Dose and Timing Limitations in the Use of Bone Marrow-Derived Mesenchymal Stem Cells for the Treatment of Experimental Arthritis.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2326-2326
Author(s):  
Evangelia Yannaki ◽  
Anastasia Papadopoulou ◽  
Minas Yiangou ◽  
Evangelia Athanasiou ◽  
Argyrw Paraskeva ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
T Cells ◽  
Mesenchymal Stem Cells ◽  
T Cell ◽  
Broad Spectrum ◽  
Synovial Membrane ◽  
Cell Contact ◽  
Dependent Manner

Abstract The recently recognized potential of mesenchymal stem cells (MSCs) to differentiate into a broad spectrum of tissues and to act as immune regulators beyond the barriers of embryonic germ layers and major histocombatibility comlex (MHC) restriction, has emerged intense research interest on their possible use in a broad spectrum of clinical entities. Although the immunoregulatory potential of MSCs has been shown to effectively control GvHD in several preclinical and clinical studies, their role in autoimmune diseases has not been extensively explored in animal models. The goal of this study was to investigate the in vitro effect of rat bone marrow-derived MSCs on cultured fibrobIast-like synoviocytes (FLS) and T-cells from the spleen after induction of adjuvant arthritis (AA) by FCA as well as their in vivo effect in a rat model of AA resembling human rheumatoid arthritis. MSCs were isolated from bone marrow and were characterized by CD45 negativity and CD54, CD29 positivity in FCM analysis. Differentiation assays were performed to confirm their adipogenic, osteogenic and chondrogenic potential. Culture of AA-FLS in the presence of supernatant from syngeneic (syng) or allogeneic (allo) MSCs at passage 2–3, reduced the AA-FLS (p<0.022) and the ConA-stimulated AA-T-cell (p=0.04) proliferation in a dose-dependent manner, as compared to AA-FLS or AA-T-cell proliferation in the absence of supernatant. Cell-to-cell contact by coculture of activated T-cells with syng or allo MSCs produced a stronger inhibition over the supernatant (p<0.0001), in all tested MSCs dilutions and even at the lowest MSCs :T-cell ratio of 0.05:1. The inhibitory effect of allo as compared to syng MSCs in activated AA T-cells, was stronger both by secreted agents (p=0.017) or by cell to cell contact (p=0.0001). In vivo, low doses of syng MSCs (0.5-5x10^5cell/recipient) administered iv, intrasplenic or intrabone marrow, at single or multiple infusions, didn’t significantly reduce the disease score of MSC-treated as compared to control rats. In contrast, repeated, higher dose (6x10^6cell/recipient), iv infusions of syng or allogeneic MSCs from male donors (Y+MSCs) to female recipients, before the onset of AA (d4 and d9 post AA induction) resulted in significantly lower arthritic scores when compared to control animals. MSC-treated animals preserved a rather normal joint architecture with focal synovial hyperplasia, limited pannus formation and without bone destruction or chondroplasia. In contrast, the joints of arthritic control rats, appeared with a thickened synovial membrane, erosive pannus and dense inflammatory cell infiltration, chondroplasia and osteoplasia. Reduced presence of CD3+, CD11b+, NF-kb+ cells and less intense angiogenesis (FVIII+cells) was demonstrated by immunohistochemistry in the synovium of transplanted rats as compared to the control group. No Y+MSCs were detected in the spleen, bone marrow or in cultured FLS from the synovial membrane at day30 post AA induction, by PCR (sry gene), immunohistochemistry (sry protein) or FISH (Y chromosome), suggesting that the observed benefit was mostly a result of immunomodulation not derived by MSCs homing to target tissues, or migration of MSCs to target tissues may have occured earlier. On the other hand, when the same cell dose was injected after the onset of arthritis (d13 and d20 post AA induction) no clinical benefit could be observed. Our data suggest that MSCs may represent a new therapeutic approach for autoimmune arthritis, however, due to dose and timing limitations in their use, further studies are needed to clinically exploit this potential.

