Limits...
Identification of a promising PI3K inhibitor for the treatment of multiple myeloma through the structural optimization.

Han K, Xu X, Chen G, Zeng Y, Zhu J, Du X, Zhang Z, Cao B, Liu Z, Mao X - J Hematol Oncol (2014)

Bottom Line: BENC-511 was able to inactivate PI3K and its downstream signals AKT, mTOR, p70S6K, and 4E-BP1 at 1 μM but had no effects on their total protein expression.Consistent with its effects on PI3K activity, BENC-511 induced MM cell apoptosis which was evidenced by the cleavage of Caspase-3 and PARP.Suppression of BENC-511 on MM tumor growth was associated with decreased PI3K/AKT activity and increased cell apoptosis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Cyrus Tang Hematology Center, Soochow University, Suzhou, Jiangsu Province 215123, China. xinliangmao@suda.edu.cn.

ABSTRACT

Background: We previously reported a PI3K inhibitor S14161 which displays a promising preclinical activity against multiple myeloma (MM) and leukemia, but the chiral structure and poor solubility prevent its further application.

Methods: Six S14161 analogs were designed based on the structure-activity relationship; activity of the compounds in terms of cell death and inhibition of PI3K were analyzed by flow cytometry and Western blotting, respectively; anti-myeloma activity in vivo was performed on two independent xenograft models.

Results: Among the six analogs, BENC-511 was one of the most potent compounds which significantly inhibited PI3K activity and induced MM cell apoptosis. BENC-511 was able to inactivate PI3K and its downstream signals AKT, mTOR, p70S6K, and 4E-BP1 at 1 μM but had no effects on their total protein expression. Consistent with its effects on PI3K activity, BENC-511 induced MM cell apoptosis which was evidenced by the cleavage of Caspase-3 and PARP. Notably, addition of insulin-like growth factor 1 and interleukin-6, two important triggers for PI3K activation in MM cells, partly blocked BENC-511-induced MM cell death, which further demonstrated that PI3K signaling pathway was critical for the anti-myeloma activity of BENC-511. Moreover, BENC-511 also showed potent oral activity against myeloma in vivo. Oral administration of BENC-511 decreased tumor growth up to 80% within 3 weeks in two independent MM xenograft models at a dose of 50 mg/kg body weight, but presented minimal toxicity. Suppression of BENC-511 on MM tumor growth was associated with decreased PI3K/AKT activity and increased cell apoptosis.

Conclusions: Because of its potent anti-MM activity, low toxicity (LD50 oral >1.5 g/kg), and easy synthesis, BENC-511 could be developed as a promising agent for the treatment of MM via suppressing the PI3K/AKT signaling pathway.

Show MeSH

Related in: MedlinePlus

BENC-511 displays potent inhibitory effects on AKT activation. (A) The structures of the analogs of S14161, including DQJ-610, DJY-611, WQD-612, QDF-510, BENC-511. (B) OPM2 cells were treated with 4 μM of S14161, BENC (BENC-511), QDF (QDF-510), DQJ (DQJ-610), DJY (DJY-611), or WQD (WQD-612) for 24 hours. After incubation, cells were harvested and total proteins were isolated. Expression of p-AKT (S473) and total AKT were measured by immunoblotting. (C) RPMI-8226 (8226), JJN3, OCI-MY5 (MY5), U266, LP1, OPM2 cells were treated with 4 μM of BENC-511 or DMSO for 24 hours followed by the analysis of the expression of p-AKT (S473) and total AKT. (D) RPMI-8226, LP1 and OPM2 cells were treated with increasing concentration of BENC-511 for 24 hours. Expression of p-AKT (S473 and T308), and total AKT were measured by immunoblotting. (E) RPMI-8226 and OPM2 cells were treated with increasing concentration of BENC-511 for 12 hours followed by the analysis of AKT activation. GAPDH was used as a loading control. T-AKT: total AKT.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC3924225&req=5

