bluuebunny/crossref_metadata_2025 huggingface dataset:
Pegfilgrastim Provides Safe and Rapid Neutrophil Support in Patients with Acute Myeloid Leukemia (AML) Undergoing Myelosuppressive Chemotherapy.
Authors: Ranze Oliver , Arland Malte , Hoeffkes Heinz-Gert
Published: April 2020

Background: Administration of recombinant G-CSF has found extensive use in the support of chemotherapy-induced neutropenia in patients undergoing myelosuppressive chemotherapy. We retrospectively analyzed patients with acute myeloid leukemia (AML) who received either pegfilgrastim or lenograstim in this setting.

Patients and Methods: Unselected patients with AML according to FAB, age >18 years, performance status ECOG < 3, excluding pregnancy received at least one cycle of chemotherapy. On day 1 after chemotherapy has ended G-CSF treatment was initiated with pegfilgrastim 6 mg once per cycle or lenograstim 5 micg/kg b.w. until leucocyte counts (LC) raised > 1,000/micL. Primary objective was the difference in the time to elevating LC > 1,000/micL in each G-CSF treatment groups if LC nadired < 1,000/micL and the difference in the substitution of packed red blood (PRB) and platelets (PLT).

Results: Twenty-eight patients, 12 males and 16 females, aged median 61.4 years (range: 27–75 years) received 46 cycles. Six patients died due to the underlying disease. In four patients (two patients in the pegfilgrastim and two patients in the lenograstim group) G-CSF treatment failed to elevate LC > 1,000/micL. There was a difference in the time-to-recovery of leucocyte counts between pegfilgrastim 11.3 ± 3.8 days (range: 6 to 28 days) and lenograstim 13.3 ± 3.3 days (range: 6 to 22 days). The frequency of transfusions of PRB was 3.3 ± 2.6 in pegfilgrastim treated patients compared with 5.0 ± 2.9 in patients treated with lenograstim and the frequency of transfusions of PLT was 4.8 ± 2.5 vs. 6.0 ± 4.5. These differences received no statistical significance.

Conclusion: In this retrospective analysis unselected patients received myelodepressive chemotherapy because of acute myeloid leukemia. Both G-CSF showed adequate recovery of leucocyte counts with a faster increase in those patients to whom pegfilgrastim was administered. No statistical difference in the frequency of donation of human blood products was recognized. This retrospective study was limited because of the small number of patients included, thus these promising results need to be confirmed in a larger, prospective trial.
Combination Therapy of Humanized Chimeric-Anti-Human VLA4 Monoclonal Antibodies and Anti-Cancer Drugs for AML.
Authors: Matsunaga Takuya , Kato Kazunori , Tanaka Maki , Masuta Yukari , Kuribayashi Kageaki , Takemoto Naofumi , Tanaka Ikuta , Fujimi Akihito , Akiyama Takehide , Hamada Hirofumi , Niitsu Yoshiro
Published: April 2020

Bone marrow (BM) minimal residual disease causes relapse after chemotherapy in AML. We have previously reported that VLA4-positive leukemic cells acquired resistance to drug-induced apoptosis through the PI-3K/AKT/Bcl-2 signaling pathway, which is activated by the interaction of VLA4 and fibronectin on BM stromal cells. This resistance was negated by mouse-anti-human VLA4Ab (mouse VLA4Ab). In human leukemia SCID mouse model, we demonstrated a 100% survival rate with combination of mouse VLA4 Ab and AraC, while with AraC alone, only slight prolongation of survival was attained. In clinical study, overall survival at 5 years was 90% for 10 VLA4− patients and 25% for 15 VLA4+ patients (Matsunaga T et al, Nature Med 2003, 9, 1158–1165). In the present study, to perform the translational research, we first examined the myelosuppressive effect of the combination of rat-anti-mouse VLA4 Ab and AraC in C57/BL6 mice, and found that CBC data were almost the same as those of the mice treated with AraC alone. We next produced humanized chimeric-anti-human VLA4 Ab (chimeric-VLA4Ab), and examined its efficacy in combination with anti-cancer drugs in vitro and in vivo (human leukemia SCID mouse model). Chimeric-VLA4Abs were produced as follows: (i) total RNA of mouse VLA4 Abs were extracted from two hybridomas (SG/17 and SG/73), (ii) cDNA were synthesized by reverse transcriptase, (iii) variable region gene of mouse VLA4 Abs were amplified by 5′RACE method, (iv) TA cloning of amplified gene was performed, (v) sequence of mouse VLA4Abs gene was determined, (v) cloned variable region gene of mouse VLA4 Abs and constant region gene of human IgG1 were inserted into expression vector, and the expression vector was transfected into 293T cells, (vi) supernatant of the 293T cells was collected and purified to obtain chimeric-VLA4 Abs. The effects of chimeric-VLA4 Abs thus obtained in in vitro and in vivo (human leukemia SCID mouse model) were comparable to those of mouse VLA4 Abs. To perform the clinical study, we are presently producing the GMP-graded chimeric-VLA4 Abs.
Predictive Value of Bcl-2 and Bcl-6 Protein Expression in Diffuse Large B-Cell Lymphoma Patients from a Single Institution.
Authors: Paszkiewicz-Kozik Ewa , Mioduszewska Olga , Federowicz Irena , Kraszewska Ewa , Kulczycka Elzbieta , Domanska-Czyz Katarzyna , Ostrowska Beata , Romejko-Jarosinska Joanna , Osowiecki Michal , Walewski Jan A.
Published: April 2020

