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Therapy-induced tumour secretomes promote resistance and tumour progression.

Obenauf AC, Zou Y, Ji AL, Vanharanta S, Shu W, Shi H, Kong X, Bosenberg MC, Wiesner T, Rosen N, Lo RS, Massagué J - Nature (2015)

Bottom Line: Drug resistance invariably limits the clinical efficacy of targeted therapy with kinase inhibitors against cancer.Here we show that targeted therapy with BRAF, ALK or EGFR kinase inhibitors induces a complex network of secreted signals in drug-stressed human and mouse melanoma and human lung adenocarcinoma cells.The tumour-promoting secretome of melanoma cells treated with the kinase inhibitor vemurafenib is driven by downregulation of the transcription factor FRA1.

View Article: PubMed Central - PubMed

Affiliation: Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, New York 10065, USA.

ABSTRACT
Drug resistance invariably limits the clinical efficacy of targeted therapy with kinase inhibitors against cancer. Here we show that targeted therapy with BRAF, ALK or EGFR kinase inhibitors induces a complex network of secreted signals in drug-stressed human and mouse melanoma and human lung adenocarcinoma cells. This therapy-induced secretome stimulates the outgrowth, dissemination and metastasis of drug-resistant cancer cell clones and supports the survival of drug-sensitive cancer cells, contributing to incomplete tumour regression. The tumour-promoting secretome of melanoma cells treated with the kinase inhibitor vemurafenib is driven by downregulation of the transcription factor FRA1. In situ transcriptome analysis of drug-resistant melanoma cells responding to the regressing tumour microenvironment revealed hyperactivation of several signalling pathways, most prominently the AKT pathway. Dual inhibition of RAF and the PI(3)K/AKT/mTOR intracellular signalling pathways blunted the outgrowth of the drug-resistant cell population in BRAF mutant human melanoma, suggesting this combination therapy as a strategy against tumour relapse. Thus, therapeutic inhibition of oncogenic drivers induces vast secretome changes in drug-sensitive cancer cells, paradoxically establishing a tumour microenvironment that supports the expansion of drug-resistant clones, but is susceptible to combination therapy.

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The secretome of RAF and ALK inhibitor treated tumour cells increases proliferation and migration of drug-resistant cells and supports the survival of drug-sensitive cellsa, Schematic (left) and representative BLI images (right) after 7 days of co-culture. Average fold change (FC) of BLI signal from A375R-TGL cells in vemurafenib treated wells relative to vehicle treated control wells is depicted on the right (n = 4 biological replicates). b, c, Conditioned media (CM) was derived from drug-sensitive cells, treated with vehicle, vemurafenib, or crizotinib. Drug resistant cells were grown in this CM and the cell number was determined on day 3. Drug-sensitive and drug-resistant cell lines and drugs used to generate CM as indicated. b, (n = 3 biological replicates). c, (n = 6 biological replicates). d, Schematic diagram of the migration assay (upper panel) and relative migration of A375R cells towards CM from different sources as indicated (lower panel, n = 10 FOV). P values were calculated using a two-tailed Mann-Whitney test (**** p<0.0001). e, Survival assay of drug-sensitive A375 cells cultured in CM and treated with vemurafenib, assessed on day 3(n = 3 biological replicates). f, Apoptosis rate of A375 cells cultured in CM and treated with vemurafenib (3μM) (n = 3 biological replicates). Data are presented as average; error bars represent s.e.m.
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Figure 2: The secretome of RAF and ALK inhibitor treated tumour cells increases proliferation and migration of drug-resistant cells and supports the survival of drug-sensitive cellsa, Schematic (left) and representative BLI images (right) after 7 days of co-culture. Average fold change (FC) of BLI signal from A375R-TGL cells in vemurafenib treated wells relative to vehicle treated control wells is depicted on the right (n = 4 biological replicates). b, c, Conditioned media (CM) was derived from drug-sensitive cells, treated with vehicle, vemurafenib, or crizotinib. Drug resistant cells were grown in this CM and the cell number was determined on day 3. Drug-sensitive and drug-resistant cell lines and drugs used to generate CM as indicated. b, (n = 3 biological replicates). c, (n = 6 biological replicates). d, Schematic diagram of the migration assay (upper panel) and relative migration of A375R cells towards CM from different sources as indicated (lower panel, n = 10 FOV). P values were calculated using a two-tailed Mann-Whitney test (**** p<0.0001). e, Survival assay of drug-sensitive A375 cells cultured in CM and treated with vemurafenib, assessed on day 3(n = 3 biological replicates). f, Apoptosis rate of A375 cells cultured in CM and treated with vemurafenib (3μM) (n = 3 biological replicates). Data are presented as average; error bars represent s.e.m.

Mentions: Tumours consist of a complex microenvironment composed of immune, stromal, and cancer cells21. Soluble mediators from this microenvironment can foster cancer growth and therapy resistance13,14,22–24. Considering that drug-sensitive cancer cells are the main population affected by targeted therapy, we hypothesized that signals derived from sensitive cancer cells in response to kinase inhibitors drive the outgrowth of drug-resistant cells. To test this hypothesis, we established an in vitro co-culture system and monitored the growth of TGL-expressing resistant cells (A375R, H2030) in the absence or presence of sensitive cells treated with kinase inhibitors or vehicle (Fig. 2a). Mimicking our in vivo findings, co-culture with vemurafenib-, crizotinib-, or erlotinib-treated sensitive cells significantly enhanced the growth of resistant cancer cells (Fig. 2a, Extended Data Fig. 2a–c).