scholarly journals Human Umbilical Cord Mesenchymal Stem Cells Inhibit the Function of Allogeneic Activated Vγ9Vδ2 T Lymphocytes In Vitro

2015 ◽  
Vol 2015 ◽  
pp. 1-10 ◽  
Author(s):  
Xiaohuan Liu ◽  
Ting Feng ◽  
Tianxiang Gong ◽  
Chongyang Shen ◽  
Tingting Zhu ◽  
...  
Keyword(s):  
Stem Cells ◽  
Flow Cytometry ◽  
T Cells ◽  
Mesenchymal Stem Cells ◽  
T Cell ◽  
Umbilical Cord ◽  
Interleukin 2 ◽  
Cell Contact ◽  
Human Umbilical Cord ◽  
Fas L

Background. Human umbilical cord mesenchymal stem cells (UC-MSCs) can regulate the function of immune cells. However, whether and how UC-MSCs can modulate the function of Vγ9Vδ2 T cells has not been fully understood. Methods. The PBMCs or Vγ9Vδ2 T cells were activated and expanded with pamidronate (PAM) and interleukin-2 (IL-2) with or without the presence UC-MSCs. The effects of UC-MSCs on the proliferation, cytokine expression, and cytotoxicity of Vγ9Vδ2 T cells were determined by flow cytometry. The effects of UC-MSCs on Fas-L, TRAIL-expressing Vγ9Vδ2 T cells, and Vγ9Vδ2 T cell apoptosis were determined by flow cytometry. Results. UC-MSCs inhibited Vγ9Vδ2 T cell proliferation in a dose-dependent but cell-contact independent manner. Coculture with UC-MSCs reduced the frequency of IFNγ+ but increased granzyme B+ Vγ9Vδ2 T cells. UC-MSCs inhibited the cytotoxicity of Vγ9Vδ2 T cells against influenza virus H1N1 infected A549 cells and also reduced the frequency of Fas-L+, TRAIL+ Vγ9Vδ2 T cells but failed to modulate the apoptosis of Vγ9Vδ2 T cells. Conclusions. These results indicated that UC-MSCs efficiently suppressed the proliferation and cytotoxicity of Vγ9Vδ2 T cells and modulated their cytokine production. Fas-L and TRAIL were involved in the regulation. Cell contact and apoptosis of Vγ9Vδ2 T cells were not necessary for the inhibition.

Co-Stimulatory Blockade With CTLA4-Ig Permits Transplantation Of Human Hematopoietic Stem Cells and HLA Incompatible T Cells In NOD/SCID γ Null (NSG) Mouse Model

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 1999-1999
Author(s):  
Annie L. Oh ◽  
Dolores Mahmud ◽  
Benedetta Nicolini ◽  
Nadim Mahmud ◽  
Elisa Bonetti ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Nsg Mice ◽  
Human Cd34 ◽  
Cd34 Cells

