Targeting insulin resistance in type 2 diabetes via immune modulation of cord blood-derived multipotent stem cells (CB-SCs) in stem cell educator therapy: phase I/II clinical trial

Yong Zhao, Zhaoshun Jiang, Tingbao Zhao, Mingliang Ye, Chengjin Hu, Huimin Zhou, Zhaohui Yin, Yana Chen, Ye Zhang, Shanfeng Wang, Jie Shen, Hatim Thaker, Summit Jain, Yunxiang Li, Yalin Diao, Yingjian Chen, Xiaoming Sun, Mary Beth Fisk, Heng Li, Yong Zhao, Zhaoshun Jiang, Tingbao Zhao, Mingliang Ye, Chengjin Hu, Huimin Zhou, Zhaohui Yin, Yana Chen, Ye Zhang, Shanfeng Wang, Jie Shen, Hatim Thaker, Summit Jain, Yunxiang Li, Yalin Diao, Yingjian Chen, Xiaoming Sun, Mary Beth Fisk, Heng Li

Abstract

Background: The prevalence of type 2 diabetes (T2D) is increasing worldwide and creating a significant burden on health systems, highlighting the need for the development of innovative therapeutic approaches to overcome immune dysfunction, which is likely a key factor in the development of insulin resistance in T2D. It suggests that immune modulation may be a useful tool in treating the disease.

Methods: In an open-label, phase 1/phase 2 study, patients (N=36) with long-standing T2D were divided into three groups (Group A, oral medications, n=18; Group B, oral medications+insulin injections, n=11; Group C having impaired β-cell function with oral medications+insulin injections, n=7). All patients received one treatment with the Stem Cell Educator therapy in which a patient's blood is circulated through a closed-loop system that separates mononuclear cells from the whole blood, briefly co-cultures them with adherent cord blood-derived multipotent stem cells (CB-SCs), and returns the educated autologous cells to the patient's circulation.

Results: Clinical findings indicate that T2D patients achieve improved metabolic control and reduced inflammation markers after receiving Stem Cell Educator therapy. Median glycated hemoglobin (HbA1C) in Group A and B was significantly reduced from 8.61%±1.12 at baseline to 7.25%±0.58 at 12 weeks (P=2.62E-06), and 7.33%±1.02 at one year post-treatment (P=0.0002). Homeostasis model assessment (HOMA) of insulin resistance (HOMA-IR) demonstrated that insulin sensitivity was improved post-treatment. Notably, the islet beta-cell function in Group C subjects was markedly recovered, as demonstrated by the restoration of C-peptide levels. Mechanistic studies revealed that Stem Cell Educator therapy reverses immune dysfunctions through immune modulation on monocytes and balancing Th1/Th2/Th3 cytokine production.

Conclusions: Clinical data from the current phase 1/phase 2 study demonstrate that Stem Cell Educator therapy is a safe approach that produces lasting improvement in metabolic control for individuals with moderate or severe T2D who receive a single treatment. In addition, this approach does not appear to have the safety and ethical concerns associated with conventional stem cell-based approaches.

Trial registration: ClinicalTrials.gov number, NCT01415726.

Figures

Figure 1
Figure 1
Improvement of metabolic control by stem cell educator therapy. (A) Twelve-week follow-up of HbA1C levels in T2D subjects. (B) Analysis of insulin sensitivity by HOMA-IR C-peptide at four weeks post-treatment with Stem Cell Educator therapy. (C) 56-week follow-up C-peptide levels in Group C T2D subjects with impaired islet β cell function. (D) Analysis of islet β cell function by HOMA-B C-peptide at 12-week follow-up post-treatment with Stem Cell Educator therapy in Group C T2D subjects.
Figure 2
Figure 2
Anti-inflammatory effects of stem cell educator therapy. (A) Up-regulation of plasma levels of TGF-β1 in T2D patients at baseline and four weeks after Stem Cell Educator therapy. (B) Flow analysis of intra-cellular cytokines demonstrating differential effects on key interleukins at four weeks post-treatment. (C) Down-regulation percentage of CD86+CD14+monocytes in T2D patients at baseline and four weeks after Stem Cell Educator therapy. (D) Flow Analysis of CD4+CD25+Foxp3+ Tregs demonstrating no change in the percentage of Tregs at four weeks post-treatment.
Figure 3
Figure 3
In vitro study of the immune modulation of CB-SCs on monocytes. (A) Phase contrast microscopy shows the co-culture of CB-SC with monocytes (bottom left panel) for 18 hrs. CB-SCs co-culture with lymphocytes (top right panel) served as the control. The impaired CB-SCs after co-culture with monocytes were restored to expansion and became 90 to approximately 100% confluence after 7 to 10 days (bottom right). Original magnification, × 100. (B) Apoptotic analysis of floating cells from the co-culture of CB-SCs with monocytes for 18 hrs. (C) Western blotting shows the expression of the cellular inhibitor of apoptosis protein (cIAP) 1, not cIAP2, in four preparations of CB-SCs. (D) Western blotting shows the expression of tumor necrosis factor receptor II (TNF-RII), not TNF-RI, in four preparations of CB-SCs. (E) TNFα suppresses the proliferation of CB-SCs in a dose–response manner. Cell proliferation was evaluated using CyQUANTR Cell Proliferation Assay Kit [25]. (F) The blocking experiment with iNOS inhibitor 1400W demonstrates that CB-SC-derived nitric oxide (NO) contributes to the immune modulation of CB-SCs on monocytes. Monocytes were initially stimulated with lipopolysaccharide (LPS, 10 μg/ml) for 8 hrs, and then co-cultured with CB-SCs at ratio 1:5 of CB-SCs:monocytes for 48 hrs in the presence or absence of 1400W (100 nM), followed by real time PCR analysis by using Human Th17 for Autoimmunity and Inflammation PCR Array kit (SABiosciences, Valencia, CA, USA).

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