VEGF-A/VEGFR Inhibition Restores Hematopoietic Homeostasis in … · Lin-cKit þSca1 (LKS)...

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VEGF-A/VEGFR Inhibition RestoresHematopoietic Homeostasis in the Bone Marrowand Attenuates Tumor GrowthRebekah K. O'Donnell1,2, Beverly Falcon3, Jeff Hanson3,Whitney E. Goldstein1,2,Carole Perruzzi3, Shahin Rafii4,William C. Aird2,5, and Laura E. Benjamin3

Abstract

Antiangiogenesis–based cancer therapies, specifically thosetargeting the VEGF-A/VEGFR2 pathway, have been approved forsubsets of solid tumors. However, these therapies result in anincrease in hematologic adverse events.We surmised that both thebone marrow vasculature and VEGF receptor–positive hemato-poietic cells could be impacted by VEGF pathway–targeted ther-apies. We used a mouse model of spontaneous breast cancer todecipher the mechanism by which VEGF pathway inhibitionalters hematopoiesis. Tumor-bearing animals, while exhibitingincreased angiogenesis at the primary tumor site, showed signs ofshrinkage in the sinusoidal bone marrow vasculature accompa-nied by an increase in the hematopoietic stem cell–containing

Lin-cKitþSca1þ (LKS) progenitor population. Therapeutic inter-vention by targeting VEGF-A, VEGFR2, and VEGFR3 inhibitedtumor growth, consistent with observed alterations in the primarytumor vascular bed. These treatments also displayed systemiceffects, including reversal of the tumor-induced shrinkage ofsinusoidal vessels and altered population balance of hematopoi-etic stem cells in the bone marrow, manifested by the restorationof sinusoidal vessel morphology and hematopoietic homeo-stasis. These data indicate that tumor cells exert an aberrantsystemic effect on the bone marrow microenvironmentand VEGF-A/VEGFR targeting restores bone marrow function.Cancer Res; 76(3); 517–24. �2015 AACR.

IntroductionCancer has local (juxtacrine/paracrine) and distal

(paracrine/endocrine) effects. The study of its local effects,particularly upon the proliferative and poorly functional vas-culature, has led to its description as a wound that does notheal (1). Although cancer can have significant pathologicsystemic effects, such as anemia and cachexia, little is knownabout its influence upon distal vasculature. One particularlyimportant non-tumor vascular bed is the sinusoidal vascula-ture of the bone marrow, due to its importance in hemato-poiesis (2). Hematopoietic stem cells (HSC) in the bonemarrow exist primarily in a quiescent state, but can be com-pelled to enter the bloodstream by agents that interfere withthe interactions between HSCs and their microenvironments(3). HSCs are a subset of the hematopoietic stem/progenitorcell (HSPC) bone marrow compartment, which includesmultiple subsets with varying differentiative capabilities.HSPC-derived cells can be recruited to distant organs where

they promote metastasis, as well as into primary tumors (4).Systemic conditions such as diabetes and estradiol treatmentcan target the subset of bone marrow HSPCs capable ofreconstituting the hematopoietic system (5, 6), and recentwork shows that tumor burden increases bone marrow HSPCnumbers (7), raising the possibility that an important sourceof HSPC-derived cells recruited by tumors may be the niches ofthe bone marrow.

In general, HSPC-derived hematopoietic cells enter bodytissues via VEGFR2-expressing blood vessels and exit viaVEGFR2- and VEGFR3-expressing lymphatic vessels. The sinu-soidal vasculature is unique in that it mediates traffic in bothdirections, serving as a gatekeeper to virtually all fluid andcellular elements that enter and exit the bone marrow. Incontrast to many other vascular beds, it lacks a thick basementmembrane, is virtually unsupported by pericytes, and expressesVEGFR2, VEGFR3, and VEGFR1 (8, 9). We wondered whetherthe sinusoidal vessels, which serve as a niche for HSPCs, wouldbe affected by increased circulating levels of VEGF-A (as seen incancer) or anti-VEGF pathway inhibitors, a classification thatincludes many cancer drugs both approved and in developmentdue to the important role angiogenesis plays in the tumormicroenvironment. In addition to their well-documentedeffects on local tumor vasculature, antiangiogenic therapiescould also affect the bone marrow sinusoidal bed in its roleas the vascular niche, potentially affecting hematopoietichomeostasis and contributing to the increase in adversehematopoietic events with these therapies (10, 11). In addition,previous studies have shown that some hematopoietic cellsexpress VEGF receptors, and of special interest to the biology ofHSPC-derived cells we considered the impact of anti-VEGFR1on VEGFR1-expressing LKS HSPCs.