Bcl-2 and Bcl-6 protein expression has been reported to predict progression-free (PFS) and overall (OS) survival of patients with Diffuse Large B-cell Lymphoma (DLBCL) in retrospective and prospective clinical studies. Reports on Bcl-2 expression frequency and predictive value have been conflicting. In the current study, paraffin-embedded tissue specimens from 79 newly diagnosed DLBCL patients subsequently treated with CHOP-like chemotherapy without rituximab were studied retrospectively using immunohistochemistry to evaluate expected influence of Bcl-2 and Bcl-6 on clinical outcome. Bcl-2 and Bcl-6 positivity was defined as staining in any of lymphoma cells and it was demonstrated in 78% of Bcl-2 and 58% of Bcl-6 patients. Frequency of Bcl-2 protein expression in our study was much higher then reported by others. Combinations of possible positive/negative staining results: Bcl2+/Bcl6+, Bcl-2+/Bcl-6-, Bcl-2-/Bcl-6+, Bcl-2-/Bcl-6- were found in 46%, 33%, 13%, and 9% of cases, respectively. Complete remission (CR) after initial chemotherapy was achieved in 54 of 74 evaluable patients (73%) and 6 patients (8%) had progressive disease (PD). CR rate was higher and relapse and death rate was lower in patients with Bcl-6+ tumor irrespective of Bcl-2 status. In patients with Bcl-6 negative lymphoma CR rate was higher when Bcl-2 was positive, but relapse and death rate was not affected by Bcl-2. Median OS and PFS were not reached after 21.3 months of follow-up in this group of patients. In the analysis using multivariate proportional hazards regression model, advanced clinical stage (CS III and IV vs. CS I and II) was the only significant factor predicting OS (p<0.002). Risk of relapse or PD was significantly lower in patients with IPI score of 0 or 1 vs. > 1 (p<0.001), and in patients with both Bcl-2 and Bcl-6 positive lymphoma (p=0.04) as compared to patients with all other protein expression combinations. When analyzed separately, Bcl-6 positivity alone predicted PFS (p=0.025) while Bcl-2 was not significant (p=0.98). These results support predictive value of Bcl-6 protein expression for response to CHOP and PFS in DLBCL patients and are in agreement with the recent findings from the E4494 trial group (Blood102:101a, Abstr. 345, 2003). Unlike the findings of the GELA study (Blood101: 4279, 2003), our data fail to confirm predictive role of Bcl-2 protein.

Figure Figure
The Expression of Protein Kinase C Beta II (PKCβ) by Immunocytochemistry Is Predictive of the Disease Free Survival (DFS) in Diffuse Large B-Cell Lymphoma (DLBCL).
Authors: Schaffel Rony , Biasoli Irene , Scheliga Adriana , Morais Jose C. , Romano Sergio , Pulcheri Wolmar , Spector Nelson
Published: April 2020

PKCβ is a regulatory factor involved in the nuclear factor kappa B pathway. PKCβ gene expression was pinpointed through microarray studies by Shipp et al as one of thirteen, among over six thousand studied genes, to be associated with the probability of cure in DLBCL. The authors also reported a good correlation between PKCβ gene expression and protein expression, and added that in their hands PKCβ protein expression was closely associated with the clinical outcome. Therefore, we analyzed the PKCβ protein expression in 71 samples of DLBCL (44 nodal) by immunocytochemistry. A case was considered positive only if the antigen was unequivocally expressed in the large cells. PKCβ expression was found in 10/71 (14%) of the cases. Some cases had PKCβ expression only in the small lymphocytes, and were considered negative, since this antibody intensely stained mantle zone cells. Most samples had positive internal controls, as both plasma cells and residual mantle zone cells were strongly stained by the antibody. The patients’ clinical and biological features were similar in PKCβ positive and negative cases (median age 51 vs. 58 years; male gender 50% vs. 67%; extranodal disease 30% vs. 39%; high IPI 10% vs. 27%; Bcl-2 expression 50% vs. 41%, and CD10 expression 20% vs. 23%). None of these differences were statistically significant. The presence of PKCβ expression was not related to 7-year overall survival (56% vs. 48%, p=0.8). The 5-year event free survival was 54% and 33% in PKCβ negative and positive cases, respectively (p=0,7). A complete remission after an anthracycline based first line chemotherapy was obtained in 75% and 90% of PKCβ negative and positive cases, respectively (p=0,4). However, the 5-year DFS was 80% and 37%, for PKCβ negative and positive cases, respectively (p<0,03). In conclusion, in our cohort of patients the prognostic impact of PKCβ expression was restricted to the DFS.
Combined Chemotherapy and Radiation Is Associated with Improved Outcomes in Stage I Follicular Lymphoma.
Authors: Ganti Apar Kishor , Bociek R. Gregory , Bierman Philip J. , Lynch James C. , Bast Martin A. , Vose Julie M. , Armitage James O.
Published: April 2020

Follicular lymphoma (FL) is the second-most common subtype of non-Hodgkin’s lymphoma. However, data addressing the management of early-stage FL is sparse. Hence we conducted this study in order to try to assess the influence of initial treatment on outcome in patients with stage I follicular lymphoma. One hundred patients, 46 males and 54 females diagnosed with stage I FL and treated on various Nebraska Lymphoma Study Group protocols have been included in this retrospective study.

