scholarly journals Long-term bone marrow culture in persons with Fanconi anemia and bone marrow failure

Blood ◽  
1994 ◽  
Vol 83 (2) ◽  
pp. 336-339
Author(s):  
A Butturini ◽  
RP Gale
Keyword(s):  
Bone Marrow ◽  
Fanconi Anemia ◽  
Autosomal Recessive ◽  
Autosomal Recessive Disease ◽  
Bone Marrow Failure ◽  
Myelogenous Leukemia ◽  
Acute Myelogenous ◽  
Nonadherent Cells

Fanconi anemia is an autosomal recessive disease characterized by a high risk of developing bone marrow (BM) failure and acute myelogenous leukemia. We studied growth of hematopoietic progenitor cells in long- term BM culture (LTBMC) in 8 persons with Fanconi anemia and BM failure. Although LTBMC were initiated with very few BM cells, an adherent layer formed in cultures from 7 persons. In these cultures, the number of nonadherent cells increased for 10 to 15 days. Cell growth continued until cultures were terminated at day 35 to 40. During the first 2 weeks of culture, most nonadherent cells were differentiated myeloid cells. By days 35 to 40, the adherent layer contained cells able to initiate secondary LTBMCs. These data indicate that hematopoietic precursors cells able to proliferate and differentiate in vitro are present in the BM of persons with Fanconi anemia and BM failure. They suggest that mechanisms other than absent precursor cells are responsible for BM failure in Fanconi anemia.

scholarly journals Long-term bone marrow culture in persons with Fanconi anemia and bone marrow failure

Blood ◽  
1994 ◽  
Vol 83 (2) ◽  
pp. 336-339 ◽  
Author(s):  
A Butturini ◽  
RP Gale
Keyword(s):  
Bone Marrow ◽  
Fanconi Anemia ◽  
Autosomal Recessive ◽  
Autosomal Recessive Disease ◽  
Bone Marrow Failure ◽  
Myelogenous Leukemia ◽  
Acute Myelogenous ◽  
Nonadherent Cells

Abstract Fanconi anemia is an autosomal recessive disease characterized by a high risk of developing bone marrow (BM) failure and acute myelogenous leukemia. We studied growth of hematopoietic progenitor cells in long- term BM culture (LTBMC) in 8 persons with Fanconi anemia and BM failure. Although LTBMC were initiated with very few BM cells, an adherent layer formed in cultures from 7 persons. In these cultures, the number of nonadherent cells increased for 10 to 15 days. Cell growth continued until cultures were terminated at day 35 to 40. During the first 2 weeks of culture, most nonadherent cells were differentiated myeloid cells. By days 35 to 40, the adherent layer contained cells able to initiate secondary LTBMCs. These data indicate that hematopoietic precursors cells able to proliferate and differentiate in vitro are present in the BM of persons with Fanconi anemia and BM failure. They suggest that mechanisms other than absent precursor cells are responsible for BM failure in Fanconi anemia.

scholarly journals Fanconi anemia - learning from children

2011 ◽  
Vol 3 (2s) ◽  
pp. 8 ◽  
Author(s):  
Johanna Svahn ◽  
Carlo Dufour
Keyword(s):  
Bone Marrow ◽  
Fanconi Anemia ◽  
Bone Marrow Failure ◽  
Myelogenous Leukemia ◽  
High Incidence ◽  
Acute Myelogenous ◽  
Progressive Pancytopenia ◽  
Very High ◽  
Head Neck ◽  
Crosslinking Agents

Fanconi Anemia (FA) is a rare autosomic recessive and X-linked disease with chromosomal instability after exposure to crosslinking agents as the hallmark. Clinical features of FA are somatic malformations, progressive bone marrow failure and cancer proneness, however there is wide clinical heterogeneity. The symptom most frequently and early associated with morbidity and mortality is progressive pancytopenia in the first decade of life although acute myelogenous leukemia (AML) or myelodysplastic syndrome (MDS) can appear before aplastic anemia. Squamous cell carcinoma (SCC) of the head-neck, intestinal or genital tract has a very high incidence in FA and can appear at young age. This paper will focus on treatment of bone marrow failure in FA.

