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Carcinoembryonic antigen (CEA) is a tumor-specific antigen overexpressed in multiple cancers. CEA is expressed as a membrane protein, a part of which is cleaved from the cell membrane and secreted into blood. The soluble form of CEA (sCEA) has been shown to accelerate the clearance of anti-CEA antibody, which limits the antibody distribution in the tumor. To overcome this issue, we developed an anti-CEA monoclonal antibody, 15-1-32, which shows a strong affinity for membrane-bound CEA (mCEA) and relatively weak affinity for sCEA. In this study, we compared the effect of sCEA on the pharmacokinetics of 15-1-32 in mice with that of another anti-CEA monoclonal antibody, labetuzumab, showing less selectivity to mCEA than 15-1-32. As expected, the effect of sCEA on the serum concentration of 15-1-32 was much smaller than that of labetuzumab. The decrease in the area under the curve (AUC) of serum concentration was 22.5% for 15-1-32 when it was coadministered with sCEA, while that of labetuzumab was 79.9%. We also compared the pharmacokinetics of these two antibodies in CEA-positive tumor-bearing mice. The AUCs of 15-1-32 and labetuzumab were decreased in tumor-bearing mice compared with non-tumor-bearing mice to a similar extent (approximately 40% decrease). These results suggested that mCEA also contributes to the clearance of anti-CEA antibodies in CEA-positive tumor-bearing mice. Although the increased selectivity to mCEA minimized the effect of sCEA on the pharmacokinetics of 15-1-32, it may be insufficient to improve the pharmacokinetics in CEA-positive cancer patients. SIGNIFICANCE STATEMENT: Because previous studies reported the rapid clearance of anti-CEA antibodies mediated by soluble CEA, we obtained a monoclonal antibody, 15-1-32, selective to membrane-bound CEA and evaluated the effects of CEA on its pharmacokinetics. Although the effect of soluble CEA on the serum concentration of 15-1-32 was very small, the clearance of 15-1-32 in CEA-positive tumor-bearing mice was still rapid, suggesting membrane-bound CEA also contributes to the clearance of anti-CEA antibodies. These results indicated that increasing selectivity to membrane-bound CEA is not enough to improve the pharmacokinetics of anti-CEA antibody.

Photodynamic therapy (PDT) utilize a photosensitizing agent and light for cancer therapy. It exerts anti-cancer effect mainly by inducing vascular occlusion at the irradiated site. By controlling the irradiation area, PDT can be used in a tumor-specific manner. However, the non-specific cellular damage in the surrounding normal tissue is still a serious concern. Photoimmunotherapy (PIT) is a new type of targeted cancer therapy that uses an antibody-photon absorber conjugate (APC). The superiority of PIT to PDT is the improved target specificity, thereby reducing the damage to normal tissues. Here, we developed a novel APC targeting epithelial cell adhesion molecule (EpCAM) as well as a negative control APC that does not bind to the EpCAM antigen. Our in vitro analysis of APC cytotoxicity demonstrated that the EpCAM APC, but not the negative control, was cytotoxic to EpCAM expressing COLO 205 cells after photoirradiation, suggesting that the cytotoxicity is antigen-dependent. However, in our in vivo analysis using a mouse xenograft tumor model, decreased volume of the tumors was observed in all the mice treated with irradiation, regardless of whether they were treated with the EpCAM APC or the negative control. Detailed investigation of the mechanism of these in vivo reveal that both APCs induce vascular occlusion at the irradiation site. Furthermore, the level of vascular occlusion was correlated with the blood concentration of APC, not the tumor concentration. These results imply that, similar to PDT, PIT can also induce non-targeted vascular occlusion and further optimization is required before widespread clinical use.

Carcinoembryonic antigen (CEA) is a classic tumor‐specific antigen that is overexpressed in several cancers, including gastric cancer. Although some anti‐CEA antibodies have been tested, to the best of our knowledge, there are currently no clinically approved anti‐CEA antibody therapies. Because of this, we have created the novel anti‐CEA antibody, 15‐1‐32, which exhibits stronger binding to membrane‐bound CEA on cancer cells than existing anti‐CEA antibodies. 15‐1‐32 also shows poor affinity for soluble CEA; thus, the binding activity of 15‐1‐32 to membrane‐bound CEA is not influenced by soluble CEA. In addition, we constructed a 15‐1‐32‐monomethyl auristatin E conjugate (15‐1‐32‐vcMMAE) to improve the therapeutic efficacy of 15‐1‐32. 15‐1‐32‐vcMMAE showed enhanced antitumor activity against gastric cancer cell lines. Unlike with existing anti‐CEA antibody therapies, antitumor activity of 15‐1‐32‐vcMMAE was retained in the presence of high concentrations of soluble CEA.

