AUGUST 1, 2000
` % a
`
`nas
`1G translocation
`and B-cell
`
`VOLUME 96
`
`NUMBER 3
`
`G23258
`Wi
`
`87
`iy
`=y
`
`ees
`
`12262
`
`(
`
`malignancy
`
`ce aa
`
`erythropoietin
`and iron
`
`|
`+
`
`CD4i-modified
` T cells in AIDS
`
`GM-CSF receptor-—
`associated protein
`
`reas Vette}
`and chromosomal
`
`positioning
`
`Miltenyi Ex. 1004 Page1
`
`Miltenyi Ex. 1004 Page 1
`
`

`

`blood
`
`JOURNAL OF
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`THE AMERICAN
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`SOCIETY OF
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`Miltenyi Ex. 1004 Page 2°".
`
`Miltenyi Ex. 1004 Page 2
`
`

`

`blood
`
`JOURNAL OF
`
`THE AMERICAN
`
`SOCIETY OF
`
`HEMATOLOGY
`
`CHEMOKINES
`
`CLINICAL OBSERVATIONS,
`INTERVENTIONS, AND
`THERAPEUTIC TRIALS
`
`1 AUGUST2000 - VOLUME 96, NUMBER 3
`
`CONTENTS
`
`COVER FIGURE
`
`Thesites of mutation in hemophilia A are mapped on the structures of the C1 and
`C2 domainsof factor VIII. From thearticle by Liu et al, beginning on page 979.
`
`PLENARY PAPERS
`
`785
`
`794
`
`800
`
`Prolonged survival andtissue trafficking following adoptive transfer of CD4¢
`gene-modified autologous CD4* and CD8* T cells in human immunodeficiency
`virus-infected subjects
`R. T. Mitsuyasu, P. A. Anton, S. G. Deeks, D. T. Scadden, E. Connick, M. T. Downs, A. Bakker,
`M.R. Roberts, C, H. June, S. Jalali, A. A. Lin, R. Pennathur-Das, and K. M. Hege
`
`Molecular characterization of a granulocyte macrophage—colony-stimulating factor
`receptor « subunit-associated protein, GRAP
`J. Tu, N. Karasavvas, M. L. Heaney, J. C. Vera, and D. W. Golde
`
`a-Thalassemia resulting from a negative chromosomalposition effect
`V. M. Barbour, C. Tufarelli, J. A. Sharpe, Z. E. Smith, H. Ayyub, C.A. Heinlein,
`J. Sloane-Stanley, K. Indrak, W. G. Wood, and D. R. Higgs
`
`REVIEW ARTICLES
`
`808
`
`823
`
`834
`
`840
`
`846
`
`852
`
`Therole of immunoglobulin translocations in the pathogenesis
`of B-cell malignancies
`T. G. Willis and M. J. S. Dyer
`
`Erythropoietin, iron, and erythropoiesis
`L. T. Goodnough, B. Skikne, and C. Brugnara
`
`Induction of monocyte— and T-cell—attracting chemokines in the lung during the
`generation of idiopathic pneumonia syndromefollowing allogeneic
`murine bone marrow transplantation
`A. Panoskaltsis-Mortari, R. M. Strieter, J. R. Hermanson, K. V. Fegeding, W. J. Murphy,
`C. L. Farrell, D. L. Lacey, and B. R. Blazar
`
`LECinduces chemotaxis and adhesion by interacting with CCR1 and CCR8
`O. M. Z. Howard, H. F. Dong, A.-K. Shirakawa, and J. J. Oppenheim
`
`Heparin-induced thrombocytopenia with thromboembolic complications:
`meta-analysis of 2 prospective trials to assess the value of parenteral
`treatment with lepirudin andits therapeutic aPTT range
`A. Greinacher, P. Eichler, N. Lubenow, H. Kwasny, and M. Luz
`
`Prognostic factors in Waldenstrém macroglobulinemia: a report on 232 patients
`with the description of a new scoring system andits validation on 253 other patients
`P. Morel, M. Monconduit, D. Jacomy, P. Lenain, B. Grosbois, C. Bateli, T. Facon, I. Dervite,
`F. Bauters, A. Najman, A. de Gramont, and E. Wattel
`Miltenyi Ex. 1004 Page 3
`
`Miltenyi Ex. 1004 Page 3
`
`

