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Heterogeneous Mechanisms of Secondary Resistance and Clonal Selection in Sarcoma during Treatment with Nutlin.

Laroche A, Tran-Cong K, Chaire V, Lagarde P, Hostein I, Coindre JM, Chibon F, Neuville A, Lesluyes T, Lucchesi C, Italiano A - PLoS ONE (2015)

Bottom Line: Further, secondary resistance to nutlin was associated with deregulation of apoptosis-related genes and marked productive autophagy, the inhibition of which resulted in significant restoration of nutlin-induced cell death.Collectively, our findings argue that secondary resistance to nutlin in STS involved heterogeneous mechanisms resulting from clonal evolution and several biological pathways.Alternative dosing regimens and combination with other targeted agents are needed to achieve successful development of nutlin in the clinical setting.

View Article: PubMed Central - PubMed

Affiliation: INSERM U916, Institut Bergonié, Bordeaux, France; Sarcoma Unit, Institut Bergonié, Bordeaux, France.

ABSTRACT
Nutlin inhibits TP53-MDM2 interaction and is under investigation in soft-tissue sarcomas (STS) and other malignancies. Molecular mechanisms of secondary resistance to nutlin in STS are unknown. We performed whole-transcriptome sequencing (RNA-seq) on three pretreatment and secondary resistant STS cell lines selected based on their high primary sensitivity to nutlin. Our data identified a subset of cancer gene mutations and ploidy variations that were positively selected following treatment, including TP53 mutations in 2 out of 3 resistant cell lines. Further, secondary resistance to nutlin was associated with deregulation of apoptosis-related genes and marked productive autophagy, the inhibition of which resulted in significant restoration of nutlin-induced cell death. Collectively, our findings argue that secondary resistance to nutlin in STS involved heterogeneous mechanisms resulting from clonal evolution and several biological pathways. Alternative dosing regimens and combination with other targeted agents are needed to achieve successful development of nutlin in the clinical setting.

No MeSH data available.


Related in: MedlinePlus

Antiproliferative activity of RG7388 (A) and activation of the p53 (B, C) in human soft-tissue sarcoma (STS) cell lines.(A) IC 50 (μM) of RG-7388 for 11 STS cells, IB111, IB115, IB128, IB114 and IB126 are P53 wild type, the other cell lines are P53-mutated. The experiments presented are representative of at least 3 experiments. Immunoblots are represented on the left (B) and densitometry of the immunoblots on the right (C). Sensitive cells untreated (NT) or exposed to IC50 of RG-7388 (RG) were immunoblotted for MDM2, TP53 and P21 expression.
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pone.0137794.g001: Antiproliferative activity of RG7388 (A) and activation of the p53 (B, C) in human soft-tissue sarcoma (STS) cell lines.(A) IC 50 (μM) of RG-7388 for 11 STS cells, IB111, IB115, IB128, IB114 and IB126 are P53 wild type, the other cell lines are P53-mutated. The experiments presented are representative of at least 3 experiments. Immunoblots are represented on the left (B) and densitometry of the immunoblots on the right (C). Sensitive cells untreated (NT) or exposed to IC50 of RG-7388 (RG) were immunoblotted for MDM2, TP53 and P21 expression.

Mentions: As predicted by the mechanistic model of TP53 regulation, the nutlin compound RG7388 inhibited significantly the proliferation of 5 out of 11 STS cell lines with no TP53 mutations as assessed by Sanger sequencing (IC50: 2–50 nM), but not of the 6 out of 11 cell lines with TP53 mutations (Fig 1A). The most sensitive cell lines were the DDLPS cell lines IB111 and IB115 characterized by an amplification of the MDM2 gene and the extraskeletal osteosarcoma cell line IB128 characterized with no alteration of the MDM2 gene copy numbers. In agreement with the mechanism of action of nutlins, treatment of wild-type TP53 (Sanger sequencing) STS cell lines with RG7388 showed an accumulation of the TP53 protein and its targets, P21 and MDM2, as revealed by Western blotting (Fig 1B and 1C).