scholarly journals Cellular maturation in human preleukemia

Blood ◽  
1978 ◽  
Vol 52 (2) ◽  
pp. 355-361
Author(s):  
HP Koeffler ◽  
DW Golde
Keyword(s):  
Bone Marrow ◽  
Cell Differentiation ◽  
Liquid Culture ◽  
Cultured Cells ◽  
Bone Marrow Cells ◽  
Myelogenous Leukemia ◽  
Bone Marrow Cultures ◽  
Acute Myelogenous ◽  
Marrow Cultures

Bone marrow cells from three preleukemic patients with prominent marrow karyotypic abnormalities were studied in liquid culture to determine if the neoplastic clones were capable of maturation. Parallel cytogenetic and cytologic studies were performed in sequentially harvested bone marrow cultures. Maturation, albeit delayed, occurred in cultures from all three patients. By 14 days of culture in vitro, morphologic, cytochemical, and functional evidence of maturation was observed in about 70% of the cells. By day 21, 85% of the cells were mature by these criteria. All but 2 of 249 metaphases from the cultured cells contained the cytogenetic abnormality of the neoplastic clone. We conclude that some preleukemic cells identified by a chromosomal abnormality can mature in vitro. Preleukemia may be viewed as a syndrome of “early leukemia” in which the neoplastic clone is established and manifested functionally as ineffective hematopoiesis. Hematopoietic cell differentiation becomes progressively abnormal with termination in the nearly complete maturational block characteristic of acute myelogenous leukemia.

scholarly journals APTO-253 Induces KLF4 to Promote Potent in Vitro Pro-Apoptotic Activity in Hematologic Cancer Cell Lines and Antitumor Efficacy As a Single Agent and in Combination with Azacitidine in Animal Models of Acute Myelogenous Leukemia (AML)

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4813-4813 ◽  
Author(s):  
William G Rice ◽  
Avanish Vellanki ◽  
Yoon Lee ◽  
Jeff Lightfoot ◽  
Robert Peralta ◽  
...  
Keyword(s):  
Bone Marrow ◽  
High Risk ◽  
Animal Models ◽  
Antitumor Activity ◽  
Single Agent ◽  
Xenograft Model ◽  
Myelogenous Leukemia ◽  
Acute Myelogenous ◽  
Xenograft Models

