8224035
`
`U'.'IITED STATES DEPARTMENT OF COMMERCE
`
`United Stntcs J>n(c,nt nnd Trndemnrk Office
`
`March 11, 2022
`
`THIS IS TO CERTIFY THAT ANNEXED IS A TRUE COPY FROM THE
`RECORDS OF THIS OFFICE OF THE FILE WRAPPER AND CONTENTS
`OF:
`
`APPLICATION NUMBER: 61/502,649
`FILING DATE: June 29, 2011
`
`Certified by
`
`Under Secretary of Commerce
`for Intellectual Property
`and Oirector of the United States
`Patent and l'rademark Office
`
`Miltenyi Ex. 1019 Page 1
`
`

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`PTO/SB/16
`0MB 0651-0032
`U.S. Patent and Trademark Office; U.S. DEPARTMENT OF COMMERCE
`PROVISIONAL APPLICATION FOR PATENT COVER SHEET
`This is a request for filing a PROVISIONAL APPLICATION FOR PATENT under 37 CPR l.53(c).
`Electronicall Filed
`
`Given Name (first and middle rif anyl)
`Carl
`
`JUNE
`
`INVENTOR(S)
`
`Family Name or Surname
`
`Residence
`(City and either State or ForeiW1 Countty)
`Merion Station, Pennsylvania
`
`□ Additional inventors are being named 011 the _ _ separately numbered sheets attached hereto
`TITLE OF THE INVENTION (500 characters max)
`COMPOSITIONS AND METHODS FOR TREATMENT OF CHRONIC
`L YMPHOCYTIC LEUKEMIA
`CORRESPONDENCE ADDRESS
`
`Direct all correspondence to:
`
`II Customer Number
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`10872
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`I
`OR
`0 Firm or Individual Name Riverside Law LLP
`Address
`300 Four Falls Corporate Center, Suite 710
`Address
`300 Conshohocken State Road
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`West Conshohocken I State
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`I Telephone
`I 215-268-3871
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`215-268-3888
`ENCLOSED APPLICATION PARTS (check all that aooly)
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`II Drawing(s) Numbero/Sheets 15
`□ Application Data Sheet. See 37 CFR 1.76
`METHOD OF PAYMENT OF FILING FEES FOR THIS PROVISIONAL APPLICATION FOR PATENT
`[8] Applicant claims small entity status. See 37 CFR 1.27.
`D A check of money order is enclosed to cover the filing fees
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`credit any overpayment to Deposit Account No. 50-5366
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`Filing Fee Amount ($110.00):
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`The invention was made by an agency of the United States Government or under a contract with an agency of the United States
`Government.
`□ No.
`t contract number arc: This invention was made with
`[8:lycs, the name of the U.S. Government agency and the Governme
`government support under K24 CA 11787901 and RO 1 CA120409 awarded by the National Institutes of Health
`(NIH).
`
`0n
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`1K:~~---- }- /-+~-,-----
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`Signature
`Typed Name: ~-Bennis L. Hna's, Ph.D., J.D.
`
`Date: b ZLl I I I
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`I
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`Registration No. 65,315
`Docket No. 46483-6001-P2-US.600881
`Telephone No. 215.268.3888
`USE ONLY FOR FILING A PROVISIONAL APPLICATION FOR PATENT
`
`Miltenyi Ex. 1019 Page 2
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`

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`Attorney Docket No. 046483-600 l-P2-US .60088 I
`
`TITLE OF THE INVENTION
`
`COMPOSITIONS AND METHODS FOR TREATMENT OF CHRONIC
`
`L YMPHOCYTIC LEUKEMIA
`
`STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR
`
`DEVELOPMENT
`
`This invention was made with government support under K24
`CAI 1787901 and ROI CA120409 awarded by the National Institutes of Health (NIH).
`The government has certain rights in the invention.