Thirty-eight percent of patients had grade 1, 22% had grade 2 and 40% had grade 3 FL. Sixty percent of patients were older than 60 years of age, 3% had a poor performance status (Karnofsky score <80), 7% had an elevated LDH level, 19% had bulky disease (tumor size ≥5 cm) and 5% had B-symptoms at presentation. Twenty-five percent of patients were treated with radiation alone, 37% with chemotherapy alone and 37% received both chemotherapy and involved-field radiation. Patients who received chemotherapy alone were more likely to have extranodal disease (p<0.01), but all other patient characteristics were similar across the three treatment groups. At a median follow-up of 8.4 years (range, 2.3 – 18.3 years), 45 patients are alive and disease-free, 39 patients have died while 16 patients are alive after progression. Of the 39 deaths that occurred, 13 were due to lymphoma progression, 4 were related to therapy, 11 patients died of unrelated causes (mainly coronary artery disease and its complications), while the cause of death was unknown in 11 patients. The median event-free survival (EFS) for this group of patients is 8.09 years and the median overall survival (OS) is 11.7 years. By multivariate analyses, only treatment with combination chemotherapy and radiation (HR: 0.3, 95% CI: 0.1 – 0.5, p<0.001) as opposed to either modality alone, and surprisingly, presence of B-symptoms at presentation (HR: 0.2, 95% CI: 0.04 – 0.8, p=0.02) predicted for a better EFS. Older age (≥60 years) predicted for an adverse OS (HR: 2.9, 95% CI: 1.3 – 6.3, p<0.01), while combination chemotherapy and radiation predicted for a better OS (HR: 0.4, 95% CI: 0.1– 0.9, p=0.02). However, there was no difference in the relapse rates between the older and younger patients (p=0.42), thereby suggesting that that poor OS in older patients was due to factors other than the disease itself. Patients who received both chemotherapy and radiation had a median EFS of 13.2 years and median OS was not yet reached in these patients. In contrast patients who received chemotherapy or radiation alone had an EFS of 5.54 years and 3.32 years respectively and an OS of 9.95 years and 10.7 years respectively. Thus, after accounting for known confounding factors, combined modality treatment with chemotherapy and radiation was associated with better EFS and OS than either modality alone in patients with Stage I follicular lymphoma.
Single Dose Per Cycle Pegfilgrastim Successfully Supports Full Dose CHOP-21 in Patients with Post-Transplant Lymphoproliferative Disorders (PTLD).
Authors: Papp-Vary Matthias , Oertel Stephan H.K. , Maschmeyer Georg , Riess Hanno
Published: April 2020

Background: PTLD is a life-threatening complication after transplantation of organs. There exists no common strategy for the treatment of patients with PTLD. Some patients benefit from reduction of immunsuppression or virostatic therapy, but most patients are treated by adriamycin based chemotherapy. Administration of chemotherapy is complicated by immunosuppressive therapy which induces bone marrow and renal insufficiency. Cytotoxic chemotherapy is frequently accompanied by febrile neutropenia and even lethal courses. Previous studies with CHOP 21 has demonstrated the need for growth factor (Filgrastim) to conduct chemotherapy in these patients. The aim of this study was to assess the ability of Pegfilgrastim, a pegylated form of Filgrastim, to support the 21 day CHOP chemotherapy. Pegfilgrastim has a sustained-duration effect relative to filgrastim, as a result of decreased renal clearance of the pegfilgrastim molecule.

Methods: Chemotherapy consisted of cyclophosphamide 750mg/m2, doxorubicin 50mg/m2 and vincristine 1,4mg/m2 at day 1 and prednisone 100mg day 1–5 and was administered at 21 days intervals for 4 cycles. Pegfilgrastim was administered by a single dose (6 mg s.c.) on day 3 of each cycle of chemotherapy

Results: In total 5 patients (4 after kidney- and 1 after liver-transplantation) were included in this study. The mean age of patients is 54 years (19–75) with two females and three males. A total of 15 cycles of CHOP has been administered. 14 of 15 cycles of CHOP could regularly be administered at day 21. In three patients five episodes of grade 4 neutropenia occurred, but none of the patients experienced infectious complications. Three patients experienced grade 3 neutropenia and two patients three episodes of grade 2 neutropenia. The lowest granulocyte count was 0.327 x 109/l. The rate of serious adverse events was low and no episode of febrile neutropenia occurred.

Conclusion: These results support the use of once-per-cycle pegfilgrastim (6 mg s.c.) for safe and effective on time delivery of a regimen with CHOP-21 in patients with Post-transplant Lymphoproliferative Disorders (PTLD).
A Stratified Risk-Adapted Approach to Lymphoma Salvage in an Outpatient Setting.
Authors: Spencer Andrew , Catalano John , Grigg Andrew , Leahy Michael , Underhill Craig , Ganju Vinod , Arthur Christopher
Published: April 2020

Aims: Interim analysis of the safety and efficacy of a risk-adjusted approach to lymphoma salvage therapy. Methods: Patients were stratified - Group 1 (G1) (good risk - first relapse following durable CR1); Group 2 (G2) (poor risk - primary refractory, >1 relapse, or non-durable CR1); or Group 3 (G3) (relapse post-ASCT). Two regimens were evaluated: VGF (vinorelbine 25mg/sqm days 1 and 8, gemcitabine 1000mg/sqm days 1 and 8, pegfilgrastim 6mg SC day 9) (G1/3) and F-GIV (VGF plus ifosfamide 3000mg/sqm day 1) (G2). Following 2 cycles all patients were re-staged. Responsive patients (>50% reduction in disease and functional imaging negative) received 2 further cycles of the same therapy, the remainder ‘escalated’ therapy to F-GIV (G1/3) or IVAC (G2) (inpatient ifosfamide, VP-16 and Ara-C). Results: 45 of a planned 90 patients, median age 56 years, are evaluable (G1 = 16, G2 = 23, G3 = 6). Diagnoses were Hodgkin’s lymphoma n = 9 and NHL, n = 36 (DLC = 24, follicular = 6, others = 6). To date G1 and G2 have received 127 cycles of VGF or F-GIV, with grades 3/4 neutropenia or thrombocytopenia in 31% and 17% (VGF) and 74% and 78% (F-GIV) of patients, respectively. Febrile neutropenia, admission, treatment delay or dose-reductions occurred with 13%, 28%, 12%, 6% of cycles, respectively. Based on ITT the ORR is 59% (CR 33%). Conclusion: VGF and F-GIV can be safely administered on an outpatient basis and show significant activity against advanced lymphoma.
Rituximab (Rituxan) Combined with ESHAP Chemotherapy Is Highly Active in Relapsed/ Refractory Aggressive Non-Hodgkin’s Lymphoma.
Authors: Venugopal Parameswaran , Gretory Stephanie A. , Showel John , Shammo Jamile , Enschede Sari , Larson Melissa , Obrien Teresa , Sokolovsky Tatiana D. , Means Aaron
Published: April 2020

