Deregulated FASN Expression in BRAF Inhibitor-Resistant Melanoma Cells Unveils New Targets for Drug Combinations

Metabolic changes promoting cell survival are involved in metastatic melanoma progression and in the development of drug resistance. In BRAF-inhibitor resistant melanoma cells, we explored the role of FASN, an enzyme involved in lipogenesis overexpressed in metastatic melanoma. Resistant melanoma cells displaying enhanced migratory and pro-invasive abilities increased sensitivity to the BRAF inhibitor PLX4032 upon the molecular targeting of FASN and upon treatment with the FASN inhibitor orlistat. This behavior was associated with a marked apoptosis and caspase 3/7 activation observed for the drug combination. The expression of FASN was found to be inversely associated with drug resistance in BRAF-mutant cell lines, both in a set of six resistant/sensitive matched lines and in the Cancer Cell Line Encyclopedia. A favorable drug interaction in resistant cells was also observed with U18666 A inhibiting DHCR24, which increased upon FASN targeting. The simultaneous combination of the two inhibitors showed a synergistic interaction with PLX4032 in resistant cells. In conclusion, FASN plays a role in BRAF-mutated melanoma progression, thereby creating novel therapeutic opportunities for the treatment of melanoma.

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Targeting CDC7 sensitizes resistance melanoma cells to BRAFV600E-specific inhibitor by blocking the CDC7/MCM2-7 pathway

Scientific Reports ◽

10.1038/s41598-019-50732-w ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 3

Author(s):

Shaimaa A. Gad ◽

Hamdy E. A. Ali ◽

Rofaida Gaballa ◽

Rania M. Abdelsalam ◽

Mourad Zerfaoui ◽

...

Keyword(s):

Drug Resistance ◽

Metastatic Melanoma ◽

Clinical Effect ◽

Specific Inhibitor ◽

Melanoma Cells ◽

Development Of Resistance ◽

Dual Targeting ◽

Low Concentrations ◽

Resistant Cells

Abstract Although the utilization of selective BRAFV600E inhibitors is associated with improved overall survival in patients with metastatic melanoma, a growing challenge of drug resistance has emerged. CDC7 has been shown to be overexpressed and associated with poor prognosis in various cancers including melanoma. Thus, we aimed to elucidate the biological role of CDC7 in promoting Vemurafenib resistance and the anticipated benefits of dual targeting of BRAFV600E and CDC7 in melanoma cells. We performed exosomes-associated microRNA profiling and functional assays to determine the role of CDC7 in drug resistance using Vemurafenib-sensitive and resistant melanoma cells. Our results demonstrated that Vemurafenib-resistant cells exhibited a persistent expression of CDC7 in addition to prolonged activity of MCM2 compared to drug-sensitive cells. Reconstitution of miR-3613-3p in resistant cells downregulated CDC7 expression and reduced the number of colonies. Treatment of cells with low concentrations of CDC7 inhibitor TAK-931 sensitized resistant cells to Vemurafenib and reduced the number of cell colonies. Taken together, CDC7 overexpression and downregulation of miR-3613-3p were associated with Vemurafenib resistance in BRAFV600E- bearing melanoma cells. Dual targeting of CDC7 and BRAFV600E reduced the development of resistance against Vemurafenib. Further studies are warranted to investigate the clinical effect of targeting CDC7 in metastatic melanoma.

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BRAF inhibitor resistance confers increased sensitivity to mitotic inhibitors

10.1101/2021.04.15.439990 ◽

2021 ◽

Author(s):

Sean Misek ◽

Thomas Dexheimer ◽

Maisah Akram ◽

Susan E Conrad ◽

Jens Christopher Schmidt ◽

...