`
`BACKGROUND OF THE INVENTION
`
`The large majority of patients having B-cell malignancies, including
`chronic lymphocytic leukemia (CLL), will die from their disease. One approach to
`treating these patients is to genetically modify T cells to target antigens expressed on
`tumor cells through the expression of chimeric antigen receptors (CARs). CARs are
`antigen receptors that are designed to recognize cell surface antigens in a human
`leukocyte antigen-independent manner. Attempts in using genetically modified cells
`expressing CARs to treat these types of patients have met with very limited success. See
`for example, Brentjens et al., 2010, Molecular Therapy, 18:4, 666-668; Morgan et al.,
`2010, Molecular Therapy, published online Febrnary 23, 2010, pages 1-9; and, Till et al.,
`2008, Blood, 112:2261-2271.
`
`In most cancers, tumor-specific antigens are not yet well defined, but in B
`cell malignancies, CD 19 is an attractive tumor target. Expression of CD 19 is restricted to
`normal and malignant B cells (Uckun, et al. Blood, 1988, 71: 13-29), so that CD 19 is a
`widely accepted target to safely test CARs. While CARs can trigger T-cell activation in a
`manner similar to an endogenous T-cell receptor, a major impediment to the clinical
`application of this technology to date has been limited in vivo expansion of CAR+ T
`cells, rapid disappearance of the cells after infusion, and disappointing clinical activity
`(Jena, et al., Blood, 2010, I 16: 1035-1044; Uckun, et al. Blood, 1988, 71: 13-29).
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`5
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`10
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`15
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`20
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`25
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`30
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`Miltenyi Ex. 1019 Page 3
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`Thus, there is an urgent need in the aii for compositions and methods for
`
`treatment of CLL. The present invention addresses this need.
`
`BRIEF DESCRIPTION OF THE DRAWINGS
`
`5
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`The following detailed description of preferred embodiments of the
`
`invention will be better understood when read in conjunction with the appended
`
`drawings. For the purpose of illustrating the invention, there are shown in the drawings
`
`embodiments which are presently preferred. It should be understood, however, that the
`
`invention is not limited to the precise arrangements and instrumentalities of the
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`10
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`embodiments shown in the drawings.
`
`Figure 1, comprising Figures IA through IC, depicts a schematic
`
`representation of the gene-transfer vector and transgene, gene modified T cell
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`manufacturing and clinical protocol design. Figure 1 A depicts the lentiviral vectors and
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`transgene that show the major functional elements. A vesicular stomatitis vims protein G
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`15
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`pseudotyped clinical grade lentiviral vector ( designated pELPs 19BBz) directing
`
`expression of anti-CD19 scFv derived from FMC63 murine monoclonal antibody, human
`
`CD8a hinge and transmembrane domain, and human 4-lBB and CD3zeta signaling
`
`domains was produced. Constitutive expression of the trans gene was directed by
`
`inclusion of an EF-la (elongation factor-la promoter); LTR, long terminal repeat; RRE,
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`20
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`rev response element. ( cPPT) and the central termination sequence (CTS). Figure is not
`
`to scale. Figure 1B depicts T cell manufacturing. Autologous cells were obtained via an
`
`apheresis, and T cells were enriched by mononuclear cell elutriation, washed and residual
`
`leukemia cells depleted by addition of anti-CD3/CD28 coated paramagnetic beads for
`
`positive selection and activation ofT cells. Lentiviral vector was added at the time of
`
`25
`
`cell activation and was washed out on day 3 post culture initiation. Cells were expanded
`
`on a rocking platform device (WAVE Bioreactor System) for 8-12 days. On the final day
`
`of culture the beads were removed by passage over a magnetic field and the CART 19 T
`
`cells harvested and cryopreserved in infusible medium. Figure 1 C depicts the clinical
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`protocol design. Patients were given lymphodepleting chemotherapy as described,
`
`30
`
`followed by CART19 infusion #1 by i.v. gravity flow drip over a period of 15-20
`
`minutes. The infusion was given using a split dose approach over 3 days (10%, 30%,
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`2
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`Miltenyi Ex. 1019 Page 4
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`60%) beginning 1 to 5 days after completion of chemotherapy. Endpoint assays were
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`conducted on study week 4. At the conclusion of active monitoring, subjects were
`
`transferred to a destination protocol for long term follow up as per FDA guidance.
