Cells resistant to HTI-286 do not overexpress P-glycoprotein but have reduced drug accumulation and a point mutation in alpha-tubulin.
HTI-286, a synthetic analogue of hemiasterlin, depolymerizes microtubules and is proposed to bind at the Vinca peptide site in tubulin. It has excellent in vivo antitumor activity in human xenograft models, including tumors that express P-glycoprotein, and is in phase II clinical evaluation. To identify potential mechanisms of resistance induced by HTI-286, KB-3-1 epidermoid carcinoma cells were exposed to increasing drug concentrations. When maintained in 4.0 nmol/L HTI-286, cells had 12-fold resistance to HTI-286. Cross-resistance was observed to other Vinca peptide-binding agents, including hemiasterlin A, dolastatin-10, and vinblastine (7- to 28-fold), and DNA-damaging drugs, including Adriamycin and mitoxantrone (16- to 57-fold), but minimal resistance was seen to taxanes, epothilones, or colchicine (1- to 4-fold). Resistance to HTI-286 was retained when KB-HTI-resistant cells were grown in athymic mice. Accumulation of [(3)H]HTI-286 was lower in cells selected in intermediate (2.5 nmol/L) and high (4.0 nmol/L) concentrations of HTI-286 compared with parental cells, whereas accumulation of [(14)C]paclitaxel was unchanged. Sodium azide treatment partially reversed low HTI-286 accumulation, suggesting involvement of an ATP-dependent drug pump. KB-HTI-resistant cells did not overexpress P-glycoprotein, breast cancer resistance protein (BCRP/ABCG2/MXR), MRP1, or MRP3. No mutations were found in the major beta-tubulin isoform. However, 4.0 nmol/L HTI-286-selected cells had a point mutation in alpha-tubulin that substitutes Ser for Ala(12) near the nonexchangeable GTP-binding site of alpha-tubulin. KB-HTI-resistant cells removed from drug became less resistant to HTI-286, no longer had low HTI-286 accumulation, and retained the Ala(12) mutation. These data suggest that HTI-286 resistance may be partially mediated by mutation of alpha-tubulin and by an ATP-binding cassette drug pump distinct from P-glycoprotein, ABCG2, MRP1, or MRP3.
Oncology Research, Wyeth Research, 401 North Middletown Road, 200/4709, Pearl River, NY 10965, USA. loganzf@wyeth.com
Frank
F
Loganzo
Malathi
M
Hari
Tami
T
Annable
Xingzhi
X
Tan
Daniel B
DB
Morilla
Sylvia
S
Musto
Arie
A
Zask
Joshua
J
Kaplan
Albert A
AA
Minnick
Michael K
MK
May
Semiramis
S
Ayral-Kaloustian
Marianne S
MS
Poruchynsky
Tito
T
Fojo
Lee M
LM
Greenberger
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters / Q000096:biosynthesis; D000255:Adenosine Triphosphate / Q000737:chemistry; D000409:Alanine / Q000737:chemistry; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D003062:Codon; D004249:DNA Damage; D018076:DNA, Complementary / Q000378:metabolism; D047630:Depsipeptides; D019281:Dimerization; D004317:Doxorubicin / Q000494:pharmacology; D019008:Drug Resistance, Neoplasm; D006801:Humans; D051379:Mice; D008819:Mice, Nude; D008942:Mitoxantrone / Q000494:pharmacology; D008958:Models, Molecular; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000096:biosynthesis; D009154:Mutation; D009363:Neoplasm Proteins / Q000096:biosynthesis; D009368:Neoplasm Transplantation; D009842:Oligopeptides / Q000494:pharmacology*; D017354:Point Mutation; D011487:Protein Conformation; D017422:Sequence Analysis, DNA; D019810:Sodium Azide / Q000494:pharmacology; D013997:Time Factors; D014404:Tubulin / Q000737:chemistry / Q000378:metabolism*; D014747:Vinblastine / Q000494:pharmacology
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; D000255:Adenosine Triphosphate; D000409:Alanine; D000970:Antineoplastic Agents; D003062:Codon; D018076:DNA, Complementary; D047630:Depsipeptides; D004317:Doxorubicin; D008942:Mitoxantrone; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D009363:Neoplasm Proteins; D009842:Oligopeptides; D019810:Sodium Azide; D014404:Tubulin; D014747:Vinblastine; C116614:ABCG2 protein, human; C473896:HTI-286; C103465:hemiasterlin A; C114462:multidrug resistance-associated protein 3; C064570:dolastatin 10; C421109:multidrug resistance-associated protein 1