doi:10.1182/blood-2013-12-5382492014 123: 310-311   

 Sakura Hosoba and Edmund K. Waller New molecule for mobilizing marrow stem cells

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recurrent miscarriage. N Engl J Med. 2010;362(17):1586-1596.

8. Clark P, Walker ID, Langhorne P, et al; ScottishPregnancy Intervention Study (SPIN) collaborators. SPIN(Scottish Pregnancy Intervention) study: a multicenter,randomized controlled trial of low-molecular-weightheparin and low-dose aspirin in women with recurrentmiscarriage. Blood. 2010;115(21):4162-4167.

9. Rey E, Garneau P, David M, et al. Dalteparin for theprevention of recurrence of placental-mediated complications ofpregnancy inwomenwithout thrombophilia: a pilot randomizedcontrolled trial. J Thromb Haemost. 2009;7(1):58-64.

10. Bates SM, Greer IA, Middeldorp S, et al. VTE,thrombophilia, antithrombotic therapy, and pregnancy:Antithrombotic Therapy and Prevention of Thrombosis,9th ed: American College of Chest Physicians Evidence-Based Clinical Practice Guidelines. Chest. 2012;141(suppl2):e691S-e736S.

11. An J, Waitara MS, Bordas M, et al. Heparin rescuesfactor V Leiden-associated placental failure independent ofanticoagulation in a murine high-risk pregnancy model.Blood. 2013;121(11):2127-2134.

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l l l TRANSPLANTATION

Comment on Zhang et al, page 428

New molecule for mobilizingmarrow stem cells-----------------------------------------------------------------------------------------------------

Sakura Hosoba1 and Edmund K. Waller1 1EMORY UNIVERSITY

In this issue of Blood, Zhang et al describe an exciting new small-molecule antagonistof CXCL12-CXCR4 binding with a potent ability to mobilize hematopoieticstem cells (HSCs).1 Further clinical development of this new drug, Me6TREN, mayhave broad applications in stem-cell mobilization for cancer and in regenerativemedicine.

The field of HSC transplantation has beenstrongly impacted by the clinical approval

of plerixafor, a CXCR4 antagonist that has beenshown to be effective in mobilizing stem cellsfrom the bone marrow into circulationin the blood.2 The clinical use of plerixaforin combination with 5 days of granulocytecolony-stimulating factor (G-CSF)administration has allowed collection ofCD341 stem cells from patients withlymphoma and myeloma undergoingautologous stem cell transplantation in fewerdays and in higher numbers than had beenpossible whenmobilizing patients with G-CSFalone.3,4 A limitation in the clinical use ofplerixafor is the need to combine it withG-CSF, thus subjecting potential donors totoxicities of 2 drugs. In addition, while theaddition of plerixafor to the armamentarium ofdrugs for stem-cell mobilization has decreasedthe incidence of mobilization failure from 10%to 40% to,7%,5 there are still patients inwhom attempts at stem-cell collection fail,and these patients are unable to proceed toautologous stem-cell transplantation. Thus,scientists studying hematopoiesis and stem-celltransplant physicians are interested in newagents that could increase the efficiency of

stem-cell mobilization and collection. Toaddress this need, Zhang et al identified a smallmolecule, Me6TREN, that potently mobilizedHSCs frombonemarrow to peripheral blood inmice. Using flow cytometry and in vitro colonyassays, they found that a single Me6TRENinjection mobilized phenotypically definedlineage2 sca1 c-kit1 (LSK) stem cells andhematopoietic colony-forming cells, and thatthe blood content of circulating progenitor cellsremained elevated for 4 days after Me6TRENtreatment.

