- Email: [email protected]
900 Multipotent Mesenchymal Stromal Cells (MSC) - Evolving Role in Cancer and Leukemia Therapy
Abstracts Figure 1 Serial Monitoring of Phⴙ Cells, T315I Cells and BCR-ABL1/ABL1 Ratio
100 F359I Mutation Start KW-2449 +
Ph Cells T315I Cells (%) BCR-ABL1 Ratio
80 Dasatinib + IFN-α
80
60 60
40
40
BCR-ABL1 Ratio
Percentage of Cells (Ph+ or T315I Mutated)
100
20
20
0 0
5
17.3
18.8
20.2
21.7
24
26.2
32.6
44.5
0
Months
Special Topics [Stem Cell Transplant, Supportive Care, etc]
900 Multipotent Mesenchymal Stromal Cells (MSC) Evolving Role in Cancer and Leukemia Therapy Michael Andreeff,1,2 Venkata Lokesh Battula,1 Rui-yu Wang,1 Ye Chen,1 Xiaoyan Ling,1 Rodrigo Jacamo,1 Erika Spaeth,2 Sergej Konoplev,3 Jared Burks,1 Amy Hazen,4 Wendy Schober,1 Hongbo Lu,1 Duncan Mak,1 Teresa McQueen,1 Yuexi Shi,1 Twee Tsao,2 Liran Zhou,1 Vivian Ruvolo,2 Peter Ruvolo,1 Juliana Benito,1 Gautam Borthakur,1 Seshagiri Duvvuri,2 Ann Klopp,5 Sendurai Mani,6 Frederick Lang,7 Zhihong Zeng,1 Bing Carter,1 Steven Kornblau,1 Marina Konopleva,1,2 Frank Marini2 1
Section of Molecular Hematology & Therapy, Department of
Leukemia, The University of Texas MD Anderson Cancer Center, 2
Department of Stem Cell Transplantation and Cellular Therapy, The
University of Texas MD Anderson Cancer Center, 3Department of Hematopathology, The University of Texas MD Anderson Cancer Center, 4The University of Texas Health Science Center at Houston, 5
Department of Radiation Oncology, The University of Texas MD
Anderson Cancer Center, 6Department of Molecular Pathology, The University of Texas MD Anderson Cancer Center, 7Department of Neurosurgery, The University of Texas MD Anderson Cancer Center
S134
Clinical Lymphoma, Myeloma & Leukemia June 2011
MSC are present in most organs, but are best investigated in bone marrow and adipose tissues. They contain subsets of cells with different biological properties, but their defining feature is the ability to differentiate into chondrocytes, adipocytes, osteoblasts and perhaps neuronal cells. Most recently, they were shown to contain a population of cells capable of differentiating into cell types of the three germ layers, establishing their true multipotency (Kuroda, PNAS 2010). Our group was first to identify a role for MSC in the tumor microenvironment and to utilize their tumor homing ability to function as gene therapy delivery vehicles (Studeny M, Cancer Res 2002, 2004, 2006). Their innate homing response is related to the increased production of inflammatory mediators and is enhanced by radiation (Klopp AH, Cancer Res 2007) or chemotherapy. Bone marrowderived MSC evolve into tumor-associated fibroblasts (TAFs) (Spaeth EL, PLoS One 2009) and contribute to vasculogenesis and tumor progression. In leukemias, MSC form an important component of the hypoxic hematopoietic stem cell niche. They produce SDF-1, the ligand for the chemokine receptor CXCR4: leukemic stem cells migrate towards this SDF-1 gradient into an expanded hypoxic microenvironment that protects them from chemotherapy and signal transduction inhibitors (Zeng Z, Blood 2009; Fiegl M, Blood 2009). Similar mechanisms are operative in solid tumors and the interactions of SDF-1/CXCR4, HA/CD44 and VLA-4 provide novel therapeutic targets. Strategies of disrupting these interactions with peptides, small molecule inhibitors and soluble decoy receptors in leukemias and breast cancer will be discussed. Because transplanted MSC home only infrequently to the murine bone marrow, we have developed an ectopic, extramedullary in vivo