Patients with relapsed/refractory aggressive lymphoma have limited options for achieving long-term remission other than stem cell transplantation. ESHAP is active in aggressive NHL and has the added advantage of being a non-anthracycline regimen. However, the response rate and remission duration with ESHAP chemotherapy alone are low in this group of patients. Encouraged by the increased efficacy of chemo-immunotherapy combinations like CHOP + Rituximab (Rituxan) in untreated aggressive lymphoma, and the in vitro synergy of Rituximab with cisplatin, cyclophosphamide and steroids, we studied the safety and efficacy of ESHAP + Rituximab combination in this group of patients. Standard ESHAP regimen (Etoposide 40 mg/m2/d IV/2h day1–4; Methylprednisolone 500 mg/m2 IV days 1–4; Cytarabine 2 g/m2 IV/3h day 5 and Cisplatin 25 mg/m2/d CIV days 1–4) was administered. Rituximab 375 mg/m2 IV was administered in the following manner: 2 doses separated by one week prior to the first cycle of ESHAP and 2 doses administered similarly after completion of the sixth cycle of ESHAP; one dose of Rituximab each was also given 48 hours prior to the third and fifth cycle of ESHAP. Treatment cycles were repeated every 28 days x 6. GMCSF (Leukine) and Erythropoietin (Procrit) were used at the discretion of the investigator. 14 patients were enrolled in the study. Median age was 55.5 years (range 37–70); there were 12 males and 2 females. 11/14 (71%) of patients had advanced disease (stage III/IV). Extranodal disease sites included kidney, lungs and adrenals. Histology was Diffuse Large Cell: 13 (93%) and Mantle Cell Lymphoma: 1 (7%). All patients had prior therapy: 1st relapse 11, 2nd relapse 1 and 3rd relapse 2. Prior therapies included CHOP, modified CHOP, Fludarabine-based regimen, Rituxan and Bexxar. 7 patients (50%) achieved CR, 2 PR, 1 stable disease and 4 PD. Among the 6 patients who received all 6 cycles of therapy, 4/6 (67%) achieved CR and 2/6 (33%) PR, (ORR 100%). Duration of response in patients who received all 6 cycles: All Responders: 26.3 months (1–53.3) and Patients who achieved CR: 35 months (24.1–53.3). Significant toxicity included grade III/IV hematologic in 43% and grade III/IV infection/febrile neutropenia in 21%.

Conclusion: ESHAP combined with Rituximab appears to be more active than ESHAP alone with no additional toxicity other than infusion-related adverse events in patients with relapsed/refractory aggressive lymphoma. Patients who achieved CR with this regimen appear to have longer remissions. Prospective, controlled trials are warranted.
Combined Effects of As4S4 and Imatinib on Chronic Myeloid Leukemia Cells and BCR-ABL Oncoprotein.
Authors: Yin Tong , Wu Ying-Li , Sun Hui-Ping , Sun Guan-Lin , Zhang Ji , Chen Guo-Qiang , Chen Sai-Juan , Chen Zhu
Published: April 2020

Imatinib is a tailored drug for chronic myeloid leukemia (CML), which has very good effects on patients at chronic phase (CP), but not on those at accelerated phase or blast phase. In addition, even among patients at CP, Imatinib seems unable to eradicate the malignant progenitors and a significant portion of patients develops drug resistance after long time use. Arsenic compounds were known as ancient remedies for CML with certain efficacy. The aim of this study was to investigate the potential benefit of combination therapy with Imatinib and arsenic sulfide (As4S4) on BCR-ABL+ K562 cells and fresh CD34+ hematopoietic progenitor cells isolated from CML patients and non-leukemic donors. Analysis of cell proliferation and clonogenic ability showed that As4S4 and Imatinib exerted synergistic effects on both K562 cells and fresh CML cells. The effective concentrations on fresh CML cells were pharmacokinetically available in vivo but had much less inhibitory effect on CD34+ cells from the non-leukemia donors. The synergistic effect of Imatinib/As4S4 combination in terms of anti-proliferation might be connected with their distinct but complementary roles in interfering with the cell cycle progression. Our data showed that Imatinib induced G1 arrest of K562 cells, while As4S4 induced G2/M arrest. In addition, Imatinib induces significant down-regulation of phosphorylated Rb and CDK1, which is in agreement with the G1/S but not G2/M arrest under this drug. However, As4S4 shows no obvious effect on these proteins in spite of a visible effect on G2/M block. Using a number of parameters such as morphology, Annexin V/PI, mitochondrial transmembrane potential, caspase3 activity and Fas/Fas-L, the synergistic effects were revealed on induction of cell apoptosis, largely through mitochondrial pathway. What’s more, the two drugs also exhibited synergistic effect in targeting BCR-ABL protein. While As4S4 triggered its degradation and Imatinib inhibited its tyrosine kinase activity, combined use of the two led to lower protein/enzymatic activity levels of BCR-ABL. In conclusion, our study suggests As4S4 and Imatinib have synergistic effects in inhibiting proliferation, inducing apoptosis of cells and reduction the tyrosine kinase activity of BCR-ABL. Our in vitro data thus strongly suggest a potential clinical application of Imatinib/As4S4 combination on CML.
PEG-Interferon for Chronic Phase CML - Still an Option in the Era of Imatinib?.
Authors: Scheid Christof , Draube Andreas , Holtick Udo , Cornely Oliver , Hacker Ulrich , Fuchs Michael , Berger Ute , Hochhaus Andreas , Hehlmann Rüdiger , Hallek Michael
Published: April 2020

Objectives

PEG-Interferon alpha 2b (PEG-IFN) is a new formulation of Interferon with a prolonged half-life and can therefore be given once weekly. It has shown efficacy in patients refractory or intolerant to IFN. Within a German multicenter study (SHG CMLIIIA) PEG-IFN (PEG-Intron, Essex) was used in our centre for patients with CML in first chronic phase.

Methods:

17 patients with newly diagnosed bcr-abl positive CML in chronic phase were included in our centre as part of the CML IIIA study. Median age was 42 yrs (range 24 – 78 yrs). After initial cytoreduction with hydroxyurea patients received 3–6 μg/kg PEG-IFN subcutaneously once a week. One patient received additional cytarabine at a dose of 20 mg/m2 10 days per month.