`

`GENE THERAPY
`
`HEMATOPOIESIS
`
`870
`
`878
`
`885
`
`894
`
`902
`
`910
`
`917
`
`925
`
`933
`
`941
`
`The immunophenotype of 177 adults with acute myeloid leukemia:
`proposal of a prognostic score
`O. Legrand,J.-Y. Perrot, M. Baudard, A. Cordier, R. Lautier, G. Simonin, R. Zittoun,
`N. Casadevall, and J.-P. Marie
`
`In vivo generation of human dendritic cell subsets by Flt3 ligand
`E. Maraskovsky, E. Daro, E. Roux, M. Teepe, C. R. Maliszewski, J. Hoek,
`D. Caron, M. E. Lebsack, and H. J. McKenna
`
`Highlyefficient gene transfer in naive humanT cells with a murine leukemia
`virus-based vector
`
`V. Dardalhon,S. Jaleco, C. Rebouissou, C. Ferrand, N. Skander, L. Swainson,P. Tiberghien,
`H. Spits, N. Noraz, and N. Taylor
`
`Inactivation of a GFP retrovirus occurs at multiple levels in long-term repopulating
`stem cells and their differentiated progeny
`C. A, Klug, S. Cheshier, and I. L. Weissman
`
`Enforced P-glycoprotein pump function in murine bone marrowcells results in
`expansion of side population stem cells in vitro and repopulatingcells in vivo
`K. D. Bunting, S. Zhou, T. Lu, and B. P. Sorrentino
`
`GATAfactor transgenes under GATA-J locus control rescue germline GATA-1
`mutant deficiencies
`S. Takahashi, R. Shimizu, N. Suwabe, T. Kuroha, K. Yoh, J. Ohta, S. Nishimura, K.-C. Lim,
`J. D. Engel, and M. Yamamoto
`
`Expression of connexin 43 (Cx43)is critical for normal hematopoiesis
`E. Montecino-Rodriguez, H. Leathers, and K. Dorshkind
`
`Inhibition of c-kit receptor tyrosine kinase activity by STI 571, a selective tyrosine
`kinase inhibitor
`
`M.C. Heinrich, D.J. Griffith, B. J. Druker, C. L. Wait, K. A. Ott, and A. J. Zigler
`
`JNK and p38 are activated by erythropoietin (EPO) but are not inducedin
`apoptosis following EPO withdrawal in EPO-dependent HCD57cells
`S. M. Jacobs-Helber, J. J. Ryan, and S. T. Sawyer
`
`A memberof Forkhead family transcription factor, FKHRL1,is one of the
`downstream molecules of phosphatidylinositol 3-kinase-Akt activation
`pathwayin erythropoietin signal transduction
`Y. Kashii, M. Uchida, K. Kirito, M. Tanaka, K. Nishijima, M. Toshima, T. Ando, K. Koizumi,
`T. Endoh, K.-i. Sawada, M. Momoi, Y. Miura, K. Ozawa, and N. Komatsu
`Miltenyi Ex. 1004 Page 4 -
`
`Miltenyi Ex. 1004 Page 4
`
`