Therapy-induced tumour secretomes promote resistance and tumour progression.

Obenauf AC, Zou Y, Ji AL, Vanharanta S, Shu W, Shi H, Kong X, Bosenberg MC, Wiesner T, Rosen N, Lo RS, Massagué J - Nature (2015)

The secretome of RAF and ALK inhibitor treated tumour cells increases proliferation and migration of drug-resistant cells and supports the survival of drug-sensitive cellsa, Schematic (left) and representative BLI images (right) after 7 days of co-culture. Average fold change (FC) of BLI signal from A375R-TGL cells in vemurafenib treated wells relative to vehicle treated control wells is depicted on the right (n = 4 biological replicates). b, c, Conditioned media (CM) was derived from drug-sensitive cells, treated with vehicle, vemurafenib, or crizotinib. Drug resistant cells were grown in this CM and the cell number was determined on day 3. Drug-sensitive and drug-resistant cell lines and drugs used to generate CM as indicated. b, (n = 3 biological replicates). c, (n = 6 biological replicates). d, Schematic diagram of the migration assay (upper panel) and relative migration of A375R cells towards CM from different sources as indicated (lower panel, n = 10 FOV). P values were calculated using a two-tailed Mann-Whitney test (**** p<0.0001). e, Survival assay of drug-sensitive A375 cells cultured in CM and treated with vemurafenib, assessed on day 3(n = 3 biological replicates). f, Apoptosis rate of A375 cells cultured in CM and treated with vemurafenib (3μM) (n = 3 biological replicates). Data are presented as average; error bars represent s.e.m.
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Figure 2: The secretome of RAF and ALK inhibitor treated tumour cells increases proliferation and migration of drug-resistant cells and supports the survival of drug-sensitive cellsa, Schematic (left) and representative BLI images (right) after 7 days of co-culture. Average fold change (FC) of BLI signal from A375R-TGL cells in vemurafenib treated wells relative to vehicle treated control wells is depicted on the right (n = 4 biological replicates). b, c, Conditioned media (CM) was derived from drug-sensitive cells, treated with vehicle, vemurafenib, or crizotinib. Drug resistant cells were grown in this CM and the cell number was determined on day 3. Drug-sensitive and drug-resistant cell lines and drugs used to generate CM as indicated. b, (n = 3 biological replicates). c, (n = 6 biological replicates). d, Schematic diagram of the migration assay (upper panel) and relative migration of A375R cells towards CM from different sources as indicated (lower panel, n = 10 FOV). P values were calculated using a two-tailed Mann-Whitney test (**** p<0.0001). e, Survival assay of drug-sensitive A375 cells cultured in CM and treated with vemurafenib, assessed on day 3(n = 3 biological replicates). f, Apoptosis rate of A375 cells cultured in CM and treated with vemurafenib (3μM) (n = 3 biological replicates). Data are presented as average; error bars represent s.e.m.
Mentions: Tumours consist of a complex microenvironment composed of immune, stromal, and cancer cells21. Soluble mediators from this microenvironment can foster cancer growth and therapy resistance13,14,22–24. Considering that drug-sensitive cancer cells are the main population affected by targeted therapy, we hypothesized that signals derived from sensitive cancer cells in response to kinase inhibitors drive the outgrowth of drug-resistant cells. To test this hypothesis, we established an in vitro co-culture system and monitored the growth of TGL-expressing resistant cells (A375R, H2030) in the absence or presence of sensitive cells treated with kinase inhibitors or vehicle (Fig. 2a). Mimicking our in vivo findings, co-culture with vemurafenib-, crizotinib-, or erlotinib-treated sensitive cells significantly enhanced the growth of resistant cancer cells (Fig. 2a, Extended Data Fig. 2a–c).

Bottom Line: Drug resistance invariably limits the clinical efficacy of targeted therapy with kinase inhibitors against cancer.Here we show that targeted therapy with BRAF, ALK or EGFR kinase inhibitors induces a complex network of secreted signals in drug-stressed human and mouse melanoma and human lung adenocarcinoma cells.The tumour-promoting secretome of melanoma cells treated with the kinase inhibitor vemurafenib is driven by downregulation of the transcription factor FRA1.

View Article: PubMed Central - PubMed

Affiliation: Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, New York 10065, USA.

ABSTRACT
Drug resistance invariably limits the clinical efficacy of targeted therapy with kinase inhibitors against cancer. Here we show that targeted therapy with BRAF, ALK or EGFR kinase inhibitors induces a complex network of secreted signals in drug-stressed human and mouse melanoma and human lung adenocarcinoma cells. This therapy-induced secretome stimulates the outgrowth, dissemination and metastasis of drug-resistant cancer cell clones and supports the survival of drug-sensitive cancer cells, contributing to incomplete tumour regression. The tumour-promoting secretome of melanoma cells treated with the kinase inhibitor vemurafenib is driven by downregulation of the transcription factor FRA1. In situ transcriptome analysis of drug-resistant melanoma cells responding to the regressing tumour microenvironment revealed hyperactivation of several signalling pathways, most prominently the AKT pathway. Dual inhibition of RAF and the PI(3)K/AKT/mTOR intracellular signalling pathways blunted the outgrowth of the drug-resistant cell population in BRAF mutant human melanoma, suggesting this combination therapy as a strategy against tumour relapse. Thus, therapeutic inhibition of oncogenic drivers induces vast secretome changes in drug-sensitive cancer cells, paradoxically establishing a tumour microenvironment that supports the expansion of drug-resistant clones, but is susceptible to combination therapy.

Show MeSH
Related in: MedlinePlus