Results:

After 3 months 13/17 patients had a complete hematological response, 3 had a partial hematological response, 1 patient had a hereditary thrombocytopenia and therefor did not fulfill hematological CR criteria. Cytogenetics/FISH after 6 months were available in 12 patients and showed 1 complete cytogenetic response, 3 partial cytogenetic responses, 5 minor cytogenetic reponses and no response in 3 patients. 9 patients remained on PEG-IFN for at least 12 months and showed 2 complete, 1 partial and 2 minor cytogenetic responses and no response in 4 patients. Thus the rate of major cytogenetic responses was 33% at 6 months and at 12 months. PEG-IFN treatment was stopped in 14 patients due IFN-intolerance (n=6), patient decision (n=1), blast crisis (n=2), accelerated phase (n=1) or cytogenetic non-response (n=4). Relevant toxicities included acute pancreatitis in 1 patient, a grand mal seizure in a patient with a history of subdural hematoma, exacerbation of supraventricular tachycardia in 1 patient, transient elevation of GPT in 1 patient and a local abscess at the injection site in 1 patient. All patients experienced fever and flu-like symptoms in the first 4 weeks which gradually improved.

Conclusion:

PEG-IFN at a dose of 3–6 μg/kg was effective in producing complete hematological remissions after 3 months in the majority of patients and showed major cytogenetic responses in one third of patients. However there was a considerable toxicity. Still 53% of patients were able to continue treatment for at least 12 months and 4 of these (44%) are in continuous partial or complete cytogenetic response. This response rate is comparable to the results for conventional IFN in combination with Ara-C from the recent IRIS study. However in the IRIS study only 10.8% remained on IFN compared to 53% in our cohort. For patients able to tolerate this treatment PEG-IFN appears to be a valid alternative to IFN+Ara-C or imatinib in chronic phase CML.
Effects of Imatinib Mesylate and Interferon-alpha Combined with Granulocyte Colony-Stimulating Factor on Chronic Myelogenous Leukemia Cells In Vitro.
Authors: Ota Shuichi , Musashi Manabu , Kondo Keiichi , Toyoshima Nobuyasu , Toubai Tomomi , Iwao Noriaki , Tanaka Junji , Asaka Masahiro , Imamura Masahiro
Published: April 2020

Aim. In the treatment of patients with chronic myelogenous leukemia (CML), neutropenia is a dose-limiting factor on the combination therapy of imatinib mesylate (STI571) and interferon-alpha. If STI571 combined with interferon-alpha effects on decreasing BCR-ABL-positive cells and granulocyte colony-stimulating factor (G-CSF) has an effect on increasing normal neutrophils, the combination therapy may improve current remission rates in CML. We evaluated in vitro combined effect of STI571, interferon-alpha and G-CSF on primary bone marrow cells from patients with CML in chronic phase (CP).

Material and Methods. The primary bone marrow mononuclear cells (BMMNC) from patients with CML-CP were incubated in the medium containing STI571 (1 μM) and/or interferon-alpha (100 U/ml) with or without G-CSF (100 ng/ml). The viability of the cells was evaluated by trypan blue dye exclusion. Apoptosis was detected by the flow cytometry analysis with Annexin V (AV) and propidium iodide (PI) double staining. The colony formation assays of BMMNC were performed by methylcelulose. And then, BCR-ABL and normal cell colony rates were evaluated on14 days.

Results. Treatment of BMMNC with STI571 revealed a decrease of cell number in dose-dependent and time-dependent manner. Incubation with STI571 and interferon-alpha for 24 and 48 hours decreased viable cell number by 58% and 39%, respectively. G-CSF did not stimulate the BCR-ABL-expressing cells proliferation in the combination therapy. Cytometry analysis with AV and PI showed about 70–75% of AV-positive cells in the combination therapy and monotherapy. There were no significant differences in apoptosis between STI571 plus G-CSF and interferon-alpha plus G-CSF. Colony formation of BMMNC from patients with CML-CP in combination of STI571 and interferon-alpha was more strongly suppressed than STI571alone, and G-CSF did not abrogate the suppressive effect of the combination therapy. From these observations, it is concluded that combination therapy of STI571 and interferon-alpha with G-CSF can induce apoptosis effectively in BCR-ABL-expressing cells.

Conclusion. Obtained data provide an evidence for the effective and safe combination therapy of STI571 and interferon-alpha with G-CSF against CML-CP patients. Although further studies are needed to clarify whether combination therapy really increase normal cells or not, the combined therapy with G-CSF appears not to stimulate leukemic cell proliferation.
Prediction of Response to Imatinib by Prospective Quantitation of BCR-ABL Transcript in Late Chronic Phase Chronic Myeloid Leukemia PatientsBy GIMEMA Working Party on CML.
Authors: Martinelli Giovanni , Rosti Gianantonio , Pane Fabrizio , Amabile Marilina , Bassi Simona , Izzo Barbara , Poerio Angela , Giannini Barbara , Cilloni Daniela , Soverini Simona , Terragna Carolina , Ottaviani Emanuela , Grafone Tiziana , De Vivo Antonio , Testoni Nicoletta , Trabacchi Elena , Alberti Daniele , Salvatore Francesco , Saglio Giuseppe , Baccarani Michele
Published: April 2020