1Department of Pathology, Beth Israel Deaconess Medical Center,Boston, Massachusetts. 2Center for Vascular Biology Research, BethIsrael Deaconess Medical Center, Boston, Massachusetts. 3Eli Lilly &Company, Indianapolis, Indiana. 4Department of Genetic and Regen-erative Medicine, Weill Cornell Medical College, New York City, NewYork. 5Department ofMedicine, Beth Israel DeaconessMedical Center,Boston, Massachusetts.

Corresponding Author: Laura E. Benjamin, Eli Lilly & Company, 450 East 29thStreet, 11th Floor New York, NY 10016. Phone: 908-400-1544; E-mail:[email protected]

doi: 10.1158/0008-5472.CAN-14-3023

�2015 American Association for Cancer Research.

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Materials and MethodsMice

Tumor volume of female MMTV-PyT mice was calculated as0.52�(smaller dimension)2�(larger dimension). Blood was ana-lyzed on a Hemavet (Drew Scientific). All animal studies wereapproved by the Beth Israel Deaconess Medical Center Institu-tional Animal Care and Use Committee.

ImmunohistochemistryTumors were fixed in 4% paraformaldehyde and paraffin

embedded. One femur per animal was decalcified in 10% EDTAafter fixation. Four-micron sections were baked for 1 hour at 60�F.Antigens were retrieved on deparaffinized, rehydrated slides usingproteinase K or Universal Decloaker (Biocare Medical). Slideswere washed with 0.05% Tween/TBS and blocked with ProteinBlock (Dako). Tumor and femur sections were incubated withbiotinylated antibodies to VEGFR3 (AFL4; eBioscience), CD31(Becton-Dickinson), or LYVE-1 (Abcam). Sections were subse-quently washed, blocked with hydrogen peroxide, incubatedwith streptavidin-HRP, and exposed with DAB (Dako). Afteradditional washes, the sections were counterstained withhematoxylin (Dako), dehydrated, and cleared with xylene priorto mounting with cytoseal XYL (ThermoScientific). Entire tissuesections were imaged at 20� with the Aperio ScanScopeXT.Regions of interest were defined to avoid quantification withinregions with folded tissue or high background staining andto restrict analysis to bone marrow regions of the femur.Endothelial area was defined as stained cells only. Averageendothelial area per vessel was quantified using custom-builtalgorithms in Definiens image analysis software. Results areaggregated per animal and expressed as mean per group� SEM.Statistical differences between experimental and control groupswere measured by the two-tailed Student t test.

Antibody treatmentFemaleMMTV-PyTmice with total tumor volumes of 0.4 to 0.6

cm3 were injected i.p. with 800 mg of control rat IgG (JacksonImmunoResearch), anti-VEGFR1 (MF1), anti-VEGFR2 (DC101),anti-VEGFR3 (mF4-31C1) or 400 mg of anti-VEGF-A (G6-31;ImClone Systems) three times a week for 1 week or until totaltumor volume reached approximately 1 cm3.

Flow cytometryFemurs were flushed with cold PBS, treated with lysis buffer

(Quality Biological), and counted by a hemacytometer. Cellswereblocked with anti-Fc (eBioscience) then incubated with directlyconjugated antibodies against lineage markers (B220, CD4, CD8,GR1, TER119, and CD11b), Sca-1, and cKit (eBioscience; Biole-gend). Antibodies against CD34, Flt3, CD48, and CD150 furthercharacterized the population. For cell cycle, LKS-stained cells werepermeabilized with 0.03% saponin and stained with 7-AAD andpyronin Y. Cells were acquired on a FACSCanto II and analyzedusing FACSDiva (BD Biosciences).

Competitive bone marrow transplantsFemale FVB recipients were given 9 Gy of gamma irradiation.

After 24 hours, the animals were reconstituted by tail-vein injec-tion with 5�105 cells from aGFPþ donor and varying numbers ofcells from a GFP� tumor-bearing MMTV-PyT or tumor-negativelittermate control. After 12 weeks of reconstitution, leukocytes

were isolated from the blood and stained with CD45(eBioscience). A positive competitive result was defined as apercentage of CD45þ GFPþ cells below the 95% confidence levelof control animals that received GFPþ bone marrow exclusively.Data were analyzed using a Poisson statistic, and the numberof competitive repopulating units was estimated using LCalc(StemCell Technologies).

