Pei-Chih Lee

Design, synthesis and antitumor evaluation of phenyl N-mustard-quinazoline conjugates

A series of N-mustard-quinazoline conjugates was synthesized and subjected to antitumor studies. The N-mustard pharmacophore was attached at the C-6 of the 4-anilinoquinazolines via a urea linker. To study the structure-activity relationships of these conjugates, various substituents were introduced to the C-4 anilino moiety. The preliminary antitumor studies revealed that these agents exhibited significant antitumor activity in inhibiting various human tumor cell growths in vitro. Compounds 21b, 21g, and 21h were selected for further antitumor activity evaluation against human breast carcinoma MX-1 and prostate PC-3 xenograft in animal model. These agents showed 54-75% tumor suppression with low toxicity (5-7% body-weight changes). We also demonstrate that the newly synthesized compounds are able to induce DNA cross-linking through alkaline agarose gel shift assay and inhibited cell cycle arrest at G2/M phase.

Potent antitumor bifunctional DNA alkylating agents, synthesis and biological activities of 3a-aza-cyclopenta[a]indenes.

A series of bifunctional DNA interstrand cross-linking agents, bis(hydroxymethyl)- and bis(carbamates)-8H-3a-azacyclopenta[a]indene-1-yl derivatives were synthesized for antitumor evaluation. The preliminary antitumor studies revealed that these agents exhibited potent cytotoxicity in vitro and antitumor therapeutic efficacy against human tumor xenografts in vivo. Furthermore, these derivatives have little or no cross-resistance to either Taxol or Vinblastine. Remarkably, complete tumor remission in nude mice bearing human breast carcinoma MX-1 xenograft by 13a,b and 14g,h and significant suppression against prostate adenocarcinoma PC3 xenograft by 13b were achieved at the maximum tolerable dose with relatively low toxicity. In addition, these agents induce DNA interstrand cross-linking and substantial G2/M phase arrest in human non-small lung carcinoma H1299 cells. The current studies suggested that these agents are promising candidates for preclinical studies.

Multidrug-resistant cells overexpressing P-glycoprotein are susceptible to DNA crosslinking agents due to attenuated Src/nuclear EGFR cascade-activated DNA repair activity

We synthesized several novel bifunctional alkylating derivatives of 3a-aza-cyclopenta[a]indene (BO-1012, BO-1005, BO-1099 and BO-1101) that are potent DNA interstrand crosslinking agents. In in vitro cytotoxicity assay, these compounds were more cytotoxic to multidrug-resistant (MDR) cells, such as KBvin10, KBtax50 and CEM/VBL, than their parental cells. Using a xenograft model, BO-1012, at a dose of 5 mg/kg, partially suppressed the growth of parental KB cells but completely suppressed the growth of KBvin10 cells in nude mice. In exploring the possible mechanism, we found that DNA double-strand break (DSB) repair activity in MDR cells, KBvin10 and CEM/VBL, was significantly reduced compared with their parental cells, KB and CEM. Reduced DSB repair activity in KBvin10 cells was likely due to a defect in nuclear translocation of DNA-dependent protein kinase (DNA-PK), a component of the non-homologous end-joining repair machinery. Furthermore, BO-1012-induced DNA-PK translocation from the cytosol into the nucleus in KB cells is associated with the activation of the Src/nuclear epidermal growth factor receptor (EGFR) cascade, which is defective in MDR cells. As knockdown of P-glycoprotein (P-gp) by siRNA reactivated the Src/nuclear EGFR cascade, DNA-PK translocation and DNA repair activity in MDR cells, overexpression of P-gp attenuates the activity of DNA DSB repair through suppression of Src/nuclear EGFR cascade. Therefore, DNA interstrand crosslinking agents may have potential therapeutic use against P-gp-overexpressing MDR cells.

Novel bifunctional alkylating agents, 5,10-dihydropyrrolo[1,2-b]isoquinoline derivatives, synthesis and biological activity.

