scholarly journals A Preclinical Non-Human Primate Model to Test Reversible Myeloablation Following IV Busulfan Therapy without Stem Cell Rescue

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 1868-1868
Author(s):  
Nadim Mahmud ◽  
Amit Khanal ◽  
Simona Taioli ◽  
Emre Koca ◽  
Sujata Gaitonde ◽  
...  
Keyword(s):  
Stem Cell ◽  
Myeloid Leukemia ◽  
Platelet Counts ◽  
Stem Cell Rescue ◽  
Cd34 Cells ◽  
Group A ◽  
Pre Treatment ◽  
Wbc Count ◽  
Group B ◽  
Acute Myeloid

Abstract Busulfan is a standard drug utilized in stem cell transplantation for myeloid malignancies, however it is not used in any induction or salvage chemotherapy regimen in these diseases. Non-human primates (NHP) represent a useful model for preclinical evaluation of chemotherapy-related toxicity. In order to identify what dose of IV busulfan could be tested in clinical studies for acute myeloid leukemia, we utilized NHP to determine the highest dose of IV busulfan needed to achieve reversible myelosuppression in unperturbed bone marrow (BM). Nine adult baboons (Papio anubis) were divided into three groups (n=3/group) for this study. Group A received the lowest dose of busulfan 6.4 mg/kg (1.6 mg/kg/day x 4 days), Group B received 8 mg/kg (3.2 mg/kg on day 1 and 1.6 mg/kg on day 2-4) and Group C received 9.6 mg/kg (3.2 mg/kg/day day 1-2 and 1.6 mg/kg/day day 3-4). Peripheral blood (PB) complete blood count (CBC), and BM CD34+ cells, colony forming unit (CFU) were monitored over 90 days after busulfan. Maximum suppression of WBC count, hemoglobin (Hb) and platelet counts in the PB of baboons in all 3 groups were observed around day 15. The suppression of WBC was 57 ± 4%, 58 ± 4% and 53 ±10% in group A, B and C respectively (group A: p = 0.006, group B: p = 0.03, group C: p = 0.07). The suppression of absolute neutrophil counts (ANC) in group A, B and C were 85 ± 2%, 8 4± 1% and 82± 5% respectively. Similarly the minimum post-busulfan Hb levels were 10± 0.6 g/dl, 10± 1g/dl, 10± 0.6g/dl in group A, B and C respectively. The suppression of platelet counts were 84 ± 6%, 81 ± 7% and 81 ± 9% in group A, B and C respectively. Taken together, PB CBC results indicate that at day 15, despite administering a higher dose of busulfan the maximum depletion of ANC, Hb and platelet was comparable among the three groups. At day 90, the PB WBC, Hb and platelet counts returned within normal ranges. Notably, in the group receiving the highest dose of busulfan (Group C) the PB WBC counts at day 90 displayed 76 ± 9% recovery and platelets a 79 ± 13% recovery. Extra-hematopoietic toxicity, including weight loss, was not significant in any group of animals. Then we examined CD34+ cells and CFU content of BM prior to and following busulfan therapy. Baboons receiving the lowest dose of busulfan (Group A) displayed 75 ± 19% CD34+ cell suppression, while the intermediate group B had 90 ± 9%, and the group C which received the highest dose of busulfan had 98 ± 0.6 % suppression. The CD34+ cell recovery at day 90 was 56%, 35% and 24% in group A, B and C, respectively, suggesting that the BM CD34+ cell compartment requires more time to recover despite near normal CBC numbers. Similarly, at day 15 the suppression of BM CFU in group A, B and C was 71 ± 10% , 86 ± 8% and 91 ± 5% respectively. Although the suppression in PB WBC count was not distinct the absolute number of CD34+ cells and CFU content of BM varied based on busulfan dose administered. At day 90 the recovery of BM CFU in group A, B & C were 51 ± 15% , 79 ± 15% and 53 ± 9%, respectively. Although there was no significant intergroup difference in degree of CFU plating efficiency, pre-busulfan versus day 90 CFU recovery in group C was statistically significant (p = 0.02) . These results further validate the hierarchy of CD34+ cells and CFU in contrast to more mature PB cells and the cell populations targeted by busulfan, emphasizing regulatory systems governing homeostasis are likely maintained by reserved BM precursors and progenitor cells. Despite near normalization of blood counts, the relatively primitive BM CD34+ cells and CFU content require longer time to return to pre treatment levels. BM biopsy results indicate a return of cellularity and tri-lineage hematopoiesis comparable to pre-treatment levels in all three groups by day 90. Taken together, our study showed that when busulfan is administered in a dose range of 6.4 to 9.6 mg/kg in an NHP model, it is capable of inducing reversible myeloablation with tolerable toxicity without requiring stem cell rescue or blood transfusions. Our results also indicate that about 30 to 40% BM reserve is capable of maintaining normal PB cell counts. Based on the results, we plan to design a phase 1/2 clinical trial where non-myeloablative doses of IV busulfan will be tested as salvage therapy in patients with acute myeloid leukemia. Disclosures No relevant conflicts of interest to declare.

