Use of Cell-Free Retroviral Vector Preparations for Transduction of Cells from the Marrow of Chronic Phase and Blast Crisis Chronic Myelogenous Leukemia Patients and from Normal Individuals

Abstract

Marrow cells were exposed to the LNL6 or G1N safety-modified variants of the N2 retrovirus, which contain the G418 bacterial resistance gene neo. The frequency of acquisition of the G418 resistance phenotype following exposure to LNL6 or G1N was compared among hematopoietic progenitor cells from the marrow of patients with chronic phase chronic myelogenous leukemia (CML), blast crisis CML, or from nonleukemic individuals. Under the conditions of our experiments, the myeloid committed progenitor cells from 3 of 6 nonleukemic individuals, 9 of 18 chronic-phase CML patients, and 2 of 4 blast crisis CML patients acquired resistance to at least 1 mg/ml G418 following incubation with cell-free supernatants from the PA317 LNL6 or PA317 G1N producer cell lines. Ten of the 32 colonies growing up in 0.8 mg/ml G418 from chronic-phase marrow exposed to LNL6 were shown to contain the neo gene by polymerase chain reaction (PCR) assay of DNA. These results were consistent with estimates of the transduction frequency based on acquisition of resistance to G418 as the number of colonies growing under G418 selection was always greater at 0.8 mg/ml G418 than at higher concentrations of G418 (1.0-1.4 mg/ml). The average transduction frequency at each G418 concentration (1.0, 1.2, and 1.4 mg/ml) in cells from blast crisis CML cells ranged from 2 to 14%, as measured by acquisition of G418 resistance. Chronic-phase CML showed slightly lower average frequencies of transduction (0.6-2.8% of the colonies are G418 resistant). The average transduction frequency of cells from nonleukemic marrow was as high as that seen from the marrow of chronic-phase CML individuals.(ABSTRACT TRUNCATED AT 250 WORDS)

Use of Safety-Modified Retroviruses to Introduce Chemotherapy Resistance Sequences into Normal Hematopoietic Cells for Chemoprotection During the Therapy of Breast Cancer: A Pilot Trial. M.D. Anderson Cancer Center, Houston, Texas

“…Our instkution has had experience with transplantation with the CD34 selected cells. We have seen recovery of the absolute neutrophil count to 500/cu mm in 10, 14, 14, and 17 days in breast cancer patients and 14,17,18,18,24, and 27 in advanced CML patients who have been transplanted with CD34 selected peripheral blood and martOW cells. The University of Denver has observed an average of 17 days to 500 neutrophils/cu mm in advanced breast cancer patients.…”

“…Anderson, and have at the moment three protocols approved by the NIH Recombinant DNA Advisory Committee, and one protocol FDA-approved and ongoing at the M.D. Anderson Cancer Center at this time (16)(17)(18)(19). We have shown in oitt own laboratory that we can achieve ttansduction frequencies in the 1-5% range even with the normal hematopoietic progenitor cells (17).…”

“…Anderson Cancer Center at this time (16)(17)(18)(19). We have shown in oitt own laboratory that we can achieve ttansduction frequencies in the 1-5% range even with the normal hematopoietic progenitor cells (17). Malcolm Brenner of St. Jude Children's Research Hospital has shown that these vectors, when used to modify the manow of AML patients in remission, lead to a hematopoietic population following ttansplant which is positive for the rettovttal ttansgenome at a frequency after ttansplant which equaled the frequency of the manow cells following in vitto expositte to the MDR vimses before ttansplant.…”

Retroviral gene transfer into cord blood stem/progenitor cells using purified vector stocks

“…Numerous in vivo studies have used retroviral transduction in conjunction with bone marrow transplantation in murine models [1,[10][11][12][13], and in larger animal models including dogs [14][15][16][17] and Rhesus monkeys [18,19]. Results of human gene therapy clinical tri-als for adenosine deaminase deficiency [20][21][22] and gene marking in cancer patients [23][24][25] have also been reported.…”

Antisense therapeutics in chronic myeloid leukaemia: the promise, the progress and the problems

“…61 ODN have also been introduced into cells by retroviral strategies. [62][63][64][65] Our group has demonstrated that following the introduction …”