Abstract APTO-253, a small molecule that mediates anticancer activity through induction of the Krüppel-like factor 4 (KLF4) tumor suppressor, is being developed clinically for the treatment of acute myelogenous leukemia (AML) and high risk myelodysplastic syndromes (MDS). APTO-253 was well tolerated in a Phase I study in patients with solid tumors using a dosing schedule of days 1, 2, 15, 16 of a 28 day cycle (2T-12B-2T-12B), but recent scientific observations guided APTO-253 toward AML and high risk MDS. Indeed, suppression of KLF4 was reported as a key driver in the leukemogenesis of AML and subsets of other hematologic diseases. The vast majority (~90%) of patients with AML aberrantly express the transcription factor CDX2 in human bone marrow stem and progenitor cells (HSPC) (Scholl et al., J Clin Invest. 2007, 117(4):1037-48). The CDX2 protein binds to CDX2 consensus sequences within the KLF4 promoter, thereby suppressing KLF4 expression in HSPC (Faber et al., J Clin Invest. 2013, 123(1):299-314). Based on these observations, the anticancer activity of APTO-253 was examined in AML and other hematological cancers. APTO-253 showed potent antiproliferative activity in vitro against a panel of blood cancer cell lines, with ηM IC50values in AML (6.9 - 305 ηM), acute lymphoblastic leukemia and chronic myeloid leukemia (39 – 250 ηM), non-Hodgkin’s lymphoma (11 – 190 ηM) and multiple myeloma (72 – 180 ηM). To explore in vivo efficacy, dose scheduling studies were initially conducted in the H226 xenograft model in mice. In the H226 model, APTO-253 showed improved antitumor activity when administered for two consecutive days followed by a five day break from dosing (2T-5B) each week, i.e. on days 1,2, 8,9, 15,16, 22,23, compared to the 2T-12B-2T-12B schedule. The 2T-5B schedule was used to evaluate antitumor activity of APTO-253 in several AML xenograft models in mice. In Kasumi-1 AML and KG-1 AML xenograft models, APTO-253 showed significant antitumor activity (p = 0.028 and p=0.0004, respectively) as a single agent when administered using the 2T-5B schedule each week for four weeks compared to control animals. Mice treated with APTO-253 had no overt toxicity based on clinical observations and body weight measurements. Mice bearing HL-60 AML xenograft tumors were treated with APTO-253 for one day or two consecutive days per week for three weeks, either as a single agent or combined with azacitidine, or with azacitidine alone twice per week (on days 1,4, 8, 11, 15 and 18). APTO-253 as a single agent inhibited growth of HL-60 tumors to approximately the same extent as azacitidine. Furthermore, both once weekly and twice weekly dosing of APTO-253 in combination with azacitidine resulted in significantly enhanced antitumor activity relative to either single agent alone (p = 0.0002 and p = 0.0006 for 1X and 2X weekly APTO-253 treatment, respectively, compared to control). Likewise, using a THP-1 AML xenograft model, APTO-253 administered as a single agent using the 2T-5B per week schedule showed significant efficacy, similar to that of azacitidine, while the combination of APTO-253 and azacitidine demonstrated greatly improved antitumor effects relative to either drug alone. APTO-253 was effective and well tolerated as a single agent or in combination with azacitidine in multiple AML xenograft models, plus APTO-253 does not cause bone marrow suppression in animal models or humans. Taken together, our results indicate that APTO-253 may serve as a targeted agent for single agent use and may provide enhanced efficacy to standard of care chemotherapeutics for AML and other hematological malignancies. Disclosures Rice: Lorus Therapeutics Inc.: Employment. Vellanki:Lorus Therapeutics Inc.: Employment. Lee:Lorus Therapeutics Inc.: Employment. Lightfoot:Lorus Therapeutics Inc.: Employment. Peralta:Lorus Therapeutics Inc.: Employment. Jamerlan:Lorus Therapeutics Inc.: Employment. Jin:Lorus Therapeutics Inc.: Employment. Lum:Lorus Therapeutics Inc.: Employment. Cheng:Lorus Therapeutics Inc.: Employment.

scholarly journals Accelerated myeloblast destruction and abnormal lysosome disruption in cultured bone marrow: association with indolent acute myelogenous leukemia

Blood ◽  
1976 ◽  
Vol 48 (1) ◽  
pp. 23-32 ◽  
Author(s):  
RF Branda ◽  
HS Jacob ◽  
SD Douglas ◽  
CF Moldow ◽  
RR Puumala
Keyword(s):  
Bone Marrow ◽  
Acute Myelogenous Leukemia ◽  
Myelogenous Leukemia ◽  
Leukemic Cells ◽  
Electron Microscopic ◽  
Granule Formation ◽  
Microscopic Studies ◽  
Acute Myelogenous ◽  
Primary Granules

Abstract Despite no chemotherapy and a marrow morphologically typical of frank relapse, an acute myelogenous leukemia (AML) patient survived for nearly 1 yr. During this time she remained asymptomatic and maintained nearly normal levels of platelets and hemoglobin. Cytochemical and electron microscopic studies of her bone marrow in liquid culture revealed on several occasions a unique maturational sequence in that leukemic cells differentiated to form morphologically abnormal primary granules which appeared to rupture and cause cytolysis of these cells. In these cultures, blasts rapidly disappeared and were replaced by more mature granulocytes, in contrast to observations in cultures derived from five other patients with AML in relapse which showed persistently elevated blast counts with no evidence of maturation in vitro. These findings support the concept that in AML cell maturation is regularly impaired and in some cases also aberrant. In addition, the abnormal granule formation with autolysis of the leukemic cells observed in one patient may explain both the early cell death in vitro and this patient's relatively indolent clinical course. Similar in vitro studies may help predict atypical clinical courses in patients with AML and facilitate design of appropriate chemotherapy.

