Missense mutations in PML-RARA are critical for the lack of responsiveness to arsenic trioxide treatment

Emi Goto, Akihiro Tomita, Fumihiko Hayakawa, Akihide Atsumi, Hitoshi Kiyoi, Tomoki Naoe

Research output: Contribution to journalArticle

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Abstract

Arsenic trioxide (As2O3) is a highly effective treatment for patients with refractory/relapsed acute promyelocytic leukemia (APL), but resistance to As2O3 has recently been seen. In the present study, we report the findings that 2 of 15 patients with refractory/relapsed APL treated with As2O3 were clinically As2O3 resistant. Leukemia cells from these 2 patients harbored missense mutations in promyelocytic leukemia gene - retinoic acid receptor-α gene (PML-RARA) transcripts, resulting in amino acid substitutions of A216V and L218P in the PML B2 domain. When wild-type or mutated PML-RARA (PR-WT and PR-B/L-mut, respectively) were overexpressed in HeLa cells, immunoblotting showed SUMOylated and/or oligomerized protein bands in PR-WT but not in PR-B/L-mut after As2O3 treatment. Protein-localization analysis indicated that PR-WT in the soluble fraction was transferred to the insoluble fraction after treatment with As2O 3, but PR-B/L-mut was stably detected in fractions both with and without As2O3. Immunofluorescent microscopy analysis showed PR-WT localization as a microgranular pattern in the cytoplasm without As2O3 and as a macrogranular pattern with As 2O3. PR-B/L-mut was diffusely observed in the cytoplasm with and without As2O3. Nearly identical localization patterns were observed in patients' primary cells. Therefore, B2 domain mutations may play an important role in aberrant molecular responses to As 2O3 and may be critical for As2O3 resistance in APL.

Original languageEnglish
Pages (from-to)1600-1609
Number of pages10
JournalBlood
Volume118
Issue number6
DOIs
Publication statusPublished - 11-08-2011

Fingerprint

Missense Mutation
Acute Promyelocytic Leukemia
Therapeutics
Refractory materials
Cytoplasm
Leukemia
Genes
arsenic trioxide
Retinoic Acid Receptors
Amino Acid Substitution
HeLa Cells
Immunoblotting
Microscopy
Microscopic examination
Proteins
Substitution reactions
Amino Acids
Mutation

All Science Journal Classification (ASJC) codes

  • Hematology
  • Biochemistry
  • Cell Biology
  • Immunology

Cite this

Goto, Emi ; Tomita, Akihiro ; Hayakawa, Fumihiko ; Atsumi, Akihide ; Kiyoi, Hitoshi ; Naoe, Tomoki. / Missense mutations in PML-RARA are critical for the lack of responsiveness to arsenic trioxide treatment. In: Blood. 2011 ; Vol. 118, No. 6. pp. 1600-1609.
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Missense mutations in PML-RARA are critical for the lack of responsiveness to arsenic trioxide treatment. / Goto, Emi; Tomita, Akihiro; Hayakawa, Fumihiko; Atsumi, Akihide; Kiyoi, Hitoshi; Naoe, Tomoki.

In: Blood, Vol. 118, No. 6, 11.08.2011, p. 1600-1609.

Research output: Contribution to journalArticle

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AU - Goto, Emi

AU - Tomita, Akihiro

AU - Hayakawa, Fumihiko

AU - Atsumi, Akihide

AU - Kiyoi, Hitoshi

AU - Naoe, Tomoki

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N2 - Arsenic trioxide (As2O3) is a highly effective treatment for patients with refractory/relapsed acute promyelocytic leukemia (APL), but resistance to As2O3 has recently been seen. In the present study, we report the findings that 2 of 15 patients with refractory/relapsed APL treated with As2O3 were clinically As2O3 resistant. Leukemia cells from these 2 patients harbored missense mutations in promyelocytic leukemia gene - retinoic acid receptor-α gene (PML-RARA) transcripts, resulting in amino acid substitutions of A216V and L218P in the PML B2 domain. When wild-type or mutated PML-RARA (PR-WT and PR-B/L-mut, respectively) were overexpressed in HeLa cells, immunoblotting showed SUMOylated and/or oligomerized protein bands in PR-WT but not in PR-B/L-mut after As2O3 treatment. Protein-localization analysis indicated that PR-WT in the soluble fraction was transferred to the insoluble fraction after treatment with As2O 3, but PR-B/L-mut was stably detected in fractions both with and without As2O3. Immunofluorescent microscopy analysis showed PR-WT localization as a microgranular pattern in the cytoplasm without As2O3 and as a macrogranular pattern with As 2O3. PR-B/L-mut was diffusely observed in the cytoplasm with and without As2O3. Nearly identical localization patterns were observed in patients' primary cells. Therefore, B2 domain mutations may play an important role in aberrant molecular responses to As 2O3 and may be critical for As2O3 resistance in APL.

AB - Arsenic trioxide (As2O3) is a highly effective treatment for patients with refractory/relapsed acute promyelocytic leukemia (APL), but resistance to As2O3 has recently been seen. In the present study, we report the findings that 2 of 15 patients with refractory/relapsed APL treated with As2O3 were clinically As2O3 resistant. Leukemia cells from these 2 patients harbored missense mutations in promyelocytic leukemia gene - retinoic acid receptor-α gene (PML-RARA) transcripts, resulting in amino acid substitutions of A216V and L218P in the PML B2 domain. When wild-type or mutated PML-RARA (PR-WT and PR-B/L-mut, respectively) were overexpressed in HeLa cells, immunoblotting showed SUMOylated and/or oligomerized protein bands in PR-WT but not in PR-B/L-mut after As2O3 treatment. Protein-localization analysis indicated that PR-WT in the soluble fraction was transferred to the insoluble fraction after treatment with As2O 3, but PR-B/L-mut was stably detected in fractions both with and without As2O3. Immunofluorescent microscopy analysis showed PR-WT localization as a microgranular pattern in the cytoplasm without As2O3 and as a macrogranular pattern with As 2O3. PR-B/L-mut was diffusely observed in the cytoplasm with and without As2O3. Nearly identical localization patterns were observed in patients' primary cells. Therefore, B2 domain mutations may play an important role in aberrant molecular responses to As 2O3 and may be critical for As2O3 resistance in APL.

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