`
`Figure 2, comprising Figures 2A through 2F, is a series of images
`
`5
`
`demonstrating sustained in vivo expansion and persistence in blood and marrow of
`
`CART19 cells. DNA isolated from whole blood as depicted in Figure 2A through 2C or
`
`marrow as depicted in Figure 2D through 2F, samples obtained from UPN 01 as depicted
`
`in Figure 2A and 2D, UPN 02 as depicted in Figure 2B and 2E and VPN 03 as depicted
`
`in Figure 2C and 2F was subjected in bulk to Q-PCR analysis using a qualified assay to
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`10
`
`detect and quantify CART19 sequences, Each data point represents the average of
`
`triplicate measurements on 100-200 ng genomic DNA, with maximal % CV less than
`
`1.56%, Pass/fail parameters for the assay included pre-established ranges for slope and
`
`efficiency of amplification, and amplification of a reference sample. The lower limit of
`
`quantification for the assay established by the standard curve range was 2 copies
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`15
`
`transgene/microgram genomic DNA; sample values below that number are considered
`estimates and presented if at least 2/3 replicates generated a Ct value with % CV for the
`
`values 15%. CART19 cells were infused at day 0, 1, and 2 for VPN 01 and UPN 03, and
`
`days 0, 1, 2 and 11 for UPN 02.
`
`Figure 3, comprising Figures 3A through 3D, is a series of images
`
`20
`
`demonstrating serum and bone marrow cytokines before and after CART cell infusion;
`
`longitudinal measurements of changes in serum cytokines, chemokines and cytokine
`
`receptors in VPN O 1 as depicted in Figure 3A, UPN 02 as depicted in Figure 3B and UPN
`
`03 as depicted in Figure 3C, on the indicated day after CART19 cell infusion and serial
`
`assessments of the same analytes in the bone marrow from VPN 03 as depicted in Figure
`
`25
`
`3D. Samples were subjected to bulk to multiplex analysis using Luminex bead array
`
`technology and pre-assembled and validated multiplex kits. Analytes with a >=3 fold
`
`change are indicated, and plotted as relative change from baseline as depicted in Figure
`
`3A through 3C or as absolute values as depicted in Figure 3D. Absolute values for each
`
`analyte at each time-point were derived from a recombinant protein-based standard curve
`
`30
`
`over a 3-fold 8-point dilution series, with upper and lower limits of quantification
`
`(ULOQ, LLOQ) determined by the 80-120% observed/expected cutoff values for the
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`3
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`Miltenyi Ex. 1019 Page 5
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`standard curves. Each sample was evaluated in duplicate with average values calculated
`
`and% CV in most cases less than 10%. To accommodate consolidated data presentation
`in the context of the wide range for the absolute values, data are presented as fold-change
`over the baseline value for each analyte. In cases where baseline values were not
`
`5
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`detectable, half of the lowest standard curve value was used as the baseline value.
`Standard curve ranges for analytes and baseline (day 0) values (listed in parentheses
`
`sequentially for UPNOI, 02 and 03), all in pg/ml: ILl-Ra: 35.5-29,3 I 8 (689, 301, 287);
`IL-6: 2.7-4,572 (7, IO. I, 8.7); IFN-y: 11.2-23,972 (2.8, ND, 4.2 ); CXCLIO: 2.1-5,319
`
`(481,115,287); MIP-lp: 3.3-7,233 (99.7, 371,174); MCP-1: 4.8-3,600 (403,560,828);
`CXCL9: 48.2-3,700 (1,412, 126, 177); IL2-Ra: 13.4-34,210 (4,319, 9,477,610); IL-8:
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`10
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`2.4-5,278 (15.3, 14.5, 14.6); IL-10: 6.7-13,874 (8.5, 5.4, 0.7); MIP-la: 7.1-13,778 (57.6,
`
`57.3, 48.1).
