clonal-analysis-embryonic-hematopoietic-stem-cell-precursors-using

Clonal Analysis of Embryonic Hematopoietic Stem Cell Precursors Using Single Cell Index Sorting Combined with Endothelial Cell Niche Co-culture

The ability to study hematopoietic stem cell (HSC) genesis during embryonic development has been limited by the rarity of HSC precursors in the early ...

Fred Hutchinson Cancer Research Center

Gemtuzumab ozogamicin (GO) is a novel immunoconjugate therapy for acute myeloid leukemia (AML). P-glycoprotein (Pgp) confers resistance to GO and is associated with a worse clinical response. To address whether multidrug resistance protein (MRP) affects GO susceptibility, we characterized Pgp, MRP1, and MRP2 expression in CD33+ cell lines and CD33+ AML samples and analyzed the effect of the Pgp inhibitor cyclosporine (CSA) and the MRP inhibitor MK-571 on GO-induced cytotoxicity. MRP1, but not MRP2, expression correlated with MRP activity. MK-571 enhanced GO-induced cytotoxicity in Pgp-negative/MRP-positive NB4 and HL-60 cells. CSA, but not MK-571 alone, restored GO susceptibility in Pgp-positive/MRP-positive TF1 cells; however, MK-571 enhanced cytotoxicity in the presence of CSA. All patient samples exhibited MRP activity, and 17 of 23 exhibited Pgp activity. CSA increased GO-induced cytotoxicity in 12 Pgp-positive samples, whereas MK-571 alone was effective in only one sample with minimal Pgp activity. In 3 Pgp-positive/MRP-positive samples, MK-571 enhanced GO-induced cytotoxicity in the presence of CSA. Thus, MRP1 may attenuate susceptibility to GO. This effect was comparatively less than that for Pgp and required the inhibition of Pgp for detection in cells that coexpressed both transporters. Because MK-571 and CSA failed to affect cytotoxicity in a portion of Pgp-positive/MRP-positive AML samples, additional resistance mechanisms are likely important.

Multidrug resistance protein attenuates gemtuzumab ozogamicin-induced cytotoxicity in acute myeloid leukemia cells.

Gemtuzumab ozogamicin (GO) contains an anti-CD33 antibody to facilitate uptake of a toxic calicheamicin-gamma(1) derivative. While recent in vitro data demonstrated a quantitative relationship between CD33 expression and GO cytotoxicity, previous correlative studies failed to identify a significant association between CD33 expression and clinical outcome. Studying patients undergoing GO monotherapy for relapsed acute myeloid leukemia (AML), we now find that AML blasts of responders have a significantly higher mean CD33 level and lower P-glycoprotein (Pgp) activity compared with nonresponders. CD33 expression and Pgp activity are inversely correlated. While both variables are associated with outcome, Pgp remains significantly associated with outcome even after adjusting for CD33, whereas CD33 does not show such an association after adjusting for Pgp. The inverse relationship between CD33 and Pgp suggests a maturation-stage-dependent expression of both proteins, and offers the rationale for using cell differentiation-promoting agents to enhance GO-induced cytotoxicity.

CD33 expression and P-glycoprotein-mediated drug efflux inversely correlate and predict clinical outcome in patients with acute myeloid leukemia treated with gemtuzumab ozogamicin monotherapy.

SIRT1 is an NAD(+)-dependent histone deacetylase implicated in the establishment of the primitive hematopoietic system during mouse embryonic development. However, investigation of the role of SIRT1 in adult hematopoiesis has been complicated by the high perinatal mortality of SIRT1-deficient mice (SIRT1(-/-)). We performed a comprehensive in vivo study of the hematopoietic stem cell (HSC) compartment in adult SIRT1(-/-) mice and show that, apart from anemia and leukocytosis in older mice, the production of mature blood cells, lineage distribution within hematopoietic organs, and frequencies of the most primitive HSC populations are comparable to those of wild-type littermate controls. Furthermore, we show that SIRT1-deficient BM cells confer stable long-term reconstitution in competitive repopulation and serial transplantation experiments. The results of the present study rule out an essential physiologic role for cell-autonomous SIRT1 signaling in the maintenance of the adult HSC compartment in mice.

Article	Year
Multidrug resistance protein attenuates gemtuzumab ozogamicin-induced cytotoxicity in acute myeloid leukemia cells. Blood\| PubMed ID: 12689934	2003
CD33 expression and P-glycoprotein-mediated drug efflux inversely correlate and predict clinical outcome in patients with acute myeloid leukemia treated with gemtuzumab ozogamicin monotherap... Blood\| PubMed ID: 17227830	2007
SIRT1 is dispensable for function of hematopoietic stem cells in adult mice. Blood\| PubMed ID: 22219225	2012
Wild-type microglia do not reverse pathology in mouse models of Rett syndrome. Nature\| PubMed ID: 25993969	2015

David Flowers

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