497 A Recombinant Dimeric Ad3 Fiber Protein Triggers Opening of Epithelial Junctions in Solid Tumors and Improves Trastuzumab/Herceptin and Cetuximab/Erbitux Therapy in Mouse Models Molecular Therapy[.]
Trang 1Molecular Therapy Volume 19, Supplement 1, May 2011
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CANCER - IMMUNOTHERAPY II Cancer - Immunotherapy II
495 Effective Eradication of Established
Ovarian Murine Tumors with MUC16 Targeted T
Cells Expressing IL-12 Gene
Alena A Chekmasova,1 Samith Sandadi,2 Yan Nikhamin,1 David
R Spriggs,1 Renier J Brentjens.1,3
1 Department of Medicine, Memorial Sloan Kettering Cancer
Center, New York, NY; 2 Department of Surgery, Memorial Sloan
Kettering Cancer Center, New York, NY; 3 The Center for Cell
Engineering, Memorial Sloan Kettering Cancer Center, New York,
NY.
Despite multimodality therapy with surgery and chemotherapy,
most patients with ovarian carcinomas have a poor prognosis
For this reason, alternative approaches to treating this disease are
urgently needed Autologous T cells may be genetically modifi ed to
target tumor antigens through retroviral transduction with chimeric
antigen receptors (CARs) We have constructed SFG retroviral vectors
encoding fi rst (4H11mz) and second (4H11m28mz) generation CARs
as well as IL-12 modifi ed CAR (4H11m28mzIRESmIL12) targeted
to the retained extra-cellular domain of MUC16, termed MUC-CD
This antigen is over-expressed on most ovarian carcinoma tumor
cells Previously we have shown the effi cacy of MUC-CD targeted
T cells via effi cient in vitro cytolytic activity against MUC-CD
expressing cell lines and eradication of established MUC-CD+
tumors in SCID-Beige mice In order to mimic the clinical setting and
generate the hostile tumor microenvironment, we utilized a syngeneic
tumor model using ID8 mouse ovarian tumor cells retrovirally
transduced to overexpress MUC-CD antigen The C57BL6 mice
were intraperitoneally (i.p.) injected with 1x107 ID8(MUC-CD)
tumor cells If left untreated, these mice developed marked ascites
and multiple nodular peritoneal tumors by 8 weeks following tumor
cell injection In this series of experiments, 5x106 MUC-CD targeted
transduced mouse T cells were injected i.p on day 1 and/or 7 post
tumor injection We found a statistically enhanced long-term survival
of mice treated with 4H11mz+ or 4H11m28mz+ transduced T cells
injected 1 day after tumor injection when compared to untreated
mice or mice treated with T cells modifi ed to express an irrelevant
control CAR (4H11mz compared to 19m28mz with 37% survival
up to 120 days, p=0.0036 and 4H11m28mz compared to 19m28mz
with 80% survival up to 120 days, p=0.0001) However, treatment of
mice with advanced tumors (7 days after i.p injection of
ID8(MUC-CD) tumor cells) with 4H11m28mz+ T cells demonstrated only 27%
long term survival (p=0.005, 4H11m28mz compared to 19m28mz),
compared to IL-12 secreting 4H11m28mz T cells treated mice (100%
long term survival up to 160 days, p=0.0028) For negative controls,
tumor bearing mice were treated with T cells modifi ed to express the
irrelevant CD19-targeted 19m28mz or 19m28mzIRESIL12 CARs
To assess whether 4H11m28mzIRESmIL12+ T cells eradicate more
clinically relevant tumor burdens, we next treated C57BL6 mice 14
days post i.p ID8(MUC-CD) tumor injection At this point, there is
evidence of overt disease as assessed by FACS No gross evidence
of disease was found on day 120 after the mice were sacrifi ced
Additionally, FACS analysis of peritoneal washes was negative for
the presence of MUC-CD+ tumor cells This data supports the further
translation of these pre-clinical studies to the clinical setting in the
form of a phase I clinical trial in patients with persistent or relapsed
ovarian carcinomas
the Adoptive Immunotherapy of HER2-Positive Malignancies
Nabil Ahmed,1,2,3 Vita Salsman,1,2,3 Oumar Diouf,1,2,3 Aidin Ashoori,1,2,3 Claudia Gerken,1,2,3 Lisa Wang,1,2 Peter Anderson,4 Benjamin Musher,1 Shanda Blackmon,3 Teresa Hayes,1 John Hicks,1,2 Hao Liu,1 Winfried Wels,5 Catherine Bollard,1,2,3 Cliona Rooney,1,2,3 Gianpietro Dotti,1,2,3 Adrian Gee,1,2,3 Malcolm Brenner,1,2,3 Helen Heslop,1,2,3 Stephen Gottschalk.1,2,3
1 Baylor College of Medicine, Houston, TX; 2 Texas Children’s Hospital, Houston, TX; 3 The Methodist Hospital, Houston, TX;
4 MD Anderson Cancer Center, Houston, TX; 5 Georg-Speyer-Haus Chemotherapeutische Forschungsinstitut, Frankfurt, Germany.
