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Novel agents that downregulate EGFR, HER2, and HER3 in parallel.

Ferreira RB, Law ME, Jahn SC, Davis BJ, Heldermon CD, Reinhard M, Castellano RK, Law BK - Oncotarget (2015)

Bottom Line: These compounds disrupt disulfide bonds and are termed Disulfide Bond Disrupting Agents (DDAs).DDA RBF3 exhibits anticancer efficacy in vivo at 40 mg/kg without evidence of toxicity.DDAs may complement existing EGFR-, HER2-, and HER3-targeted agents that function through alternate mechanisms of action, and combination regimens with these existing drugs may overcome therapeutic resistance.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of Florida, Gainesville, FL 32611, USA.

ABSTRACT
EGFR, HER2, and HER3 contribute to the initiation and progression of human cancers, and are therapeutic targets for monoclonal antibodies and tyrosine kinase inhibitors. An important source of resistance to these agents arises from functional redundancy among EGFR, HER2, and HER3. EGFR family members contain conserved extracellular structures that are stabilized by disulfide bonds. Compounds that disrupt extracellular disulfide bonds could inactivate EGFR, HER2, and HER3 in unison. Here we describe the identification of compounds that kill breast cancer cells that overexpress EGFR or HER2. Cell death parallels downregulation of EGFR, HER2, and HER3. These compounds disrupt disulfide bonds and are termed Disulfide Bond Disrupting Agents (DDAs). DDA RBF3 exhibits anticancer efficacy in vivo at 40 mg/kg without evidence of toxicity. DDAs may complement existing EGFR-, HER2-, and HER3-targeted agents that function through alternate mechanisms of action, and combination regimens with these existing drugs may overcome therapeutic resistance.

No MeSH data available.


Related in: MedlinePlus

EGFR/HER2/HER3-targeted Compounds Induce Cell Death and Suppress Response to EGFA. MDA-MB-468 cells treated as indicated for 24 hours were analyzed by immunoblot for levels of EGFR and EGFR phosphorylation. B. MDA-MB-468 cells treated as indicated for 24 hours were analyzed by immunoblot for PARP cleavage. C. MDA-MB-468 cells were either left untreated, or treated with 20 μM NSC624205 for 24 hours. NSC624205-treated cells were then washed and incubated for the indicated periods in the absence of drug. EGFR electrophoretic mobility was analyzed by immunoblot. D. MDA-MB-468 cells were pretreated with 25 μM NSC624205 or vehicle for 15 hours and then either left untreated or stimulated for 15 minutes with 20 ng/ml EGF, after which cell extracts were analyzed by immunoblot. E. MDA-MB-468 cells were treated as indicated for 24 hours and analyzed by immunoblot.
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Figure 2: EGFR/HER2/HER3-targeted Compounds Induce Cell Death and Suppress Response to EGFA. MDA-MB-468 cells treated as indicated for 24 hours were analyzed by immunoblot for levels of EGFR and EGFR phosphorylation. B. MDA-MB-468 cells treated as indicated for 24 hours were analyzed by immunoblot for PARP cleavage. C. MDA-MB-468 cells were either left untreated, or treated with 20 μM NSC624205 for 24 hours. NSC624205-treated cells were then washed and incubated for the indicated periods in the absence of drug. EGFR electrophoretic mobility was analyzed by immunoblot. D. MDA-MB-468 cells were pretreated with 25 μM NSC624205 or vehicle for 15 hours and then either left untreated or stimulated for 15 minutes with 20 ng/ml EGF, after which cell extracts were analyzed by immunoblot. E. MDA-MB-468 cells were treated as indicated for 24 hours and analyzed by immunoblot.

Mentions: As mentioned above, we initially hypothesized that sulfinate compounds may be useful in destabilizing EGFR-family members; therefore, we examined the effects of NSC624205 on the levels and phosphorylation of EGFR in MDA-MB-468 cells. NSC624205 induced a concentration-dependent increase in EGFR electrophoretic mobility that correlated with a decrease in EGFR phosphorylation detected using a phospho-specific antibody (Figure 2A). NSC624205 also caused a concentration-dependent increase in PARP cleavage, consistent with the induction of apoptosis (Figure 2B). To examine the reversibility of NSC624205 actions, MDA-MB-468 cells were treated for 24 hours with NSC624205 and then the compound was washed out and the cells were allowed to recover for various periods of time. This experiment revealed that at 24 hours post-treatment, EGFR electrophoretic mobility was restored to near control levels, indicating that the effects of this compound are slowly reversible (Figure 2C). To examine whether NSC624205 can suppress cellular responses to EGF, cells were stimulated with EGF in the presence or absence of NSC624205. NSC624205 decreased both overall EGF-induced cellular tyrosine phosphorylation and EGFR tyrosine phosphorylation on Tyr845 (Figure 2D). Comparison of NSC624203 with AG490 or an EGFR/HER2 kinase inhibitor (Calbiochem #324673) showed that NSC624203 was more effective at decreasing Akt phosphorylation, increasing PARP cleavage and reducing EGFR tyrosine phosphorylation and overall levels of cellular tyrosine phosphorylation (Figure 2E). Interestingly, combining NSC624203 with the EGFR/HER2 inhibitor blocked Erk phosphorylation more effectively than either drug alone.


