Lentiviral gene transfer of TCIRG1 into peripheral blood CD34+ cells restores osteoclast function in infantile malignant osteopetrosis

Bone ◽  
2013 ◽  
Vol 57 (1) ◽  
pp. 1-9 ◽  
Author(s):  
Ilana Moscatelli ◽  
Christian Schneider Thudium ◽  
Carmen Flores ◽  
Ansgar Schulz ◽  
Maria Askmyr ◽  
...  
Keyword(s):  
Gene Transfer ◽  
Peripheral Blood ◽  
Peripheral Blood Cd34 ◽  
Cd34 Cells ◽  
Malignant Osteopetrosis ◽  
Osteoclast Function ◽  
Infantile Malignant Osteopetrosis

Gene Transfer to CD34+ Cells From Peripheral Blood Restores Osteoclast Function in Infantile Malignant Osteopetrosis

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2058-2058
Author(s):  
Ilana Moscatelli ◽  
Christian Thudium ◽  
Maria Askmyr ◽  
Ansgar S Schulz ◽  
Oscar Porras ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Gene Transfer ◽  
Peripheral Blood ◽  
Bone Marrow Failure ◽  
Transcriptional Level ◽  
Hematopoietic Stem ◽  
Cd34 Cells ◽  
Malignant Osteopetrosis ◽  
Infantile Malignant Osteopetrosis ◽  
Hsc Transplantation

Abstract Abstract 2058 PURPOSE: Infantile malignant osteopetrosis (IMO) is a rare, lethal, autosomal recessive disorder characterized by nonfunctional osteoclasts. More than 50% of the patients have mutations in the TCIRG1 gene, encoding for the a3 subunit of a proton pump used by the osteoclast to acidify the resorption area. As a consequence of the lack of resorption, remodeling of bone is severely hampered, which results in dense and fragile bone. This, in turn, causes bone marrow failure followed by anemia and hepatosplenomegaly. The only curative treatment for IMO is HSC transplantation, but this form of therapy is associated with high mortality, especially when an HLA-identical donor is not available. IMO is thus a candidate disease for development of gene therapy because of its fatal outcome early in life if treatment with HSC transplantation is not possible. We have previously shown that the murine oc/oc disease model of osteopetrosis can be rescued by gene therapy targeting hematopoietic stem cells (Johansson et al, Blood 2007). The aim of the present study was to rescue the phenotype of human IMO osteoclasts by lentiviral mediated gene transfer of the TCIRG1 cDNA. METHODS AND RESULTS: CD34+ cells from peripheral blood of three IMO patients were isolated without need for mobilization as they have high levels (around 3%) of circulating blood progenitors (Steward et al, Biol Blood Marrow Transplant. 2005). These cells were cultured in SFEM medium with 50 ng/ml M-CSF, 30 ng/ml GM-CSF, 10 ng/ml IL-6, 200 ng/ml SCF and 50 ng/ml Flt3L for 2 weeks. During culture the cells expanded 500 fold and gradually lost CD34 expression while 50% became positive for CD14, a marker for osteoclast precursors. The cells were transduced with SIN lentiviral vectors expressing either endogenous or codon optimized TCIRG1, plus GFP, under a SFFV promotor. The transduction efficiency was approximately 40% at 2 weeks. Cells were then differentiated to mature osteoclasts by culturing for 10 days on bone slices with α-MEM containing 10% serum, 50 ng/ml M-CSF and 50 ng/ml RANKL. Expression of GFP was retained throughout differentiation. qPCR analysis and western blot revealed increased mRNA and protein levels of TCIRG1 compared to controls. Interestingly the protein appeared only at the end of the differentiation protocol suggesting regulation at the post-transcriptional level, a phenomenon that is under further investigation. Vector-corrected IMO osteoclasts generated increased Ca2+ release and bone degradation products such as C-telopeptide of type 1 collagen (CTX-1) into the media, while non-corrected IMO osteoclasts failed to resorb bone. Resorption per osteoclast (CTX-1/TRAP ratio) was 20–50% of that of osteoclasts derived from normal CD34+ cord blood cells and about 2–6 fold higher than that of osteoclasts derived from non-transduced IMO CD34+ cells. CONCLUSION: In conclusion we provide the first in vitro evidence of lentiviral-mediated correction of a genetic disease involving the osteoclast lineage, supporting further development of gene therapy of IMO and other diseases affecting these cells. Disclosures: Richter: Novartis: Honoraria; Bristol-Myers Squibb: Honoraria.