Abstracts model of human bone marrow that allows the investigation of genes critical for normal and leukemic hematopoiesis by genetic manipulation of its components (Chen Y, unpublished). Furthermore, evolving evidence from genomic studies suggests that MSC in leukemias frequently carry genomic abnormalities, a finding that could change our understanding of leukemia biology with potentially significant clinical implications. The availability of genetically controllable models of the BM microenvironment would facilitate these studies. MSC can be genetically modified to produce high levels of anticancer agents such as interferon alpha or beta, TRAIL, IL-24 or oncolytic adenovirus, resulting in pronounced anti-tumor activity, suppression of metastases, control of tumor growth and increased survival in syngenic or xenograft murine models of blastic transformation of CML, breast cancer, ovarian cancer and gliomas. This concept will shortly be tested in clinical trials. To better understand the origin of different stromal elements in tumors, transgenic mice with reporter genes were constructed (“rainbow stroma mice”). These investigations reveal that portions of the tumor stroma originate from bone marrow-derived MSC, while neovascular endothelial cells in tumor neo-vasculature originate from fat-derived MSC (Kidd S. . . Marini F, submitted). Interestingly, another source of tumor stroma may be the tumor itself: Mani et al (Cell 2008) reported that mammalian cells, after epithelial-mesenchymal transition (EMT), emerge as stem cells and we now characterize these cells as MSC (Battula L et al, Stem Cells 2010). Hence it may be possible that the tumor provides components of its own microenvironment, and/or of the pre-metastatic niche. In summary, our improved understanding of tumor- microenvironment interactions is yielding a more comprehensive, less tumor cell-centric view of malignancies that could affect a paradigm shift in cancer research and is already leading to the development of transformational concepts for cancer and leukemia therapy. (Supported by grants from NCI (AML P01, CML P01, Leukemia, Lymphoma and Breast Cancer, Ovarian Cancer and Glioma SPORES and RO1 and R21 grants to MA, MK, FL, FM).
903 Bone Marrow Derived Human Mesenchymal Stem Cells have the Capacity to Differentiate into B-Cells in Vivo Rui-yu Wang,1 Yuexi Shi,1 Zhihong Zeng,1 Wendy Schober,1 Jeffrey Tarrand,2 Teresa McQueen,1 Vicki Hopwood,3 Sergej Konoplev,4 Borys Korchin,1 Bingliang Fang,5 Marina Konopleva,1 Michael Andreeff1 1
Department of Leukemia, The University of Texas MD Anderson 2
Cancer Center, Clinical Laboratory, The University of Texas MD Anderson Cancer Center, 3School of Health Sciences, The University of Texas MD Anderson Cancer Center, 4
Hematopathology, The University of Texas MD Anderson Cancer
Brief Abstract: Cytogenetically diploid Human Mesenchymal Stem Cells derived from AML and healthy-donor bone marrows have the capacity to differentiate into CD19⫹ B-lineage cells in NOG mice in vivo. This unexpected finding suggests that bone marrow stroma cells may have an unexpected differentiation capacity into lymphoid hematopoietic cells. Full Abstract: Human mesenchymal stem cells (MSCs) derived from bone marrows are characterized by high proliferative potential and pluripotentiality to differentiate into multiple lineages such as osteo-, chondro-, and adipogenic cells. MSC express CD105, CD73 and CD90, but not CD45, CD34, CD33 or CD19 surface molecules. In this study, we observed that MSC derived from the bone marrows of AML patients and healthy-donors differentiated into B-cells with NOD/SCID/IL2Rgamma-/- engraftment potential. MSC cell lines were established by culturing adherent cells from newly diagnosed AML (n⫽6) and healthy donors (n⫽3) in alphaDMEM medium supplement with 20% fetal bovine serum. All MSCs were EBV (Epstein-Barr virus) negative. Surface antigen phenotype analysis and G-banding karyotype analysis were performed in