Keyword(s):

Cell Line ◽

Single Agent ◽

Braf Inhibitor ◽

Cyclin B1 ◽

Melanoma Cells ◽

Resistant Cell Line ◽

Braf Mutations ◽

Increased Sensitivity ◽

Mitotic Inhibitors ◽

Resistant Cells

Single agent and combination therapy with BRAFV600E/K and MEK inhibitors have remarkable efficacy against melanoma tumors with activating BRAF mutations, but in most cases resistance eventually develops. The purpose of this study is to uncover pharmacological vulnerabilities of BRAFi-resistant melanoma cells, with the goal of identifying new therapeutic options for patients whose tumors have developed resistance to BRAFi/MEKi therapy. We screened a well-annotated compound library against a panel of isogenic pairs of parental and BRAFi-resistant melanoma cell lines to identify classes of compounds that selectively target BRAFi-resistant cells over their BRAFi-sensitive counterparts. Two distinct patterns of increased sensitivity to classes of pharmacological inhibitors emerged. In two cell line pairs, BRAFi resistance conferred increased sensitivity to compounds that share the property of cell cycle arrest at M-phase, including inhibitors of aurora kinase (AURK), polo-like kinase (PLK), tubulin, and kinesin. Live cell microscopy used to track mitosis in real time revealed that parental, but not BRAFi-resistant, melanoma cells were able to exit from compound-induced mitotic arrest through mitotic slippage, thus escaping death. Consistent with the key role of Cyclin B1 levels in regulating mitosis at the spindle checkpoint, in arrested cells we found higher Cyclin B1 levels in parental over BRAFi-resistant melanoma cells, suggesting that altered Cyclin B1 expression levels may explain why these BRAFi resistant cells have gained increased vulnerability to mitotic inhibitors. Another BRAFi-resistant cell line showed increased sensitivity to Chk1/2 inhibitors, possibly due to an accumulation of DNA damage, resulting in mitotic failure. This study shows that BRAFi-resistance in melanoma cells confers vulnerability to pharmacological disruption of mitosis and suggests a targeted synthetic lethal approach to treat BRAF-mutant melanomas that have become resistant to BRAF/MEK-directed therapies.

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The role of FoxP3+ regulatory T cells and IDO+ immune and tumor cells in malignant melanoma – an immunohistochemical study

BMC Cancer ◽

10.1186/s12885-021-08385-4 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Satu Salmi ◽

Anton Lin ◽

Benjamin Hirschovits-Gerz ◽

Mari Valkonen ◽

Niina Aaltonen ◽

...

Keyword(s):

T Cells ◽

Prognostic Factors ◽

Regulatory T Cells ◽

Tumor Cells ◽

Immune Cells ◽

Positive Association ◽

Tumor Stage ◽

Melanoma Cells ◽

Melanoma Progression

Abstract Background FoxP3+ Regulatory T cells (Tregs) and indoleamine-2,3-dioxygenase (IDO) participate in the formation of an immunosuppressive tumor microenvironment (TME) in malignant cutaneous melanoma (CM). Recent studies have reported that IDO expression correlates with poor prognosis and greater Breslow’s depth, but results concerning the role of FoxP3+ Tregs in CM have been controversial. Furthermore, the correlation between IDO and Tregs has not been substantially studied in CM, although IDO is known to be an important regulator of Tregs activity. Methods We investigated the associations of FoxP3+ Tregs, IDO+ tumor cells and IDO+ stromal immune cells with tumor stage, prognostic factors and survival in CM. FoxP3 and IDO were immunohistochemically stained from 29 benign and 29 dysplastic nevi, 18 in situ -melanomas, 48 superficial and 62 deep melanomas and 67 lymph node metastases (LNMs) of CM. The number of FoxP3+ Tregs and IDO+ stromal immune cells, and the coverage and intensity of IDO+ tumor cells were analysed. Results The number of FoxP3+ Tregs and IDO+ stromal immune cells were significantly higher in malignant melanomas compared with benign lesions. The increased expression of IDO in melanoma cells was associated with poor prognostic factors, such as recurrence, nodular growth pattern and increased mitotic count. Furthermore, the expression of IDO in melanoma cells was associated with reduced recurrence˗free survival. We further showed that there was a positive correlation between IDO+ tumor cells and FoxP3+ Tregs. Conclusions These results indicate that IDO is strongly involved in melanoma progression. FoxP3+ Tregs also seems to contribute to the immunosuppressive TME in CM, but their significance in melanoma progression remains unclear. The positive association of FoxP3+ Tregs with IDO+ melanoma cells, but not with IDO+ stromal immune cells, indicates a complex interaction between IDO and Tregs in CM, which demands further studies.