`

`HEMOSTASIS,
`THROMBOSIS, AND
`VASCULAR BIOLOGY
`
`IMMUNOBIOLOGY
`
`950
`
`958
`
`966
`
`973
`
`979
`
`988
`
`996
`
`1006
`
`1013
`
`1021
`
`1030
`
`1039
`
`Mouse Jagged?is differentially expressed in hematopoietic progenitors and
`endothelial cells and promotes the survival and proliferation of hematopoietic
`progenitors by direct cell-to-cell contact
`S. Tsai, J. Fero, and $. Bartelmez
`
`A novel cause of mild/moderate hemophilia A: mutations scattered in the factor VIII
`Cl domain reducefactor VIII binding to von Willebrand factor
`M. Jacquemin, R. Lavend’homme,A. Benhida, B. Vanzieleghem, R. d’Oiron,J.-M. Lavergne,
`H. H. Brackmann, R. Schwaab, T. VandenDriessche, M. K. L. Chuah, M. Hoylaerts,
`J. G. G, Gilles, K. Peerlinck, J. Vermylen, and J.-M. R. Saint-Remy
`
`Induction of monocyte tissue factor expression by homocysteine: a possible
`mechanism for thrombosis
`A. Khajuria and D. S. Houston
`
`A topological study of the human y-glutamyl carboxylase
`J. Tie, S.-M. Wu, D.Jin, C. V. Nicchitta, and D. W.Stafford
`
`Hemophilic factor VIII C1- and C2-domain missense mutations and their
`modeling to the 1.5-angstrom human C2-domain crystal structure
`M.-L. Liu, B. W. Shen, S. Nakaya, K.P. Pratt, K. Fujikawa, E. W. Davie,
`B. L. Stoddard, and A. R. Thompson
`
`The factor XIII V34L polymorphism accelerates thrombin activation of factor XTII
`and affects cross-linked fibrin structure
`
`R. A. S. Ariéns, H. Philippou, C. Nagaswami, J. W. Weisel, D. A. Lane, and P. J. Grant
`
`Pharmacologic properties of Psz/P2X, receptor characterized in murine dendritic
`cells: role on the induction of apoptosis
`O. K. Nihei, A. C. Campos de Carvalho, W. Savino, and L. A. Alves
`
`Interleukin-15 redirects the outcomeofa tolerizing T-cell stimulus from apoptosis
`to anergy
`H. Dooms,T. Van Belle, M. Desmedt, P. Rottiers, and J. Grooten
`
`A-myb rescues murine B-cell lymphomas from IgM-receptor-mediated apoptosis
`through c-myc transcriptional regulation
`M. Arsura, C. §. Hofmann, J. Golay, M. Introna, and G. E. Sonenshein
`
`CD3¢ and CD28 down-modulation on CD8T cells during viral infection
`L. A. Trimble, L. W. Kam, R. S. Friedman, Z. Xu, and J. Lieberman
`
`Caspase-independent commitmentphase to apoptosis in activated blood
`T lymphocytes: reversibility at low apoptotic insult
`C. Dumont, A. Diirrbach, N. Bidére, M. Rouleau, G. Kroemer, G. Bernard, F. Hirsch,
`B. Charpentier, S$. A. Susin, and A. Senik
`
`The PI3 kinase, p38 SAP kinase, and NF-«B signal transduction pathways are
`involved in the survival and maturation of lipopolysaccharide-stimulated
`human monocyte-derived dendritic cells
`K. M.Ardeshna, A. R. Pizzey, S. Devereux, and A. Khwaja
`
`Miltenyi Ex. 1004 Page 5
`
`Miltenyi Ex. 1004 Page 5
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`

`

`1064
`
`B-cell-autonomous somatic mutation deficit following bone marrow transplant
`A. M. Glas, E. H. N. van Montfort, J. Storek, E.-G. N. Green, R. P. M. Drissen, V. J. Bechtold,
`J. Z. Reilly, M. A. Dawson, and E. C. B. Milner
`
`NEOPLASIA
`
`
`
`1070
`
`1080
`
`1087
`
`1094
`
`1100
`
`1106
`
`Selection and characterization of BCR-ABL positive cell lines with differential
`sensitivity to the tyrosine kinase inhibitor STI571: diverse mechanismsof resistance
`F. X. Mahon, M. W.N.Deininger, B. Schultheis, J. Chabrol, J. Reiffers, J. M. Goldman,
`and J. V. Melo
`
`Plasmalevels of the differentiation inhibitory factor nm23-H1 protein and their
`clinical implications in acute myelogenous leukemia
`N.Niitsu, J. Okabe-Kado, M. Nakayama, N. Wakimoto, A. Sakashita, N. Maseki, K. Motoyoshi,
`M. Umeda, and Y. Honma
`
`Molecular features responsible for the absence of immunoglobulin heavy chain
`protein synthesis in an IgH~ subgroup of multiple myeloma
`T. Szezepanski, M. B. van “t Veer, I. L. M. Wolvers-Tettero, A. W. Langerak,
`and J. J. M. van Dongen
`
`TEL/AML1 genefusionis related to in vitro drug sensitivity for L-asparaginase
`in childhood acute lymphoblastic leukemia
`N. L. Ramakers-van Woerden,R. Pieters, A. H. Loonen, I. Hubeek, E. van Drunen,
`H. B. Beverloo, R. M. Slater, J. Harbott, J. Seyfarth, E. R. van Wering, K. Hihlen,
`K. Schmiegelow, G. E. Janka-Schaub, and A. J. P. Veerman
`
`LSP1 modulates the locomotion of monocyte-differentiated U937 cells
`Y. Li, Q. Zhang, R. Aaron, L. Hilliard, and T. H. Howard
`
`Molecularanalysis of 9 new families with chronic granulomatous disease caused by
`mutations in CYBA, the gene encoding p22?!
`J. Rae, D. Noack, P. G. Heyworth, B. A. Ellis, J. T. Curnutte, and A. R. Cross
`
`PHAGOCYTES
`
`RED CELLS
`
`
`
`1113
`
`1119
`
`1125
`
`The molecular defect in hypotransferrinemic mice
`C. C. Trenor II, D. R. Campagna,V. M.Sellers, N. C. Andrews, and M. D. Fleming
`
`Fetal hemoglobinin sickle cell disease: relationship to erythrocyte
`phosphatidylserine exposure and coagulation activation
`B.N.Y. Setty, S. Kulkarni, A. K. Rao, and M. J. Stuart
`
`Wild-type HFE protein normalizes transferrin iron accumulation in macrophages
`from subjects with hereditary hemochromatosis
`G, Montosi, P. Paglia, C. Garuti, C. A. Guzman,J. M. Bastin, M. P. Colombo,and A.Pietrangelo
`Miltenyi Ex. 1004 Page 6
`
`Miltenyi Ex. 1004 Page 6
`
`