Novel agents that downregulate EGFR, HER2, and HER3 in parallel.

Ferreira RB, Law ME, Jahn SC, Davis BJ, Heldermon CD, Reinhard M, Castellano RK, Law BK - Oncotarget (2015)

EGFR/HER2/HER3-targeted Compounds Induce Cell Death and Suppress Response to EGFA. MDA-MB-468 cells treated as indicated for 24 hours were analyzed by immunoblot for levels of EGFR and EGFR phosphorylation. B. MDA-MB-468 cells treated as indicated for 24 hours were analyzed by immunoblot for PARP cleavage. C. MDA-MB-468 cells were either left untreated, or treated with 20 μM NSC624205 for 24 hours. NSC624205-treated cells were then washed and incubated for the indicated periods in the absence of drug. EGFR electrophoretic mobility was analyzed by immunoblot. D. MDA-MB-468 cells were pretreated with 25 μM NSC624205 or vehicle for 15 hours and then either left untreated or stimulated for 15 minutes with 20 ng/ml EGF, after which cell extracts were analyzed by immunoblot. E. MDA-MB-468 cells were treated as indicated for 24 hours and analyzed by immunoblot.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4496366&req=5

Figure 2: EGFR/HER2/HER3-targeted Compounds Induce Cell Death and Suppress Response to EGFA. MDA-MB-468 cells treated as indicated for 24 hours were analyzed by immunoblot for levels of EGFR and EGFR phosphorylation. B. MDA-MB-468 cells treated as indicated for 24 hours were analyzed by immunoblot for PARP cleavage. C. MDA-MB-468 cells were either left untreated, or treated with 20 μM NSC624205 for 24 hours. NSC624205-treated cells were then washed and incubated for the indicated periods in the absence of drug. EGFR electrophoretic mobility was analyzed by immunoblot. D. MDA-MB-468 cells were pretreated with 25 μM NSC624205 or vehicle for 15 hours and then either left untreated or stimulated for 15 minutes with 20 ng/ml EGF, after which cell extracts were analyzed by immunoblot. E. MDA-MB-468 cells were treated as indicated for 24 hours and analyzed by immunoblot.
Mentions: As mentioned above, we initially hypothesized that sulfinate compounds may be useful in destabilizing EGFR-family members; therefore, we examined the effects of NSC624205 on the levels and phosphorylation of EGFR in MDA-MB-468 cells. NSC624205 induced a concentration-dependent increase in EGFR electrophoretic mobility that correlated with a decrease in EGFR phosphorylation detected using a phospho-specific antibody (Figure 2A). NSC624205 also caused a concentration-dependent increase in PARP cleavage, consistent with the induction of apoptosis (Figure 2B). To examine the reversibility of NSC624205 actions, MDA-MB-468 cells were treated for 24 hours with NSC624205 and then the compound was washed out and the cells were allowed to recover for various periods of time. This experiment revealed that at 24 hours post-treatment, EGFR electrophoretic mobility was restored to near control levels, indicating that the effects of this compound are slowly reversible (Figure 2C). To examine whether NSC624205 can suppress cellular responses to EGF, cells were stimulated with EGF in the presence or absence of NSC624205. NSC624205 decreased both overall EGF-induced cellular tyrosine phosphorylation and EGFR tyrosine phosphorylation on Tyr845 (Figure 2D). Comparison of NSC624203 with AG490 or an EGFR/HER2 kinase inhibitor (Calbiochem #324673) showed that NSC624203 was more effective at decreasing Akt phosphorylation, increasing PARP cleavage and reducing EGFR tyrosine phosphorylation and overall levels of cellular tyrosine phosphorylation (Figure 2E). Interestingly, combining NSC624203 with the EGFR/HER2 inhibitor blocked Erk phosphorylation more effectively than either drug alone.

Bottom Line: These compounds disrupt disulfide bonds and are termed Disulfide Bond Disrupting Agents (DDAs).DDA RBF3 exhibits anticancer efficacy in vivo at 40 mg/kg without evidence of toxicity.DDAs may complement existing EGFR-, HER2-, and HER3-targeted agents that function through alternate mechanisms of action, and combination regimens with these existing drugs may overcome therapeutic resistance.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of Florida, Gainesville, FL 32611, USA.

ABSTRACT
EGFR, HER2, and HER3 contribute to the initiation and progression of human cancers, and are therapeutic targets for monoclonal antibodies and tyrosine kinase inhibitors. An important source of resistance to these agents arises from functional redundancy among EGFR, HER2, and HER3. EGFR family members contain conserved extracellular structures that are stabilized by disulfide bonds. Compounds that disrupt extracellular disulfide bonds could inactivate EGFR, HER2, and HER3 in unison. Here we describe the identification of compounds that kill breast cancer cells that overexpress EGFR or HER2. Cell death parallels downregulation of EGFR, HER2, and HER3. These compounds disrupt disulfide bonds and are termed Disulfide Bond Disrupting Agents (DDAs). DDA RBF3 exhibits anticancer efficacy in vivo at 40 mg/kg without evidence of toxicity. DDAs may complement existing EGFR-, HER2-, and HER3-targeted agents that function through alternate mechanisms of action, and combination regimens with these existing drugs may overcome therapeutic resistance.

No MeSH data available.


Related in: MedlinePlus