Similarities in long-term cultures of blood and bone marrow from patients with acute myelogenous leukemia

1991 ◽  
Vol 9 (5) ◽  
pp. 461-473 ◽  
Author(s):  
Hector Mayani ◽  
Gerald G. Miller ◽  
Anna Janowska-Wieczorek ◽  
A. Robert Turner ◽  
Andrew R. Belch ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Acute Myelogenous Leukemia ◽  
Myelogenous Leukemia ◽  
Acute Myelogenous

Xeroderma Pigmentosum as a Model for Treatment of Bone Marrow Failure in Fanconi Anemia and Aplastic Anemia.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4235-4235
Author(s):  
W. Clark Lambert ◽  
Santiago A. Centurion
Keyword(s):  
Cell Cycle ◽  
Bone Marrow ◽  
Fanconi Anemia ◽  
Xeroderma Pigmentosum ◽  
Bone Marrow Failure ◽  
S Phase ◽  
White Female ◽  
Cross Linking

Abstract We have previously shown that the primary cell cycle defect in the inherited, cancer-prone, bone marrow failure associated disease, Fanconi anemia (FA), is not in the G2 phase of the cell cycle, as had been thought for many years, but rather in the S phase. FA cells challenged with the DNA cross-linking agent, psoralen coupled with long wavelength, ultraviolet (UVA) radiation (PUVA), fail to slow their progression through the S phase of the subsequent cell cycle, as do normal cells. FA cells are extremely sensitive to the cytotoxic and clastogenic effects of DNA cross-linkers, such as PUVA, so much so that the diagnosis of FA is based on an assay, the “DEB test”, in which cells are examined for clastogenic and cytotoxic effects of diepoxybutane (DEB), a DNA cross-linking agent. More recently, we have shown that artificially slowing the cell cycle of FA cells exposed to PUVA by subsequent treatment with agents which slow their progression through S phase leads to markedly increased viability and reduced chromosome breakage in vitro. We now show that similar results can be obtained in vivo in patients with another DNA repair deficiency disease, xeroderma pigmentosum (XP), a recessively inherited disorder associated with defective repair of sunlight induced adducts in the DNA of sun-exposed tissues followed by development of numerous mutations causing large numbers of cancers in these same tissues. We treated two patients with XP, a light complected black male and a white female, both 14 years of age, in sun-exposed areas with 5-fluorouracil, an inhibitor of DNA synthesis, daily for three months. In contrast to normal patients, who only show clinical results if an inflammatory response is invoked, marked improvement in the clinical appearance of the skin was seen with no inflammation observed. This effect was confirmed histologically by examining epidermis adjacent to excised lesions in sun-exposed areas and further verified by computerized image analysis. Treatment with agents that slow progression through S phase, such as hydroxyurea, may similarly improve clinical outcomes in patients with FA or others who are developing bone marrow failure.

Suppression of Cyclin D 1 (CD1) by on 01910.Na Is Associated with Decreased Survival or Trisomy 8 Myelodysplastic Bone Marrow: A Potential Targetted Therapy for Trisomy 8 MDS.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1651-1651
Author(s):  
Aarthi Shenoy ◽  
Loretta Pfannes ◽  
Francois Wilhelm ◽  
Manoj Maniar ◽  
Neal Young ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cyclin D1 ◽  
Cultured Cells ◽  
Bone Marrow Failure ◽  
Refractory Anemia ◽  
Trisomy 8 ◽  
Monosomy 7 ◽  
Diploid Cells