Gene transfer to CD34+ blood cells restores osteoclast function in infantile malignant osteopetrosis

Bone ◽  
2012 ◽  
Vol 50 ◽  
pp. S37 ◽  
Author(s):  
C.S. Thudium⁎ ◽  
I. Moscatelli ◽  
M. Askmyr ◽  
A. Schulz ◽  
O. Porras ◽  
...  
Keyword(s):  
Gene Transfer ◽  
Blood Cells ◽  
Malignant Osteopetrosis ◽  
Osteoclast Function ◽  
Infantile Malignant Osteopetrosis

High Level of Retrovirus-Mediated Gene Transfer into Dendritic Cells Derived from Cord Blood and Mobilized Peripheral Blood CD34+ Cells

1999 ◽  
Vol 10 (2) ◽  
pp. 175-187 ◽  
Author(s):  
Mojgan Movassagh ◽  
Claude Baillou ◽  
Francois Loic Cosset ◽  
David Klatzmann ◽  
Martine Guigon ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Gene Transfer ◽  
Cord Blood ◽  
Peripheral Blood ◽  
Peripheral Blood Cd34 ◽  
Cd34 Cells ◽  
High Level ◽  
Mobilized Peripheral Blood

Successful transfer of ADA gene in vitro into human peripheral blood CD34+cells by transfecting EBV-based episomal vectors

FEBS Letters ◽  
1998 ◽  
Vol 441 (1) ◽  
pp. 39-42 ◽  
Author(s):  
Etsuko Satoh ◽  
Hideyo Hirai ◽  
Tohru Inaba ◽  
Chihiro Shimazaki ◽  
Masao Nakagawa ◽  
...  
Keyword(s):  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Peripheral Blood Cd34 ◽  
Cd34 Cells ◽  
Episomal Vectors ◽  
Successful Transfer

scholarly journals Lower Hematopoietic Progenitor Cell Counts and Yields at Subsequent Donations Is Influenced By a Shorter Inter-Donation Interval between the First and Subsequent Mobilizations

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1962-1962
Author(s):  
Sandhya R. Panch ◽  
Brent R. Logan ◽  
Jennifer A. Sees ◽  
Bipin N. Savani ◽  
Nirali N. Shah ◽  
...  
Keyword(s):  
Stem Cell ◽  
Board Of Directors ◽  
Peripheral Blood ◽  
Research Funding ◽  
Time Interval ◽  
Advisory Committees ◽  
Hematopoietic Stem ◽  
Peripheral Blood Cd34 ◽  
Cell Counts ◽  
Cd34 Cells

Introduction: Approximately 7% of unrelated hematopoietic stem cell (HSC) donors are asked to donate a subsequent time to the same or different recipient. In a recent large CIBMTR study of second time donors, Stroncek et al. incidentally found that second peripheral blood stem cell (PBSC) collections had lower total CD34+ cells, CD34+ cells per liter of whole blood processed, and CD34+ cells per kg donor weight. Based on smaller studies, the time between the two independent PBSC donations (inter-donation interval) as well as donor sex, race and baseline lymphocyte counts appear to influence CD34+ cell yields at subsequent donations. Our objective was to retrospectively evaluate factors contributory to CD34+ cell yields at subsequent PBSC donation amongst NMDP donors. Methods. The study population consisted of filgrastim (G-CSF) mobilized PBSC donors through the NMDP/CIBMTR between 2006 and 2017, with a subsequent donation of the same product. evaluated the impact of inter-donation interval, donor demographics (age, BMI, race, sex, G-CSF dose, year of procedure, need for central line) and changes in complete blood counts (CBC), on the CD34+ cell yields/liter (x106/L) of blood processed at second donation and pre-apheresis (Day 5) peripheral blood CD34+ cell counts/liter (x106/L) at second donation. Linear regression was used to model log cell yields as a function of donor and collection related variables, time between donations, and changes in baseline values from first to second donation. Stepwise model building, along with interactions among significant variables were assessed. The Pearson chi-square test or the Kruskal-Wallis test compared discrete variables or continuous variables, respectively. For multivariate analysis, a significance level of 0.01 was used due to the large number of variables considered. Results: Among 513 PBSC donors who subsequently donated a second PBSC product, clinically relevant decreases in values at the second donation were observed in pre-apheresis CD34+ cells (73.9 vs. 68.6; p=0.03), CD34+cells/L blood processed (32.2 vs. 30.1; p=0.06), and total final CD34+ cell count (x106) (608 vs. 556; p=0.02). Median time interval between first and second PBSC donations was 11.7 months (range: 0.3-128.1). Using the median pre-apheresis peripheral blood CD34+ cell counts from donation 1 as the cut-off for high versus low mobilizers, we found that individuals who were likely to be high or low mobilizers at first donation were also likely to be high or low mobilizers at second donation, respectively (Table 1). This was independent of the inter-donation interval. In multivariate analyses, those with an inter-donation interval of >12 months, demonstrated higher CD34+cells/L blood processed compared to donors donating within a year (mean ratio 1.15, p<0.0001). Change in donor BMI was also a predictor for PBSC yields. If donor BMI decreased at second donation, so did the CD34+cells/L blood processed (0.74, p <0.0001). An average G-CSF dose above 960mcg was also associated with an increase in CD34+cells/L blood processed compared to donors who received less than 960mcg (1.04, p=0.005). (Table 2A). Pre-apheresis peripheral blood CD34+ cells on Day 5 of second donation were also affected by the inter-donation interval, with higher cell counts associated with a longer time interval (>12 months) between donations (1.23, p<0.0001). Further, independent of the inter-donation interval, GCSF doses greater than 960mcg per day associated with higher pre-apheresis CD34+ cells at second donation (1.26, p<0.0001); as was a higher baseline WBC count (>6.9) (1.3, p<0.0001) (Table 2B). Conclusions: In this large retrospective study of second time unrelated PBSC donors, a longer inter-donation interval was confirmed to be associated with better PBSC mobilization and collection. Given hematopoietic stem cell cycling times of 9-12 months in humans, where possible, repeat donors may be chosen based on these intervals to optimize PBSC yields. Changes in BMI are also to be considered while recruiting repeat donors. Some of these parameters may be improved marginally by increasing G-CSF dose within permissible limits. In most instances, however, sub-optimal mobilizers at first donation appear to donate suboptimal numbers of HSC at their subsequent donation. Disclosures Pulsipher: CSL Behring: Membership on an entity's Board of Directors or advisory committees; Miltenyi: Research Funding; Bellicum: Consultancy; Amgen: Other: Lecture; Jazz: Other: Education for employees; Adaptive: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Medac: Honoraria. Shaw:Therakos: Other: Speaker Engagement.