scholarly journals Safety and Cost Effectiveness of a 10 × 109/L Trigger for Prophylactic Platelet Transfusions Compared With the Traditional 20 × 109/L Trigger: A Prospective Comparative Trial in 105 Patients With Acute Myeloid Leukemia

Blood ◽  
1998 ◽  
Vol 91 (10) ◽  
pp. 3601-3606 ◽  
Author(s):  
Hannes Wandt ◽  
Markus Frank ◽  
Gerhard Ehninger ◽  
Christiane Schneider ◽  
Norbert Brack ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Blood Cell ◽  
Red Blood Cell ◽  
Myeloid Leukemia ◽  
Platelet Transfusion ◽  
Bleeding Complications ◽  
Group A ◽  
Red Blood Cell Transfusions ◽  
Group B ◽  
Acute Myeloid

Abstract In 105 consecutive patients with de novo acute myeloid leukemia (French-American-British M3 excluded), we compared prospectively the risk of bleeding complications, the number of platelet and red blood cell transfusions administered, and the costs of transfusions using two different prophylactic platelet transfusion protocols. Two hundred sixteen cycles of induction or consolidation chemotherapy and 3,843 days of thrombocytopenia less than 25 × 109/L were evaluated. At the start of the study, each of the 17 participating centers decided whether they would use a 10 × 109/L prophylactic platelet transfusion trigger (group A/8 centers) or a 20 × 109/L trigger (group B/9 centers). Bleeding complications (World Health Organization grade 2-4) during treatment cycles were comparable in the two groups: 20 of 110 (18%) in group A and 18 of 106 (17%) in group B (P = .8). Serious bleeding events (grade 3-4) were generally not related to the patient's platelet count but were the consequence of local lesions and plasma coagulation factor deficiencies due to sepsis. Eighty-six percent of the serious bleeding episodes occurred during induction chemotherapy. No patient died of a bleeding complication. There were no significant differences in the number of red blood cell transfusions administered between the two groups, but there were significant differences in the number of platelet transfusions administered per treatment cycle: pooled random donor platelet concentrates averaged 15.4 versus 25.4 (P < .01) and apheresis platelets averaged 3.0 versus 4.8 (P < .05) for group A versus group B, respectively. This resulted in the cost of platelet therapy being one third lower in group A compared with group B without any associated increase in bleeding risk.

Birth Order and Outcome in HLA-Identical Sibling Donor Hematopoietic Stem Cell Transplants. Impact of a Sequential Fetomaternal - Maternofetal Cell Transfer?.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2034-2034
Author(s):  
Christoph M. Bucher ◽  
Dominik Heim ◽  
Andreas Buser ◽  
Jakob R. Passweg ◽  
Alois Gratwohl
Keyword(s):  
Stem Cell ◽  
Birth Order ◽  
Myeloid Leukemia ◽  
Cell Transfer ◽  
Black Line ◽  
Cell Transplants ◽  
Group A ◽  
Grade Ii ◽  
Stem Cell Transplants ◽  
Group B

Abstract Fetomaternal and maternofetal cell transfer have been described. Their clinical relevance is unknown. We hypothesized that firstborn siblings could tolerize their siblings born thereafter through sequential fetomaternal-maternofetal cell transfer. Hence, stem cell transplants within a family from a firstborn sibling (group A) should result more graft-versus-host disease (GvHD) and worse overall survival than transplants to a firstborn donor (group B). Results of a retrospective single center cohort analysis of 321 HLA-identical sibling donor hemopoietic stem cell transplants showed a survival of 48.3% (+/−10.9%) at 10 years in the group with firstborn donors (group A, 110 patients) as compared to 63.2 (+/−10.6%) in the group with firstborn recipients (group B, 105 patients; p<0.02) and a RR of death after adjustment for other risk factors of 2.6 (CI 1.45–4.66; p<0.001) for the group with firstborn donors. These results support the concept of a clinically relevant tolerizing effect of birth order, possibly mediated by fetomaternal cell transfer in man. Patients characteristics and outcomes Birth Order Donor First Sibling Recipient First Sibling n 110 105 Donor Age median (range) 30.1 (4.9–68) 25.4 (0.4–59.1) p=0.005 Recipient Age median (range) 25.4(3.2–62.1) 30.8 (2.2–63.0) Number of Siblings 2 67 69 n.s. 3 26 22 >3 17 14 Diagnosis n.s. Acute myeloid leukemia n 33 26 Acute lymphoblastic leukemia n 24 24 Chronic myeloid leukemia n 23 23 Lymphoproliferative disorders n 14 13 Severe aplastic anemia n 11 12 Myelodysplastic syndromes n 5 7 Stem Cell Source n.s. Bone Marrow n 71 73 PBSCT n 39 32 Conditioning n.s. With totoal body irradiation n 18 22 Without total body irradiation n 92 83 Outcomes Survival at 10 yrs % 48.3 63.2 p<0.02 Relapse at 3 yrs. % 26 20 n.s. Acute GvHD p=0.017 < Grade II n 46 61 >= Grade II n 64 44 Chronic GvHD n.s. none n 28 30 n.a. n 29 17 Limited n 32 30 Extensive n 21 23 Figure 1: Kaplan Meyer estimate of cumulative survival of groups A(firstborn donor: gray line) and B (firstborn recipient: black line). + indicates censored patient. Figure 1:. Kaplan Meyer estimate of cumulative survival of groups A(firstborn donor: gray line) and B (firstborn recipient: black line). + indicates censored patient.