A series of linear pyrrolo[1,2-b]isoquinoline derivatives was synthesized for antitumor evaluation. The preliminary antitumor studies reveal that both bis(hydroxymethyl) and their bis(alkylcarbamate) derivatives show significant antitumor activity in inhibiting various human tumor cell growth in vitro. 1,2-Bis(hydroxymethyl)-3-methyl-5,10-dihydropyrrolo[1,2-b]isoquinoline (20a) was selected for antitumor studies in animal models. The results show that this agent can induce complete tumor remission or significant suppression in nude mice bearing human breast (MX-1) xenograft and ovarian (SK-OV-3) xenografts, respectively. Alkaline agarose gel shifting assay showed that 20a is able to cross-link with DNA. Studies on the cell cycle inhibition revealed that this agent induces cell arrest at G2/M phase. The results warrant further antitumor investigation against other human tumor growth in animal models.

Abstract C31: Derivatives of 3a-aza-cyclopenta[a]indene effectively suppress the growth of P-glycoprotein overexpressing cells in in vitro and in vivo systems.

We synthesized several novel bifunctional alkylating derivatives of 3a-aza-cyclopenta[ a ]indene (BO-1012, BO-1005, BO-1099, and BO-1101), which are potent DNA interstrand crosslinking agents. In in vitro cytotoxicity assay, these compounds were more cytotoxic to multidrug resistant (MDR) cells, such as KBvin10, KBtax50, and CEM/VBL, than their parental cells. Using a xenograft model, BO-1012, at a dose of 5 mg/kg, partially suppressed the growth of parental KB cells but completely suppressed the growth of KBvin10 cells in nude mice. In exploring the possible mechanism, we found that DNA double-strand break (DSB) repair activity in MDR cells, KBvin10 and CEM/VBL, was significantly reduced compared to their parental cells, KB and CEM. Reduced DSB repair activity in KBvin10 cells was likely due to a defect in nuclear translocation of DNA-PK, a component of the non-homologous end-joining repair machinery. Furthermore, BO-1012-induced DNA-PK translocation from the cytosol into the nucleus in KB cells is associated with activation of the Src/nuclear EGFR cascade, which is defective in MDR cells. Since knockdown of P-glycoprotein (P-gp) by siRNA reactivated the Src/nuclear EGFR cascade, DNA-PK translocation, and DNA repair activity in MDR cells, overexpression of P-gp attenuates the activity of DNA DSB repair via suppression of SRc/nuclear EGFR cascade. Therefore, DNA interstrand crosslinking agents may have potential therapeutic use against P-gp-overexpressing MDR cells. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr C31.

Abstract 1722: Multidrug resistant P-glycoprotein mediates through Src-EGFR cascade to attenuate DNA-PK-mediated DNA repair

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL We have synthesized several novel bifunctional alkylating derivatives of 3a-aza-cyclopenta[a]inden (BO-1012, BO-1005, BO-1099, and BO-1101) and demonstrated that they were more cytotoxic to multidrug resistant (MDR) cells, such as KBvin10, KBtax50, and CEM/VBL, than their parental cells. Using xenograft model, we confirmed that BO-1012 was more effective to suppress the growth of MDR KBvin10 cells in nude mice than to the parental KB cells. To explore the possible mechanism, we revealed that KBvin10 and CEM/VBL MDR cells have significantly lower repair activity than KB and CEM parental cells. Our results also showed that reduced repair activity in KBvin10 cells was likely due to the defective in translocation of DNA-PK, a component of repair machinery of non-homologous end-joining, from cytosol into nucleus. Furthermore, we found that BO-1012 could activate Src kinase and trigger nuclear transport of EGFR in parental cells but not in MDR cells. Inhibition of Src resulted in suppression of BO-1012-induced EGFR translocation and consequently sensitized the parental cells to BO-1012. On the contrary, Src activation induced by BO-1012 was revived in MDR cells by using siRNA to reduce the expression of P-glycoprotein (P-gp). Meanwhile, P-gp silencing also facilitated the EGFR translocation and DNA repair in MDR cells treated with BO-1012. Taken together, our results demonstrated that overexpression of P-gp in MDR cells mediates through inhibition of Src-EGFR pathway to attenuate the DNA-PK-mediated repair. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1722. doi:10.1158/1538-7445.AM2011-1722