 14 (39%) patients, and 16 episodes (15%) of grade 3-4 infections were observed. Grade 3-4 thrombocytopenia or anemia was seen in 9 patients (25%) and 10 patients (28%), respectively. The results of our study show that the CMC regimen is effective salvage therapy with acceptable toxicity in heavily pretreated patients with NHL including MCL and DLBCL.
    Leukemia and Lymphoma 07/2007; 48(6):1092-101. · 2.61 Impact Factor
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    ABSTRACT: Inherited differences in xenobiotic transport and metabolism may play an important role in the development of adult acute myeloid leukemia (AML) and response to the chemotherapy. An ATP-binding cassette (ABC) family transporter P-glycoprotein (P-gp or ABCB1), encoded by ABCB1 (MDR1) gene, is involved in the protection against xenobiotics and multi-drug resistance. The aim of this study was to investigate the potential involvement of the ABCB1 gene exon 26 3435C>T single nucleotide polymorphism (SNP) in the genetic susceptibility to AML and regulation of P-gp expression and activity in AML cells. A total of 180 adult AML patients and 180 sex-matched controls were genotyped using PCR-RFLP method. Moreover, in 40 AML patients ABCB1 gene expression was studied by real-time RT-PCR and P-gp expression and activity were assessed by flow cytometry assays. The prevalence of 3435C>T ABCB1 polymorphism was similar in patient and control cohorts (P = 0.16). Furthermore, the carriers of different ABCB1 genotypes did not differ significantly according to ABCB1 gene expression (P = 0.99), P-gp expression (P = 0.42) and P-gp activity (P = 0.83) in leukemic cells. The authors conclude that isolated 3435C>T ABCB1 SNP is not a major factor of the genetic susceptibility to adult AML, and that genotyping of this polymorphism does not allow predicting P-gp expression or activity in AML cells.
    Therapeutic Drug Monitoring 11/2006; 28(5):707-11. · 2.23 Impact Factor
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    ABSTRACT: Combination of vincristine, doxorubicin and dexamethasone (VAD regimen) is a widely-used, effective and economic initial treatment of multiple myeloma (MM). However, between 30 and 50% of MM patients do not respond to VAD, and exposure to VAD may induce multi-drug resistance. Therefore, in the era of emerging novel therapies for MM, there is a need for pre-treatment identification of potential responders to VAD. Importantly, all three VAD components have common steps in their pathways of transport and metabolism, thus even modest inherited alteration caused by single nucleotide polymorphisms (SNP) may influence VAD efficacy an toxicity. The aim of this study was to investigate whether analysis of common functional SNP in drug transporter P-glycoprotein gene (MDR1 or ABCB1), phase I cytochrome P450A metabolizing enzymes genes (CYP3A4 and CYP3A5), phase II glutathione S-transferase P metabolizing enzyme gene (GSTP1) as well as in glucocorticoid receptor gene NR3C1 can be useful to predict response to VAD in MM. Fifty-two uniformly treated MM patients were investigated for 8 common SNPs including MDR1 exon 12 C1236T, MDR1 exon 21 G2677T/A, MDR1 exon 26 C3435T, CYP3A4*1B, CYP3A5*3, GSTP1 A313G as well as NR3C1 bcl I and NR3C1 N363S polymorphisms. Genotypes were identified using direct sequencing or RFLP methods. The response to VAD was estimated after administration of 3 courses of VAD, and patients who achieved complete or partial remission of MM were assigned as responders to VAD. In the cohort of included MM patients there were 39/52 (75%) responders to VAD. The analysis of the prognostic impact of the genotyped SNPs showed that carriers of P-glycoprotein gene MDR1 2677G allele had a significantly greater probability of achievement of the response to VAD as compared to the non-carriers, odds ratio(OR)=8.0, 95% confidence interval (95%CI)= 1.2–52.9, Fisher exact test p=0.031). Moreover, the predictive effect of MDR1 2677G allele was further increased by the presence of glucocorticoid receptor NR3C1 gene 363N allele. The carriers of both MDR1 2677G and NR3C1 363N alleles had 8.4-fold greater probability to achieve complete or partial remission with 3 courses of VAD (OR=8.40, 95% CI=1.8–40.0, p=0.008). Taken together, these data support the hypothesis that analysis of common functional polymorphisms in drug transporters, metabolizing enzymes and receptors may be useful for individualization of the initial therapy of MM. Particularly MDR1 G2677T/A and NR3C1 N363S SNPs can be considered as determinants of response to VAD therapy.
    47th Congress of the American Society of Hematology, Atlanta, USA; 12/2005
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    XL Zjazd Polskiego Towarzystwa Biochemicznego, Lublin, Polska; 09/2005