Bacterial, Viral and Fungal Infections in Patients after Allogeneic Stem Cell or Bone Marrow Transplantation - A Comparison between Patients with Related and Patients with HLA-Matched Non-Related Stem Cell Donors.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4978-4978
Author(s):  
Christina T. Rieger ◽  
Johanna Tischer ◽  
Helmut Ostermann
Keyword(s):  
Bone Marrow ◽  
Candida Albicans ◽  
Stem Cell ◽  
Stem Cell Transplantation ◽  
Myeloid Leukemia ◽  
Fungal Infections ◽  
Stem Cell Source ◽  
Cell Source ◽  
Group A ◽  
Group B

Abstract Bacterial, viral and fungal pathogens frequently cause severe, life-threatening infections in immunocompromised patients after allogeneic stem cell transplantation (SCT). We investigated whether patients with related stem cell donors (group A) developed infections less frequently than patients with HLA-matched, non-related donors (group B). Fifty-nine consecutive patients treated at our transplantation unit between April 2004 and January 2005 were included into the analysis. We documented demographic and clinical characteristics at baseline, treatment, clinical course, microbiological examinations, clinical and radiological signs of infection and mortality. Of the total 59 patients analyzed, 22 received stem cells from related and 37 from HLA-matched non-related donors. Both groups were well balanced regarding age and weight. 50% of the patients in group A and 60% in group B were male. Most frequent diagnoses were acute myeloid leukemia (30 of 59 patients [50.8%]; group A: 68.2%; group B: 40.5%), multiple myeloma (15.2%), acute lymphoblastic leukemia (11.9%) and chronic myeloid leukemia (10.2%). Bone marrow was more often the stem cell source in group A (45.5%/ 10 patients) than in group B (10.8%/ 4 patients), peripheral stem cell transplantation respectively was predominant in the unrelated group (86.5%/ 32 patients) versus the family donor group (54.5%/ 12 patients), cord blood was used as unrelated stem cell source in1 patient (2.7%). Clinically documented infections occurred in 6% in group A and in 14% in group B. Pulmonary infiltrates were observed more frequently in group A (11 patients/ 50%) than in group B (16 patients/ 43.2%). The predominant findings were atypical infiltrates (total 16 patients), followed by signs of fungal (total 7 patients) and bacterial pulmonary infiltration (total 4 patients). Microbiologically documented infections were detected in all patients. The average number of pathogens was equal in both groups. Detected pathogens were HHV-6 (48 patients), coagulase-negative Staphylocci (17 patients), EBV (14 patients) and CMV (11 patients). Three fungal infections were detected by microbiological approaches in group A (2 × Candida albicans, 1 × Pitysporum ovale) compared to nine fungal infections in group B (5 × Candida albicans, 1 × Candida glabrata, 1 × Candida parapsilosis, 2 × Geotrichum capitatum). Two years after transplantation, 55.9% of patients were alive (group A: 68.2%; group B: 48.6%). Patients with AML had a two-year survival of 50% (group A: 53.3%; group B: 46.7%). In our study, we observed no clear relation between frequency of infection and donor type, yet there was a trend towards more invasive fungal infections in the unrelated group (13% group A vs. 24% group B).

Evidence for CD34+ Hematopoietic Progenitor Cell Involvement in Acute Myeloid Leukemia with NPM1 Gene Mutation: Implications for the Cell of Origin

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 307-307 ◽  
Author(s):  
Maria Paola Martelli ◽  
Valentina Pettirossi ◽  
Elisabetta Bonifacio ◽  
Federica Mezzasoma ◽  
Nicla Manes ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Myeloid Leukemia ◽  
Hox Genes ◽  
Leukemic Cells ◽  
Mutant Protein ◽  
Cell Of Origin ◽  
Hematopoietic Progenitor ◽  
Cd34 Cells ◽  
Acute Myeloid