The natural question, when faced witha new drug, is whether it is more effectivethan existing drugs. Based upon theirchemical structures, it would appear that thesedrugs might have different mechanisms ofaction, because plerixafor is a 1,19-[1,4-phenylenebis(methylene)]bis[1,4,8,11-tetraazacyclotetradecane] molecule withbilateral symmetry, whereas Me6TREN isa tris[2-(dimethylamino)ethyl]amine moleculewith trivalent symmetry (see figure). Tocompare the activity of Me6TREN withG-CSF and plerixafor as single agents for HSCmobilization, the authors measured the contentof high-proliferative–potential colony-formingunits (HPP-CFU) in the blood of mice 1 hour

following plerixafor administration, 12 hoursafter Me6TREN administration, and after5 days of G-CSF administration andperformed competitive repopulation assays bytransplanting mobilized blood from mice indifferent treatment groups into irradiatedrecipients in combination with a fixed numberof congenic bone marrow cells. In theseexperiments, mice treated withMe6TREN hadsignificantly moreHPP-CFU in their blood andsuperior competitive repopulating activity whentransplanted into irradiatedmice compared withblood mobilized with plerixafor or G-CSF.Me6TREN-mobilized progenitors contributedto long-termdurablemultilineagehematopoieticreconstitution in recipient mice after secondarytransplantation of marrow from mice engraftedwith mobilized cell, thus fulfilling the criteriafor mobilization of a self-renewing HSC.

Some questions regarding this interestingnew compound remain. Although Me6TRENantagonized SDF-1–induced migrationof murine and human hematopoieticprogenitors, antagonistic binding of Me6TRENto CXCR4 has not been established. Theschedule of plerixafor administration usedby Zhang et al was based upon work fromBroxmeyer et al, who showed that single-agent plerixafor resulted in a 5- to 10-foldmobilization of CFU-GEM into blood 1 hourafter drug administration.2 In contrast to theschedule of plerixafor used in these murineexperiments, our clinical experience withplerixafor indicates that optimal mobilizationof HSCs in humans usually occurs.8 hoursand up to 17 hours following a subcutaneousplerixafor injection.6 Although plerixaforappears to be a pure CXCR4 antagonist and thusacts directly on CXCR4/CXCL12 binding, theauthors show that Me6TREN activates thephospho-Akt, mitogen-activated protein kinase,and phospho-extracellular signal-regulatedkinase pathways and inducesMMP9 expression.Using radiation chimeras engrafted withMMP9-knockout bone marrow, they showthat much of the mobilization induced byMe6TREN is dependent upon expressionof MMP9 by hematopoietic cells. Localproduction of metalloproteases is implicated indisruption of adhesion molecules that tetherHSCs to stromal cells, including VLA4 (seefigure). It would be of interest to test whetherMe6TREN acts indirectly on the CXCR4/CXCL12 axis by modulating stromal-cellexpression of messenger RNA for CXCL12, theligand for CXCR4. Thus, differences in

310 BLOOD, 16 JANUARY 2014 x VOLUME 123, NUMBER 3

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mechanism of action of plerixafor andMe6TREN could be responsible for theobserved differences in the pharmacodynamicsof stem-cell mobilization following drugadministration. AlthoughMe6TREN appears tobe quite potent in mobilizing HSCs in thesemurine models, its absolute superiority overplerixafor as a single agent for stem-cellmobilization has not been confirmed, and theoptimal schedule of this new agent in humansremains to be defined.

Beyond the mobilization of hematopoieticprogenitors for autologous transplantationas studied in this report, promising datapublished by Devine et al suggest thatsingle-agent plerixafor can mobilize anallogeneic blood stem cell graft that mayhave favorable properties for donor-derivedhematopoietic and immune reconstitutioncompared with G-CSF–mobilized products.7

It is unknown whether the use of Me6TRENin a setting of mobilization allogeneicdonors would result in a graft with similarfavorable immunological properties, but thediscovery and preclinical development ofthis drug make that application an exciting new

avenue for research in the field oftransplantation. Finally, the availability ofa new small-molecule drug with potentsingle-agent ability to mobilize HSCs opensthe door to new clinical applications ofbone marrow progenitors in nontransplantsettings. We have previously shown thatintermittent dosing of granulocyte-macrophage colony-stimulating factor(GM-CSF) mobilizes CD341 cells into bloodand improves endothelial dysfunction andexercise capacity in patients with severeperipheral arterial disease.8 Seiler et aldemonstrated with a randomized, double-blind, placebo-controlled test that intracoronaryand subcutaneous injection of GM-CSF couldimprove collateral flow patients with extensivecoronary artery disease not eligible for coronaryartery bypass surgery.9 In addition, a highercontent of CD341 cells in the blood of patientswho are at high risk for myocardial infarction anddeath is associated with improved event-freesurvival.10 Thus, new drugs that mobilizeprogenitor cells from the blood to the bonemarrow as single agents could be used ona chronic or intermittent basis in patients