The human epidermal growth factor receptor 2 (HER2) is expressed
in a broad range of adult as well as pediatric malignancies The majority of these cancers express HER2 at low levels, rendering HER2 monoclonal antibodies like trastuzumab ineffective Genetic modifi cation of T cells is one attractive approach to overcome this limitation and we have constructed a 2nd generation HER2-specifi c chimeric antigen receptor (CAR) containing the HER2-specifi c
costimualtory/signaling domain (HER2-CAR) T cells expressing HER2-CARs (HER2-CAR T cells) had potent antitumor activity
in preclinical animal models Based on these preclinical fi ndings
we developed three Phase I/II clinical studies for patients with lung cancer, osteosarcoma, and glioblastoma multiforme Due to the adverse event reported after the infusion of 1010 HER2-CAR T cells after nonmyeloablative conditioning in a patient with colon cancer, we designed our trials without lymphodepleting chemotherapy starting
at T-cell doses of 104 cells/m2 or 106 cells/m2 Our clinical studies for patients with osteosarcoma and lung cancer have recently opened for accrual and we have infused so far 6 patients with HER2-CAR T cells Patients received cell doses between 104/m2 to 1x105/m2cells Infusions were well tolerated without systemic side effects, and
HER2-CAR T cells were detectable for up to 5 weeks post infusion and localized to sites of disease In conclusion, HER2-CAR T cells have shown promising antitumor activity in preclinical animal models We are currently evaluating the safety and effi cacy of HER2-CAR T cells
in three Phase I/II clinical studies Initial safety data is encouraging, warranting further active exploration of HER2-CAR T-cell based therapies for cancer
Triggers Opening of Epithelial Junctions in Solid Tumors and Improves Trastuzumab/Herceptin and Cetuximab/Erbitux Therapy in Mouse Models
Ines Beyer,1 Hongjie Wang,1 ZongYi Li,1 Roma Yumul,1 Andre Lieber.1
1 Division of Medical Genetics, University of Washington, Seattle, WA.
The effectiveness of cancer therapeutics is decreased by the blocking of intratumoral diffusion and/or physical masking of target receptors on malignant cells In immunohistochemical studies of tumor sections from breast cancer or non-small cell lung cancer xenografts,
we observed colocalization of epithelial junction protein with Her2/ neu or with EGFR - tumor-associated antigens that are the targets for monoclonal antibodies i.e trastuzumab (Herceptin) and cetuximab (Erbitux), respectively In epithelial cells, adenovirus serotype 3 binding to the junction protein desmoglein 2 (DSG2) triggers events reminiscent of epithelial-to-mesenchymal transition, leading to the transient opening of intercellular junctions This opening improves access to receptors, e.g Her2/neu and EGFR1, that are trapped
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CANCER - IMMUNOTHERAPY II
in these intercellular junctions In addition to complete virions,
penton recombinant dodecahedral particles (PtDds) and dimeric
Ad3 fi ber domains (Ad3K-K), can trigger DSG2-mediated opening
of intercellular junctions In in vitro studies we show a signifi cant
improvement of trastuzumab dependent cell killing of the Her2/neu
positive breast cancer cell line BT474M1 subsequent to treatment
with the dimeric Ad3K fi ber We also tested whether the application
of Ad3K-K would result in the improvement of trastuzumab or
cetuximab therapy in vivo In mouse models with xenograft breast
cancer tumors (BT474-M1, HCC 1954) and xenograft lung cancer
tumors (A549), Ad3K-K improved the effi ciency of the monoclonal
antibody used and enabled control of tumor growth in subcutaneous
tumor models We are currently testing a combination of Ad3K-K and
cetuximab therapy in an orthotropic lung metastasis model, where
we expect to see a benefi t in the overall survival and control of tumor
growth in the group that received the co-therapy Our results have
potential implication for cancer therapy with T-cells or encapsulated
chemotherapy drugs
Specifi city for Safe and Effective Adoptive
Immunotherapy
Pietro Genovese,1,2 Elena Provasi,2,3 Angelo Lombardo,1,2 Zulma
Magnani,3 Pei-Qi Liu,4 Andreas Reik,4 Victoria Chu,4 David E
Paschon,4 Lei Zhang,4 Jurgen Kuball,5 Attilio Bondanza,3 Giulia
Casorati,3 Fabio Ciceri,3 Claudio Bordignon,3 Philip D Greenberg,5
Michael C Holmes,4 Philip D Gregory,4 Luigi Naldini,1,2 Chiara
Bonini.2,3
1 HSR-Telethon Institute for Gene Therapy, Milan, Italy; 2 Vita
Salute San Raffaele University, Milan, Italy; 3 San Raffaele
Scientifi c Institute, Milan, Italy; 4 Sangamo BioSciences, Richmond;
