학술논문
Activity and rational combinations of a novel, engineered chimeric, TRAIL-based ligand in diffuse large B-cell lymphoma.
Document Type
Academic Journal
Author
Piechna K; Department of Experimental Hematology, Institute of Hematology and Transfusion Medicine, Warsaw, Poland.; Żołyniak A; Department of Experimental Hematology, Institute of Hematology and Transfusion Medicine, Warsaw, Poland.; Jabłońska E; Department of Experimental Hematology, Institute of Hematology and Transfusion Medicine, Warsaw, Poland.; Noyszewska-Kania M; Department of Experimental Hematology, Institute of Hematology and Transfusion Medicine, Warsaw, Poland.; Szydłowski M; Department of Experimental Hematology, Institute of Hematology and Transfusion Medicine, Warsaw, Poland.; Żerek B; Department of Drug Discovery, Adamed Pharma S.A. Pienkow, Czosnow, Poland.; Kulecka M; Department of Genetics, Maria Sklodowska-Curie National Institute of Oncology, Warsaw, Poland.; Department of Gastroenterology, Hepatology and Clinical Oncology, Centre for Postgraduate Medical Education, Warsaw, Poland.; Rumieńczyk I; Department of Genetics, Maria Sklodowska-Curie National Institute of Oncology, Warsaw, Poland.; Mikula M; Department of Genetics, Maria Sklodowska-Curie National Institute of Oncology, Warsaw, Poland.; Juszczyński P; Department of Experimental Hematology, Institute of Hematology and Transfusion Medicine, Warsaw, Poland.
Source
Publisher: Frontiers Research Foundation] Country of Publication: Switzerland NLM ID: 101568867 Publication Model: eCollection Cited Medium: Print ISSN: 2234-943X (Print) Linking ISSN: 2234943X NLM ISO Abbreviation: Front Oncol Subsets: PubMed not MEDLINE
Subject
Language
English
ISSN
2234-943X
Abstract
Background: TRAIL (TNF-related apoptosis inducing ligand) exhibits selective proapoptotic activity in multiple tumor types, while sparing normal cells. This selectivity makes TRAIL an attractive therapeutic candidate. However, despite encouraging activity in preclinical models, clinical trials with TRAIL mimetics/death receptor agonists demonstrated insufficient activity, largely due to emerging resistance to these agents. Herein, we investigated the cytotoxic activity of a novel, TRAIL-based chimeric protein AD-O51.4 combining TRAIL and VEGFA-derived peptide sequences, in hematological malignancies. We characterize key molecular mechanisms leading to resistance and propose rational pharmacological combinations sensitizing cells to AD-O51.4.
Methods: Sensitivity of DLBCL, classical Hodgkin lymphoma, (cHL), Burkitt lymphoma (BL) and acute myeloid leukemia (AML) to AD-O51.4 was assessed in vitro with MTS assay and apoptosis tests (Annexin V/PI staining). Markers of apoptosis were assessed using immunoblotting, flow cytometry or fluorogenic caspase cleavage assays. Resistant cell lines were obtained by incubation with increasing doses of AD-O51.4. Transcriptomic analyses were performed by RNA sequencing. Sensitizing effects of selected pathway modulators (BCL2, dynamin and HDAC inhibitors) were assessed using MTS/apoptosis assays.
Results: AD-O51.4 exhibited low-nanomolar cytotoxic activity in DLBCL cells, but not in other lymphoid or AML cell lines. AD-O51.4 induced death-receptor (DR) mediated, caspase-dependent apoptosis in sensitive DLBCL cells, but not in primary resistant cells. The presence of DRs and caspase 8 in cancer cells was crucial for AD-O51.4-induced apoptosis. To understand the potential mechanisms of resistance in an unbiased way, we engineered AD-O51.4-resistant cells and evaluated resistance-associated transcriptomic changes. Resistant cells exhibited changes in the expression of multiple genes and pathways associated with apoptosis, endocytosis and HDAC-dependent epigenetic reprogramming, suggesting potential therapeutic strategies of sensitization to AD-O51.4. In subsequent analyses, we demonstrated that HDAC inhibitors, BCL2 inhibitors and endocytosis/dynamin inhibitors sensitized primary resistant DLBCL cells to AD-O51.4.
Conclusions: Taken together, we identified rational pharmacologic strategies sensitizing cells to AD-O51.4, including BCL2, histone deacetylase inhibitors and dynamin modulators. Since AD-O51.4 exhibits favorable pharmacokinetics and an acceptable safety profile, its further clinical development is warranted. Identification of resistance mechanisms in a clinical setting might indicate a personalized pharmacological approach to override the resistance.
Competing Interests: BŻ is an ADAMED S.A. employee. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
(Copyright © 2022 Piechna, Żołyniak, Jabłońska, Noyszewska-Kania, Szydłowski, Żerek, Kulecka, Rumieńczyk, Mikula and Juszczyński.)
Methods: Sensitivity of DLBCL, classical Hodgkin lymphoma, (cHL), Burkitt lymphoma (BL) and acute myeloid leukemia (AML) to AD-O51.4 was assessed in vitro with MTS assay and apoptosis tests (Annexin V/PI staining). Markers of apoptosis were assessed using immunoblotting, flow cytometry or fluorogenic caspase cleavage assays. Resistant cell lines were obtained by incubation with increasing doses of AD-O51.4. Transcriptomic analyses were performed by RNA sequencing. Sensitizing effects of selected pathway modulators (BCL2, dynamin and HDAC inhibitors) were assessed using MTS/apoptosis assays.
Results: AD-O51.4 exhibited low-nanomolar cytotoxic activity in DLBCL cells, but not in other lymphoid or AML cell lines. AD-O51.4 induced death-receptor (DR) mediated, caspase-dependent apoptosis in sensitive DLBCL cells, but not in primary resistant cells. The presence of DRs and caspase 8 in cancer cells was crucial for AD-O51.4-induced apoptosis. To understand the potential mechanisms of resistance in an unbiased way, we engineered AD-O51.4-resistant cells and evaluated resistance-associated transcriptomic changes. Resistant cells exhibited changes in the expression of multiple genes and pathways associated with apoptosis, endocytosis and HDAC-dependent epigenetic reprogramming, suggesting potential therapeutic strategies of sensitization to AD-O51.4. In subsequent analyses, we demonstrated that HDAC inhibitors, BCL2 inhibitors and endocytosis/dynamin inhibitors sensitized primary resistant DLBCL cells to AD-O51.4.
Conclusions: Taken together, we identified rational pharmacologic strategies sensitizing cells to AD-O51.4, including BCL2, histone deacetylase inhibitors and dynamin modulators. Since AD-O51.4 exhibits favorable pharmacokinetics and an acceptable safety profile, its further clinical development is warranted. Identification of resistance mechanisms in a clinical setting might indicate a personalized pharmacological approach to override the resistance.
Competing Interests: BŻ is an ADAMED S.A. employee. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
(Copyright © 2022 Piechna, Żołyniak, Jabłońska, Noyszewska-Kania, Szydłowski, Żerek, Kulecka, Rumieńczyk, Mikula and Juszczyński.)