Figure 1: BENC-511 displays potent inhibitory effects on AKT activation. (A) The structures of the analogs of S14161, including DQJ-610, DJY-611, WQD-612, QDF-510, BENC-511. (B) OPM2 cells were treated with 4 μM of S14161, BENC (BENC-511), QDF (QDF-510), DQJ (DQJ-610), DJY (DJY-611), or WQD (WQD-612) for 24 hours. After incubation, cells were harvested and total proteins were isolated. Expression of p-AKT (S473) and total AKT were measured by immunoblotting. (C) RPMI-8226 (8226), JJN3, OCI-MY5 (MY5), U266, LP1, OPM2 cells were treated with 4 μM of BENC-511 or DMSO for 24 hours followed by the analysis of the expression of p-AKT (S473) and total AKT. (D) RPMI-8226, LP1 and OPM2 cells were treated with increasing concentration of BENC-511 for 24 hours. Expression of p-AKT (S473 and T308), and total AKT were measured by immunoblotting. (E) RPMI-8226 and OPM2 cells were treated with increasing concentration of BENC-511 for 12 hours followed by the analysis of AKT activation. GAPDH was used as a loading control. T-AKT: total AKT.

Mentions: We previously reported S14161 as a novel PI3K inhibitor [11], to improve its physical and chemical properties, we designed two classes of analogs of S14161. Class I members WQD-612, DJY-611, DQJ-610 contained the 2-phenyl ring, but the flouro substituent was replaced with a hydrogen atom or an electron-withdrawing cyano group or electron-donating methoxy substituent at the para position (Figure 1A). Class II members QDF-510 and BENC-511 were designed with a simplified structure by removing the 4-fluorophenyl group at the 2-position of the chromene core (Figure 1A). In the analysis of the effects of these analogs on PI3K activity, we treated the PTEN-negative MM cell line OPM2 with 4 μM of each compound for 24 hours. Western blotting analyses revealed that all class I compounds except DJY-611 showed no significant inhibitory effects on AKT phosphorylation, while the class II compounds effectively suppressed AKT activation (Figure 1B). We next evaluated the inhibitory effects of these compounds on OPM2 cell growth. Over a 72-hour treatment, BENC-511 was found to be the most potent one in inhibiting OPM2 cell proliferation by a measurement of viable cells using MTT assay (Additional file 1: Figure S1). BENC-511 was then applied for evaluation of AKT phosphorylation levels in a panel of MM cell lines, including RPMI-8226, JJN3, OCI-MY5, U266, LP1, and OPM2. As shown in Figure 1C, BENC-511 inhibited AKT activation in all cell lines examined.


Identification of a promising PI3K inhibitor for the treatment of multiple myeloma through the structural optimization.

Han K, Xu X, Chen G, Zeng Y, Zhu J, Du X, Zhang Z, Cao B, Liu Z, Mao X - J Hematol Oncol (2014)

BENC-511 displays potent inhibitory effects on AKT activation. (A) The structures of the analogs of S14161, including DQJ-610, DJY-611, WQD-612, QDF-510, BENC-511. (B) OPM2 cells were treated with 4 μM of S14161, BENC (BENC-511), QDF (QDF-510), DQJ (DQJ-610), DJY (DJY-611), or WQD (WQD-612) for 24 hours. After incubation, cells were harvested and total proteins were isolated. Expression of p-AKT (S473) and total AKT were measured by immunoblotting. (C) RPMI-8226 (8226), JJN3, OCI-MY5 (MY5), U266, LP1, OPM2 cells were treated with 4 μM of BENC-511 or DMSO for 24 hours followed by the analysis of the expression of p-AKT (S473) and total AKT. (D) RPMI-8226, LP1 and OPM2 cells were treated with increasing concentration of BENC-511 for 24 hours. Expression of p-AKT (S473 and T308), and total AKT were measured by immunoblotting. (E) RPMI-8226 and OPM2 cells were treated with increasing concentration of BENC-511 for 12 hours followed by the analysis of AKT activation. GAPDH was used as a loading control. T-AKT: total AKT.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3924225&req=5