`
`Figure 4, comprising Figures 4A through 4D, is a series of images
`
`depicting prolonged surface CART19 expression and establishment of functional
`
`15 memory CARs in vivo. Figure 4A depicts detection of CAR-expressing CD3+
`
`lymphocytes and absence of B cells in periphery and marrow. Freshly processed
`
`peripheral blood or marrow mononuclear cells obtained from UPN 03 at day 169 post(cid:173)
`
`CART19 cell infusion were evaluated by tlow-cytometry for surface expression of
`
`CARI 9 (top) or presence of B cells (bottom); as a control, PBMC obtained from a
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`20
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`healthy donor ND365 were stained. The gating strategy for the CD3+ and B cell
`
`populations is presented in Figure 9. To evaluate CAR19 expression in CD3+
`lymphocytes, samples were co-stained with antibodies to CD14-PE-Cy7 and CD16-PE(cid:173)
`
`Cy7 (dump channel) and CD3-FITC, positively gated on CD3+, and evaluated for
`
`CAR19 expression in the CDS+ and CDS-lymphocyte compartments by co-staining with
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`25
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`CD8a-PE and the anti-CARI 9 idiotype antibody conjugated to Alexa-64 7. Data in plots
`
`are gated on the dump channel-negative/CD3-positive cell population. To evaluate the
`
`presence of B cells, samples were co-stained with antibodies to CD14-APC and CD3-
`
`FITC (dump channels) and evaluated for the presence of B cells in the dump channel(cid:173)
`
`negative fraction by co-staining with antibodies to CD20-PE and CD19-PE-Cy-7. In all
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`30
`
`cases, negative gate quadrants were established on no-stain controls as depicted in
`
`Figures 4B and 4C. T cell immunophenotyping of CD4+ (panel b) and CDS+ (panel c) T
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`cell subsets. Frozen peripheral blood samples from UPN 03 obtained by apheresis at day
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`56 and 169 post T cell infusion were rested overnight in culture medium with no added
`
`factors, washed, and subjected to multi-parametric immunophenotyping for expression of
`
`markers of T cell memory, activation, and exhaustion. The gating strategy, as depicted in
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`5
`
`Figure 8, involved an initial gating on dump channel (CD14, CD16, Live/Dead Aqua)(cid:173)
`
`negative and CD3-positive cells, followed by positive gates on CD4+ and CDS+ cells.
`
`Gates and quadrants were established using FMO controls (CAR, CD45RA, PD-1, CD25,
`
`CD127, CCR?) or by gating on positive cell populations (CD3, CD4, CDS) and clearly
`
`delineated subsets (CD27, CD28, CD57); data were displayed after bi-exponential
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`10
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`transformation for objective visualization of events. Figure 4D depicts functional
`
`competence of persisting CAR cells. Frozen peripheral blood samples from UPN 03
`
`obtained by apheresis at day 56 and 169 post T cell infusion were rested overnight in
`
`culture medium with no added factors, washed, and evaluated directly ex-vivo for the
`
`ability to recognize CD 19-expressing target cells using CD 107 de granulation assays.
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`15
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`Following a two-hour incubation in the presence of anti-CD2S, anti-CD49d, and CD 107-
`
`FITC, cell mixtures were harvested, washed, and subjected to multi-parametric flow
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`cytometric analysis to evaluate the ability of CART19 cells to de-granulate in response to
`
`CD 19-expressing targets. The gating strategy involved an initial gate on dump channels
`
`(CD14-PE-Cy7, CD16-PE-Cy7, Live/Dead Aqua)-negative and CD3-PE-positive cells,
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`20
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`followed by gating on CDS-PE-Texas Red-positive cells; presented data is for the CDS+
`
`gated population. In all cases, negative gate quadrants were established on no-stain
`
`controls.
`
`Figure 5, comprising Figures 5A through 5C, is series of images depicting
`
`the results of experiments evaluating clinical responses after infusion ofCART19 cells.
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`25
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`Figure 5A depicts that UPN 02 was treated with two cycles ofrituximab and
`
`bendamustine with minimal response (RIB, arrow). CART19 T cells were infused
`
`beginning 4 days after bendamustine only (B, arrow). The rituximab and bendamustine(cid:173)
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`resistant leukemia was rapidly cleared from blood, as indicated by a decrease in the
`
`absolute lymphocyte count (ALC) from 60,600/itl to 200/µl within 18 days of the
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`30
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`infusion. Corticosteroid treatment was staiied on day 18 post infusion due to malaise and
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`non-infectious febrile syndrome. The reference line ( dotted) indicates upper limit of
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`normal for ALC. Figure 5B depicts the results of example experiments staining
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`sequential bone marrow biopsy or clot specimens from patient UPN 01 and 03 for CD20.