Abstract Our previous studies have shown the ability of human CD34+ cells to stimulate T cell alloproliferative responses in-vitro. Here, we investigated anti-CD34 T cell alloreactivity in-vivo by co-transplanting human CD34+ cells and allogeneic T cells of an incompatible individual into NSG mice. Human CD34+ cells (2x105/animal) were transplanted with allogeneic T cells at different ratios ranging from 1:50 to 1:0.5, or without T cells as a control. No xenogeneic GVHD was detected at 1:1 CD34:T cell ratio. Engraftment of human CD45+ (huCD45+) cells in mice marrow and spleen was analyzed by flow cytometry. Marrow engraftment of huCD45+ cells at 4 or 8 weeks was significantly decreased in mice transplanted with T cells compared to control mice that did not receive T cells. More importantly, transplantation of T cells at CD34:T cell ratios from 1:50 to 1:0.5 resulted in stem cell rejection since >98% huCD45+ cells detected were CD3+. In mice with stem cell rejection, human T cells had a normal CD4:CD8 ratio and CD4+ cells were mostly CD45RA+. The kinetics of human cell engraftment in the bone marrow and spleen was then analyzed in mice transplanted with CD34+ and allogeneic T cells at 1:1 ratio and sacrificed at 1, 2, or 4 weeks. At 2 weeks post transplant, the bone marrow showed CD34-derived myeloid cells, whereas the spleen showed only allo-T cells. At 4 weeks, all myeloid cells had been rejected and only T cells were detected both in the bone marrow and spleen. Based on our previous in-vitro studies showing that T cell alloreactivity against CD34+ cells is mainly due to B7:CD28 costimulatory activation, we injected the mice with CTLA4-Ig (Abatacept, Bristol Myers Squibb, New York, NY) from d-1 to d+28 post transplantation of CD34+ and allogeneic T cells. Treatment of mice with CTLA4-Ig prevented rejection and allowed CD34+ cells to fully engraft the marrow of NSG mice at 4 weeks with an overall 13± 7% engraftment of huCD45+ marrow cells (n=5) which included: 53±9% CD33+ cells, 22±3% CD14+ monocytes, 7±2% CD1c myeloid dendritic cells, and 4±1% CD34+ cells, while CD19+ B cells were only 3±1% and CD3+ T cells were 0.5±1%. We hypothesize that CTLA4-Ig may induce the apoptotic deletion of alloreactive T cells early in the post transplant period although we could not detect T cells in the spleen as early as 7 or 10 days after transplant. Here we demonstrate that costimulatory blockade with CTLA4-Ig at the time of transplant of human CD34+ cells and incompatible allogeneic T cells can prevent T cell mediated rejection. We also show that the NSG model can be utilized to test immunotherapy strategies aimed at engrafting human stem cells across HLA barriers in-vivo. These results will prompt the design of future clinical trials of CD34+ cell transplantation for patients with severe non-malignant disorders, such as sickle cell anemia, thalassemia, immunodeficiencies or aplastic anemia. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Mechanisms behind the Immunoregulatory Dialogue between Mesenchymal Stem Cells and Th17 Cells

Cells ◽  
2020 ◽  
Vol 9 (7) ◽  
pp. 1660 ◽  
Author(s):  
Claudia Terraza-Aguirre ◽  
Mauricio Campos-Mora ◽  
Roberto Elizondo-Vega ◽  
Rafael A. Contreras-López ◽  
Patricia Luz-Crawford ◽  
...  
Keyword(s):  
Stem Cells ◽  
T Cells ◽  
Mesenchymal Stem Cells ◽  
Lupus Erythematosus ◽  
Th17 Cells ◽  
Cell Contact ◽  
T Helper 17 ◽  
Reduced Frequency

Mesenchymal stem cells (MSCs) exhibit potent immunoregulatory abilities by interacting with cells of the adaptive and innate immune system. In vitro, MSCs inhibit the differentiation of T cells into T helper 17 (Th17) cells and repress their proliferation. In vivo, the administration of MSCs to treat various experimental inflammatory and autoimmune diseases, such as rheumatoid arthritis, type 1 diabetes, multiple sclerosis, systemic lupus erythematosus, and bowel disease showed promising therapeutic results. These therapeutic properties mediated by MSCs are associated with an attenuated immune response characterized by a reduced frequency of Th17 cells and the generation of regulatory T cells. In this manuscript, we review how MSC and Th17 cells interact, communicate, and exchange information through different ways such as cell-to-cell contact, secretion of soluble factors, and organelle transfer. Moreover, we discuss the consequences of this dynamic dialogue between MSC and Th17 well described by their phenotypic and functional plasticity.