Imatinib mesylate (STI571), a specific Bcr-Abl inhibitor, has shown a potent antileukemic activity in clinical studies of chronic myeloid leukemia (CML) patients. Early prediction of response to imatinib cannot be anticipated. We used a standardized quantitative reverse-transcriptase polymerase chain reaction (QRT-PCR) for bcr-abl transcripts on 191 out of 200 late-chronic phase CML patients enrolled in a phase II clinical trial with imatinib 400 mg/day. Bone marrow samples were collected before treatment, after 3, 6 and 12 months or at the end of study treatment (12 months) while peripheral blood samples were obtained after 2, 3, 6, 10, 14, 20 and 52 weeks of therapy. The amount of Bcr-Abl transcript was expressed as the ratio of Bcr-Abl to β2-microglobulin (β2M). We show that, following initiation of imatinib, the early Bcr-Abl level trends in both bone marrow and peripheral blood samples made it possible to predict the subsequent cytogenetic outcome after 6 and 12 months of treatment, and that these early trends were also predictive of progression-free survival.
Generating Human Immune Responses to Mutations in Bcr-Abl Kinase Selected by Imatinib.
Authors: Pinilla Javier , May Rena , Gomez Marta , Korontsvit Tanya , Barry Kappel , Zakhaleva Victoriya , Dao Tao , Scheinberg David A.
Published: April 2020

Imatinib mesylate is now standard treatment for chronic myelogenous leukemia. In spite of its high efficacy in controlling the disease, residual bcr-abl cells measured by RT-PCR are still detectable in more than 95% of the patients treated with this drug. In addition, recent reports describe the emergence of resistance to Imatinib mesylate. A major mechanism for the appearance of resistance, are point mutations changing amino acids in the bcr-abl tyrosine kinase that make imatinib ineffective. We hypothesized that some bcr-abl kinase mutations selected by imatinib would generate new immunogenic peptide-epitopes that could be recognized by the human immune system. For this purpose, we selected with predictive computer algorithms potentially immunogenic mutated amino acid sequences and synthetic analogues of mutated bcr-abl kinase sequences to study for immunogenicity and anti-leukemia activity in an in vitro human system. Initially we designed a number of synthetic peptides derived from the most frequent bcr-abl kinase mutations induced by imatinib in which single amino acid substitution was introduced at different MHC-binding positions. We investigated the most common mutations: E255V, E255K, Y253H, Y253F, F311L, T315I, M351T and H396P. Specific single and combined mutations improved the predicted binding of peptides to HLA A0201. Further additional single amino-acid substitutions on mutated peptides generated high predicted binding. Selected sequences were evaluated for eliciting MHC-restricted, peptide-specific CTL responses in an in vitro model using T-cells from different HLA A0201 healthy donors after stimulation with monocyte-derived DC. CTL lines were assessed by either IFN gamma ELISPOT or by a chromium51 release assay. Mutated derived peptides from E255K, E255V, Y253F and T315I were shown to be immunogenic in vitro by the specific release of IFN gamma. Furthermore, single amino-acid substitutions in key residues improved the immunogenicity of these peptides. Finally, T cell lines generated with optimized peptides derived from E255K and T315I (the most common imatinib-induced mutations) were able to kill peptide pulsed targets; more important, these T cell lines cross-reacted with the mutated peptides (a heteroclitic response). In conclusion, Imatinib-selected mutations of peptides from bcr-abl kinase have a better predicted binding than native kinase sequences. Selected peptides from the most common mutations are immunogenic in vitro in a HLA A0201 restricted manner. T cells generated with optimized peptides from these mutations are able to elicit heteroclitic peptide-specific cytotoxicity.
Philadelphia-Negative Clonal Hematopoiesis Following Imatinib Therapy in Patients with Chronic Myeloid Leukemia (CML).
Authors: Lin Yulia , Bruyere Helene , Horsman Douglas E. , Barnett Michael J. , Hogge Donna E. , Nevill Thomas J. , Nantel Stephen H. , Sutherland Heather J. , Toze Cynthia L. , Shepherd John D. , Lavoie Julye C. , Song Kevin W. , Smith Clayton A. , Forrest Donna L.
Published: April 2020

There is an increasing number of reports of Philadelphia-negative (Ph-) clonal hematopoiesis developing in patients (pts) with CML treated with imatinib. The significance of these abnormalities remains unclear. To examine the incidence, natural history and risk factors for the development of Ph- clones, we undertook a retrospective review of 142 consecutive pts with CML treated with imatinib and followed at VGH or BCCA between June 1999 and July 2004. Four pts were excluded due to incomplete records. Out of 138 pts, there were 76 (55%) males. The median age at diagnosis was 48.6 yrs (range 19.3–82.8). At the time of initiation of imatinib, 93 (67%) pts were in stable phase, 33 (24%) in accelerated phase and 12 (9%) in blast phase. 126 pts had received prior therapy with hydroxyurea (81%), interferon (53%), hematopoietic stem cell transplantation (17%), cytarabine (11%), or other therapy (17%). The median number of prior therapies was 2 (range 0–4). The median time from diagnosis to treatment with imatinib was 12.7 mos (range 0–240.1) and the median duration of imatinib therapy was 15.2 mos (range 0.6–45.1). Best responses to imatinib were complete hematologic response (96%), major cytogenetic response (53%) and complete cytogenetic response (43%). The event-free survival (EFS) at 3 yrs from the start of imatinib was 59% (95%CI 47–70%). Of the 138 pts, 7 (5%) pts developed Ph- clonal abnormalities: −7 and +8 (2 pts), +8 (2 pts), −7 (1 pt), −X and −22 (1 pt) and t(12;16) (1 pt). One pt had −7 and 1 pt had +8 in both Ph- and Ph+ cells. For the pts who developed Ph- clonal abnormalities, the median time from diagnosis to the start of imatinib was 28.4 mos (range 0.7–143.9). Six of these 7 pts had received prior therapy. The median time from the start of imatinib therapy to the development of the Ph- clone was 11.9 mos (range 2.7–23.5). All but two of the Ph- clones were transient and none was associated with myelodysplasia. Two of the 7 pts had progressed on imatinib. There was no difference in EFS or overall survival between pts with or without Ph- clones. In univariate analysis comparing characteristics of pts with and without Ph- clones, no predictive factors could be identified. In conclusion, the development of Ph- clonal hematopoiesis in pts with CML treated with imatinib occurred in ~5% of this series and did not appear to confer a worse prognosis. The majority of the Ph- clones included −7 and/or +8 and were transient without associated myelodysplasia. Finally, the observation of −7 (to our knowledge the first such case reported) or +8 in both Ph- and Ph+ cells in the same patient could be explained by the existence of a Ph- monoclonal state prior to the BCR/ABL mutation.
Chronic Myeloid Leukemia Patients Treated with Imatinib Show Increased IFN-Gamma Synthesis in T Cells and Hypogammaglobulinemia.
Authors: Gulbas Zafer , Akay Meltem O. , Sahin Fezan , Gunduz Eren
Published: April 2020

