Suppression of Mir-93 May Regulate Anti-Oxidant Metabolism in Mesenchymal Stromal Cells Derived From Acute Myeloid Leukemia Patients.
Authors: Ruvolo Peter P. , Jacamo Rodrigo , McQueen Teresa , Pan Rongqing , Samudio Ismael , Konopleva Marina , Kornblau Steven M. , Garzon Ramiro , Croce Carlo M. , Andreeff Michael 
Published: October 2019

Abstract
Abstract 2354
Though still in its infancy, increasing interest is focused on the role of metabolism in cancers and leukemias. At present, most of the studies addressing this question have centered on the malignant cells themselves and not on the cells that comprise the tumor microenvironment. The leukemia microenvironment in the bone marrow has been found to provide survival advantage to AML stem cells, and to protect against chemotherapy. The underlying mechanisms, however, are not well understood. Furthermore, it is not known how the leukemic microenvironment in AML patients differs from the hematopoietic microenvironment of healthy individuals. A better understanding of leukemia will be achieved when we determine if such differences exist and importantly, how these differences could impact the microenvironment's ability to render cells resistant to therapy and possibly also support leukemogenesis. Mesenchymal stromal cells (MSC) are a key component of the bone marrow and play a key regulatory role in the leukemic niche. We have previously shown that co-cultured MSC can alter metabolism of leukemic cell lines by a mechanism involving mitochondrial uncoupling induced by the activation of Uncoupling Protein 2, to promote the Warburg Effect (Samudio et al Cancer Research, 2008; 68: 5198–5205). However, the metabolic nature of MSC from the leukemic microenvironment has not been investigated. In the present study, analysis of 352 known metabolites was performed on MSC from normal healthy donors (N = 15) and AML-derived MSC (N = 14) using Liquid Chromatography/Mass Spectrometry (LC/MS) and Gas Chromatography/Mass Spectrometry (GC/MS) (Metabolon, Durham, NC). Results indicated that metabolites involved in signaling and anti-oxidant rather than in energy regulation showed significant differences. AML derived MSC had significantly elevated levels of both reduced and oxidized glutathione compared to healthy donor MSC. Homocysteine, 5-methyltetrahydrofolate, and opthalmate levels were higher in AML MSC which is consistent with increased glutathione synthesis. The antioxidants α- and γ-tocopherol (components of vitamin E) were also significantly increased in AML MSC. These changes in key components of cellular redox homeostasis may reflect a cellular response that leads to the upregulation of glutathione synthesis and tocopherol uptake to protect the MSC cells from a more oxidative environment. In addition to metabolic analysis, we conducted miR profiling on MSC from normal healthy donors (N = 15) and AML derived MSC (N = 28). Eighteen miRs showed significant differences in expression between the 2 groups. AML derived MSC were found to express 2.3 fold less miR-93 compared to MSC from healthy donors. Previous analysis of miRs in AML cells by our groups indicated that miR-93 levels were lower in AML blast cells compared to normal counterparts (Garzon et al Blood, 2008; 111: 3183–3189). Mir-93 is a member of the miR-106b-25 cluster that comprises a group of three miRNAs on human chromosome 7q22. A recent report by Li and colleagues (Mechanisms of Ageing and Development, 2011; 132: 75–85) demonstrated in a mouse model that the aging liver is subject to damage by oxidative stress due to increased levels of miR-93. MiR-93 targets microsomal glutathione S-transferase 1 (MGST1) and the potential increase in MGST1 and the higher abundance of gluthathione support a model in which leukemia MSC could have increased xenobiotic (chemotherapy) detoxifying capacity. Future studies will include analysis of MGST1 protein expression to determine if this enzyme is indeed elevated in AML-derived MSC. In conclusion, AML-derived MSC exhibit lower expression of miR-93 that could support production of anti-oxidant metabolites to protect the cells from oxidative stress damage in the leukemic microenvironment.

Disclosures:
No relevant conflicts of interest to declare.