`

`1130
`
` Triose phosphate isomerase deficiency in 3 French families: two novel null alleles,
`a frameshift mutation (TPI Alfortville) and an alteration in the initiation
`codon (TPIParis)
`C. Valentin, S. Pissard, J. Martin, D. Héron, P. Labrune, M.-O. Livet, M. Mayer, T. Gelbart,
`A. Schneider, I. Max-Audit, and M. Cohen-Solal
`
`1136
`
`The human ankyrin-1 geneisselectively transcribed in erythroid cell lines despite
`the presence of a housekeeping-like promoter
`P. G. Gallagher, M. Romana,W.T. Tse, S. E. Lux, and B. G. Forget
`
`TRANSFUSION MEDICINE
`
`1144
`
`Recipient elimination of allogeneic lymphoid cells: donor CD4* cells are effective
`alloantigen-presenting cells
`L. D. Fast
`
`RECEIVED
`UCF LIBRARY
`
`TRANSPLANTATION
`1150
` Graft-versus-host disease and donor-directed hemagglutinin titers e
`after ABO-mismatched related and unrelated marrowallografts:
`evidence for a graft-versus-plasmacell effect
`M.Mielcarek, W. Leisenring, B. Torok-Storb, and R. Storb
`
`S e rl a | S
`
`1157 Molecular modification of a recombinant anti-CD3e-directed immunotoxin by
`inducing terminal cysteine bridging enhances anti-GVHD efficacy
`and reduces organtoxicity in a lethal murine model
`D. A.Vallera, D. W. Kuroki, A. Panoskaltsis-Mortari, D. J. Buchsbaum,B. E. Rogers,
`and B. R. Blazar
`
`1166 Anovel application of cyclosporine A in nonmyeloablative pretransplant host
`conditioning for allogeneic BMT
`B. Nikolic, G. Zhao, K. Swenson, and M. Sykes
`
`BRIEF REPORTS
`
`1173
`
`Therapeutic levels of human factor VIII and IX using HIV-1-based
`lentiviral vectors in mouse liver
`F. Park, K. Ohashi, and M. A. Kay
`
`1177.
`
`a-Fibrinogen Thr312Ala polymorphism and venous thromboembolism
`A. M.Carter, A. J. Catto, H. P. Kohler, R. A. S. Ariéns, M. H. Stickland, and P. J. Grant
`
`1180
`
`Splenectomyselectively affects the distribution and mobility of the recirculating
`lymphocyte pool
`T. J. Seabrook, W. R. Hein, L. Dudler, and A. J. Young
`
`1184
`
`Pure red cell aplasia due to parvovirus B19 in a patient treated with rituximab
`V.R. Sharma, D. R. Fleming, and S. P. Slone
`
`1187
`
`1191
`
`Ligation of the CD44 adhesion molecule inhibits drug-induced apoptosis
`in human myeloid leukemiacells
`M.Allouche, R. S. Charrad, A. Bettaieb, C. Greenland, C. Grignon, and F. Smadja-Joffe
`
`Familial thrombophilia associated with fibrinogen Paris V: Dusart syndrome
`T. Tarumi, D, Martincic, A. Thomas, R. Janco, M. Hudson, P. Baxter, and D. Gailani
`Miltenyi Ex. 1004 Page 7
`
`Miltenyi Ex. 1004 Page 7
`
`