Bone marrow mesenchymal stem cells inhibit the response of naive and memory antigen-specific T cells to their cognate peptide

Blood ◽  
2003 ◽  
Vol 101 (9) ◽  
pp. 3722-3729 ◽  
Author(s):  
Mauro Krampera ◽  
Sarah Glennie ◽  
Julian Dyson ◽  
Diane Scott ◽  
Ruthline Laylor ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Cell Proliferation ◽  
T Cells ◽  
Mesenchymal Stem Cells ◽  
T Cell ◽  
Memory T Cells ◽  
T Cell Response ◽  
Target Antigen ◽  
Mhc Molecules

Mesenchymal stem cells (MSCs) have been recently shown to inhibit T-cell proliferation to polyclonal stimuli. We characterized the effect of MSCs of bone marrow origin on the T-cell response of naive and memory T cells to their cognate antigenic epitopes. The immune response to murine male transplantation antigens, HY, was selected because the peptide identity and major histocompatibility complex (MHC) restriction of the immunodominant epitopes are known. C57BL/6 female mice immunized with male cells were the source of memory T cells, whereas C6 mice transgenic for HY-specific T-cell receptor provided naive T cells. Responder cells were stimulated in vitro with male spleen cells or HY peptides in the presence or absence of MSCs. MSCs inhibited HY-specific naive and memory T cells in a dose-dependent fashion and affected cell proliferation, cytotoxicity, and the number of interferon γ (IFN-γ)–producing HY peptide-specific T cells. However, the MSC inhibitory effect did not selectively target antigen-reactive T cells. When MSCs were added to the T-cell cultures in a Transwell system or MSCs were replaced by MSC culture supernatant, the inhibitory activity was abrogated. T-cell reactivity was also restored if MSCs were removed from the cultures. The expression of MHC molecules and the presence in culture of antigen-presenting cells (APCs) or of CD4+/CD25+ regulatory T cells were not required for MSCs to inhibit. We conclude that MSCs inhibit naive and memory T-cell responses to their cognate antigens. Overall our data suggest that MSCs physically hinder T cells from the contact with APCs in a noncognate fashion.

scholarly journals The m6A “reader” YTHDF1 promotes osteogenesis of bone marrow mesenchymal stem cells through translational control of ZNF839

2021 ◽  
Vol 12 (11) ◽  
Author(s):  
Tao Liu ◽  
Xinfeng Zheng ◽  
Chenglong Wang ◽  
Chuandong Wang ◽  
Shengdan Jiang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Translational Control ◽  
Zinc Finger Protein ◽  
Dependent Manner ◽  
Marrow Mesenchymal Stem Cells ◽  
Rna Immunoprecipitation

AbstractN6-methyladenosine (m6A) is required for differentiation of human bone marrow mesenchymal stem cells (hBMSCs). However, its intrinsic mechanisms are largely unknown. To identify the possible role of m6A binding protein YTHDF1 in hBMSCs osteogenesis in vivo, we constructed Ythdf1 KO mice and showed that depletion of Ythdf1 would result in decreased bone mass in vivo. Both deletion of Ythdf1 in mouse BMSCs and shRNA-mediated knockdown of YTHDF1 in hBMSCs prevented osteogenic differentiation of cells in vitro. Using methylated RNA immunoprecipitation (Me-RIP) sequencing and RIP-sequencing, we found that ZNF839 (a zinc finger protein) served as a target of YTHDF1. We also verified its mouse homolog, Zfp839, was translationally regulated by Ythdf1 in an m6A-dependent manner. Zfp839 potentiated BMSC osteogenesis by interacting with and further enhancing the transcription activity of Runx2. These findings should improve our understanding of the mechanism of BMSC osteogenesis regulation and provide new ideas for the prevention and treatment of osteoporosis.