Abstract Acute myeloid leukemia expressing mutated NPM1 gene and cytoplasmic nucleophosmin (NPMc+ AML) [Falini B et al, NEJM2005;352:254–266] is a new entity of WHO classification that shows distinctive biological and clinical features, including a unique molecular signature characterized by downregulation of CD34 and upregulation of most HOX genes [Falini B et al, Blood2007;109:874–885]. Involvement of HOX genes in the maintenance of the stem-cell phenotype strongly suggest that AML with mutated NPM1 originates from a multipotent hematopoietic progenitor (HSC). This view is also supported by immunohistological findings showing that AML with mutated NPM1 frequently displays multilineage involvement [Pasqualucci L et al, Blood2006;108:4146–4155]. On the other hand, the frequent negativity of NPMc+ AML for the HSC-associated antigen CD34 raises the question of whether the mutation event occurs in a CD34-negative HSC (these cells have been identified in mice) or whether a minimal pool of CD34-positive NPM1-mutated leukemic cells does exist. Currently, the hierarchical level of stem cell involvement in NPMc+ AML is unknown. To address this issue, we purified CD34+ cells from NPMc+ AML patients and detected NPM1 mutant protein in the sorted population by Western blot with anti-NPM mutant specific antibodies [Martelli MP et al, Leukemia 2008] (Figure 1A). We investigated 6 NPMc+ AML patients presenting at diagnosis with 0.12%, 0.14%, 0.38%, 5%, 22%, and 28% of CD34+ cells in the peripheral blood. In all cases, CD34+ fractions (purity &gt;90%) harboured NPM1 mutant protein, indicating they belong to the leukemic clone (Figure 1B). The percentage of most undifferentiated CD34+/CD38− cells in the CD34+ fractions ranged from 5 to 97%. Notably, in at least one case, all CD34+ NPM1-mutated leukemic cells were CD38−negative. Moreover in all cases, CD34+ NPM1-mutated leukemic cells appeared to express CD123 (IL-3 receptor), considered a marker of the leukemic stem cell and target of potential therapy. Double staining of bone marrow biopsies with anti-CD34 and anti-NPM antibodies revealed that the rare CD34+ cells expressed NPM1 aberrantly in the cytoplasm. Inoculation of CD34+ NPM1-mutated AML cells into sublethally irradiated NOD/SCID mice resulted into leukemia engrafment in various body sites, especially bone marrow, spleen, lung and liver. Preliminary results showed that CD34+ leukemic cells reacquired the same leukemic phenotype as the original patient’s, including CD34-negativity of the leukemic bulk in spite of any lack of differentiation. This finding suggests that NPM1 mutant protein may be involved in downregulation of CD34 antigen, while keeping a gene expression profile typical of the hematopoietic stem cell. These findings suggest the CD34+ fraction contains the SCID-leukemia initiating cells (SL-IC) and point to CD34+/CD38− HSC as the cell of origin of AML with mutated NPM1. Figure Figure

A Randomized Clinical Trial Comparing Cytokine Administration Sites for Mobilization of Peripheral Hematopoietic Progenitor Cells for Patients with Hematological Malignancies Undergoing Autologous Stem Cell Transplantation.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4230-4230
Author(s):  
Heather Renfroe ◽  
Edmund K. Waller ◽  
Mike Arnold ◽  
Louette Vaughn ◽  
R. Donald Harvey ◽  
...  
Keyword(s):  
Stem Cell ◽  
Injection Site ◽  
Research Funding ◽  
Randomized Study ◽  
Peripheral Blood Stem Cells ◽  
Cd34 Cells ◽  
Blood Stem Cells ◽  
Group A ◽  
Cell Mobilization ◽  
Group B

Abstract Abstract 4230 Background The optimal injection site for cytokine administration when used to mobilize peripheral blood stem cells for collection is unclear. There are known differences in the pharmacokinetics of subcutaneously injected drugs based upon site adiposity. We hypothesized that injection in lower adipose-tissue-containing sites in the extremities would result in a reduced reservoir effect leading to lower exposures of granulocyte colony stimulating factor (G-CSF) and therefore reduced stem cell collection following cytokine mobilization. Methods We completed a prospective single institution IRB-approved randomized study to determine the efficiency and tolerability of different injection sites among patients with multiple myeloma or lymphoma undergoing stem cell mobilization and apheresis. The primary end-points were the total number of CD34+ cells collected and the number of days of apheresis required to collect target numbers (5 × 10E6 CD34+ cells/kg for patients with lymphoma; 10 × 10E6 CD34+ cells/kg for patients with myeloma). Forty patients were randomized to receive cytokine injections in their abdomen (group A) or extremities (group B). Randomization was stratified based upon diagnosis (myeloma; N=29 vs. lymphoma; N=11), age (≤50; N=13 vs. >50; N=27), and mobilization strategy (cytokines alone; N=27 vs. chemomobilization; N=13). Both group A and B were balanced with respect to the stratification criteria. Filgrastim was planned at a dose of 10 ug/kg/day for patients undergoing chemomobilization or 15 ug/kg/day for patients undergoing cytokine-only mobilization. Actual mean cytokine doses were 11.78 ug/kg/day using chemomobilization and 12.96 ug/kg/day using cytokines alone due to rounding to nearest vial size. Patients recorded the injection site for G-CSF and symptoms daily. Results Of those enrolled, 90% were evaluable with 18 patients in each group. Four were deemed non-evaluable due to failure to proceed to the planned mobilization procedure (1 in group A and 2 in group B) or lack of consistent injection site (1 patient). In addition, one patient in group B received a non-protocol specified injection of plerixafor due to poor mobilization and collected a total of 13.62 × 10E6 CD34+ cells/kg in 2 days of apheresis. Among the 36 evaluable subjects, 1 subject in each group failed collection with a total of < 2.0 × 10E6 CD34+ cells/kg collected. Mean BMI at the time of mobilization was not different between groups A and B (27.25 ± 4.7 versus 29.39 ± 5.7, respectively; p=NS). Mean numbers of CD34+ cells (±SD) collected were not different between groups A and B (9.15 ± 4.7 versus 9.85 ± 5 × 106/kg, respectively; p=NS). The mode and median duration of apheresis was 2 days for both groups. Subjects from both groups reported similar toxicities of pain and discomfort at the injection site. Conclusions Based upon the analysis, G-CSF administration site (extremities versus abdomen), does not affect the number of CD34+ cells collected by apheresis or the duration of apheresis needed to reach the target cell dose. Disclosures: Lonial: Celgene: Consultancy; Millennium: Consultancy, Research Funding; BMS: Consultancy; Novartis: Consultancy; Gloucester: Research Funding.