Quantitative real-time PCRRNA was isolated from bone marrow and lungs using TRIzol

(Invitrogen) and cDNA synthesis performedwith theHighCapac-ity cDNA Reverse Transcription Kit (Applied Biosystems). RNAwas quantified using Sybr Green (Applied Biosystems) on a 7500Fast Real-Time PCR System (Applied Biosystems) using primersagainst PyT (F50-AGCCCGATGACAGCATATCC-30, R50-GGTCT-TGGTCGCTTTCTGGA-30; Integrated DNA Technologies). Valueswere normalized against 18S.

ELISABlood was collected in EDTA-containing tubes (BD Bio-

sciences) and spun for 20 minutes at 2,000 rpm at 4�C. Con-centrations of SDF-1/CXCL12, SCF, G-CSF, VEGF-A, VEGF-C, andVEGF-D (R&D Systems) in plasma and/or serum were analyzedaccording to the manufacturer's instructions. GM-CSF, IL6, andTNFa were assessed with the Multi-Analyte ELISArray Kit(SABiosciences).

Results and DiscussionTo examine the impact of cancer and antiangiogenic therapy

on the bone marrow sinusoidal vessels and HSPC population,we used the MMTV-PyT model of triple-negative breast cancer.The spontaneous tumors arising in this model are metastaticand display comparable morphology to human disease (12).Because some but not all tumors contain VEGFR3þ bloodvessels (13), we first profiled the expression of VEGFR2 andVEGFR3 in our model. VEGFR3 was widely expressed through-out primary MMTV-PyT tumors, primarily on blood vesselsthat were identified by lack of LYVE-1 positivity and presence ofluminal erythrocytes (Fig. 1A). We examined tumor growth inresponse to selective neutralizing antibodies to all three VEGFreceptors. As expected, anti-VEGFR2 and anti-VEGF-A antibo-dies resulted in a significant (and comparable) delay in tumorgrowth, while administration of anti-VEGFR1 antibodies hadno such effect. This latter result is consistent with previousreports showing failure of VEGFR1 inhibition to block tumorangiogenesis and growth (14). Surprisingly, anti-VEGFR3 alsoresulted in delayed tumor growth (Fig. 1B) without inducingnecrosis like anti-VEGFR2 treatment (Fig. 1C). Antibody treat-ment did not reduce the density of CD31þ vessels overall,although nonspecific staining of necrotic tissue may havecamouflaged anti-VEGFR2 effects. However, the inhibitoryeffects of anti-VEGFR2 and anti-VEGFR3 antibodies on tumorprogression were associated with significant shrinkage of endo-thelial area per vessel (Fig. 1C and D). These findings suggestthat anti-VEGFR3 antibodies inhibit tumor growth in thismodel, at least in part, by attenuating vascular endothelial cellsignaling in tumor blood vessels (15, 16).

We next asked whether the tumor per se and/or its treatmentwith anti-VEGFR2 and anti-VEGFR3 antibodies influences themorphology of the bone marrow sinusoids. Compared with

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Cancer Res; 76(3) February 1, 2016 Cancer Research518




Figure 1.Morphologic and growth effects of anti-VEGFR2/3 treatment on MMTV-PyT primary tumor. A, VEGFR3þ vessels in the primary tumor were identified as membersof the blood vasculature through similarity to CD31þ vessels, lack of LYVE-1 positivity, and presence of circulating blood cells (�20; scale bar, 100 mm). B,animals bearing tumors with total volume of 0.4 to 0.6 cm3 received antibody injections 3 times per week; animals were sacrificed when total tumor volumereached approximately 1.0 cm3. Inhibition of VEGF-A, VEGFR2, or VEGFR3 significantly delayed primary tumor growth, while VEGFR1 inhibition had no effect(n ¼ 4–12). C and D, treatment with anti-VEGFR2/3 did not change CD31þ vessel density, but resulted in decreases in per-vessel endothelial area (�20; n ¼ 3–5).

Antiangiogenesis Restores Bone Marrow Vascular Niche

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healthy controls, the sinusoid vessels of tumor-bearing animalsappeared subtly smaller and more uniform (Fig. 2A) with asignificant decrease in per-vessel endothelial area (Fig. 2B). Treat-

ment with anti-VEGFR3 decreased the number of vessels, indi-cating an increased dependence on VEGFR3 signaling in the bonemarrow compared with the primary tumor (Fig. 2B). However,

Figure 2.Morphologic effect of tumor burden and anti-VEGFR2/3 treatment on bone marrow vasculature. A, bone marrow from control and MMTV tumor–bearinganimals treated with IgG, anti-VEGFR2, or anti-VEGFR3 was stained against VEGFR3 to identify the sinusoid vasculature (�20). B, quantification showedsignificant decreases in VEGFR3þ vessel density with VEGFR3 inhibition and partial rescue of per-vessel endothelial area with anti-VEGFR2/3 treatment (n¼ 3–5).