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`Pretreatment infiltration with leukemia present in both patients was absent on post
`
`treatment specimens accompanied by normalization of cellularity and trilineage
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`5
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`hematopoiesis. UPN 0 1 has not had any CLL cells detected as assessed by flow
`
`cytometry, cytogenetics and fluorescence in~situ hybridization or normal B cells detected
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`by flow cytometry in bone marrow or blood. UPN 03 had 5% residual normal CDS(cid:173)
`
`negative B cells confirmed by flow cytometry on d +23, which also showed them to be
`
`polyclonal; no normal B cells were detected at day + 176. Figure SC depicts the results of
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`10
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`experiments using sequential CT imaging to assess the rapid resolution of chemotherapy(cid:173)
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`resistant generalized lymphadenopathy. Bilateral axillary masses resolved by 83 (UPN
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`01) and 31 (UPN 03) days post infusion, as indicated by arrows and circle.
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`Figure 6, comprising Figures 6A through 6C, is a series of images
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`depicting absolute lymphocyte counts and total CART19+ cells in circulation for UPN
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`15
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`01, 02, 03. The total number oflymphocytes (Total normal and CLL cells) vs. Total
`
`CART19+ cells in circulation is plotted for all 3 subjects using the absolute lymphocyte
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`count from CBC values, and assuming a 5.0 L volume of blood, The total number of
`
`CART19 cells in circulation was calculated by using the tandem CBC values with
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`absolute lymphocyte counts and the Q-PCR marking values as depicted in Figure 2,
`converting copies/µg DNA to average % marking as described elsewhere herein. The Q(cid:173)
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`20
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`PCR % marking was found to correlate closely ( <2 fold variation) with the flow
`
`cytometric characterization of the infusion products (see Methods) and with data from
`
`samples where concomitant flow cytometry data was available to directly enumerate
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`CART19 cells by staining.
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`25
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`Figure 7, comprising Figures 7 A through 7D is a series of images
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`depicting experiments involving the direct ex vivo detection of CART19-positive cells in
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`UPN-01 PBMC 71 days post-T cell infusion. UPN-01 PBMC collected either fresh post(cid:173)
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`apheresis on day71 day post infusion, or frozen at the time of apheresis for manufacture
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`of the T cell product(baseline) and viably thawed prior to the staining, were subjected to
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`30
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`flow-cytometric analysis to detect the presence of CAR Tl 9 cells that express the CARI 9
`
`moiety on the surface. To evaluate the expression of CAR19 in lymphocytes, samples
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`were co-stained with CD3-PE and the anti-CAR19 idiotype antibody conjugated to
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`Alexa-647, or co-stained with CD3-PE alone (FMO for CAR19). Figure 7A depicts that
`
`an initial lymphocyte gate was established based on forward and side scatter (FSC vs
`SSC), followed by gating on CD3+ cells. Figure 7B depicts CD3+ lymphocyte gate;
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`5
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`Figure 7C depicts CAR idiotype stain; Figure 7D depicts CAR idiotype FMO. The
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`CAR19-positive gate was established on the CAR19 FMO samples.
`
`Figure 8, comprising Figures 8A through 8C, is a series of images
`
`depicting the gating strategy to identify CART19 expression by using polychromatic
`
`flow cytometry in UPN 03 blood specimens. The gating strategy for Figure 8C is shown
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`10
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`for the UPN 03 Day 56 sample and is representative of the strategy used on the UPN 03
`
`Day 169 sample. Figure 8A depicts primary gate: Dump (CD14, CD16, LIVE/dead
`
`Aqua) negative, CD3-positive. Figure 8B depicts secondary gates: CD4-positive, CD8-
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`positive. Figure 8C depicts tertiary gates: CARI 9-positive and CARI 9-negative,
`
`established on CAR FMO samples (right-most panels).
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`15
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`Figure 9 depicts the gating strategy to directly identify CART19
`
`expression and B cells in blood and marrow specimens. The gating strategy for Figure
`
`4A, which shows detection of CAR-expressing CD3+ lymphocytes and absence of B
`
`cells in periphery and marrow: Leftplot: Cell gate; Upper panel: positive gate for CD3+
`cells, Lower panel: negative gate (CD14-negative, CD3-negative) for B cells. NC365,
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`20
`
`peripheral blood control cells from a healthy donor
`
`Figure 10 is an image summarizing the patient demographics and
`
`response.