`

`CONTENTS(continued)
`
`CORRESPONDENCE
`
`1194 Cytomegalovirus infection following transplantation of autologous CD34-selected
`progenitorcells
`G.J. Schiller, R. Vescio, and J. Berenson
`
`1194
`
`1196
`
`1197
`
`1199
`
`1200
`
`Cyclosporine (CSP) or CSP plus methylprednisolone for graft-versus-host-disease
`prophylaxis in patients with high-risk lymphohemopoietic malignancies: long-term
`follow-up of a randomizedtrial
`H.J. Deeg, M. E. D. Flowers, W. Leisenring, F. R. Appelbaum, P. J. Martin, and R. F. Storb
`
`Thelimitations of site-directed mutagenesis in the localization of Rh D epitopes
`T. Y. Chang and D.L. Siegel
`
`Response:limitations of prediction of epitope structure from corresponding
`antibody sequence
`N. D. Avent, W.Liu, J. W. Jones, M. L. Scott, and D. Voak
`
`A potential role for leukemia inhibitory factor in the increased clonogenicityof
`humanfetal progenitorcells
`P. Kurre and S. Burdach
`
`Response: leukemia inhibitory factor does not have a direct effect on the expansion
`of transplantable human hematopoietic stem cells
`C.-C. Shih and S, J. Forman
`
`SPECIAL DEPARTMENTS
`
`xix
`
`Author guide
`
`xxxii
`
`Future articles
`
`807
`
`863
`
`987
`
`Erratum in Bruggeret al. Positively selected autologous blood CD34* cells and
`unseparated peripheral blood progenitor cells mediate identical hematopoietic
`engraftment after high-dose VP16, ifosfamide, carboplatin, and epirubicin. Blood.
`1994;84:1421-1426.
`
`Erratum in Banget al. Unique processing pathways within recipient antigen-
`presenting cells determine IgG immunity against donor platelet MCH antigens.
`Blood. 2000;95:1735-1742.
`
`Erratum in Alcobiaet al. Spatial associations of centromeres in the nuclei of
`hematopoietic cells: evidence forcell-type-specific organizational patterns. Blood.
`2000:95:1608-1615.
`
`1202
`
`Announcements
`
`Manuscripts to be considered for publication in Blood should be sent to our new address:
`
`Kenneth Kaushansky, MD
`Editor-in-Chief,
`Blood, Journalof the
`AmericanSociety of Hematology
`1900 M Street, NW,Suite 200
`Washington, DC 20036
`
`Forother information, call (202) 776-0548. All manuscripts submitted to Blood and
`subsequently acceptedfor publication will be assessed a $50 charge per published page.
`This charge is necessary to offset the increasing costof publication.
`Miltenyi Ex. 1004 Page 8
`
`Miltenyi Ex. 1004 Page 8
`
`