Background: In patients with untreated chronid myeloid leukemia(CML), lymphopoesis has been affected and Th1 cytokine levels are decreased. Imatinib tretment for CML has been shown to induce clinical and cytogenetic remission and increases plasmocytoid dendritic cells, affects lymphopoesis. Flow cytometry has been used to detect Th1 IFN-gamma, IL-2 and Th2 IL-4 cytokines in T cells.

Methods: 10 patients with CML before imitinib treatment, 29 patients after at least 2 months imatinib treatment and 10 healthy controls were tested to detect Th1 IFN-gamma, IL-2 and Th2 IL-4 cytokines in T cells using FITC, PE, PerCP monoclonal antibodies by flow cytometry(FacsCalibur,B.D.). Also; IgG, IgM, IgA levels by nepholometry and lymphocytes subpopulations by flow cytometry were detected.

Results: 10 patients before imatinib treatment showed decreased Th1 IFN-gamma, IL-2 and Th2 IL-4 cytokines in T cells according to the controls (p<0.05). 29 patients after at least 2 months imatinib treatment showed increasing statistically important Th1 IFN-gamma cytokine but statistically insignificant increase IL-2 cytokine and Th2 IL-4 cytokines in T cells (Table 1). Hypogammaglobulinemia were developed 17% of the patients. No correlation was found imatinib treatment period and Th1 IFN-gamma, IL-2 and Th2 IL-4 cytokines in T cells.

Conclusions: Imatinib treatment increases Th1 IFN-gamma producing T cells and also causes hypogammaglogulinemia. This suggest that enhanced Th 1 cell function is achievable with imatinib treatment in patients with CML.

Percantage of Th1 IFN-gamma, IL-2 and Th2 IL-4 cells in CML patients and controls

before imitinib after imitinib healthy control CD4+IL-2+(%) 15+/− 13 25+/−14 44+/−12 CD4+ IFN-gamma+(%) 7+/− 5 11+/−7 15+/−2 CD4+ IL-4+(%) 1+/− 1 2+/− 1 4+/− 1
Prevalence of MDS in Shanghai, China Using the WHO Classification.
Authors: Irons Richard D. , Bao Liming , Wang Xiao Qin , Ryder John , Ji Jeirong , Du Xinyu , Gross Sherilyn A. , Chen Hui , Chen Yan , Kerzic Patrick J. , Fu Hua , Lin Guowei
Published: April 2020

The prevalence of subgroups of myelodysplastic syndrome (MDS) was determined in an unselected series of 92 patients presenting at Shanghai hospitals over a 10 month period. Diagnosis was established in a single laboratory, analyzing morphologic, immunophenotypic, fluorescence in situ hybridization (FISH) and cytogenetic data, according to the World Health Organization (WHO) revised classification. The frequencies of the major MDS subgroups were: refractory anemia (RA) 9.7%, refractory anemia with ringed sideroblasts (RARS) 1%, refractory anemia with excess blasts (RAEB) 15.2%, refractory cytopenia with multilineage dysplasia (RCMD) 52.6%, and MDS-unclassifiable (MDS-u) 7.6%. The subgroup, myelodysplastic/myeloproliferative diseases, unclassifiable (MDS/MPD-u) (9.7%) was also included for comparison with other studies. The median age at diagnosis for all cases was 57 yr. Males slightly predominated (53%). The overall incidence of clonal cytogenetic abnormalities was 24.7%. The most frequently encountered lesions were trisomy 8, 20q-, 5q- and 7q-. The prevalence of clonal cytogenetic abnormalities in each subgroup differed significantly from none in RA to 31% in RCMD. Deletions involving 5q- and 7q- were found predominantly in RAEB and RCMD subgroups. The incidence of MDS in China has been reported to be less than that in the West. However, information on the frequency of different subgroups of MDS in Shanghai previously has not been available. These cases presented at 24 hospitals serving a population of approximately 18 million. These results suggest major differences in the presentation and frequency of different subgroups of MDS between China and the West.
Predictive Progenitor Profiling in Chronic Myelomonocytic Leukemia.
Authors: Jamieson Catriona , Gotlib Jason , Coutre Steve , Li Kevin , Weissman Irving
Published: April 2020

Chronic myelomonocytic leukemia (CMML) is a unique myeloproliferative disease characterized by marrow dysplasia and an increase in monocytes. The median survival of patients with CMML is short, in part, because CMML is frequently resistant to therapy. In order to provide more effective CMML targeted therapies, a better understanding of the basic biology of CMML progenitors is required. We used FACS analysis and recently identified cell surface markers to identify phenotypic and functional differences between normal and CMML (n=14) bone marrow hematopoietic stem cells and myeloid progenitors. CMML marrow was typified by a reduction in CD34+CD38−CD90+(Thy1)Lin− hematopoietic stem cells and an expansion of CD34+CD38−CD90−Flk2+Lin- cells relative to normal bone marrow. In addition, there was a two-fold expansion in common myeloid progenitors (CMPs) and a corresponding decrease in megakaryocyte-erythroid progenitors (MEPs) suggesting that there was a skew in differentiation toward the myeloid lineage. In contrast to normal bone marrow derived CMPs, CMML CMPs gave rise to myeloid but not erythroid colonies. Moreover, real time quantitative RT-PCR analysis of highly purified FACS-sorted CMML CMPs demonstrated increased expression of two key regulators of myelomonocytic differentiation, PU.1 and c-jun, compared with normal bone marrow. A more detailed understanding of the basic biology of CMML myeloid progenitors and the genes that work in concert to expand them may aid in identifying novel molecular targets for CMML therapy.
Perforin Expressing CD4+T Cells Respond to CMV Antigens in CMV Seropositive Patients with B Cell Chronic Lymphocytic Leukaemia (B-CLL).
Authors: Walton James , Lawson Keirissa , Manoussaka Maria S. , Nathwani Amit , Emory Vincent , Lydyard Peter M. , Porakishvili Nino
Published: April 2020