`
`DETAILED DESCRIPTION
`
`25
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`The present invention relates generally to the treatment of a patient having
`
`Chronic Lymphocytic Leukemia (CLL) or at risk of having CLL using lymphocyte
`
`infusion. Preferably, autologous lymphocyte infusion is used in the treatment.
`
`Autologous PBMCs are collected from a patient in need of treatment and T cells are
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`activated and expanded using the methods described herein and known in the art and then
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`30
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`infused back into the patient.
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`Miltenyi Ex. 1019 Page 9
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`In some instances, the T cells are genetically modified prior to
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`administering to a patient in need thereof, Preferably, the cell can be genetically
`
`modified to stably express an antibody binding domain on its surface, conferring novel
`
`antigen specificity that is MHC independent. Chimeric antigen receptors (CAR) are an
`
`5
`
`application of this approach that combines an antigen recognition domain of a specific
`
`antibody with an intracellular domain of the CD3-zeta chain or FcyRI protein into a
`
`single chimeric protein.
`
`In one embodiment, the invention includes autologous cells that are
`
`transfected with a vector comprising an aCD 19 CAR transgene. Preferably, the vector is
`
`10
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`a retroviral vector. More preferably, the vector is a self-inactivating lentiviral vector as
`
`described elsewhere herein. For example, chime1ic antigen receptors (CAR) are an
`
`application of this approach that combines an antigen recognition domain of a specific
`
`antibody with an intracellular domain of the CD3-zeta chain or FcyRI protein into a
`
`single chimeric protein.
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`15
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`In one embodiment, the CAR T cells further express a 4-1 BB signaling
`
`domain. For example, the CART cells of the invention can be generated by introducing
`
`a lentiviral vector comprising aCD19, CD8a hinge and transmembrane domain, and
`
`human 4-lBB and CD3zeta signaling domains into the cells. CAR-modified T cells of
`
`the invention are able to replicate in vivo resulting in long-term persistence that can lead
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`20
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`to sustained tumor control.
`
`In various embodiments, the T cells administered to the patient, or their
`
`progeny, persist in the patient for at least four months, five months, six months, seven
`
`months, eight months, nine months, ten months, eleven months, twelve months, thirteen
`
`months, fourteen month, fifteen months, sixteen months, seventeen months, eighteen
`
`25 months, nineteen months, twenty months, twenty-one months, twenty-two months,
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`twenty-three months, two years, three years, four years, or five years after administration
`
`of the T cell to the patient.
`
`In yet another embodiment, the invention relates generally to the treatment
`
`of a patient at risk of developing CLL. For example, treating a malignancy or an
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`30
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`autoimmune disease in which chemotherapy and/or immunotherapy in a patient results in
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`significant immunosuppression in the patient that raises the risk of the patient of
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`developing CLL.
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`Definitions
`
`5
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`Unless defined otherwise, all technical and scientific terms used herein
`
`have the same meaning as commonly understood by one of ordinary skill in the art to
`
`which the invention pertains. Although any methods and materials similar or equivalent
`
`to those described herein can be used in the practice for testing of the present invention,
`
`the preferred materials and methods are described herein. In describing and claiming the
`
`10
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`present invention, the following terminology will be used.
`
`It is also to be understood that the terminology used herein is for the
`
`purpose of describing particular embodiments only, and is not intended to be limiting.
`
`The at1icles "a" and "an" are used herein to refer to one or to more than
`
`one (i.e., to at least one) of the grammatical object of the article. By way of example, "an
`
`15
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`element" means one element or more than one element.
`
`"About" as used herein when referring to a measurable value such as an
`
`amount, a temporal duration, and the like, is meant to encompass variations of ±20% or
`
`±10%, more preferably ±5%, even more preferably ±1 %, and still more preferably ±0.1 %
`
`from the specified value, as such variations are appropriate to perform the disclosed
`
`20 methods.