with vascular diseases. With progress inunderstanding the biology of stem-cellmobilization and with newer drugs that appeareffective as single agents in mobilizing progenitorcells, research in stem cells and hematopoiesisbegins to have wide clinical applications inmedicine beyond stem-cell transplantation.Conflict-of-interest disclosure:The authors declare

no competing financial interests. n

REFERENCES1. Zhang J, Ren X, Shi W, et al. Small moleculeMe6TREN mobilizes hematopoietic stem/progenitor cellsby activating MMP-9 expression and disrupting SDF-1/CXCR4 axis. Blood. 2014;123(3):428-441.

2. Broxmeyer HE, Orschell CM, Clapp DW, et al. Rapidmobilization of murine and human hematopoietic stem andprogenitor cells with AMD3100, a CXCR4 antagonist.J Exp Med. 2005;201(8):1307-1318.

3. DiPersio JF, Micallef IN, Stiff PJ, et al; 3101Investigators. Phase III prospective randomized double-blind placebo-controlled trial of plerixafor plus granulocytecolony-stimulating factor compared with placebo plusgranulocyte colony-stimulating factor for autologous stem-cell mobilization and transplantation for patients withnon-Hodgkin’s lymphoma. J Clin Oncol. 2009;27(28):4767-4773.

4. DiPersio JF, Stadtmauer EA, Nademanee A, et al; 3102Investigators. Plerixafor and G-CSF versus placebo andG-CSF to mobilize hematopoietic stem cells for autologousstem cell transplantation in patients with multiplemyeloma. Blood. 2009;113(23):5720-5726.

5. Li J, Hamilton E, Vaughn L, et al. Effectivenessand cost analysis of “just-in-time” salvage plerixaforadministration in autologous transplant patients with poorstem cell mobilization kinetics. Transfusion. 2011;51(10):2175-2182.

6. Harvey RD, Kaufman JL, Johnson HR, et al. Temporalchanges in plerixafor administration and hematopoieticstem cell mobilization efficacy: results of a prospectiveclinical trial in multiple myeloma. Biol Blood MarrowTransplant. 2013;19(9):1393-1395.

7. Devine SM, Vij R, Rettig M, et al. Rapid mobilizationof functional donor hematopoietic cells without G-CSFusing AMD3100, an antagonist of the CXCR4/SDF-1interaction. Blood. 2008;112(4):990-998.

8. Poole J, Mavromatis K, Binongo JN, et al. Effect ofprogenitor cell mobilization with granulocyte-macrophagecolony-stimulating factor in patients with peripheral arterydisease: a randomized clinical trial [published online aheadof print November 8, 2013]. JAMA. doi:10.1001/jama.2013.282540.

9. Seiler C, Pohl T, Wustmann K, et al. Promotionof collateral growth by granulocyte-macrophage colony-stimulating factor in patients with coronary artery disease:a randomized, double-blind, placebo-controlled study.Circulation. 2001;104(17):2012-2017.

10. Patel RS, Eapen DJ, Li Q, et al. Hematopoieticprogenitor cells expressing CD341 and CD1331 surfacemarkers are predictors of adverse cardiovascular outcomesin a population with coronary artery disease circulation[abstract]. Circulation. 2012;126:A17150.

© 2014 by The American Society of Hematology

Mechanism of release of HSCs from the bone marrow microenvironment using G-CSF, plerixafor, or Me6TREN as

mobilization agents. G-CSF induces synthesis of proteases elastase, cathepsin, and MMP9 by neutrophils and osteoclasts

after binding to G-CSF receptors (G-CSFR). Proteases degrade adhesion molecules that tether HSCs to stromal

cells in the bone-marrow microenvironment. As a secondary effect mediated through osteoclasts, G-CSF binding

leads to downregulation of CXCL12 on bone-marrow stromal cells. Plerixafor antagonizes binding of CXCR4 to CXCL12,

whereas Me6TREN has reported activity in antagonizing CXCR4/CXCL12 binding as well as inducing synthesis of MMP9.

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