Heterogeneous Mechanisms of Secondary Resistance and Clonal Selection in Sarcoma during Treatment with Nutlin.

Laroche A, Tran-Cong K, Chaire V, Lagarde P, Hostein I, Coindre JM, Chibon F, Neuville A, Lesluyes T, Lucchesi C, Italiano A - PLoS ONE (2015)

Antiproliferative activity of RG7388 (A) and activation of the p53 (B, C) in human soft-tissue sarcoma (STS) cell lines.(A) IC 50 (μM) of RG-7388 for 11 STS cells, IB111, IB115, IB128, IB114 and IB126 are P53 wild type, the other cell lines are P53-mutated. The experiments presented are representative of at least 3 experiments. Immunoblots are represented on the left (B) and densitometry of the immunoblots on the right (C). Sensitive cells untreated (NT) or exposed to IC50 of RG-7388 (RG) were immunoblotted for MDM2, TP53 and P21 expression.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4591276&req=5

pone.0137794.g001: Antiproliferative activity of RG7388 (A) and activation of the p53 (B, C) in human soft-tissue sarcoma (STS) cell lines.(A) IC 50 (μM) of RG-7388 for 11 STS cells, IB111, IB115, IB128, IB114 and IB126 are P53 wild type, the other cell lines are P53-mutated. The experiments presented are representative of at least 3 experiments. Immunoblots are represented on the left (B) and densitometry of the immunoblots on the right (C). Sensitive cells untreated (NT) or exposed to IC50 of RG-7388 (RG) were immunoblotted for MDM2, TP53 and P21 expression.
Mentions: As predicted by the mechanistic model of TP53 regulation, the nutlin compound RG7388 inhibited significantly the proliferation of 5 out of 11 STS cell lines with no TP53 mutations as assessed by Sanger sequencing (IC50: 2–50 nM), but not of the 6 out of 11 cell lines with TP53 mutations (Fig 1A). The most sensitive cell lines were the DDLPS cell lines IB111 and IB115 characterized by an amplification of the MDM2 gene and the extraskeletal osteosarcoma cell line IB128 characterized with no alteration of the MDM2 gene copy numbers. In agreement with the mechanism of action of nutlins, treatment of wild-type TP53 (Sanger sequencing) STS cell lines with RG7388 showed an accumulation of the TP53 protein and its targets, P21 and MDM2, as revealed by Western blotting (Fig 1B and 1C).

Bottom Line: Further, secondary resistance to nutlin was associated with deregulation of apoptosis-related genes and marked productive autophagy, the inhibition of which resulted in significant restoration of nutlin-induced cell death.Collectively, our findings argue that secondary resistance to nutlin in STS involved heterogeneous mechanisms resulting from clonal evolution and several biological pathways.Alternative dosing regimens and combination with other targeted agents are needed to achieve successful development of nutlin in the clinical setting.

View Article: PubMed Central - PubMed

Affiliation: INSERM U916, Institut Bergonié, Bordeaux, France; Sarcoma Unit, Institut Bergonié, Bordeaux, France.

ABSTRACT
Nutlin inhibits TP53-MDM2 interaction and is under investigation in soft-tissue sarcomas (STS) and other malignancies. Molecular mechanisms of secondary resistance to nutlin in STS are unknown. We performed whole-transcriptome sequencing (RNA-seq) on three pretreatment and secondary resistant STS cell lines selected based on their high primary sensitivity to nutlin. Our data identified a subset of cancer gene mutations and ploidy variations that were positively selected following treatment, including TP53 mutations in 2 out of 3 resistant cell lines. Further, secondary resistance to nutlin was associated with deregulation of apoptosis-related genes and marked productive autophagy, the inhibition of which resulted in significant restoration of nutlin-induced cell death. Collectively, our findings argue that secondary resistance to nutlin in STS involved heterogeneous mechanisms resulting from clonal evolution and several biological pathways. Alternative dosing regimens and combination with other targeted agents are needed to achieve successful development of nutlin in the clinical setting.

No MeSH data available.


Related in: MedlinePlus