Abstract CD34 positive cells from patients with trisomy 8 myelodysplastic syndrome (MDS) have pronounced expression of early apoptotic markers compared to normal hematopoietic cells. However, trisomy 8 clones persist in patients with bone marrow failure and expand following immunosuppression (Sloand EM et al; Blood2005; 106(3):841). We have demonstrated up-regulation of c-myc, survivin, and CD1 in CD34 cells of patients with trisomy 8 (Sloand et al; Blood2007; 109(6):2399). Employing siRNA mediated knockdown of the anti-apoptotic protein survivin, we demonstrated a decrease in trisomy 8 cell growth and postulated that increased Cyclin D1 caused the upregulation of survivin resulting in resistance of these cells to apoptosis. Using fluorescent in situ hybridization (FISH) we showed that the novel styryl sulfone, ON 01910.Na (Vedula MS et al; European Journal of Medicinal Chemistry2003;38:811), inhibits cyclin D1 accumulation and is selectively toxic to trisomy 8 cells while promoting maturation of diploid cells. Flow cytometry of cultured cells demonstrated increased proportions of mature CD15 positive myeloid cells and decreased number of immature CD33+ cells or CD34+ blasts (Sloand EM et al; Blood2007;110:822). These encouraging in vitro data led to a phase I/II trial of ON 01910.Na in MDS patients with refractory anemia with excess blasts who had IPSS =/> int-2. This study was designed to assess the safety, and activity of escalating doses of ON 01910.Na (800 mg/m2/day × 3 days, 800 mg/m2/day × 5 days, 1500 mg/m2/day × 5 days, 1800 mg/m2/day × 5 days every 2 weeks) in MDS patients. To date five MDS patients have been treated with ON 01910.Na for 4 to 16 weeks in the first two dose cohorts. Two patients had isolated trisomy 8, two had complex cytogenetic abnormalities including trisomy 8 in all aneuploid cells, and one had monosomy 7. Three and five day infusions were well tolerated. Pharmakokinetic analysis showed that the half life of the drug is 1.3 ± 0.5 hours without signs of drug accumulation. Four of five patients demonstrated a rapid and significant decrease in the number of peripheral blasts and aneuploid cells after 4 weeks of therapy (see below), concomitantly with increases in neutrophil and/or platelet counts in four patients. All four patients exhibiting a biological effect of drug treatment had trisomy 8 in their aneuploid clone prior to therapy. One monosomy 7 patient, previously refractory to EPO became responsive to Darbopoietin and another trisomy 8 patient became platelet-transfusion independent. In this early safety trial, ON 01910.Na demonstrates efficacy at early timepoints with respect to improved cytopenias and decreased blast counts. Continued enrollment and long term follow-up will further detail clinical efficacy and impact on the long term prognosis of high risk MDS patients treat with this drug. Figure Figure

Clonal Evolution In Fanconi Anemia; The Role of HOXA9

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 126-126
Author(s):  
Grover C Bagby ◽  
Jane Yates ◽  
Praveen Anur ◽  
Keaney Rathbun ◽  
Christina Harrington ◽  
...  
Keyword(s):  
Fanconi Anemia ◽  
Bone Marrow Failure ◽  
Clonal Evolution ◽  
Ectopic Expression ◽  
Gene Signature ◽  
Inhibitory Response ◽  
Myelogenous Leukemia ◽  
Analytical Approaches ◽  
Marrow Cells