5 Fred Hutchinson Cancer Research Center, Seattle.
Transfer of a T cell receptor (TCR) from a high-avidity
tumor-specific T cell (CTL) to polyclonal T cells may overcome the
challenges of expanding rare tumor-specifi c CTLs under conditions
that preserve function and prevent exhaustion Unfortunately, the
full potential of this approach is limited by the co-expression in the
same lymphocyte of the endogenous and exogenous TCR chains,
resulting in competition for surface expression and acquisition of
novel unpredicted specifi cities due to TCR mispairing between
the 4 chains, which might lead to dangerous autoreactivity Thus,
we developed two sets of Zinc Finger Nucleases (ZFNs) to target
by the induced Non Homologous End Joining (NHEJ) repair process
We used a stimulation protocol based on anti-CD3/CD28
antibody-conjugated beads and culture with low doses of IL-7/IL-15 to preserve
cells with an early T cell differentiation phenotype Upon stimulation,
delivery of either ZFN set by Adenoviral vectors (Ad5/F35) resulted in
functional inactivation of the target genes in primary T cells (>45%),
as indicated by the generation of cells that lack surface expression of
the CD3/TCR complex (CD3neg) Once sorted, CD3neg cells proved
stable in culture for more than 50 days, maintaining an early T cell
phenotype (CD62L+ CD28+ CD27+ IL7Ra+), and were permissive
to further lentiviral vector (LV) transduction For complete editing of
T cell specifi city, we selected a codon-optimized, cystein-modifi ed
TCR specifi c for the Wilm’s Tumor Antigen 1 (WT1), involved in
oncogenic transformation in several tumors, and developed a strategy
for sequential disruption of each endogenous TCR chain followed by
LV transfer of the respective WT1-specifi c chain Using this approach
we obtained a population of TCR-edited lymphocytes, carrying only
the tumor-specifi c TCR that, in the absence of competition, was
robustly expressed at physiological levels Genetic editing did not
affect the proliferative potential and the clonability of engineered cells
nor their capacity to differentiate into effector and effector memory T
cells edited lymphocytes were superior to conventional TCR-transferred cells in WT1 pentamer binding, specifi c recognition and lysis of WT1pos targets, including primary acute myeloid leukemia blasts Importantly, these cells showed sharply reduced non-specifi c reactivity, including alloreactivity, demonstrating the advantage
of preventing TCR mispairing Overall, our results demonstrate that full genetic editing of T cell specifi city is feasible, and allows the rapid generation of effective and safe T cells for adoptive immunotherapy
To Express a PD1-CD28 Chimeric Receptor Are Co-Stimulated through the Exploitation of Tumor Expressed PD-L1
Megan E Prosser,1 Christine E Brown,1 Michael C Jensen.2
1 Department of Cancer Immunotherapeutics and Tumor Immunology, Beckman Research Institute of the City of Hope, Duarte, CA; 2 Center for Immunity and Immunotherapies, Seattle Childrens Research Institute, Seattle, WA.
Adoptive immunotherapy is a promising therapeutic approach for the treatment of malignancies However, several challenges must be addressed to enhance therapeutic effi cacy, including the loss of effector function of adoptively transferred cells in the tumor microenvironment Tumors elicit multiple mechanisms for immunosuppression of transferred cells including defi ciency of costimulation and expression of inhibitory ligands T cell activation requires both antigenic stimulation through the T cell receptor as well
as a costimulatory second signal This secondary costimulatory signal
is commonly defi cient within the tumor microenvironment resulting
in T cell anergy In addition, tumors often express the inhibitory ligand Programmed Death Ligand 1 (PD-L1) which interacts with Programmed Death 1 (PD-1) expressed on the T cell surface, resulting
in T cell exhaustion, characterized inhibition of TcR signaling, and enhanced apoptosis We hypothesize that the development
of a PD1-CD28 chimera could exploit the PD-L1 tumor derived inhibitory mechanism to result in tumor-induced costimulation and maintenance of effector function of adoptively transferred CD8+ T cells, thereby addressing the two aforementioned immunosuppressive mechanisms Preliminary assessment of this PD1-CD28 chimera in transformed cells reveals that fusion of the PD1 extracellular domain
to the intracellular signaling domain of CD28 does not alter PD-L1 binding to the extracellular PD-1 moiety Furthermore, the chimera results in increased phosphorylation of ERK and AKT suggesting increased signaling through CD28- and TCR- signaling pathways in transformed cells upon ligand binding In addition, this PD1-CD28 chimera results in increased costimulatory cytokine production in both transformed T cell lines as well as primary human CD8+ T cells The PD1-CD28 chimera also provides a proliferative advantage, as well as increased accumulation of cytotoxic granules for CD8+ T cells This work suggests that genetically engineering T cells to express the PD1-CD28 chimera may exploit an intrinsic tumor immunosuppressive mechanism to result in costimulation and maintenance of effector function of adoptively transferred cells
500 CD56-Specifi c T Cells Can Distinguish between Allogeneic and Autologous CD56+ Targets
Denise L Kellar,1 Sonny Ang,1 Simon Olivares,1 Helen Huls,1 Laurence J N Cooper.1
1 Division of Pediatrics, UT MD Anderson Cancer Center, Houston, TX.
Some candidate tumor-associated antigens (TAAs) are also expressed on T cells limiting the use of targeted T-cell therapy CD56 is an attractive TAA with expression on many malignancies, yet CD56 upregulation on a subset of activated T cells could lead to