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Abstract 1070: miR-579-3p is a novel master regulator of melanoma progression and drug resistance in metastatic melanoma

10.1158/1538-7445.am2016-1070 ◽

2016 ◽

Cited By ~ 2

Author(s):

Luigi Fattore ◽

Mario Acunzo ◽

Giulia Romano ◽

Alessandro Laganà ◽

Debora Malpicci ◽

...

Keyword(s):

Drug Resistance ◽

Metastatic Melanoma ◽

Master Regulator ◽

Melanoma Progression

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Abstract C03: Role of human endogenous retrovirus-K in phenotype-switching of metastatic melanoma cells during microenvironment alterations

10.1158/1538-7445.tme16-c03 ◽

2016 ◽

Author(s):

Ayele Argaw-Denboba ◽

Emanuela Balestrieri ◽

Annalucia Serafino ◽

Ilaria Bucci ◽

Chiara Cipriani ◽

...

Keyword(s):

Metastatic Melanoma ◽

Melanoma Cells ◽

Endogenous Retrovirus ◽

Human Endogenous Retrovirus ◽

Phenotype Switching

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Nongenetic Mechanisms of Drug Resistance in Melanoma

Annual Review of Cancer Biology ◽

10.1146/annurev-cancerbio-030419-033533 ◽

2020 ◽

Vol 4 (1) ◽

pp. 315-330

Author(s):

Vito W. Rebecca ◽

Meenhard Herlyn

Keyword(s):

Drug Resistance ◽

Metastatic Melanoma ◽

Metastatic Disease ◽

Resistance Mechanisms ◽

Melanoma Cells ◽

Therapeutic Strategies ◽

Therapeutic Resistance ◽

Primary Resistance ◽

Future Design ◽

The Future

Resistance to targeted and immune-based therapies limits cures in patients with metastatic melanoma. A growing number of reports have identified nongenetic primary resistance mechanisms including intrinsic microenvironment- and lineage plasticity–mediated processes serving critical functions in the persistence of disease throughout therapy. There is a temporally shifting spectrum of cellular identities fluidly occupied by therapy-persisting melanoma cells responsible for driving therapeutic resistance and metastasis. The key epigenetic, metabolic, and phenotypic reprogramming events requisite for the manifestation and maintenance of so-called persister melanoma populations remain poorly understood and underscore the need to comprehensively investigate actionable vulnerabilities. Here we attempt to integrate the field's observations on nongenetic mechanisms of drug resistance in melanoma. We postulate that the future design of therapeutic strategies specifically addressing therapy-persisting subpopulations of melanoma will improve the curative potential of therapy for patients with metastatic disease.

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CD133 Is Associated with Increased Melanoma Cell Survival after Multikinase Inhibition

Journal of Oncology ◽

10.1155/2019/6486173 ◽

2019 ◽

Vol 2019 ◽

pp. 1-19 ◽

Cited By ~ 3

Author(s):

Cynthia M. Simbulan-Rosenthal ◽

Anirudh Gaur ◽

Hengbo Zhou ◽

Maryam AbdusSamad ◽

Qing Qin ◽

...