Acute Myeloid Leukemia with Myelodysplasia-Related Changes (MRC) Did Not Show Inferior Outcomes Compared to Those without MRC,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3563-3563
Author(s):  
Jee Hyun Kong ◽  
Hyun Ae Jung ◽  
Hee Kyung Ahn ◽  
Silvia Park ◽  
Hee-Jin Kim ◽  
...  
Keyword(s):  
Overall Survival ◽  
Acute Myeloid Leukemia ◽  
Induction Chemotherapy ◽  
Myeloid Leukemia ◽  
Risk Group ◽  
Univariate Analysis ◽  
Group A ◽  
Group B ◽  
Group D ◽  
Acute Myeloid

Abstract Abstract 3563 The distinctive features of the World Health organization (WHO) classification compared to French-America-British Co-operative group (FAB) classification of acute myeloid leukemia is the new morphological entity “AML with multilineage dysplasia (MLD)”, and now this subgroup has been renamed as 'AML with myelodysplasia-related change (MRC)”. It generally accepted that dysplasia was most frequently noted in older individual, is often associated with an unfavorable cytogenetic profiles and unfavourable response to therapy. However it is still controversial. Therefore, we evaluated the impact of MRC on overall survival (OS) and leukemia free survival (LFS) in acute myeloid leukemia patients. A total of 644 adult AML patients diagnosed at Samsung Medical Center (SMC) between Sep.1994 and Oct. 2010 were enrolled. We reviewed their medical histories, clinical parameters, hemogram data, bone marrow aspirate and cytogenetic studies, and reclassified them into AML with of MRC and without MRC groups. Of 664 patients, 543 patients were received induction chemotherapy, among them, 84 patients demonstrated MRC and 451 patients did not. Median age was 50 (15–88) years old, and 57.1% of patients were male. Median follow up period was 77.3 [0–191] months. AML without MRC group had more favorable cytogenetic risk, higher WBC counts and LDH levels than those with MRC. However, other variable such as age, sex, hemoglobin level, absolute neutrophil, and peripheral blast count, induction chemotherapy regimen, hematopoietic stem cell transplantation, CR1 (complete response after induction chemotherapy), CRp (complete recovery of platelet), and relapse rate were not different between two groups. Since FLT3-ITD and NPM1 tests were introduced into laboratory work after 2005, results of these tests were available only in 158 and 75 patients respectively, and these were not different between two groups. In univariate analysis, advanced age (>65 years) predicted worse LFS (median LFS [95% C.I.]; ° Â65 years vs >65 years; 9.3[7.2–11.4] vs 5.9[4.6–7.2] months, p =0.014). In terms of OS, young age (p=0.000), female (p=0.000), favorable cytogenetic risk (p=0.000), CR1 (p=0.000), CRp (p=0.000), absence of relapse (p=0.000), and HSCT (p=0.000) showed a higher probability of longer OS (Table 1). The presence of MRC, FLT3-ITD, and NPM1 did not affect OS (Table 1).Table 1.Summary of univariate analysis for overall survival.Median OS (months) [95% C.I.] or mean OS ± SD (months)pAge°Â65 years47.9 [17.1 – 78.6]0.000>65 years13.1 [5.9 – 20.4]SexMale23.2 [16.5 – 30.0]0.000¢Female112.2 [ – ]Cytogenetic risk¢Favorable107.0±7.40.000Intermediate28.0 [19.0 – 36.9]Unfavorable10.8 [7.8 – 13.7]Unknown17.5 [6.8 – 28.1]MRCAbsence35.9 [6.6 – 65.2]0.081Presence19.0 [6.9 – 31.0]CR1¢Yes112.2 [–]0.000No3.6 [1.1 – 6.1]CRp¢Yes70.9±4.40.000No54.8 [18.5 – 91.1]RelapseYes21.5 [17.2 – 25.9]0.000No17.5 [–]HSCT¢Auto86.2±5.60.000¢Allo83.8±6.5Not done17.5 [10.8 – 24.1]FLT3-ITDPositive8.2 [0–26.1]0.595Negative29.5 [20.9–38]NPM1¢Positive104.1±11.00.978¢Negative78.9±15.0¢“median survival not reached Next, we analyzed MRC effect in each variable to OS. The presence MRC did not affect OS of each group which divided according to the age (Figure 1. A and B), sex, cytogenetic risk groups (Figure 1. C and D), relapse, CR1, HSCT, and FLT3-ITD, though AML with MRC group had tendancy to have poor survival rate in intermediate cytogenetic risk group (Figure 1. C). However in patients who did not acheived CRp or showed NPM1, the presence of MRC correlated with shorter OS.Figure 1.Overall survival (OS) according to the presence of MRC in age ° Â65 group (A), age>65 group (B), intermediate cytogenetic risk group (C)), and unfavorable cytogenetic risk group (D).Figure 1. Overall survival (OS) according to the presence of MRC in age ° Â65 group (A), age>65 group (B), intermediate cytogenetic risk group (C)), and unfavorable cytogenetic risk group (D). In this study, patients with MRC did not show inferior outcomes than those without MRC. Therefore it is not necessary to decide different treatment strategy according to the presence of MRC Disclosures: No relevant conflicts of interest to declare.