`
`The term "antibody," as used herein, refers to an immunoglobulin
`
`molecule which specifically binds with an antigen. Antibodies can be intact
`
`immunoglobulins derived from natural sources or from recombinant sources and can be
`
`immunoreactive portions of intact immunoglobulins. Antibodies are typically tetramers
`
`25
`
`of immunoglobulin molecules. The antibodies in the present invention may exist in a
`
`variety of forms including, for example, polyclonal antibodies, monoclonal antibodies,
`
`Fv, Fab and F(ab)2, as well as single chain antibodies (scFv) and humanized antibodies
`(Harlow et al., 1999, In: Using Antibodies: A Laboratory Manual, Cold Spring Harbor
`
`Laboratory Press, NY; Harlow et al., 1989, In: Antibodies: A Laboratory Manual, Cold
`
`30
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`Spring Harbor, New York; Houston et al., 1988, Proc. Natl. Acad. Sci. USA 85:5879-
`
`5883; Bird et al., 1988, Science 242:423-426).
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`The term "antigen" or "Ag" as used herein is defined as a molecule that
`
`provokes an immune response. This immune response may involve either antibody
`
`production, or the activation of specific immunologically-competent cells, or both. The
`
`skilled artisan will understand that any macromolecule, including virtually all proteins or
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`5
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`peptides, can serve as an antigen. Furthermore, antigens can be derived from
`
`recombinant or genomic DNA. A skilled artisan will understand that any DNA, which
`
`comprises a nucleotide sequences or a partial nucleotide sequence encoding a protein that
`
`elicits an immune response therefore encodes an "antigen" as that term is used herein.
`
`Furthermore, one skilled in the art will understand that an antigen need not be encoded
`
`10
`
`solely by a full length nucleotide sequence of a gene. It is readily apparent that the
`
`present invention includes, but is not limited to, the use of partial nucleotide sequences of
`
`more than one gene and that these nucleotide sequences are arranged in various
`
`combinations to elicit the desired immune response. Moreover, a skilled artisan will
`
`understand that an antigen need not be encoded by a "gene" at all. It is readily apparent
`
`15
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`that an antigen can be generated synthesized or can be derived from a biological sample.
`
`Such a biological sample can include, but is not limited to a tissue sample, a tumor
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`sample, a cell or a biological fluid.
`
`The term "anti-tumor effect" as used herein, refers to a biological effect
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`which can be manifested by a decrease in tumor volume, a decrease in the number of
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`20
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`tumor cells, a decrease in the number of metastases, an increase in life expectancy, or
`
`amelioration of various physiological symptoms associated with the cancerous condition.
`
`An "anti-tumor effect" can also be manifested by the ability of the peptides,
`
`polynucleotides, cells and antibodies of the invention in prevention of the occurrence of
`
`tumor in the first place.
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`25
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`As used herein, the term "autologous" is meant to refer to any material
`
`derived from the same individual to which it is later to be re-introduced into the
`
`individual.
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`species.
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`"Allogeneic" refers to a graft derived from a different animal of the same
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`30
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`"Xenogeneic" refers to a graft derived from an animal of a different
`
`species.
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`10
`
`Miltenyi Ex. 1019 Page 12
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`

`

`The term "cancer" as used herein is defined as disease characterized by the
`
`rapid and uncontrolled growth of aberrant cells. Cancer cells can spread locally or
`
`through the bloodstream and lymphatic system to other parts of the body. Examples of
`
`various cancers include but are not limited to, breast cancer, prostate cancer, ovarian
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`5
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`cancer, cervical cancer, skin cancer, pancreatic cancer, colorectal cancer, renal cancer,
`liver cancer, brain cancer, lymphoma, leukemia, lung cancer and the like.
`
`"Encoding" refers to the inherent prope1iy of specific sequences of
`
`nucleotides in a polynucleotide, such as a gene, a cDNA, or an mRNA, to serve as
`
`templates for synthesis of other polymers and macromolecules in biological processes
`having either a defined sequence of nucleotides (i.e., rRNA, tRNA and mRNA) or a
`
`10
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`defined sequence of amino acids and the biological prope1iies resulting therefrom. Thus,
`a gene encodes a protein if transcription and translation of mRNA corresponding to that
`
`gene produces the protein in a cell or other biological system, Both the coding strand, the
`
`nucleotide sequence of which is identical to the mRNA sequence and is usually provided
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`15
`
`in sequence listings, and the non-coding strand, used as the template for transcription of a
`
`gene or cDNA, can be referred to as encoding the protein or other product of that gene or
`
`cDNA.