passage 2-4. FACS-sorted CD90⫹/CD45- cells were then intravenously injected into NOD/SCID/IL-2Rgamma-/- (NOG) mice via tail vein (AML-BM-MSCs n⫽19, Normal-BM-MSCs n⫽11) or directly into the murine bone marrows (n⫽3). Circulating cells were analyzed for CD19, CD33, CD34, and CD90 expression on days 45, 60, 75 after injection of MSC. Results: 1) G-banding showed normal karyotype in all MSC; 2) Injected MSC engrafted and differentiated in NOG mice. Surprisingly, human CD19⫹ cells were found in all samples starting on day 45 and increasing on days 60 and 75 (from D45:0.7⫾0.5%, D60:2.6 ⫾ 1.07% and D75:6.2 ⫾ 1.0% for AML-MSC and D45:4.6⫾1.1%, D60:5.78⫾ 1.2% and D75: 3.99 ⫾ 1.43% for NL-MSC. (D45, P⫽0.007, D60, p⫽0.01, D75, p⫽0.16 respectively.); 3) CD19⫹ cells were found in peripheral blood on day45 (0.7%), day 60 (1.5%) and day75 (9.9%) in mice with AML-MSC Injected into the bone marrow; 4) CD90⫹ cells were found on day 45 (range from 0.1-8.7% and decreased to 0.10.5% on day 75). A low percentage of CD33 (D45:0.2-0.8% and D60:0.1-2.5%) and CD34⫹ cells (D45:0.3-1.9% and D60:0.22.4%) were observed before day 60 but were undetectable by day 75 in experiments with both, normal and AML-derived MSC. Conclusion: Cytogenetically diploid Human MSC derived from AML and healthy-donor bone marrows have the capacity to differentiate into CD19⫹ B-lineage cells in NOG mice in vivo. This unexpected finding suggests that bone marrow stroma cells may have an unexpected differentiation capacity into lymphoid-hematopoietic cells.
904 A Comparison of Races and Leukemia Subtypes among Patients in Different Cancer Survivorship Phases Devesh Pandya,1 Sukeshi Patel,1 Norma Ketchum,1 Brad Pollock,1 Swaminathan Padmanabhan1
Center, 5Thoracic & Cardio Surgery, The University of Texas MD
1
Anderson Cancer Center
of Hematology and Medical Oncology
University of Texas Health Science Center at San Antonio, Division
Clinical Lymphoma, Myeloma & Leukemia June 2011
S135
100 F359I Mutation Start KW-2449 +
Ph Cells T315I Cells (%) BCR-ABL1 Ratio
80 Dasatinib + IFN-α
80
60 60
40
40
BCR-ABL1 Ratio
Percentage of Cells (Ph+ or T315I Mutated)
100
20
20
0 0
5
17.3
18.8
20.2
21.7
24
26.2
32.6
44.5
0
Months
Special Topics [Stem Cell Transplant, Supportive Care, etc]
900 Multipotent Mesenchymal Stromal Cells (MSC) Evolving Role in Cancer and Leukemia Therapy Michael Andreeff,1,2 Venkata Lokesh Battula,1 Rui-yu Wang,1 Ye Chen,1 Xiaoyan Ling,1 Rodrigo Jacamo,1 Erika Spaeth,2 Sergej Konoplev,3 Jared Burks,1 Amy Hazen,4 Wendy Schober,1 Hongbo Lu,1 Duncan Mak,1 Teresa McQueen,1 Yuexi Shi,1 Twee Tsao,2 Liran Zhou,1 Vivian Ruvolo,2 Peter Ruvolo,1 Juliana Benito,1 Gautam Borthakur,1 Seshagiri Duvvuri,2 Ann Klopp,5 Sendurai Mani,6 Frederick Lang,7 Zhihong Zeng,1 Bing Carter,1 Steven Kornblau,1 Marina Konopleva,1,2 Frank Marini2 1
Section of Molecular Hematology & Therapy, Department of
Leukemia, The University of Texas MD Anderson Cancer Center, 2
Department of Stem Cell Transplantation and Cellular Therapy, The
University of Texas MD Anderson Cancer Center, 3Department of Hematopathology, The University of Texas MD Anderson Cancer Center, 4The University of Texas Health Science Center at Houston, 5
Department of Radiation Oncology, The University of Texas MD
Anderson Cancer Center, 6Department of Molecular Pathology, The University of Texas MD Anderson Cancer Center, 7Department of Neurosurgery, The University of Texas MD Anderson Cancer Center
S134
Clinical Lymphoma, Myeloma & Leukemia June 2011
