Contributions
Abstract: EP945
Type: E-Poster Presentation
Session title: Myeloma and other monoclonal gammopathies - Biology & Translational Research
Background
Melphalan flufenamide (melflufen) is a novel peptide–drug conjugate that rapidly delivers a cytotoxic alkylating agent into tumor cells. High lipophilicity of melflufen allows for increased cellular uptake, and once inside the cell, aminopeptidases hydrolyze melflufen leading to entrapment and increased concentration of cytotoxic alkylating agents. Previous studies demonstrated that treatment of myeloma cells with melflufen results in significantly higher intracellular concentrations and cytotoxicity than melphalan. In addition to increased intracellular concentration, the increased lipophilicity of melflufen has the potential to contribute to altered intracellular distribution and distinct mechanisms of action.
Aims
The subcellular localization of melflufen and melphalan could have consequences regarding unique mechanisms of action and overcoming resistance mechanisms. Therefore, understanding the subcellular localization patterns of melflufen and melphalan, and the specific effects within organelles, will provide insight into the activity of these agents in myeloma and other tumor cells.
Methods
Using radiolabeled melflufen and melphalan, we directly examined the uptake of melflufen compared to melphalan within the cytosol of the cell, as well as within the nucleus, mitochondria, lysosomes and endoplasmic reticulum (ER). Furthermore, we assessed the effects of melflufen with regard to inducing mitochondrial DNA damage as well as effects on mitochondrial reactive oxygen species, mitochondrial specific gene expression and membrane potential to assess mitochondrial function in response to melflufen and melphalan treatment.
Results
We confirmed the increased cellular uptake of melflufen into the cytosol of myeloma cells, and further established that unlike melphalan, melflufen rapidly accumulated within the nucleus, where DNA alkylators are expected to function. Additionally, we determined that melflufen, but not melphalan, localized significantly within lysosomes, ER, and importantly, mitochondria where resulting effects on mitochondrial stress were analyzed. We also examined mitochondrial expression of specific aminopeptidases with potential to hydrolyze melflufen to deliver the alkylating payload within the mitochondria. Finally, expression of mitochondrial specific gene transcripts was analyzed for effects unique to melflufen treatment resulting from melflufen accumulation.
Conclusion
The increased lipophilicity of melflufen functions not only to increase tumor cell intracellular exposure, but also redistributes the alkylating payload within the cell which may have implications for additional mechanisms of action, and contribute to overcoming resistance mechanisms.
Keyword(s): AL amyloidosis, Apoptosis, Melphalan
Abstract: EP945
Type: E-Poster Presentation
Session title: Myeloma and other monoclonal gammopathies - Biology & Translational Research
Background
Melphalan flufenamide (melflufen) is a novel peptide–drug conjugate that rapidly delivers a cytotoxic alkylating agent into tumor cells. High lipophilicity of melflufen allows for increased cellular uptake, and once inside the cell, aminopeptidases hydrolyze melflufen leading to entrapment and increased concentration of cytotoxic alkylating agents. Previous studies demonstrated that treatment of myeloma cells with melflufen results in significantly higher intracellular concentrations and cytotoxicity than melphalan. In addition to increased intracellular concentration, the increased lipophilicity of melflufen has the potential to contribute to altered intracellular distribution and distinct mechanisms of action.
Aims
The subcellular localization of melflufen and melphalan could have consequences regarding unique mechanisms of action and overcoming resistance mechanisms. Therefore, understanding the subcellular localization patterns of melflufen and melphalan, and the specific effects within organelles, will provide insight into the activity of these agents in myeloma and other tumor cells.
Methods
Using radiolabeled melflufen and melphalan, we directly examined the uptake of melflufen compared to melphalan within the cytosol of the cell, as well as within the nucleus, mitochondria, lysosomes and endoplasmic reticulum (ER). Furthermore, we assessed the effects of melflufen with regard to inducing mitochondrial DNA damage as well as effects on mitochondrial reactive oxygen species, mitochondrial specific gene expression and membrane potential to assess mitochondrial function in response to melflufen and melphalan treatment.
Results
We confirmed the increased cellular uptake of melflufen into the cytosol of myeloma cells, and further established that unlike melphalan, melflufen rapidly accumulated within the nucleus, where DNA alkylators are expected to function. Additionally, we determined that melflufen, but not melphalan, localized significantly within lysosomes, ER, and importantly, mitochondria where resulting effects on mitochondrial stress were analyzed. We also examined mitochondrial expression of specific aminopeptidases with potential to hydrolyze melflufen to deliver the alkylating payload within the mitochondria. Finally, expression of mitochondrial specific gene transcripts was analyzed for effects unique to melflufen treatment resulting from melflufen accumulation.
Conclusion
The increased lipophilicity of melflufen functions not only to increase tumor cell intracellular exposure, but also redistributes the alkylating payload within the cell which may have implications for additional mechanisms of action, and contribute to overcoming resistance mechanisms.
Keyword(s): AL amyloidosis, Apoptosis, Melphalan