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    10th Congress of the European Hematology Association, Stockholm, Sweden; 06/2005
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    Agnieszka Janus, Tadeusz Robak, Piotr Smolewski
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    ABSTRACT: The mammalian target of rapamycin (mTOR) is a kinase responsible for mitogen-induced cell proliferation/survival signaling. Its activation in response to mitogens leads to a cell-cycle progression from G1 to S phase. mTOR controls the activation of ribosomal protein translation and the initiation of cap-dependent translation. A role of mTOR signaling pathway dysregulation in tumourigenesis is postulated. mTOR and pathways upstream of this kinase were found to be frequently upregulated in neoplastic diseases. Therefore, it is also an attractive target for antitumour therapy. Several mTOR inhibitors were developed, including rapamycin and its analogues: CCI-779, RAD001 and AP23573. After promising phase I studies, their potential clinical significance is currently under evaluation in several phase II-III trials on patients with solid tumours and some hematological malignancies.
    Cellular & Molecular Biology Letters 02/2005; 10(3):479-98. · 1.95 Impact Factor
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    ABSTRACT: P-glycoprotein (P-gp) encoded by MDR1 gene is a xenobiotic transporter involved in protection of normal tissues against environmental toxicants and multi-drug resistance of cancer cells. Recently, a number of authors, including our group, have indicated that phenomenon of single nucleotide polymorphism (SNP) in MDR1 may influence susceptibility or prognosis in various diseases as well as pharmacokinetics of P-gp substrates. Although environmental chemicals are suspected carcinogens in multiple myeloma (MM) and P-gp transported drugs are used in the treatment of this malignancy, little data is known on the significance of MDR1 SNPs in MM. In this study, three common MDR1 SNPs: C1236T, G2677T/A and C3435T were assessed in 89 MM patients using automated sequencing and RFLP method. Moreover, in the control group of 215 healthy individuals MDR1 C3435T SNP was genotyped in all subjects and P-gp expression and activity were examined in a proportion of subjects. The haplotypes were inferred using Partition-Ligation algorithm showing highest frequency of the haplotype with wild-type alleles at all sites studied (1236C-2677G-3435C, 30%), followed by haplotype with all mutant alleles (1236T-2677T-3435T; 27%) and 1236C-2677G-3435T haplotype (12%). Carriers of a mutant allele at MDR1 1236 site showed a trend to longer overall survival as compared to wild-type homozygotes MDR1 1236CC (p=0.08), while no correlations with survival were observed regarding other polymorphisms. As compared to the control group, the MDR1 C3435T SNP allele frequency did not differ in MM patients, however the carriers of at least one T-allele (CT and TT genotypes) were significantly more frequent in the MM cohort (79% vs. 65%, p=0.02). The flow cytometry results of anti-P-gp MRK16 antibody staining and functional rhodamin123/PSC833 assay performed on peripheral blood mononuclear cells, CD19+ B-lymphocytes and CD3+ T-lymphocytes were similar among healthy individuals having MDR1 3435CC, CT and TT genotypes. In conclusion, we observed that MDR1 gene SNPs may exert some effect in genetic predisposition and prognosis in MM, which should be further examined in larger cohorts of uniformly treated patients.
    46th Annual Meeting of the American-Society-of-Hematology, San Diego; 12/2004
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    ABSTRACT: P-glycoprotein (P-gp), a membrane transporter encoded by MDR1 gene, influences pharmacokinetics of anti-cancer drugs and contributes to multi-drug resistance phenotype in adult acute lymphoblastic leukemia (ALL). In this study, we explored prognostic and functional role of single nucleotide polymorphism C3435T in MDR1 gene in 44 adult Caucasian patients with ALL. We found that the outcome of chemotherapy as well as MDR1 gene expression, P-gp expression and P-gp activity in isolated ALL blast cells were comparable among the patients carrying different MDR1 genotypes. Our results suggest that C3435T polymorphism in MDR1 gene is not a major prognosticator in adult ALL.
    Pharmacological reports: PR 57(6):882-8. · 1.97 Impact Factor

Publication Stats

231 Citations
76.84 Total Impact Points

Institutions

  • 2012
    • Università Vita-Salute San Raffaele
      Milano, Lombardy, Italy
  • 2005–2009
    • University of Lodz
      • Department of Cytobiochemistry
      Łódź, Łódź Voivodeship, Poland
    • Medical University of Łódź
      Łódź, Łódź Voivodeship, Poland