MSC are present in most organs, but are best investigated in bone marrow and adipose tissues. They contain subsets of cells with different biological properties, but their defining feature is the ability to differentiate into chondrocytes, adipocytes, osteoblasts and perhaps neuronal cells. Most recently, they were shown to contain a population of cells capable of differentiating into cell types of the three germ layers, establishing their true multipotency (Kuroda, PNAS 2010). Our group was first to identify a role for MSC in the tumor microenvironment and to utilize their tumor homing ability to function as gene therapy delivery vehicles (Studeny M, Cancer Res 2002, 2004, 2006). Their innate homing response is related to the increased production of inflammatory mediators and is enhanced by radiation (Klopp AH, Cancer Res 2007) or chemotherapy. Bone marrowderived MSC evolve into tumor-associated fibroblasts (TAFs) (Spaeth EL, PLoS One 2009) and contribute to vasculogenesis and tumor progression. In leukemias, MSC form an important component of the hypoxic hematopoietic stem cell niche. They produce SDF-1, the ligand for the chemokine receptor CXCR4: leukemic stem cells migrate towards this SDF-1 gradient into an expanded hypoxic microenvironment that protects them from chemotherapy and signal transduction inhibitors (Zeng Z, Blood 2009; Fiegl M, Blood 2009). Similar mechanisms are operative in solid tumors and the interactions of SDF-1/CXCR4, HA/CD44 and VLA-4 provide novel therapeutic targets. Strategies of disrupting these interactions with peptides, small molecule inhibitors and soluble decoy receptors in leukemias and breast cancer will be discussed. Because transplanted MSC home only infrequently to the murine bone marrow, we have developed an ectopic, extramedullary in vivo
Abstracts model of human bone marrow that allows the investigation of genes critical for normal and leukemic hematopoiesis by genetic manipulation of its components (Chen Y, unpublished). Furthermore, evolving evidence from genomic studies suggests that MSC in leukemias frequently carry genomic abnormalities, a finding that could change our understanding of leukemia biology with potentially significant clinical implications. The availability of genetically controllable models of the BM microenvironment would facilitate these studies. MSC can be genetically modified to produce high levels of anticancer agents such as interferon alpha or beta, TRAIL, IL-24 or oncolytic adenovirus, resulting in pronounced anti-tumor activity, suppression of metastases, control of tumor growth and increased survival in syngenic or xenograft murine models of blastic transformation of CML, breast cancer, ovarian cancer and gliomas. This concept will shortly be tested in clinical trials. To better understand the origin of different stromal elements in tumors, transgenic mice with reporter genes were constructed (“rainbow stroma mice”). These investigations reveal that portions of the tumor stroma originate from bone marrow-derived MSC, while neovascular endothelial cells in tumor neo-vasculature originate from fat-derived MSC (Kidd S. . . Marini F, submitted). Interestingly, another source of tumor stroma may be the tumor itself: Mani et al (Cell 2008) reported that mammalian cells, after epithelial-mesenchymal transition (EMT), emerge as stem cells and we now characterize these cells as MSC (Battula L et al, Stem Cells 2010). Hence it may be possible that the tumor provides components of its own microenvironment, and/or of the pre-metastatic niche. In summary, our improved understanding of tumor- microenvironment interactions is yielding a more comprehensive, less tumor cell-centric view of malignancies that could affect a paradigm shift in cancer research and is already leading to the development of transformational concepts for cancer and leukemia therapy. (Supported by grants from NCI (AML P01, CML P01, Leukemia, Lymphoma and Breast Cancer, Ovarian Cancer and Glioma SPORES and RO1 and R21 grants to MA, MK, FL, FM).