Enhancement of T Cell Development in Murine Bone Marrow Transplant Recipients by Transfer of the IL-7 or SCF Gene into Marrow-Derived Mesenchymal Stem Cells

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3572-3572
Author(s):  
Brile Chung ◽  
Dullei Min ◽  
Mark Krampf ◽  
Won Jong Ju ◽  
Kenneth I. Weinberg
Keyword(s):  
Stem Cells ◽  
Gene Therapy ◽  
Bone Marrow ◽  
T Cells ◽  
Mesenchymal Stem Cells ◽  
T Cell ◽  
Immune Reconstitution ◽  
T Cell Development ◽  
Cell Development ◽  
Allogeneic Bmt

Abstract The ability of the thymus to generate T cells diminishes with increasing age, the use of chemotherapy, bone marrow transplantation (BMT), anti-retroviral therapy for HIV, and graft-versus-host disease (GVHD) which can lead to a major clinical problem. Therefore, developing a clinically relevant strategy for the rapid development of T lymphocytes is crucial for treating immune deficiency. Stem cell factor (SCF: also known as kit ligand) and interleukin-7 (IL-7) are stroma–derived cytokines that induce proliferation, differentiation, and survival of developing immature T cells in the thymus. Studies have shown that administration of recombinant human IL-7 following murine BMT resulted in improved thymopoiesis and immune function. However, our previous studies have shown that that IL-7 treatment post-HSCT to enhance immune reconstitution in the allogeneic setting may have adverse effects because of the dual role of IL-7 in supporting both thymopoiesis and mature T lymphocyte expansion. Therefore it raises the question of whether IL-7 treatment after allogeneic BMT will increase the frequency or severity of GVHD. The purpose of this study was to examine whether: administration of IL-7 and SCF with infusion of mature T cell depleted (TCD) BM cells can induce enhancement of donor-derived immune reconstitution more rapidly than treatment with either cytokine alone and whether IL-7 and SCF are synergistic and partially complementary signals for the proliferation, survival, and differentiation of immature T cells. To evaluate the combinatory effect of IL-7 and SCF in T cell development following BMT, we developed a gene therapy approach using retrovirally-mediated transduction of BM-derived mesenchymal stem cells (MSC) with the human IL-7 or murine SCF gene (soluble isoform). C57BL/6J (CD45.2) recipient mice were irradiated (1300 cGy) and co-transplanted with 1 × 10 6 T cell depleted (TCD) bone marrow cells from congenic donor B6.SJL mice (CD45.1) and different doses (0.1 × 10 6 or 0.3 × 10 6) of eGFP (control), IL-7, SCF, or combination of IL-7 and SCF MSC. At day 30 following BMT, we observed that transplantation of both IL-7 and SCF MSC resulted in significantly higher numbers of donor-derived thymocytes and peripheral lymphocytes than either IL-7 or SCF MSC transplantation alone. Most noticeably, the number of donor-derived immature and mature T cells recovered from the animals receiving transplantation of 0.1 × 10 6 IL-7 MSC and 0.3 × 10 6 SCF MSC was similar to that of animals receiving 0.3 × 10 6 IL-7 MSC alone, demonstrating that the reduced proliferative signals produced by 0.1 × 10 6 IL-7 MSC can be compensated by co-transplantation of 0.3 × 10 6 SCF MSC. Moreover, transplantation of IL-7 and SCF MSC significantly increased the number of donor-derived common lymphoid progenitors (CLP [Lin-, Sca-1 low, Thy1-, c-Kit low, IL-7R+]) in the BM, suggesting that transplanted CLPs are induced to differentiate or expand more rapidly in response to IL-7 and SCF and may have contributed to increased immune reconstitution. Collectively, our findings demonstrate that IL-7 and SCF gene therapy may be a therapeutically useful method to promote enhancement of T cell development in de novo. Furthermore, the experiments resulted in important knowledge about complementary signals provided between IL-7 and SCF, and suggest various doses of IL-7 and SCF therapy may enhance development of T cells with limited expansion of mature T cells responsible for causing GVHD in allogeneic BMT setting.