A Highly Selective Anti-ROR1 Monoclonal Antibody Inhibits Human Acute Myeloid Leukemia CD34+ Cell Survival and Self-Renewal.

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2560-2560
Author(s):  
Larissa Balaian ◽  
Anil Sadarangani ◽  
George F. Widhopf ◽  
Rui-kun Zhong ◽  
Charles Prussak ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Myeloid Leukemia ◽  
The Self ◽  
Self Renewal ◽  
Leukemia Stem Cell ◽  
Cd34 Cells ◽  
Selective Expression ◽  
Dose Dependent ◽  
Acute Myeloid

Abstract Abstract 2560 The mammalian orphan receptor tyrosine kinase-1 (ROR1) is expressed in a wide-variety of tissues during early embryonic development. By the late stages of embryogenesis the expression of this developmentally important protein is greatly diminished. Although not expressed in the tissues of post-partum animals, the ROR1 protein is expressed on neoplastic cells in chronic lymphocytic leukemia (CLL), some B-cell malignancies, and a variety of different carcinomas. We examined for expression of ROR1 in primary acute myeloid leukemia (AML) cells harvested from marrow aspirates and their normal counterparts by whole transcriptome paired-end RNA sequencing and by flow-cytometric analyses. These studies revealed selective expression of ROR1 in 62 (35%) of 179 AML samples examined. Many of these samples were found to have cells that co-expressed ROR1 and CD34, suggesting that ROR1 was present on the self-renewing leukemia stem-cell population, which resides in the marrow niche and potentially accounts for resistance to many cytotoxic drugs used in therapy. We investigated the activity of a chimeric anti-ROR1 mAb found effective in clearing CLL cells (UC99961) on AML expansion, growth, and renewal in a leukemia-stem-cell supportive niche assay. Mouse marrow cells lines SL/SL and M2–10B4 (transfected to produce hSCF,hIL3 and hIL3, hG-CSF respectively) were mixed 1:1 after mitomycin-C treatment, and used as a SLM2 stromal monolayer. CD34+ cells were selected from ROR1-positive (n=6) or negative (n=4) AML primary samples. As a normal control, CD34+ cells from cord blood (CB) were used (CB, n=3). In some experiments CD34+ cells were transfected with a GLP-lentivirus prior to co-culture. At the initiation of the co-culture, 10–50 μg/ml of the chimeric anti-ROR-1 mAb (UC99961) or control hIgG were added to the cultures. Two weeks after co-culture initiation, both stromal attached and floating cells were collected and their survival investigated by colony forming assay in methylcellulose. The UC99961 mAb was not cytotoxic to CB or ROR1-negative AML samples. In contrast, the UC99961 mAb provided a dose-dependent inhibition of colony formation for all ROR-1-positive AML samples examined. These results demonstrate the in vitro anti-leukemic specificity of this anti-ROR1 mAb in down-regulating AML stem and progenitor cell populations, without effecting normal CD34+ stem cells. To analyze the effect of ROR1 ligation on AML stem cell populations exclusively, AML self-renewal assays (2-ry colonies) were performed. In these studies, ROR1–positive AML samples were divided based on their response to mAb treatment. Half of the samples (n=3; 50%) demonstrated statistically significant (up to 90%) dose-dependent decreases in colony formation. However, another half was non-responsive and no correlation was found between ROR1 expression on leukemia CD34+ cells and response to anti-ROR1 mAb treatment in the self-renewal assays. Again UC99961 mAb treatment did not negatively impact CD34+ cells from CB or ROR1-negative AML, confirming the specificity and selective toxicity of the mAb for ROR1+ AML stem cells. These studies reveal selective expression of ROR1 on leukemia-stem-cells of large subset of AML patients. Furthermore, this study demonstrates that an anti-ROR1 mAb (UC99961) can inhibit survival and self-renewal in LSC supportive niche assays. Targeted ROR1 inhibition may represent a vital component of therapeutic strategies aimed at eradicating therapeutically recalcitrant malignant stem cells in AML and potentially other refractory cancer-stem-cell-driven malignancies. Disclosures: No relevant conflicts of interest to declare.