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treatment with anti-VEGFR2 or anti-VEGFR3 antibodies was ableto partially rescue per-vessel endothelial area of the remainingvessels, suggesting that aberrant VEGFR family signaling can alterbone marrow vasculature without spurring angiogenesis (Fig. 2Aand B).

Recent work has demonstrated that the majority of HSCs arelocated in close proximity to the sinusoidal vasculature (2),which regulates HSC engraftment and hematopoiesis (9, 17).HSC subtypes can be identified by flow cytometry as part of theLineage�cKitþSca1þ(LKS) HSPC compartment. Bone marrowfrom tumor-bearing MMTV-PyT mice displayed an increase inthe percentage (data not shown) and absolute number (Fig.3A) of LKS HSPCs, consistent with recent work in the MMTV-neuOTI/OTII model (7). In contrast to the study by Sio andcolleagues (7), there was no evidence of increased extrame-dullary hematopoiesis in this model, with constant numbersof nucleated cells and LKS HSPCs in the spleen at all tumorvolumes (data not shown). The increase in bone marrow LKSHSPCs was present even at small total tumor volumes andwas independent of total bone marrow cellularity (Fig. 3A).This expanded LKS population in the bone marrow of tumor-bearing mice was hyperproliferative (Fig. 3B) and more dif-ferentiated, as shown by a relative decrease of less differenti-ated, CD34�Flt3�LKS long-term HSCs (LT-HSC) and anincrease of more differentiated, CD34�Flt3þLKS short-term

HSCs (ST-HSCs; Fig. 3C). This shift toward a more dif-ferentiated LKS population was also observed using the mar-kers CD48�CD150þ (data not shown). Competitive bonemarrow transplants revealed a decrease in functional LT-HSCs,with tumor-bearing animals displaying a significant decreaseof over twofold in competitive repopulating units (CRU;Fig. 3D).

To determine if the hyperproliferative, pro-differentiatedphenotype of LKS HSPCs could be mediated locally by metas-tasized tumor cells in the bone marrow, quantitative real-timePCR was performed on lung and bone marrow from tumor-bearing mice. This assay identified metastasized tumor cellsexpressing PyT mRNA in lung tissue even from mice bearingtumors of only 0.2 to 0.4 cm3 (Fig. 4A), a pattern similar to theincreased LKS population in the bone marrow. However, nosignificant burden of metastasized tumor cells was found in thebone marrow at any tumor volume (Fig. 4A). Therefore, weentertained the likely possibility that the effect of tumor onsinusoidal vasculature and HSPCs in the bone marrow wasmediated by soluble factors. Leading candidates that we con-sidered included VEGF-A, which promotes cycling and mobi-lization of hematopoietic progenitor cells from the bone mar-row (18, 19), and G-CSF, which is used therapeutically tomobilize HSCs from the bone marrow (3). Indeed, plasmalevels of both VEGF-A and G-CSF were significantly increased in

Figure 3.The LKS progenitor population is dysregulated in the bone marrow of tumor-bearing MMTV-PyT mice. A, bone marrow isolated from tumor-bearing MMTV-PyTanimals at various total tumor volumes was analyzed by flow cytometry. Tumor-bearing animals at total tumor volumes as small as 0.2 to 0.4 cm3 showed anincreased number (� , P < 0.05; ��, P ¼ 0.005) of LKS cells in the bone marrow. Numbers of cells were calculated by multiplying the percentage of cells bythe total number of cells in the bone marrow, as counted by hemacytometer (n ¼ 4–9). B, staining with 7-AAD and pyronin Y revealed a relative decreasein G0–G1 phases and a relative increase in S–G2–M phases within the LKS population (n ¼ 3; �P < 0.05). C, LKS cells were further characterized by staining againstCD34 and Flt3. Tumor-bearing mice showed relative decreases in the less-differentiated CD34�Flt3� LKS population and a relative increase in the more-differentiated CD34þFlt3� LKS population (n ¼ 5–6; � , P < 0.04). D, lethally irradiated animals underwent competitive reconstitution with bone marrow fromcontrol or tumor-bearing mice versus GFPþ bone marrow. After 12 weeks of reconstitution, animals were sacrificed, and the leukocyte fraction was isolatedfrom blood and stained for CD45. Increased competition from the unlabeled animals correlated with a decreased fraction of GFPþ cells. A positive competitiveresult is defined as a CD45þGFPþ fraction below the lower bound of the 95% confidence level of animals that were reconstituted with only GFPþ bone marrow.Data were analyzed using the Poisson statistic (L-Calc; � , P < 0.03).