`

`blood
`
`JOURNAL OF
`
`THE AMERICAN
`
`SOCIETY OF
`
`HEMATOLOGY
`
`EDITOR-IN-CHIEF
`
`Kenneth Kaushansky
`Seattle, WA
`
`EDITORIAL OFFICE
`Washington, DC
`
`Interim Managing Editor:
`Andrew Harmon
`
`Editorial Assistants:
`Louise Quinn
`Nicole Gray
`Helen Okeke
`Carolyn Elliott
`
`PUBLISHING OFFICE
`Washington, DC
`
`Director of Publishing:
`SabineJ. Beisler
`
`Production Manager:
`Todd D.Reitzel
`
`Production Editors:
`Karen S. Mason
`MeganE. O'Reilly
`Trish Weisman
`
`Production Assistant:
`Nichole S. Merkt
`
`Marketing Assistant:
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`Miltenyi Ex. 1004 Page 9
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`Miltenyi Ex. 1004 Page 9
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`
`This material may be protected by Copyright law (Title 17 U.S. Code)
`
`
`Prolongedsurvival andtissue trafficking following adoptive transfer of
`CD4¢ gene-modified autologous CD4* and CD8*Tcells in
`human immunodeficiency virus—infected subjects
`Ronald T. Mitsuyasu, Peter A. Anton, Steven G. Deeks, David T. Scadden, Elizabeth Connick, Matthew T. Downs, Andreas Bakker,
`MargoR. Roberts, Carl H. June, Sayeh Jalali, Andy A. Lin, Rukmini Pennathur-Das, and Kristen M. Hege
`
`We have genetically engineered CD4+
`and CD8* T cells with human immunode-
`
`ficiency virus (HIV) specificity by insert-
`ing a gene, CD4¢, containing the extracel-
`lular domain of human CD4 (which binds
`HIV env) linked to the zeta (<) chain of the
`T-cell
`receptor (which mediates T-cell
`activation). Twenty-four HIV-positive sub-
`jects received a single infusion of 2 to
`3 x 10'° autologous CD4i-modified CD4+
`and CD8+ T cells administered with
`
`(n = 11) or without (n = 13) interleukin-2
`(IL-2). Subjects had CD4 counts greater
`than 50/wLandviral loads of at least 1000
`
`copies/mL at entry. T cells were costimu-
`lated ex vivo through CD3 and CD28 and
`expanded for approximately 2 weeks.
`CD4¢ was detected in 1% to 3% of blood
`mononuclear cells at 8 weeks and 0.1%at
`
`1 yearafter infusion, and survival was not
`enhanced by IL-2. Trafficking of gene-
`modified T cells to bulk rectal
`tissue
`
`and/or isolated lamina propria lympho-
`cytes was documentedin a subsetof 5 of
`5 patients at 14 days and 2 of 3 at 1 year.A
`greater than 0.5 log mean decrease in
`rectal
`tissue—associated HIV RNA was
`
`compartmentalantiviral activity of CD4¢ T
`cells. CD4+ counts increased by 73/pL at
`8 weeksin the groupreceiving IL-2. There
`wasnosignificant mean change in plasma
`HIV RNAor blood proviral DNAin either
`treatment arm. This sustained, high-level
`persistence of gene-modified T cells dem-
`onstrates the feasibility of ex vivo T-cell
`gene therapy in HIV-infected adults and
`suggests the importance of providing
`HIV-specific T-helper function.
`(Blood.
`2000;96:785-793)
`
`observedforat least 14 days, suggesting
`
`© 2000 by The American Society of Hematology
`
`Introduction
`
`
`
`Much experimental and observational data suggest that the T-cell
`immune response plays a major role in containment of human
`immunodeficiency virus (HIV) during acute and chronic infection.
`The emergence of HIV-specific CD8* cytotoxic T lymphocytes
`(CTLs) coincides with the rapid decrease in plasma viremia during
`acute infection, and the frequency of HIV env-—specific CTLs is
`inversely correlated with plasmaviral load and the rate of decline
`in CD4* T-cell counts.!* Furthermore, a decline in HIV-specific
`CD8* T cells occurs in patients as they progress to later stages of
`the disease.? Most HIV-1—infected long-term nonprogressors have
`high circulating levels of HIV-1-specific CTL precursors,*+° and
`increased HIV-specific CTLs as well as high HIV-specific CD4~
`T-cell proliferative responses have been demonstrated in HIV-
`exposed seronegative subjects.®’ The most direct evidence of the
`role of CD8* T cells in HIV infection comes from recentstudies of
`CD8* T-cell depletion in acute and chronically infected macaques
`with simian immunodeficiency virus (SIV).
`In the setting of
`chronic SIV infection, CD8* T-cell depletion led to a rapid 1- to
`4-log increase in plasma viral
`load, which returned to baseline
`coincident with recovery of CD8* T cells in the blood.* Following
`acute SIV infection, CD8* T-cell depletion led to longer persis-
`
`tence of high-level viremia and a more rapidly progressive disease
`course compared with nondepleted monkeys.’
`Animal and human experiments in chronic viral infection have
`demonstrated that antigen-specific CD4~ T-cell
`responses are
`critical for maintenance of CTLs and eradication of viral infec-
`tion.!°!! Most studies have shown HIV-specific CD4* T-cell
`proliferative responses to be absent or low in patients with chronic,
`progressive HIV infection.!*!> Following initiation of combination
`highly active antiretroviral
`therapy (HAART),
`the recovery of
`HIV-specific CD4* T-helpercell responses has been observed only
`in patients treated very early after acute infection.'* Vigorous
`HIV-specific CD47 responses have been associated with control of
`viremia in the absence of drug therapy and clinical
`long-term
`nonprogression.'+ Although recent flow cytometry—based studies
`have demonstrated the presence of HIV-specific CD4* T cells in
`the majority of actively infected individuals, functional enhance-
`ment isstill likely to be necessary for optimal defense against
`HIV.!> In total, these data suggest that both arms ofthe host cellular
`immune responseare necessary for containment of HIV.
`Immunotherapy of viral
`infection with the use of antigen-
`specific T cells has been studied in the setting of cytomegalovirus
`
`From the University of California, Los Angeles, CA; San Francisco General
`Hospital, San Francisco, CA; Massachusetts General Hospital, Boston, MA;
`University of Colorado Health Science Center, Denver, CO; Statistics Collabo-
`rative, Washington DC; Specialty Labs, Los Angeles, CA; University of Virginia,
`Charlottesville, VA; University of Pennsylvania, Philadelphia, PA; and Cell
`Genesys, Inc, Foster City, CA.
`
`Submitted December8, 1999; accepted March 12, 2000.
`
`Supported by Cell Genesys Incin collaboration with Hoechst Marion Roussel.
`P.A.A. was supported in part by Mucosal
`Immunology CORE Grant no.
`Al28697,University of California, Los Angeles. This work was carried outin part
`
`in the GeneralClinical Research Centersat the followinginstitutions: University
`of California, Los Angeles (RR-00865), San Francisco General Hospital
`(5-MO1-RR00083-37), and University of Colorado Health Science Center
`(RR-00051).
`
`Reprints: Kristen M. Hege, Cell Genesys, Inc, 342 LakesideDr, Foster City, CA
`94404; e-mail: kristenh @cellgenesys.com.
`
`The publication costs of this article were defrayed in part by page charge
`payment. Therefore, and solely to indicate this fact,
`this article is hereby
`marked “advertisement”in accordance with 18 U.S.C. section 1734.
`
`© 2000 by The American Society of Hematology
`
`BLOOD, 1 AUGUST 2000 - VOLUME 96, NUMBER 3
`
`Miltenyi Ex. 1004 Page 10
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`785
`
`Miltenyi Ex. 1004 Page 10
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`