Figure 1: BENC-511 displays potent inhibitory effects on AKT activation. (A) The structures of the analogs of S14161, including DQJ-610, DJY-611, WQD-612, QDF-510, BENC-511. (B) OPM2 cells were treated with 4 μM of S14161, BENC (BENC-511), QDF (QDF-510), DQJ (DQJ-610), DJY (DJY-611), or WQD (WQD-612) for 24 hours. After incubation, cells were harvested and total proteins were isolated. Expression of p-AKT (S473) and total AKT were measured by immunoblotting. (C) RPMI-8226 (8226), JJN3, OCI-MY5 (MY5), U266, LP1, OPM2 cells were treated with 4 μM of BENC-511 or DMSO for 24 hours followed by the analysis of the expression of p-AKT (S473) and total AKT. (D) RPMI-8226, LP1 and OPM2 cells were treated with increasing concentration of BENC-511 for 24 hours. Expression of p-AKT (S473 and T308), and total AKT were measured by immunoblotting. (E) RPMI-8226 and OPM2 cells were treated with increasing concentration of BENC-511 for 12 hours followed by the analysis of AKT activation. GAPDH was used as a loading control. T-AKT: total AKT.
Mentions: We previously reported S14161 as a novel PI3K inhibitor [11], to improve its physical and chemical properties, we designed two classes of analogs of S14161. Class I members WQD-612, DJY-611, DQJ-610 contained the 2-phenyl ring, but the flouro substituent was replaced with a hydrogen atom or an electron-withdrawing cyano group or electron-donating methoxy substituent at the para position (Figure 1A). Class II members QDF-510 and BENC-511 were designed with a simplified structure by removing the 4-fluorophenyl group at the 2-position of the chromene core (Figure 1A). In the analysis of the effects of these analogs on PI3K activity, we treated the PTEN-negative MM cell line OPM2 with 4 μM of each compound for 24 hours. Western blotting analyses revealed that all class I compounds except DJY-611 showed no significant inhibitory effects on AKT phosphorylation, while the class II compounds effectively suppressed AKT activation (Figure 1B). We next evaluated the inhibitory effects of these compounds on OPM2 cell growth. Over a 72-hour treatment, BENC-511 was found to be the most potent one in inhibiting OPM2 cell proliferation by a measurement of viable cells using MTT assay (Additional file 1: Figure S1). BENC-511 was then applied for evaluation of AKT phosphorylation levels in a panel of MM cell lines, including RPMI-8226, JJN3, OCI-MY5, U266, LP1, and OPM2. As shown in Figure 1C, BENC-511 inhibited AKT activation in all cell lines examined.

Bottom Line: BENC-511 was able to inactivate PI3K and its downstream signals AKT, mTOR, p70S6K, and 4E-BP1 at 1 μM but had no effects on their total protein expression.Consistent with its effects on PI3K activity, BENC-511 induced MM cell apoptosis which was evidenced by the cleavage of Caspase-3 and PARP.Suppression of BENC-511 on MM tumor growth was associated with decreased PI3K/AKT activity and increased cell apoptosis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Cyrus Tang Hematology Center, Soochow University, Suzhou, Jiangsu Province 215123, China. xinliangmao@suda.edu.cn.

ABSTRACT

Background: We previously reported a PI3K inhibitor S14161 which displays a promising preclinical activity against multiple myeloma (MM) and leukemia, but the chiral structure and poor solubility prevent its further application.

Methods: Six S14161 analogs were designed based on the structure-activity relationship; activity of the compounds in terms of cell death and inhibition of PI3K were analyzed by flow cytometry and Western blotting, respectively; anti-myeloma activity in vivo was performed on two independent xenograft models.

Results: Among the six analogs, BENC-511 was one of the most potent compounds which significantly inhibited PI3K activity and induced MM cell apoptosis. BENC-511 was able to inactivate PI3K and its downstream signals AKT, mTOR, p70S6K, and 4E-BP1 at 1 μM but had no effects on their total protein expression. Consistent with its effects on PI3K activity, BENC-511 induced MM cell apoptosis which was evidenced by the cleavage of Caspase-3 and PARP. Notably, addition of insulin-like growth factor 1 and interleukin-6, two important triggers for PI3K activation in MM cells, partly blocked BENC-511-induced MM cell death, which further demonstrated that PI3K signaling pathway was critical for the anti-myeloma activity of BENC-511. Moreover, BENC-511 also showed potent oral activity against myeloma in vivo. Oral administration of BENC-511 decreased tumor growth up to 80% within 3 weeks in two independent MM xenograft models at a dose of 50 mg/kg body weight, but presented minimal toxicity. Suppression of BENC-511 on MM tumor growth was associated with decreased PI3K/AKT activity and increased cell apoptosis.

Conclusions: Because of its potent anti-MM activity, low toxicity (LD50 oral >1.5 g/kg), and easy synthesis, BENC-511 could be developed as a promising agent for the treatment of MM via suppressing the PI3K/AKT signaling pathway.

Show MeSH
Related in: MedlinePlus