Abstract 32: Intravenous Mobilized Autologous Peripheral Blood CD34+ Stem Cell Transplantation for Angina

2016 ◽  
Vol 119 (suppl_1) ◽  
Author(s):  
Hadyanto Lim ◽  
Lindarto Dharma ◽  
Zein Umar ◽  
Hariaji Ilham
Keyword(s):  
Stem Cell ◽  
Angina Pectoris ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Peripheral Blood ◽  
Exercise Tolerance ◽  
Exercise Stress ◽  
Peripheral Blood Cd34 ◽  
Cd34 Cells ◽  
Angina Frequency

Background: Intramyocardial CD34+ stem cell therapy for patients with refractory angina shows that this is safe and feasible. We aimed to determine whether intravenous transplantation of mobilized autologous peripheral blood CD34+ stem cells provides beneficial effects for patients with angina pectoris. Methods: We administered granulocyte colony stimulating factor (G-CSF, 5.0 μg/kg/day) subcutaneously once a day for 4 days to 15 patients (4 women and 11 men aged 50-78 years) with intractable angina pectoris (Canadian Cardiovascular Society functional class III-IV) for mobilization of CD34+ cells into the peripheral blood. Ischemia was assessed by exercise stress testing. Leukapheresis procedure was started on the day 4 of G-CSF using the Spectra Optia cell separator. Circulating and intravenous transplantation of autologous CD34+ cells after leukapheresis were measured by flow cytometry. The effects of G-CSF on blood were measured by hematology analyzer and semi-auto chemistry analyzer. Results: Intravenous peripheral blood CD34+ cells increased after leukapheresis (from 1.12±0.48 cells/μL to 107.42±23.83 cells/μL, p<0.001) and total white blood cells count (from 7.82 ± 2.63x10 3 /μl to 37.47±15.07 x10 3 /μl, p<0.001). Indices of hsCRP, platelets, hemoglobin, alanine aminotransferase, lactic dehydrogenase, and uric acid were not changed by treatment. At week 4, angina frequency was significantly lower after intravenous CD34+ cells (from 15.07±4.03 to 3.27±1.49, p<0.001). Similarly, improvement in exercise tolerance was significantly higher by stem cell transplantation (from 5.90±2.53 minutes to 8.41±2.49 minutes, p<0.001). Most patients reported mild myalgia which were easily managed with acetaminophen. Conclusions: Intravenous autologous CD34+ stem cell transplantation improved angina frequency and exercise tolerance. The cell mobilization and leukapheresis procedures were found safe and tolerable in patients with angina pectoris.

scholarly journals Increased number of peripheral blood CD34+cells in lithium‐treated patients

1998 ◽  
Vol 100 (1) ◽  
pp. 219-221 ◽  
Author(s):  
Ami Ballin ◽  
Dan Lehman ◽  
Pinkhas Sirota ◽  
Ulia Litvinjuk ◽  
Dina Meytes
Keyword(s):  
Peripheral Blood ◽  
Peripheral Blood Cd34 ◽  
Cd34 Cells
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