Abstract Abstract 126 Bone marrow failure in patients with Fanconi anemia (FA) is often complicated by the clonal neoplasms myelodysplasia (MDS) and acute myelogenous leukemia (AML). FA stem cells (HSC) exhibit self-replicative defects and HSC and committed progenitor cells (HPC) from Fancc-deficient (Fancc −/−) mice and FA-C-deficient patients are hypersensitive to suppressive cytokines, particularly TNFα, a cytokine also known to be overproduced by Fancc−/− macrophages. In fact, in Fancc−/− mice overproduction of TNFα plays an important role in both marrow failure and evolution of TNF-resistant neoplastic clones. To test the notion that evolution of clones in patients with FA might follow a like adaptive pathway to TNF-resistance, we performed gene expression microarray analysis (Affymetrix GeneChip HG-U133A [22,283 probe sets] arrays) using RNA obtained from low-density marrow cells from; (a) normal volunteers (NV, n=11) (b) FA patients with clonal cytogenetic defects detected in at least 50% of metaphases (FA-CL, n=10) and (c) FA patients with no detectable cytogenetic defects (FA-APL, n=22). Data were normalized using the MAS 5.0 global scaling method with the target intensity of 325. The adapted nature of evolved clones was supported by unsupervised hierarchical clustering (GeneSifter) and principal component analyses (Partek) which revealed a high degree of relatedness between the NV and FA-CL samples while FA-APL samples were more distant from both NV and FA-CL in transcriptomal space. To identify genes whose differential expression might play a role in clonal evolution, two analytical approaches were taken. First, using the shrunken centroids method we identified a 30 gene signature that distinguished FA-CL and FA-APL samples. Secondly, we applied two filters to the data. The first revealed 2317 genes that were either over-expressed or under-expressed (>1.5 fold, p<0.05, adjusted for multiple comparisons using the Benjamini-Hochberg method) in a pairwise comparison of FA-APL to NV samples. The second filter utilized a pattern reading tool (GeneSifter) to identify those genes on the first list of 2317 whose expression did not differ in the comparison between NV and FA-CL samples. The majority (959 of 1063) of these genes were those whose expression was suppressed in FA-APL samples and either not-suppressed or overexpressed in the FA-CL samples. Three genes of interest were in the 30 gene signature identified using the shrunken centroids method and in those identified using the pattern reading tool. Specifically, HOXA9, a gene with known leukemogenic potential, and two genes known to enhance the expression of HOXA9 (MYB and KDM5B) were suppressed in FA-APL samples and over-expressed in the FA-CL samples. After confirming the RNA results using quantitative real time RT-PCR and subsequently determining that the differentially expressed genes in the FA-CL samples included most of the genes known to be regulated by HOXA9 expression in CD34+ marrow cells (Dorsam,S.T. et al, Blood 2004) (p<0.05, Fisher's Exact P.T.) we demonstrated the following: (a) retroviral expression of HOXA9 in normal human CD34+ marrow cells enhanced TNFα resistance in CFU-GM and BFU-E, (b) 3-day in vitro exposure of kit+/lin−/sca-1+ (KLS) cells from Fancc−/− mice to TNFα and multilineage growth factors (MGF; IL-11, Flt3L, SCF, IL-6) suppressed expression of HoxA9 and suppressed survival of KLS cells but enhanced expression of HoxA9 and enhanced survival of wild-type KLS cells, (c) ectopic expression of HOXA9 in Fancc−/− KLS cells enhanced KLS survival and blunted the inhibitory response to TNFα in 3-day suspension cultures with MGF. Taken together, these results indicate that clonal evolution in FA stem cell pools represents the ascendance of a fundamentally adapted clone and that over-expression of HOXA9 in the clonal progeny may represent at least one early step in the pathway of clonal adaptation. Disclosures: Pasquini: Novartis: Consultancy, Honoraria; Bristol-Myers Squibb: Consultancy, Honoraria.

scholarly journals Knockdown of Fanconi anemia genes in human embryonic stem cells reveals early developmental defects in the hematopoietic lineage

Blood ◽  
2010 ◽  
Vol 115 (17) ◽  
pp. 3453-3462 ◽  
Author(s):  
Asmin Tulpule ◽  
M. William Lensch ◽  
Justine D. Miller ◽  
Karyn Austin ◽  
Alan D'Andrea ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Embryonic Stem Cells ◽  
Human Embryonic Stem Cells ◽  
Fanconi Anemia ◽  
Bone Marrow Failure ◽  
Embryonic Stem ◽  
Developmental Defects ◽  
Hematopoietic Development

Abstract Fanconi anemia (FA) is a genetically heterogeneous, autosomal recessive disorder characterized by pediatric bone marrow failure and congenital anomalies. The effect of FA gene deficiency on hematopoietic development in utero remains poorly described as mouse models of FA do not develop hematopoietic failure and such studies cannot be performed on patients. We have created a human-specific in vitro system to study early hematopoietic development in FA using a lentiviral RNA interference (RNAi) strategy in human embryonic stem cells (hESCs). We show that knockdown of FANCA and FANCD2 in hESCs leads to a reduction in hematopoietic fates and progenitor numbers that can be rescued by FA gene complementation. Our data indicate that hematopoiesis is impaired in FA from the earliest stages of development, suggesting that deficiencies in embryonic hematopoiesis may underlie the progression to bone marrow failure in FA. This work illustrates how hESCs can provide unique insights into human development and further our understanding of genetic disease.

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