tumor-bearing mice (Fig. 4B). The increased levels of G-CSFgreatly affected the downstream differentiation of HSPCs inthis model, with tumor-bearing mice displaying an 8-foldincrease in circulating neutrophil concentration (data notshown). CXCL12/SDF-1 was another likely candidate given itsrole in HSC retention and mobilization (3), but the levels ofCXCL12 in plasma (Fig. 4B) and serum (data not shown) wereunchanged at even maximum total tumor volumes. Circulatinglevels of SCF, GM-CSF, IL6, and TNFa also remainedunchanged (data not shown).

We then tested the ability of anti-VEGF pathway inhibitors toreverse the hyperproliferative, pro-differentiated phenotype ofHSPCs in the bone marrow. Although VEGFR1 inhibition didnot affect primary tumor growth, VEGFR1 was includedbecause it is expressed on LKS HSPCs as well as bone marrowendothelium (9, 20). We treated MMTV-PyT animals bearingtumors of 0.4 to 0.6 cm3 total tumor volume with antibodiesagainst VEGF-A, VEGFR1, VEGFR2, VEGFR3, or control IgGthree times per week until the total tumor volume reached1.0 cm3. Anti-VEGF-A treatment reversed the tumor-inducedexpansion of the LKS HSPC compartment, evidenced by nor-malization of the cell cycle (Fig. 4C). Reduction in LT-HSCnumbers confirmed that anti-VEGF-A treatment halted theinappropriate proliferation of that subset. Anti-VEGF-A–treatedbone marrow, like tumor stimulated bone marrow, contained ahigh number of more differentiated, phenotypically marked ST-HSCs but a normal number of multipotent progenitors (MPP),a downstream subset that retains multilineage differentiationpotential (Fig. 4C), suggesting that differentiation from ST-HSCto MPP in this context is slow and not controlled by VEGF-A. Incontrast, VEGFR1 inhibition did not restore normal HPSCdifferentiation, even showing additional increases in ST-HSCsand MPPs (Fig. 4C).

The ability of VEGF-A and the inability of VEGFR1 inhibitionto rescue the hyperproliferative, pro-differentiated LKS pheno-type suggested that the sinusoidal vasculature may mediatecommunication between distant tumor and bone marrowHSPCs. We have shown that transgenic hyperactivation of Aktin endothelial cells results in an increase in bone marrowCRUs (21), and vasculature also mediates the establishmentand regression of extramedullary hematopoietic sites in aVEGF-overexpressing tumor model (22). In support of thishypothesis, treatment with anti-VEGFR2, which is expressedon endothelium but not on LKS cells, restored LKS cell-cyclebalance. However, like anti-VEGFR1 treatment, VEGFR2

Figure 4.Tumor effect on hematopoietic progenitor cells in the bone marrow ismediated by VEGF-A and can be rescued with anti-VEGF-A or anti-VEGFR3treatment. A, lungs and bone marrow of mice bearing tumors of various sizesand their littermate controls were assayed for PyT mRNA by quantitativereal-time PCR. B, plasma levels of VEGF-A, G-CSF, and CXCL12 were assayedby ELISA (n¼ 3–7). Animals with a total tumor burden of 0.4 to 0.6 cm3 wereinjected with antibodies against VEGF-A (G6), VEGFR1 (MF1), VEGFR2(DC101), VEGFR3 (mF4-31C1). Anti-VEGF-A, VEGFR2, or VEGFR3 treatmentsignificantly reduced the percentage of LKS cells in the S–M–G2 stages of thecell cycle. Evaluation of LKS or control IgG subsets by flow cytometryrevealed twopatterns of HSPC population response. C, anti-VEGF-A and anti-VEGFR3 treatments reduced CD34�Flt3� LKS LT-HSCs and LSKs overall,while anti-VEGFR1 and anti-VEGFR2 treatment increased CD34þFlt3� LKSST-HSCs and CD34þFlt3þMPPpopulations (n¼ 6–16). D, model of tumor andVEGF family inhibitor actions in the bone marrow in vivo.