A Pilot Study of Peripheral Blood Hematopoietic Stem Cells Mobilization with the Combination of Bortezomib and G-CSF in Multiple Myeloma and Non-Hodgkin's Lymphoma Patients.

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 1902-1902
Author(s):  
Divaya Bhutani ◽  
Vidya sri Kondadasula ◽  
Joseph P. Uberti ◽  
Voravit Ratanatharathorn ◽  
Lawrence G. Lum ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Stem Cell ◽  
Peripheral Blood ◽  
Median Number ◽  
Peripheral Blood Cd34 ◽  
Cd34 Cells ◽  
Group A ◽  
Cell Mobilization ◽  
Group B ◽  
Stem Cell Collection

Abstract Background: Bortezomib has become an integral part of front-line therapy of multiple myeloma in a large majority of patients. There are preliminary reports which show that addition of bortezomib can augment the peripheral blood CD34 count during stem cell mobilization. In this single center prospective trial we added bortezomib to G-CSF to evaluate the effects of bortezomib on peripheral CD34 counts and collection. Methods: Patients aged 18-70 years with diagnosis of multiple myeloma (MM) or non-hodgkin's lymphoma (NHL) who were eligible for autologous stem cell transplantation (ASCT) and had received no more than three prior chemotherapeutic regimens were eligible for the study. Patients were enrolled in two groups. Group A (N=3) received G-CSF 16mcg/kg for 5 days and proceeded to stem cell collection on D5 and then received bortezomib 1.3mg/m2 on D5 after stem cell collection and G-CSF 16mcg/kg on D6, 7, 8 and repeat stem cell collection on D6, 7, 8 till the goal was achieved. Group B (N=17) received G-CSF 16mg/kg on D1-5 and received bortezomib 1.3mg/m2 on D4 and proceeded to stem cell collection on D5. If the patient was not able to collect the predefined goal CD34, G-CSF was continued on D 6, 7, 8 and a second dose of bortezomib 1.3mg/m2 was given on D7. Mobilization procedure was stopped once the predefined goal CD34 collection (4 x 106/kg for MM and 2 x 106/kg for NHL) had been collected. Primary objectives of the study was to determine if addition of bortezomib to G-CSF will result in an increase in PBSCs by > 2-fold and to achieve median neutrophil engraftment 12 days post ASCT. Secondary objectiveswere to evaluate the collected product for co-mobilization of lymphoma or myeloma cells and to determine if the use of bortezomib increases the mobilization of immune-stimulatory Dendritic cell (DC) -1 subsets. Results: A total of 23 patients were enrolled and 20 were evaluable for the results. Only one patient with NHL was enrolled and rest had MM. Median age of pts was 57 years, M/F 8/12, median number of previous chemotherapy regimens was 1 (range 1-3). The median peripheral blood CD34 count pre and post bortezomib in all patients were 28.8 x 106/kg and 37 x 106/kg respectively. All three patients in group A had drop in peripheral blood CD34 counts on D6 post bortezomib as they had undergone stem cell collection on day 5. In part B (N=17), 15 patients had increase in peripheral blood CD 34+ve cell counts with 4 patients achieved doubling while 11 pts had less than doubling of peripheral blood CD34 count after receiving bortezomib. Two patients had minimal drop in the peripheral blood CD34 counts post bortezomib. Median number of CD34 cells collected in15 patients (part B) were 5.06 x 106 CD34 cells/kg (range 4-15.1). 18 patients proceeded to ASCT and median time to neutrophil engraftment (ANC ≥500/cumm) post transplant was 12 days (range 11-16) and platelet engraftment (Plt count ≥ 20,000/cumm) was 18 days (range 15-27). There was no significant change in DC1/DC2 ratio in both groups following treatment with bortezomib and G-CSF (Figure 1). In group A all three patients collected goal CD34 count on day 5 and 2/3 patients collected >4 x106 CD34 cells/kg on D6 post bortezomib and1/3 patients collected 2.6 x 106 on D6 post bortezomib. In group B (n=17), 2 patients were unable to collect because of low CD34 counts on D4 and D5, 11 pts collected the goal in one day (D 5) and 4 pts required two days of apheresis (D 5 and 6). None of the patients received D7 bortezomib. Conclusion: Use of bortezomib during autologous stem cell collection was safe and well tolerated. Majority of patients had increase in peripheral blood CD34 counts post bortezomib administration on D4. Future trials should explore bortezomib as an alternate strategy to chemo-mobilization in combination with growth factors. Figure 1. DC1/DC2 ratio in group A and group B at various time points. Figure 1. DC1/DC2 ratio in group A and group B at various time points. Figure 2. Figure 2. Disclosures Off Label Use: Bortezomib for stem cell mobilization. Lum:Karyopharm Therapeutics Inc: Equity Ownership; Transtarget.Inc: Equity Ownership. Deol:Bristol meyer squibb: Research Funding. Abidi:celgene: Speakers Bureau; Millenium: Research Funding.