scholarly journals Bone Marrow-Derived Mesenchymal Stem Cells Inhibit CD8+ t Cell Immune Responses Via PD-1/PD-L1 Pathway in Multiple Myeloma

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 53-54
Author(s):  
Zhaoyun Liu ◽  
Fu Mi ◽  
Mei Han ◽  
Mengyue Tian ◽  
Hui Liu ◽  
...  
Keyword(s):  
Stem Cells ◽  
Multiple Myeloma ◽  
Bone Marrow ◽  
T Cells ◽  
Mesenchymal Stem Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Conflicts Of Interest ◽  
Significant Difference ◽  
Normal Controls

High expression of the inhibitory receptor programmed death ligand 1 (PD-L1) on tumor cells and tumor stromal cells have been found play a key role in tumor immune evasion in several human malignancies. However, the expression of PD-L1 on bone marrow mesenchymal stem cells (BMSCs) and whether the PD-1/PD-L1 signal pathway is involved in the BMSCs versus T cell immune response in Multiple Myeloma (MM) remain poorly defined. In this study, we explored the expression of PD-L1 on BMSCs from newly diagnosed MM (NDMM) patients and the role of PD-1/PD-L1 pathway in BMSCs-mediated regulation of CD8+T cells. The data showed that the expression of PD-L1 on BMSCs in NDMM patients was significantly increased than that in normal controls (NC) (18.81±1.61% vs. 2.78±0.70 %; P<0.001). Furthermore, the PD-1 expression on CD8+T cells with NDMM patients was significantly higher than that in normal controls (43.22±2.98% vs. 20.71±1.08%; P<0.001). However, there was no significant difference in PD-1 expression of CD4+ T cells and NK cells between NDMM group and NC group. Additionally, the co-culture assays revealed that BMSCs significantly promoted CD8+ T cells apoptosis and suppressed CD8+ T cells function. However, PD-L1 inhibitor effectively reversed BMSCs-mediated suppression in CD8+ T cells. We also found that the combination of PD-L1 inhibitor and pomalidomide can further enhance the killing effect of CD8+ T cells on MM cells. In summary, our findings demonstrated that BMSCs in patients with MM may induce apoptosis of CD8+T cells through the PD-1/PD-L1 axis and inhibit the release of perforin and granzyme B from CD8+ T cells so as to promote the immune escape of MM. Disclosures No relevant conflicts of interest to declare.

scholarly journals The Therapeutic Effect of ICAM-1-Overexpressing Mesenchymal Stem Cells on Acute Graft-Versus-Host Disease

2018 ◽  
Vol 46 (6) ◽  
pp. 2624-2635 ◽  
Author(s):  
Bo Tang ◽  
Xue Li ◽  
Yuanlin Liu ◽  
Xiuhui Chen ◽  
Ximei Li ◽  
...  
Keyword(s):  
Stem Cells ◽  
T Cells ◽  
Mesenchymal Stem Cells ◽  
T Cell ◽  
Mouse Model ◽  
Graft Versus Host Disease ◽  
Culture Methods ◽  
Graft Versus Host