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blockade resulted in an increase in both ST-HSCs and MPPs(Fig. 4C). Treatment with anti-VEGFR3, which is also expressedon sinusoid endothelium but not on LKS cells, restored cell-cycle balance without further buildup of ST-HSCs or MPPs, andlike VEGF-A blockade reduced LT-HSC and overall LKS num-bers to control levels (Fig. 4C). In summary, we observed mostcomplete reversal of tumor-induced progenitor buildup withanti-VEGF-A and anti-VEGFR3, consistent with decreasedtumor growth, suggesting that these treatments counteracttumor-induced proliferation and differentiation of LT-HSCs(Fig. 4D). Anti-VEGFR2 treatment resulted in decreased HSPCproliferation and restoration of sinusoid structure, but alsoincreased ST-HSC and MPP populations. Similar increases inST-HSC and MPP populations with anti-VEGFR1 treatmentsuggest that expression of VEGFR1 on HSPCs and/or VEGFR1and VEGFR2 on bone marrow vasculature may be required forterminal differentiation or exit of differentiated cells from thebone marrow, causing a buildup of these more differentiatedcells in the bone marrow (Fig. 4D).

We confirm recent work showing that primary tumors areable to communicate long-distance with HSPCs in the bonemarrow (7). Although the effect of other tumor-derived factorscannot be ruled out, this communication appears to be medi-ated by the effects of circulating VEGF-A on the sinusoidalvasculature of the bone marrow, as targeting of vascular-specific VEGF receptors can reverse the aberrant entrance ofHSPCs into proliferative and differentiative phases of the cellcycle. In addition to the anti-VEGF-A (e.g., bevacizumab) andVEGFR non-selective TKIs (e.g., sunitinib, sorafenib, axitinib)approved for cancer therapy, there are multiple anticancertherapies under investigation for VEGFR1, VEGFR2, andVEGFR3 signaling. A selective fully human antibody toVEGFR2 (ramucirumab) has recently shown benefits in overall

survival in gastric (10) and lung cancers, and selective anti-bodies to VEGFR3 (IMC-3C5) and VEGFR1 (icrucumab) are inearly-phase trials for multiple solid tumors (www.clinical-trials.gov). A better understanding of the role of the sinusoidvasculature may lead to increased antitumor treatment effica-cy, specific targeting of hematopoietic malignancies that resideand proliferate in the bone marrow, improvements in HSCrecruitment techniques for stem cell transplantation, and,given the importance of bone marrow–derived cells in pre-paring the metastatic niche (4), therapies that target theformation of metastases.

Disclosure of Potential Conflicts of InterestNo potential conflicts of interest were disclosed.

Authors' ContributionsConception and design: R.K. O'Donnell, S. Rafii, W.C. Aird, L.E. BenjaminDevelopment of methodology: R.K. O'Donnell, B. Falcon, C. PerruzziAcquisition of data (provided animals, acquired and managed patients,provided facilities, etc.): R.K. O'Donnell, B. Falcon, C. PerruzziAnalysis and interpretation of data (e.g., statistical analysis, biostatistics,computational analysis): R.K. O'Donnell, B. Falcon, J. Hanson,W.E. GoldsteinWriting, review, and/or revision of themanuscript: R.K. O'Donnell, B. Falcon,J. Hanson, C. Perruzzi, W.C. Aird, L.E. BenjaminAdministrative, technical, or material support (i.e., reporting or organizingdata, constructing databases): C. PerruzziStudy supervision: W.C. Aird, L.E. Benjamin

Grant SupportThis work was supported by the NIH (grant CA131152) and Eli Lilly and

Company.

ReceivedNovember 10, 2014; revisedOctober 20, 2015; acceptedOctober 22,2015; published OnlineFirst December 30, 2015.

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2016;76:517-524. Published OnlineFirst December 30, 2015.Cancer Res Rebekah K. O'Donnell, Beverly Falcon, Jeff Hanson, et al. the Bone Marrow and Attenuates Tumor GrowthVEGF-A/VEGFR Inhibition Restores Hematopoietic Homeostasis in

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VEGF-A/VEGFR Inhibition Restores Hematopoietic Homeostasis in … · Lin-cKit þSca1 (LKS)...

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Transcript of VEGF-A/VEGFR Inhibition Restores Hematopoietic Homeostasis in … · Lin-cKit þSca1 (LKS)...