Abstract
BackgroundRelapsed T-lineage acute lymphoblastic leukemia (T-ALL) has been an incurable disease. Recent reports showed that an L-arginine depleting enzyme, pegylated arginase (BCT-100) may be effective against T-ALL cells. On the other hand, studies including ours had shown the symbiosis of ALL blasts and human mesenchymal stromal cells (hMSCs) in bone marrow microenvironment during L-asparaginase treatment. As L-asparaginase and BCT-100 both act by depleting lymphoid cells of specific amino acid, we hypothesized that hMSCs may also protect T-ALL blasts from BCT-100 treatment in co-culture and such protection may be abrogated by pre-treating hMSCs with vincristine (VCR).MethodsXTT assay was used to test sensitivities of T-ALL cell lines and hMSCs to BCT-100. Apoptosis of T-ALL cell lines with or without BCT-100 treatment were tested by annexin V / propidium iodide (AV/PI) assay using flow cytometer. Western blotting was performed to analyze the expression of ornithine transcarbamylase (OTC), an enzyme involved in L-arginine metabolism which may account for BCT-100 resistance.ResultshMSCs were resistant to BCT-100 while CCRF-CEM, Jurkat and MOLT-4 were very sensitive to it. hMSCs could protect all the three cell lines from BCT-100 treatment in transwell co-culture. All the 3 T-ALL cell lines were also found to be rescued by an L-arginine precursor citrulline, while the breakdown product of BCT-100, ornithine only had limited salvaging effect on CCRF-CEM but not Jurkat and MOLT-4. Both hMSCs and 3 T-ALL cell lines express citrulline synthesis enzyme, ornithine transcarbamylase (OTC) at basal level while only hMSCs could express OTC at relatively higher level under BCT-100 treatment. Treating hMSCs with vincristine before co-culturing with T-ALL could resume the cytotoxicity of BCT-100 to CCRF-CEM and MOLT-4 cells.ConclusionsOur results suggest a possible strategy to overcome resistance to BCT-100 from cancer microenvironments by suppressing hMSCs either in marrow or in the perivascular niche using vincristine.
Highlights
Relapsed T-lineage acute lymphoblastic leukemia (T-Acute lymphoblastic leukemia (ALL)) has been an incurable disease
We showed that human mesenchymal stromal cells (hMSCs) were resistant to BCT-100 and such differential cytotoxicity of BCT-100 to T-lineage acute lymphoblastic leukemia (T-ALL) blasts and hMSCs might imply a possible symbiosis between T-ALL and hMSCs during arginine starvation stress, similar to the B-lineage ALL (B-ALL) blasts/hMSCs symbiosis during asparagine depletion [5]
By annexin V / propidium iodide (AV/propidium iodide (PI)) assay using flow cytometry we showed that pre-treating hMSCs or hTertMSCs with VCR did not increase the percentage of T-ALL blasts apoptosis in all the three T-ALL cell lines (Figure 5a)
Summary
Relapsed T-lineage acute lymphoblastic leukemia (T-ALL) has been an incurable disease. Studies including ours had shown the symbiosis of ALL blasts and human mesenchymal stromal cells (hMSCs) in bone marrow microenvironment during L-asparaginase treatment. As L-asparaginase and BCT-100 both act by depleting lymphoid cells of specific amino acid, we hypothesized that hMSCs may protect T-ALL blasts from BCT-100 treatment in co-culture and such protection may be abrogated by pre-treating hMSCs with vincristine (VCR). It has been shown that mesenchymal stromal cells (MSCs) may contribute to L-asparaginase resistance of B-ALL blasts in bone marrow microenvironment by secreting asparagine [5]. MSCs are resistant to most of the commonly used chemotherapeutic agents for leukemia, but remain sensitive to anti-microtubule agents such as paclitaxel and vincristine (VCR) [6]. Drugs with higher efficacy against T-ALL are urgently needed
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