903 Bone Marrow Derived Human Mesenchymal Stem Cells have the Capacity to Differentiate into B-Cells in Vivo Rui-yu Wang,1 Yuexi Shi,1 Zhihong Zeng,1 Wendy Schober,1 Jeffrey Tarrand,2 Teresa McQueen,1 Vicki Hopwood,3 Sergej Konoplev,4 Borys Korchin,1 Bingliang Fang,5 Marina Konopleva,1 Michael Andreeff1 1
Department of Leukemia, The University of Texas MD Anderson 2
Cancer Center, Clinical Laboratory, The University of Texas MD Anderson Cancer Center, 3School of Health Sciences, The University of Texas MD Anderson Cancer Center, 4
Hematopathology, The University of Texas MD Anderson Cancer
Brief Abstract: Cytogenetically diploid Human Mesenchymal Stem Cells derived from AML and healthy-donor bone marrows have the capacity to differentiate into CD19⫹ B-lineage cells in NOG mice in vivo. This unexpected finding suggests that bone marrow stroma cells may have an unexpected differentiation capacity into lymphoid hematopoietic cells. Full Abstract: Human mesenchymal stem cells (MSCs) derived from bone marrows are characterized by high proliferative potential and pluripotentiality to differentiate into multiple lineages such as osteo-, chondro-, and adipogenic cells. MSC express CD105, CD73 and CD90, but not CD45, CD34, CD33 or CD19 surface molecules. In this study, we observed that MSC derived from the bone marrows of AML patients and healthy-donors differentiated into B-cells with NOD/SCID/IL2Rgamma-/- engraftment potential. MSC cell lines were established by culturing adherent cells from newly diagnosed AML (n⫽6) and healthy donors (n⫽3) in alphaDMEM medium supplement with 20% fetal bovine serum. All MSCs were EBV (Epstein-Barr virus) negative. Surface antigen phenotype analysis and G-banding karyotype analysis were performed in passage 2-4. FACS-sorted CD90⫹/CD45- cells were then intravenously injected into NOD/SCID/IL-2Rgamma-/- (NOG) mice via tail vein (AML-BM-MSCs n⫽19, Normal-BM-MSCs n⫽11) or directly into the murine bone marrows (n⫽3). Circulating cells were analyzed for CD19, CD33, CD34, and CD90 expression on days 45, 60, 75 after injection of MSC. Results: 1) G-banding showed normal karyotype in all MSC; 2) Injected MSC engrafted and differentiated in NOG mice. Surprisingly, human CD19⫹ cells were found in all samples starting on day 45 and increasing on days 60 and 75 (from D45:0.7⫾0.5%, D60:2.6 ⫾ 1.07% and D75:6.2 ⫾ 1.0% for AML-MSC and D45:4.6⫾1.1%, D60:5.78⫾ 1.2% and D75: 3.99 ⫾ 1.43% for NL-MSC. (D45, P⫽0.007, D60, p⫽0.01, D75, p⫽0.16 respectively.); 3) CD19⫹ cells were found in peripheral blood on day45 (0.7%), day 60 (1.5%) and day75 (9.9%) in mice with AML-MSC Injected into the bone marrow; 4) CD90⫹ cells were found on day 45 (range from 0.1-8.7% and decreased to 0.10.5% on day 75). A low percentage of CD33 (D45:0.2-0.8% and D60:0.1-2.5%) and CD34⫹ cells (D45:0.3-1.9% and D60:0.22.4%) were observed before day 60 but were undetectable by day 75 in experiments with both, normal and AML-derived MSC. Conclusion: Cytogenetically diploid Human MSC derived from AML and healthy-donor bone marrows have the capacity to differentiate into CD19⫹ B-lineage cells in NOG mice in vivo. This unexpected finding suggests that bone marrow stroma cells may have an unexpected differentiation capacity into lymphoid-hematopoietic cells.
904 A Comparison of Races and Leukemia Subtypes among Patients in Different Cancer Survivorship Phases Devesh Pandya,1 Sukeshi Patel,1 Norma Ketchum,1 Brad Pollock,1 Swaminathan Padmanabhan1
Center, 5Thoracic & Cardio Surgery, The University of Texas MD
1
Anderson Cancer Center
of Hematology and Medical Oncology
University of Texas Health Science Center at San Antonio, Division
Clinical Lymphoma, Myeloma & Leukemia June 2011
S135