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Anti-melanoma activity of the 9.2.27PE immunotoxin in dacarbazine resistant cells
Karianne Risberg1, Oystein Fodstad, Yvonne Andersson
1Department of Tumor Biology, Institute for Cancer Research, Oslo University Hospital Radiumhospitalet, The Norwegian Radium Hospital, University in Oslo, Norway. Karianne.Risberg@rr-research.no
Abstract:
We have earlier shown that the 9.2.27 Pseudomonas Exotoxin A (PE) immunotoxin (IT) efficiently kills melanoma cells through inhibition of protein synthesis followed by some morphologic and biochemical features of apoptosis, a different cell killing mechanism than the one caused by Dacarbazine (DTIC), a chemotherapeutic drug used to treat malignant melanoma. To examine whether induced DTIC resistance also is a determining factor for the effectiveness of 9.2.27PE IT, we developed a DTIC resistant subline, FEMX-200DR, from the DTIC sensitive cell line FEMX. The cell variants were treated with 9.2.27PE, an IT binding to the high molecular weight-melanoma associated antigen (HMW-MAA) expressed on most malignant melanoma cells. The IT was equally effective in killing the FEMX-200DR and the FEMX cells, and the cell death was primarily caused by inhibition of protein synthesis. The DNA repair enzyme and apoptotic marker PARP, a substrate of caspase-3, was inactivated, although we observed only a minor activation of caspase-3 and caspase-8, intracellular proteases involved in apoptosis. In addition to being DTIC resistant, the FEMX-200DR cells were also more resistant to apoptosis than the parent cells as a 3 times higher concentration of the apoptotic inducer Staurosporine was needed to obtain IC50. Furthermore, in early passage malignant melanoma cell lines established from lymph node metastases, the 9.2.27PE caused a time-dependent and dose-dependent decrease in cell viability independent of their DTIC sensitivity. These findings show that the 9.2.27PE IT efficiently can cause cell death in malignant melanoma cells independent of their level of resistance to apoptosis and DTIC.
Insights
The 9.2.27 Pseudomonas Exotoxin A immunotoxin effectively kills melanoma cells by inhibiting protein synthesis. This immunotoxin works even in cells resistant to Dacarbazine and apoptosis, offering a promising melanoma treatment strategy.
Area of Science:
- Oncology
- Immunology
- Molecular Biology
Background:
- Malignant melanoma is a significant health concern.
- Dacarbazine (DTIC) is a standard chemotherapy for melanoma.
- Pseudomonas Exotoxin A (PE) immunotoxins (ITs) show promise in melanoma treatment.
Purpose of the Study:
- To investigate if Dacarbazine (DTIC) resistance affects the efficacy of the 9.2.27 Pseudomonas Exotoxin A (PE) immunotoxin (IT).
- To determine if the 9.2.27 PE IT is effective against melanoma cells with acquired resistance to DTIC and apoptosis.
Main Methods:
- Developed a DTIC-resistant melanoma cell line (FEMX-200DR) from a sensitive parent line (FEMX).
- Treated both cell lines with 9.2.27 PE IT, which targets the high molecular weight-melanoma associated antigen (HMW-MAA).
- Assessed cell viability, protein synthesis inhibition, and apoptosis markers (caspase activation, PARP inactivation) in response to treatment.
Main Results:
- The 9.2.27 PE IT demonstrated equal effectiveness in killing both DTIC-sensitive (FEMX) and DTIC-resistant (FEMX-200DR) melanoma cells.
- Cell death was primarily mediated by inhibition of protein synthesis, with minimal caspase activation observed.
- The resistant cell line (FEMX-200DR) exhibited increased resistance to apoptosis induction by Staurosporine.
- Treatment of early passage melanoma cell lines from metastases showed dose- and time-dependent cell death, irrespective of DTIC sensitivity.
Conclusions:
- The 9.2.27 PE IT is effective against malignant melanoma cells regardless of their resistance to Dacarbazine (DTIC) or apoptosis.
- Protein synthesis inhibition is the primary mechanism of cell death induced by this immunotoxin.
- This immunotoxin represents a potential therapeutic strategy for melanoma, overcoming common resistance mechanisms.
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