Keyword(s):

Drug Resistance ◽

Kinase Inhibitors ◽

Braf Inhibitor ◽

Mek Inhibitor ◽

Melanoma Cells ◽

Stem Cell Marker ◽

High Dose ◽

Recurrence Rates ◽

Mapk Inhibitors

FDA-approved kinase inhibitors are now used for melanoma, including combinations of the MEK inhibitor trametinib, and BRAF inhibitor dabrafenib for BRAFV600 mutations. NRAS-mutated cell lines are also sensitive to MEK inhibitionin vitro, and NRAS-mutated tumors have also shown partial response to MEK inhibitors. However, melanoma still has high recurrence rates due to subpopulations, sometimes described as “melanoma initiating cells,” resistant to treatment. Since CD133 is a putative cancer stem cell marker for different cancers, associated with decreased survival, we examined resistance of patient-derived CD133(+) and CD133(-) melanoma cells to MAPK inhibitors. Human melanoma cells were exposed to increasing concentrations of trametinib and/or dabrafenib, either before or after separation into CD133(+) and CD133(-) subpopulations. In parental CD133-mixed lines, the percentages of CD133(+) cells increased significantly (p<0.05) after high-dose drug treatment. Presorted CD133(+) cells also exhibited significantly greater (p<0.05) IC50s for single and combination MAPKI treatment. siRNA knockdown revealed a causal relationship between CD133 and drug resistance. Microarray and qRT-PCR analyses revealed that ten of 18 ABC transporter genes were significantly (P<0.05) upregulated in the CD133(+) subpopulation, while inhibition of ABC activity increased sensitivity, suggesting a mechanism for increased drug resistance of CD133(+) cells.

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Exosomal miR-331 from Chemoresistant Osteosarcoma Cells induced Chemoresistance through Autophagy

10.21203/rs.3.rs-587973/v2 ◽

2021 ◽

Author(s):

Aiqing Zhao ◽

Yanbin Zhao ◽

Wanlin Liu ◽

Wei Feng ◽

Wenhua Xing ◽

...

Keyword(s):

Drug Resistance ◽

Resistant Cell ◽

Drug Resistant ◽

Rt Pcr ◽

Chemotherapeutic Resistance ◽

Acquire Drug Resistance ◽

And Migration ◽

Drug Resistant Cell ◽

Resistant Cells

Abstract BackgroundOsteosarcoma (OS) is a highly malignant tumor. Improving chemotherapeutic resistance is very important to improve the survival rate of OS. Exosomes and microRNAs (MiRNA) play important roles in the mechanism of chemotherapeutic resistance transmission. More and more researches focus the mechanism of miRNAs carried by exosomes in the transmission of chemotherapeutic resistance of OS. This study focused on exploring the mechanism of exosomal miR-331 in the transmission of chemoresistance in OS.MethodsWe cultured OS drug-resistant cells and extracted exosomes of these cells. The secretion and uptake of exosomes in OS drug-resistant cells and OS cells (OSCs) were confirmed by fluorescence tracking assay and transwell experiments. The differential expression of microRNA-331 (miR-331) in exosomes of OS resistant and OS cells was investigated by RT-PCR. The effects of drug-resistant exosomes on proliferation and migration of OS cells were determined by MTT assay and scratches assay. MDC staining, RT-PCR, and Western blot were used to detect the role of autophagy which regulated by drug-resistant cell-derived exosom-miR-331.ResultsWe found that the expression difference of miR-331 between MG63/CDDP and MG63 was the most significant. Drug resistant OSCs secreted exosomes and were ingested by OSCs, which then promoted OSCs to acquire drug resistance. In addition, exosomes secreted by drug-resistant OSCs promote drug resistance by carrying miRNAs. Interestingly, inhibition of miRNA resulted in reduced drug resistance transmission of exosomes. Finally, we found that the exosomes secreted by drug-resistant OSCs could induce autophagy of OSCs by carrying miR-331, thus making OSCs acquire drug resistance. Inhibition of miR-331 can effectively improve drug resistance of OSCs.ConclusionsChemoresistant OSCs-derived exosomes promote the transmission of drug resistance by carrying miR-331 and inducing autophagy. Inhibition of miR-331 could effectively alleviate drug resistance of OSCs.

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