Allogeneic Transplanst for ACUTE Myeloid Leukemia: Cut Off Levels of WT1 Expression in 207 Patients and Pre-Emptive Therapy of Relapse

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 1866-1866
Author(s):  
Carmen Di Grazia ◽  
Simona Geroldi ◽  
Raffaella Grasso ◽  
Maurizio Miglino ◽  
Nicoletta Colombo ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Cell Transplant ◽  
Allogeneic Hsct ◽  
Group A ◽  
Leukemia Relapse ◽  
Group B ◽  
Acute Myeloid ◽  
Marrow Cells

Abstract Leukemia relapse remains a significant problem in patients with AML undergoing an allogeneic stem cell transplant(HSCT). Wilms Tumour 1 (WT1) expression has been shown to be a sensitive marker of minimal residual disease (MRD), both in patients after induction chemotherapy, as well as in patients undergoing an allogeneic HSCT. Hypotheses. The present study had 2 hypotheses: (1) WT1 expression in marrow cells of AML patients post-HSCT, will predict leukemia relapse and (2) WT1 based pre-emptive immunotherapy (IT) such as abrupt cyclosporin discontinuation and/or donor lymphocyte infusion (DLI), will prevent leukemia relapse. Patients. Bone marrow WT1 expression, was monitored in 207 patients with acute myeloid leukemia (AML) before and monthly after an allogeneic HSCT, until day +150, and then at every other outpatient access. Eligible for IT were patients without acute or chronic GvHD, with increased WT1 expression and a a marrow in hematologic remission. The trigger for IT was 180 WT1 copies in a first group of 122 patients (group A): this was based on the fact that WT1 expression in normal bone marrow is up to 180 copies . In a subsequent group of 85 patients (group B) the cut off for IT, was 100 copies, due to the fact that a first analysis of group A had shown 100 copies to be an earlier predictor of relapse (BJH 2013; 160: 503). DLI were given in escalating doses, starting at 1x105 CD3+ cells/kg in alternative donor grafts and at 1x106/kg in HLA identical grafts. DLI were escalated ½ log every month, in the absence of GvHD, to a maximum dose of 1x107/kg. Sixtyfour patients were eligible for IT, but only 35 received IT: reasons for non intervention were ongoing GHD, unavailable donor and delay in WT1 results. Results-Hypothesis N.1. Following transplantation, WT1 expression, was highly predictive of leukemia relapse: 12 relapses in 99 patients with WT1 < 100 copies /104 abl (12%); 19 relapses in 55 patients with WT1 between 101 and 180 copies (35%) and 37 relapses in 53 patients with WT1 >180 copies (70%) (p<0.0001). The median interval between WT1 positivity and relapse was 75 days in group A and 60 days in group B. Results-Hypothesis N.2. 35 patients received pre-emptive immune intervention, 17 in group A and 18 in group B. The latter had more patients beyond first remission at transplant (56% vs 23%) , more myeloablative regimens (100% vs 65%) and more family haploidentical donors (72% vs 6%); age was comparable. The risk of relapse was 13/17 (76%) for group A and 3/18 (17%) for group B (p<0.001), despite the larger proportion of patients beyond CR1 at transplant. GvHD following DLI occurred in 15% of patients. DLI-related mortality was 0%. The overall 3 year survival for patients in group A and B was 69% vs 47% (p=0.3). The relapse risk in patients of group A eligible but not receiving IT (n=21) was 74%; in group B (n=8) it was 50%. In conclusion, WT1 expression post-transplant is a strong predictor of leukemia relapse in patients with AML, and can be used to trigger pre-emptive immunotherapy, in approximately 50% of eligible patients. IT triggered at a WT1 cut-off level of 100 copies in bone marrow cells, is more effective, as compared to180 copies, in preventing leukemia relapse. Disclosures No relevant conflicts of interest to declare.

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