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A high D8/D3 ratio was independently associated with better PFS in patients with NSCLC treated with gefitinib. Our findings suggest that the pharmacokinetics of gefitinib may be involved in its antitumor activity.

The outcomes of osteosarcoma patients still remain poor because of intractable pulmonary metastasis. We previously established a highly metastatic osteosarcoma cell line, LM8 from Dunn mouse osteosarcoma by in vivo selection. We herein aimed to clarify the characteristic biological features related with high metastatic potential and new target molecules to suppress pulmonary metastasis of osteosarcoma, using this syngeneic spontaneous metastatic model. LM8 cells acquired fibroblastic morphology with striking filopodia on the cell surface. Immunostaining showed faint stress fiber formation and peripherally localized integrin β1, and biochemical analyses showed the activated Cdc42 and autophosphorylation of focal adhesion kinase (FAK) in LM8 cells when compared to Dunn cells. LM8 cells had activated motility in single cell migration mode. LM8 migration was increased by a Rho-associated kinase (ROCK) inhibitor, Y-27632, while decreased by Cdc42 silencing using RNA interference system. We found that a clinically approved camptothecin analog, irinotecan suppressed the migration, Cdc42 activity, and autophosphorylation of FAK, and attenuated integrin β1 distribution selectively in LM8 cells. Daily oral administration of irinotecan significantly reduced the rate and size of pulmonary metastasis in syngeneic C3H mice. The fibroblastic morphology and activated cell migration with the dependency on Cdc42 but not Rho-ROCK signaling pathway argued that LM8 moved in mesenchymal mode of cell migration. This activated mesenchymal migration was a key component of the pulmonary metastasis of LM8 cells. The inhibition of mesenchymal migration by irinotecan, in addition to its cytotoxic effects, might be effective in preventing pulmonary metastasis of osteosarcoma.

Key words: SN-38; CPT analogue; ABCG2; SAR; DNA topoisomerase I inhibitorCamptothecin (CPT) is an antitumor alkaloid that was originally isolated from Camptotheca acuminata, a tree native to southern China. 1 CPT has been demonstrated to inhibit mammalian DNA topoisomerase I (Topo I), the nuclear enzyme that changes the topologic state of duplex DNA by single-strand breakage and resealing. Stabilization of the covalent Topo I-DNA complex (so-called cleavable complex) by CPT is a critical step in its antitumor action where Topo I-mediated DNA breaks are induced via prevention of DNA relegation. 2 Clinical and preclinical studies, however, revealed reversible bone marrow depression and hemorrhagic cystitis as the major dose-limiting toxicities of CPT. Thus, efforts have been directed at finding new CPT analogues with higher antitumor activity and less toxicity.Irinotecan (7-ethyl-10-[4-(1-piperidino)-1-piperidino]-carbonyloxycamptothecin; CPT-11) has been synthesized and developed as a new water-soluble analogue of CPT with wide-spectrum antitumor activity against many types of human tumor cells. 3,4 CPT-11 is a prodrug, the antitumor activity of which is exerted by 7-ethyl-10-hydroxycamptothecin (SN-38), the active metabolite of CPT-11. 5 Despite enormous expense and efforts spent on the development of cancer chemotherapies, acquired and intrinsic drug resistance in tumors is the major obstacle to long-term sustained patient response to chemotherapy. Hitherto several mechanisms for the resistance to SN-38 and its analogues have been proposed, e.g., mutations or decreased expression of Topo I, increased expression of the UGT1A protein or single nucleotide polymorphisms (SNPs) of the UGT1A gene, increased activity of O 6 -methylguanine-DNA-methyltransferase, a DNA repair protein, decreased activity of carboxylesterase that catalyzes the biosynthesis of SN-38 from CPT-11 in the plasma and liver and overexpression of drug export pumps. 6 -11 It has been documented that several ATP-binding cassette (ABC) transporters, such as P-glycoprotein (ABCB1/MDR1/P-gp) and multidrug resistance-associated protein 1 (ABCC1/MRP1), can cause drug resistance in tumor cells by actively extruding antitumor drugs. 12,13 Recently, a novel ABC transporter, ABCG2, also known as breast cancer-resistant protein (BCRP/MXR1/ ABCP), has been discovered in drug-resistant cell lines selected for by mitoxantrone or Topo I inhibitors. 14,15 Overexpression of ABCG2 has been shown to confer resistance to doxorubicin, mitoxantrone and various CPT analogues. 14,16,17 In a recent study with plasma membrane vesicles prepared from ABCG2-overexpressing SN-38-selected human small cell lung carcinoma cells, we found that ABCG2 transported SN-38 and its glucuronide metabolite in an ATP-dependent manner. 18 Thus, it is highly likely that ABCG2 actively extrudes SN-38 from tumor cells and thereby confers drug resistance.To circumvent ABCG2-associated drug resistance, in the present study we have synthesized a total of 14 different analogues of CPT and examine...

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