Background/Aims: Mesenchymal stem cells (MSCs) do not readily migrate to appropriate sites, and this creates a major obstacle for their use in the treatment of graft-versus-host disease (GVHD). Intercellular adhesion molecule-1 (ICAM-1) can guide the homing of various immune cells to the proper anatomical location within secondary lymphoid organs (SLOs), which are the major niches for generating immune responses or tolerance. MSCs rarely migrate to SLOs after intravenous infusion, and are constitutively low expression of ICAM-1. So in our previous work, ICAM-1 was engineered into a murine MSC line C3H10T1/2 by retrovirus transfection system (ICAM-1MSCs). Here, we hypothesized that ICAM-1highMSCs may significantly improve their immunomodulatory effect. Methods: We used different co-culture methods combined with real-time PCR and flow cytometry to evaluate ICAM-1highMSCs immunomodulatory effect on dendritic cells (DCs) and T cells in vitro and in vivo. MSCs were labeled with carboxyfluorescein diacetate succinimidylester (CFSE) to detect its distribution in mouse model. Results: Our in vitro analyses revealed ICAM-1 MSCs could suppress DCs maturation according to co-culture methods and suppress the T cell immune response according to the mixed lymphocyte response (MLR) and lymphoblast transformation test (LTT) tests. We found that infusion of ICAM-1highMSCs potently prolonged the survival of GVHD mouse model. The infused ICAM-1highMSCs migrate to SLOs in vivo, and suppressed DCs maturation, suppressed CD4+ T cell differentiation to Th1 cells, and increased the ratios of Treg cells. Conclusions: Taken together, these data demonstrate that ICAM-1highMSCs had an enhanced immunosuppressive effect on DCs and T cells, which may help explain the protective effect in a GVHD model. This exciting therapeutic strategy may improve the clinical efficacy of MSC-based therapy for GVHD.

scholarly journals A Novel Asymmetrical Anti-CLL-1×CD3 Bispecific Antibody, ABL602, Induces Potent CLL1-Specific Antitumor Activity with Minimized Sensitization of Pro-Inflammatory Cytokines

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 2234-2234
Author(s):  
Yangmi Lim ◽  
Eunhee Lee ◽  
Shinai Lee ◽  
Sumyeong Park ◽  
Hyeyoung Park ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Antitumor Activity ◽  
Cell Lines ◽  
Bispecific Antibody ◽  
Dependent Manner ◽  
Tumor Killing ◽  
Dose Dependent

Abstract Acute myeloid leukemia (AML) is a disease with high incidence of relapse that is originated and maintained from leukemia stem cells (LSCs). Expression of C-type lectin-like molecule-1 (CLL-1; also known as CLEC12A, c-type lectin domain family 12 member A) is mainly restricted to LSCs but absent in normal hematopoietic stem cells (HSCs), suggesting CLL-1 as an excellent therapeutic target for AML. This unique expression pattern paves the way to develop therapies that potentially eliminate CLL1-positive LSC while preserving CLL1-negative HSC. To re-direct T cells to AML cells, we generated IgG-based asymmetric (2+1, ABL602) bispecific antibody (BsAb) targeting CLL-1 and CD3. As a 2+1 format BsAb, ABL602 has bivalent binding to CLL-1 for target arm and monovalent binding to CD3. ABL602 exhibited higher binding activity to CLL-1-expressing AML cell lines and greater tumor-killing efficacy than 1+1 format BsAb and benchmark antibody MCLA-117 (Merus; CLEC12AxCD3 bispecific antibody). ABL602 induced potent cytotoxic activities on CLL1-expressing AML cell lines (EC 50 of 0.04~3.05pM and 0.97~16.64pM for U937 and HL-60, respectively) with concomitant T cell activation (EC 50 of 0.10~3.54pM and 0.94~4.92pM for U937 and HL-60, respectively) and cytokine/granzyme B release. Despite strong tumor-killing activity, ABL602 did not kill CLL1-negative cancer cell lines, suggesting that ABL602 induces CLL-1-dependent cytotoxicity. Moreover, ABL602 did not or minimally induce TNF-α and IL-6 in PBMC in the absence of AML cell lines, while MCLA-117 triggered high level of expression of those cytokines. In established orthotopic AML mouse model using HL-60 Luc, ABL602 demonstrated statistically significant anti-tumor activity in a dose-dependent manner. Proportions of bone marrow CD33 + AML blasts diminished in a dose-dependent manner, while CD3 + T cells more infiltrated to the bone marrow. Overall, our results indicate that ABL602, appropriately engineered 2+1 asymmetric BsAb, promotes T-cell activity specifically against CLL1-expressing AML cells and is a promising treatment strategy for AML patients by achieving the desired balance between antitumor activity and safety. Disclosures No relevant conflicts of interest to declare.

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