薬物の多くは，血漿中でアルブミンやα1-酸性糖タンパクなどのタンパク質へ結合している．アルブミンは脂溶性の高い酸性化合物，α1-酸性糖タンパクは塩基性化合物に対し高い親和性を示す．タンパクへ結合した薬物は細胞膜を透過することができないため，血漿中の遊離型薬物のみが組織に分布して薬効や毒性を発現し，代謝や排泄を受けて体内から除去される．したがって，血漿中タンパク結合は薬物の体内動態や薬効，毒性に多大な影響を及ぼす．血漿中タンパク結合には種差があり，体内動態や薬効，毒性の種差の原因となる．また，血漿中タンパク結合は病態や薬物間相互作用によっても変動し，薬効の減弱や副作用の増強など臨床上好ましくない現象を引き起こす場合がある．したがって，薬物の血漿中タンパク結合は医薬品の探索・開発を通じて評価しなければならない重要な項目の一つである．

Heparin-binding EGF-like growth factor (HB-EGF) is a member of the EGF family and is an important therapeutic target in some types of human cancers. KM3566 is a mouse anti-HB-EGF monoclonal antibody that neutralizes HB-EGF activity by inhibiting the binding of HB-EGF to its receptors. Based on the results of our pharmacokinetics study, a humanized derivative antibody, KHK2866, is rapidly cleared from serum and shows nonlinear pharmacokinetics in cynomolgus monkeys. In this study, we examined the antigen-dependent clearance of an anti-HB-EGF monoclonal antibody in vivo and in vitro in order to pharmacokinetically explain the rapid elimination of KHK2866. We revealed tumor size-dependent clearance of KM3566 in in vivo studies and obtained good fits between the observed and simulated concentrations of KM3566 based on the two-compartment with a saturable route of clearance model. Furthermore, in vivo imaging analyses demonstrated tumor-specific distribution of KM3566. We then confirmed rapid internalization and distribution to lysosome of KM3566 at a cellular level. Moreover, we revealed that the amounts of HB-EGF on cell surface membrane were maintained even while HB-EGF was internalized with KM3566. Recycled or newly synthesized HB-EGF, therefore, may contribute to a consecutive clearance of KM3566, which could explain a rapid clearance from serum. These data suggested that the rapid elimination in pharmacokinetics of KM3566 is due to antigen-dependent clearance. Given that its antigen is expressed in a wide range of normal tissue, it is estimated that the rapid elimination of KHK2866 from cynomolgus monkey serum is caused by antigen-dependent clearance.

Background: Heparin-binding EGF-like Growth Factor (HB-EGF), a member of the EGF family, exerts its biological activity through activation of the EGF receptors. HB-EGF is initially synthesized as a membrane-anchored precursor protein (proHB-EGF), and then proteolytically cleaved, resulting in the mitogenically active soluble form. HB-EGF plays pivotal roles in many physiologic and pathologic processes such as development and cell proliferation. In this study, we measured soluble HB-EGF concentrations in serum samples obtained from healthy volunteers and patients of various cancer types. Materials and methods: Soluble HB-EGF levels in human serum samples were quantified by the immuno-PCR method. Results: The mean soluble HB-EGF levels of the 20 healthy volunteers and 10 colon, breast, ovarian, head and neck, non-small cell lung, pancreatic, and small cell lung cancer patients were 5.04, 18.0, 15.1, 10.4, 6.69, 9.78, 23.6, and 5.60 pg/mL, respectively. There was a statistically significant difference between the HB-EGF concentrations of healthy volunteers and patients in 5 out of 7 cancer types. Furthermore, a trend for HB-EGF levels to increase along with disease stage was observed. Conclusion: Soluble HB-EGF may be a useful diagnostic serological biomarker for various cancer types, and a predictive and/or pharmacodynamic biomarker for HB-EGF-targeted therapeutics.