`
`Unless otherwise specified, a "nucleotide sequence encoding an amino
`
`acid sequence" includes all nucleotide sequences that are degenerate versions of each
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`20
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`other and that encode the same amino acid sequence. Nucleotide sequences that encode
`
`proteins and RNA may include intrans.
`
`"Effective amount" or "therapeutically effective amount" are used
`
`interchangeably herein, and refer to an amount of a compound, formulation, material, or
`
`composition, as described herein effective to achieve a particular biological result. Such
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`25
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`results may include, but are not limited to, the inhibition of virus infection as determined
`
`by any means suitable in the art.
`
`As used herein "endogenous" refers to any material from or produced
`
`inside an organism, cell, tissue or system.
`
`As used herein, the term "exogenous" refers to any material introduced
`
`30
`
`from or produced outside an organism, cell, tissue or system.
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`11
`
`Miltenyi Ex. 1019 Page 13
`
`

`

`The term "expression" as used herein is defined as the transcription and/or
`
`translation of a particular nucleotide sequence driven by its promoter.
`
`"Expression vector" refers to a vector comprising a recombinant
`
`polynucleotide comprising expression control sequences operatively linked to a
`
`5
`
`nucleotide sequence to be expressed. An expression vector comprises sufficient cis(cid:173)
`
`acting elements for expression; other elements for expression can be supplied by the host
`
`cell or in an in vitro expression system. Expression vectors include all those known in
`
`the aii, such as cosmids, plasmids (e.g., naked or contained in liposomes) and virnses
`
`(e.g., lentivirnses, retroviruses, adenovirnses, and adeno-associated viruses) that
`
`10
`
`incorporate the recombinant polynucleotide.
`
`"Homologous" as used herein, refers to the subunit sequence identity
`
`between two polymeric molecules, e.g., between two nucleic acid molecules, such as, two
`
`DNA molecules or two RNA molecules, or between two polypeptide molecules. When a
`
`subunit position in both of the two molecules is occupied by the same monomeric
`
`15
`
`subunit; e.g., if a position in each of two DNA molecules is occupied by adenine, then
`
`they are homologous at that position. The homology between two sequences is a direct
`
`fimction of the number of matching or homologous positions; e.g., if half (e.g., five
`
`positions in a polymer ten subunits in length) of the positions in two sequences are
`
`homologous, the two sequences are 50% homologous; if 90% of the positions (e.g., 9 of
`
`20
`
`10), are matched or homologous, the two sequences are 90% homologous.
`
`As used herein, an "instructional material" includes a publication, a
`
`recording, a diagram, or any other medium of expression which can be used to
`
`communicate the usefulness of the compositions and methods of the invention. The
`
`instrnctional mate1ial of the kit of the invention may, for example, be affixed to a
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`25
`
`container which contains the nucleic acid, peptide, and/or composition of the invention or
`
`be shipped together with a container which contains the nucleic acid, peptide, and/or
`
`composition. Alternatively, the instructional material may be shipped separately from the
`
`container with the intention that the instructional material and the compound be used
`
`cooperatively by the recipient.
`
`30
`
`"Isolated" means altered or removed from the natural state. For example,
`
`a nucleic acid or a peptide naturally present in a living animal is not "isolated," but the
`
`12
`
`Miltenyi Ex. 1019 Page 14
`
`

`

`5
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`10
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`15
`
`20
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`25
`
`same nucleic acid or peptide partially or completely separated from the coexisting
`materials of its natural state is "isolated." An isolated nucleic acid or protein can exist in
`substantially purified fonn, or can exist in a non-native environment such as, for
`example, a host cell.
`
`In the context of the present invention, the following abbreviations for the
`commonly occurring nucleic acid bases are used. "A" refers to adenosine, "C" refers to
`cytosine, "G" refers to guanosine, "T" refers to thymidine, and "U" refers to uridine.
`Unless otherwise specified, a "nucleotide sequence encoding an amino
`acid sequence" includes all nucleotide sequences that are degenerate versions of each
`other and that encode the same amino acid sequence. The phrase n