Introduction: Perforin (PF) expressing CD4+ T cells are expanded in B-CLL patients [1] and are able to induce PF-mediated apoptosis of autologous leukaemic cells in the presence of bispecific anti-CD3/CD19 antibodies [2]. However, the role of the expanded PF+CD4+ T population in B-CLL remains unclear. In this study we have examined possible involvement of this cell subset in immune responses to cytomegalovirus (CMV).

Methods: Blood mononuclear cells from 11 CMV seropositive (SP) and 6 seronegative (SN) B-CLL patients were cultured for 4 and 18 hours in presence of Downe cell lysates containing CMV-antigen or cell lysates alone (control) and brefeldin and co-stimulated with anti-CD28 and anti-CD49 monoclonal antibodies. Anti-CMV response was assessed by flow cytometry as percentages of CD69+ and IFNγ+ cells in PF+ and PF- CD4+T cell populations.

Results: CD4+ T cells from 9 of 11 SP patients showed a strong increase in percentages of IFNγ+ cells after 18h, but not 4h, in culture with CMV antigens, compared to the control lysates and SN patients (Table) and healthy age-matched SP subjects (1.78±0.45, p=0.012). In contrast, none of the 6 SN patients as well as 2 out of 11 SP patients responded to CMV by increased IFNγ expression. In responders IFNγ+ cells were proportionally distributed between PF+ and PF- CD4+ T cells. High levels of cell activation measured by CD69 expression, especially in PF+ population, were mainly due to the Downe cell lysate since expression was significantly lower in lysate free cell cultures (8.3±11.5%, p=0.0018 for SP B-CLL patients). Anti-CMV response was accompanied by a decrease in percentages of CD4+PF+ cells suggesting antigen-induced degranulation.

Conclusion: PF+CD4+ T cells respond to CMV antigens suggesting an expansion of mature CMV specific cytotoxic CD4+ T cells in the majority of CMV seropositive B-CLL patients.

IFN-γ and CD69 expression by CD4+PF+ and PF- T cells after 18h exposure to CMV+ and CMV- lysates in CMV seropositive and seronegative B-CLL patients

CD4+ PF+ CD4+ PF-CD4+ SN SP SN SP SN SP IFN- γ, % CMV+lys 0.7±0.58 9.1±7.9 2.0±3.0 p=0.013 14.4±13.6 0.61±0.56 p=0.008 9.0±8.1 CMV−lys 0.26±0.34 0.9±1.7 p=0.007 1.1±1.7 3.9±6.9 p=0.029 0.24±0.27 1.46±2.55 p=0.013 CD69+cells, % CMV+lys 16.3±14.2 21.4±13.0 41.9±35.1 51.2±30.3 14.9±13.4 20.4±13.2 CMV−lys 14.4±12.8 9.7±7.8 28.6±34.8 39.8±33.0 13.8±12.1 12.9±15.0
A Phase I/II Study of the Oral mTOR Inhibitor RAD001 in Patients with Advanced Hematologic Malignancies.
Authors: Yee Karen W.L. , Wierda William , O’Brien Susan , Thomas Deborah , Kurzrock Razelle , Fayad Luis E. , Hagemeister Fredrick B. , Cortes Jorge , Prestifilippo Judith , Szatrowski Ted P. , Kantarjian Hagop M. , Giles Francis J.
Published: April 2020

RAD001 (everolimus, Novartis) is an orally bioavailable derivative of rapamycin with demonstrated anti-proliferative activity against a broad panel of tumor cell lines and antitumor activity in experimental animal models of human cancer. RAD001 inhibits the mammalian target of rapamycin (mTOR) signaling pathway, which is involved in regulating many aspects of cell growth and cell cycle progression. Several lines of evidence implicate the importance of the PI3K/Akt/mTOR pathway in hematological malignancies. As there has been extensive experience with this agent in the solid organ transplantation setting, two dose levels (5 and 10 mg/day) were evaluated in the Phase I portion of this study to determine the maximum tolerated dose (MTD) to be used in the Phase II portion. RAD001 was administered orally once a day at the starting dose level of 5 mg. Fifteen patients have been enrolled, of which 14 (3 B-CLL, 4 MDS, 1 MF, 1 NK/T cell lymphoma/leukemia, 1 MCL, 3 AML, 1 T-PLL) were evaluable for safety and toxicity. Median age was 65 years (range, 56 to 76 years). Twelve patients (86%) had received prior therapy (median, 2 regimens; range, 0 to 6 regimens). Median time on study was 29 days (range, 12 to 105 days). No dose-limiting toxicities were observed in the 6 patients enrolled in the Phase I portion of the study (cohorts of 3 patients per dose level). The most common adverse events were grade ≥ 2 and consisted of hyperglycemia, hyperlipidemia, hypocalcemia, hypomagnesemia, hypophosphatemia and hypokalemia, transaminitis, diarrhea, dermatitis, and mucositis. Grade 3 toxicities consisted of asymptomatic hypophosphatemia (2), and hyperglycemia (3). No patient experienced grade 4 toxicities or death from RAD001. Of the 14 evaluable patients, 2 patients with B-CLL had a 33% and 65% reduction in the size of lymph nodes, respectively, and 1 patient with MDS had decreased transfusion requirements. At the time of analysis, only 1 patient had discontinued therapy due to disease progression. These preliminary findings indicate that RAD001 is well tolerated at a daily dose of 10 mg/day and may have clinical activity in patients with hematologic malignancies. Enrollment is ongoing.