`

`recovery or adoptive transfer of antigen-specific CD4* T cells was
`required for long-term maintenance of transferred CMV- or EBV-
`specific CTLs.'°° Thus, adoptive immunotherapy with antigen-
`specific T cells is likely to require co-infusion of CD4* and CD8*
`T cells to achieve optimal in vivo survival andactivity.
`Adoptive transfer of HIV-specific T cells has potential as
`immunotherapy for HIV infection. Strategies to date have focused
`on ex vivo-expanded autologous HIV-specific CD8* monoclonal
`or polyclonal T cells.*!*4 Rather than isolating and expanding rare
`T-cell clones with major histocompatibility complex (MHC)-
`restricted antigenic specificity, a method has been developed to
`generate large numbers of HIV-specific primary T cells rapidly
`using retroviral-mediated gene transfer to insert an HIV-targeting
`gene (CD4{).2° CD4{ is a genetically engineered, MHC-unre-
`stricted, chimeric immune receptor composed of the zeta (C)
`subunit of the CD3 T-cell
`receptor (the cytoplasmic domain
`involved in signal transduction) fused to the transmembrane and
`extracellular domains of human CD4 (which targets HIV env
`expressed on the surface of infected cells). The MHC-unrestricted
`nature of this chimeric receptor allows HIV-specific targeting of
`both CD4* and CD8* T cells. This may also circumvent
`the
`potential ability of HIV to evade the T-cell
`immune response
`through down-regulation of HLA molecules on the surface of
`infectedcells.*°78
`
`Preclinical studies of CD4¢ gene-modified CD8* T cells have
`demonstrated antigen-specific proliferation and cytokine produc-
`tion, cytolytic activity against HIV-infected T cells, and inhibition
`of viral replication in HIV-infected macrophages equivalentto that
`seen with naturally occurring HIV-specific CTL clones.*? Prelimi-
`nary studies of adoptive transfer of ex vivo-expanded CD4¢-
`modified syngeneic CD8* T cells in HIV-infected twin pairs
`demonstrated a rapid decline in gene-marked cells in the blood,
`suggesting a lack of HIV-specific CD4* T-cell help to maintain the
`transferred CTLs.*° Recent advances in ex vivo culture methods
`have allowed us to develop an efficient ex vivo T-cell stimulation
`and gene transfer system that yields mixed populations of gene-
`modified CD4* and CD8*T cells.?! We now report the data from a
`phaseII clinical trial of co-infusion of autologous CD4¢-modified
`CD4* and CD8* T cells administered with or without exogenous
`interleukin-2 (IL-2) in 24 HIV-infected patients with detectable