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Multi-kinase inhibitors can associate with heat shock proteins through their NH2-termini by which they suppress chaperone function

Journal Article


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Abstract


  • We performed proteomic studies using the GRP78 chaperone-inhibitor drug AR-12 (OSU-03012) as bait. Multiple additional chaperone and chaperone-associated proteins were shown to interact with AR-12, including: GRP75, HSP75, BAG2; HSP27; ULK-1; and thioredoxin. AR-12 down-regulated in situ immuno-fluorescence detection of ATP binding chaperones using antibodies directed against the NH2-termini of the proteins but only weakly reduced detection using antibodies directed against the central and COOH portions of the proteins. Traditional SDS-PAGE and western blotting assessment methods did not exhibit any alterations in chaperone detection. AR-12 altered the sub-cellular distribution of chaperone proteins, abolishing their punctate speckled patterning concomitant with changes in protein co-localization. AR-12 inhibited chaperone ATPase activity, which was enhanced by sildenafil; inhibited chaperone – chaperone and chaperone – client interactions; and docked in silico with the ATPase domains of HSP90 and of HSP70. AR-12 combined with sildenafil in a GRP78 plus HSP27 –dependent fashion to profoundly activate an eIF2α/ATF4/CHOP/Beclin1 pathway in parallel with inactivating mTOR and increasing ATG13 phosphorylation, collectively resulting in formation of punctate toxic autophagosomes. Over-expression of [GRP78 and HSP27] prevented: AR-12 –induced activation of ER stress signaling and maintained mTOR activity; AR-12 –mediated down-regulation of thioredoxin, MCL-1 and c-FLIP-s; and preserved tumor cell viability. Thus the inhibition of chaperone protein functions by AR-12 and by multi-kinase inhibitors very likely explains why these agents have anti-tumor effects in multiple genetically diverse tumor cell types.

Authors


  •   Booth, Laurence (external author)
  •   Shuch, Brian (external author)
  •   Albers, Thomas (external author)
  •   Roberts, Jane L. (external author)
  •   Tavallai, Mehrad (external author)
  •   Proniuk, Stefan (external author)
  •   Zukiwski, Alexander (external author)
  •   Wang, Dasheng (external author)
  •   Chen, Ching-Shih (external author)
  •   Bottaro, Don (external author)
  •   Ecroyd, Heath
  •   Lebedyeva, Iryna O. (external author)
  •   Dent, Paul (external author)

Publication Date


  • 2016

Citation


  • Booth, L., Shuch, B., Albers, T., Roberts, J. L., Tavallai, M., Proniuk, S., Zukiwski, A., Wang, D., Chen, C., Bottaro, D., Ecroyd, H., Lebedyeva, I. O. & Dent, P. (2016). Multi-kinase inhibitors can associate with heat shock proteins through their NH2-termini by which they suppress chaperone function. Oncotarget, 7 (11), 12975-12996.

Scopus Eid


  • 2-s2.0-84962840558

Ro Full-text Url


  • http://ro.uow.edu.au/cgi/viewcontent.cgi?article=4741&context=smhpapers

Ro Metadata Url


  • http://ro.uow.edu.au/smhpapers/3718

Has Global Citation Frequency


Number Of Pages


  • 21

Start Page


  • 12975

End Page


  • 12996

Volume


  • 7

Issue


  • 11

Place Of Publication


  • United States

Abstract


  • We performed proteomic studies using the GRP78 chaperone-inhibitor drug AR-12 (OSU-03012) as bait. Multiple additional chaperone and chaperone-associated proteins were shown to interact with AR-12, including: GRP75, HSP75, BAG2; HSP27; ULK-1; and thioredoxin. AR-12 down-regulated in situ immuno-fluorescence detection of ATP binding chaperones using antibodies directed against the NH2-termini of the proteins but only weakly reduced detection using antibodies directed against the central and COOH portions of the proteins. Traditional SDS-PAGE and western blotting assessment methods did not exhibit any alterations in chaperone detection. AR-12 altered the sub-cellular distribution of chaperone proteins, abolishing their punctate speckled patterning concomitant with changes in protein co-localization. AR-12 inhibited chaperone ATPase activity, which was enhanced by sildenafil; inhibited chaperone – chaperone and chaperone – client interactions; and docked in silico with the ATPase domains of HSP90 and of HSP70. AR-12 combined with sildenafil in a GRP78 plus HSP27 –dependent fashion to profoundly activate an eIF2α/ATF4/CHOP/Beclin1 pathway in parallel with inactivating mTOR and increasing ATG13 phosphorylation, collectively resulting in formation of punctate toxic autophagosomes. Over-expression of [GRP78 and HSP27] prevented: AR-12 –induced activation of ER stress signaling and maintained mTOR activity; AR-12 –mediated down-regulation of thioredoxin, MCL-1 and c-FLIP-s; and preserved tumor cell viability. Thus the inhibition of chaperone protein functions by AR-12 and by multi-kinase inhibitors very likely explains why these agents have anti-tumor effects in multiple genetically diverse tumor cell types.

Authors


  •   Booth, Laurence (external author)
  •   Shuch, Brian (external author)
  •   Albers, Thomas (external author)
  •   Roberts, Jane L. (external author)
  •   Tavallai, Mehrad (external author)
  •   Proniuk, Stefan (external author)
  •   Zukiwski, Alexander (external author)
  •   Wang, Dasheng (external author)
  •   Chen, Ching-Shih (external author)
  •   Bottaro, Don (external author)
  •   Ecroyd, Heath
  •   Lebedyeva, Iryna O. (external author)
  •   Dent, Paul (external author)

Publication Date


  • 2016

Citation


  • Booth, L., Shuch, B., Albers, T., Roberts, J. L., Tavallai, M., Proniuk, S., Zukiwski, A., Wang, D., Chen, C., Bottaro, D., Ecroyd, H., Lebedyeva, I. O. & Dent, P. (2016). Multi-kinase inhibitors can associate with heat shock proteins through their NH2-termini by which they suppress chaperone function. Oncotarget, 7 (11), 12975-12996.

Scopus Eid


  • 2-s2.0-84962840558

Ro Full-text Url


  • http://ro.uow.edu.au/cgi/viewcontent.cgi?article=4741&context=smhpapers

Ro Metadata Url


  • http://ro.uow.edu.au/smhpapers/3718

Has Global Citation Frequency


Number Of Pages


  • 21

Start Page


  • 12975

End Page


  • 12996

Volume


  • 7

Issue


  • 11

Place Of Publication


  • United States