Elsevier

Tissue and Cell

Volume 61, December 2019, Pages 109-114
Tissue and Cell

Oral recombinant methioninase combined with oxaliplatinum and 5-fluorouracil regressed a colon cancer growing on the peritoneal surface in a patient-derived orthotopic xenograft mouse model

https://doi.org/10.1016/j.tice.2019.09.006Get rights and content

Highlights

  • Colorectal cancer is the third most common cancer worldwide.

  • The peritoneal surface is a common site of metastasis in colorectal cancer.

  • The combination of 5-FU + OXA + o-rMETase was efficacious for colon cancer peritoneal metastasis.

  • First study to show o-rMETase inhibits peritoneal metastases.

  • o-rMETase is promising as a novel cancer therapeutic for human colon cancer.

Abstract

The aim of this study was to determine the efficacy of oral recombinant methioninase (o-rMETase) on a model of colon cancer growing on the peritoneal surface using a patients-derived orthotopic xenograft (PDOX) nude mouse model. Forty PDOX mouse models with colon cancer growing on the peritoneum were divided into 4 groups of 10 mice each by measuring the tumor size and fluorescence intensity: untreated control; 5-fluorouracil (5-FU) (50 mg/kg, once a week for two weeks, ip) and oxaliplatinum (OXA) (6  mg/kg, once a week for two weeks, ip); o-rMETase (100 units/day, oral 14 consecutive days); combination 5-FU + OXA and o-rMETase. All treatments inhibited tumor growth compared to the untreated control. The combination of 5-FU + OXA plus o-rMETase was significantly more efficacious than the control and each drug alone and was the only treatment that caused tumor regression. The present study is the first demonstrating the efficacy of o-rMETase combination therapy on a PDOX model of peritoneal colon cancer, suggesting potential clinical development of o-rMETase in a recalcitrant cancer.

Introduction

The elevated methionine (MET) requirement of cancer cells is referred to as MET dependence (Hoffman, 1984, 2015a, 2017; Stern and Hoffman, 1984; Wang et al., 2019). The elevated MET use in cancer is called the "Hoffman effect" analogous to the Warburg effect of excess glucose consumption by cancers. Comparison of radioactive MET- and glucose- PET (positron-emission tomography) imaging has shown a stronger signal with MET suggesting that the Hoffman effect is more pronounced than the Warburg effect (Hoffman, 2015a, 2017; Xu et al., 2017).

MET restriction by recombinant methioninase (rMETase) can inhibit the growth of cancer cells in vitro and in vivo (Hoffman, 2015a, 2017). rMETase has been used as a treatment strategy for various types of cancer (Hoffman, 2015a; Kreis and Hession, 1973; Tan et al., 2010; Xin et al., 2013, 2018; Yano et al., 2014, 2016).

Previous studies have shown that intra-peritoneal recombinant methioninase injection (ip-rMETase) was effective against patient-derived orthotopic xenograft (PDOX) mouse models of recalcitrant cancer (Kawaguchi et al., 2017a,2018a,c; Igarashi et al., 2018a,b]. Recently, we reported that oral recombinant methioninase (o-rMETase) was significantly more effective than intraperitoneal injection rMETase (ip-rMETase) indicating the potential widespread use of rMETase for cancer treatment (Higuchi et al., 2018; Higuchi et al., 2019; Kawaguchi et al., 2018a; Kawaguchi et al., 2017b; Kawaguchi et al., 2018d; Oshiro et al., 2019; Park et al., 2019a).

Orthotopic implantation of intact tumor tissue in appropriate mouse models can result in metastasis resembling the clinical pattern, unlike subcutaneous transplantation. Our laboratory has used surgical orthotopic implantation (SOI) to establish PDOX nude-mouse models from patient tumor specimens (Fu and Hoffman, 1993; Hiroshima et al., 2014; Hiroshima et al., 2015; Hoffman, 2015b; Igarashi et al., 2018a,2018b,c; Kawaguchi et al., 2017a,2017b; Kawaguchi et al., 2018c,d,e; Kawaguchi et al., 2016; Kiyuna et al., 2016; Metildi et al., 2014; Murakami et al., 2016; Wang et al., 1992; Yamamoto et al., 2016). The PDOX orthotopic models are much more patient-like than the ectopic subcutaneous models (Hoffman, 2015b). However, in orthotopic models, it is difficult to visualize tumor growth. To address this problem of imaging such orthotopic tumor grafts, we have recently developed the technology to introduce fluorescent protein-expressing stroma into tumors by passaging tumor grafts through transgenic nude mice expressing fluorescent protein (Suetsugu et al., 2012).

The present report demonstrates the efficacy of o-rMETase using a PDOX model of colon cancer growing on the peritoneal surface in nude mouse, with brightly labeled red fluorescent protein (RFP)-expressing stroma for imaging.

Section snippets

Mice

Athymic nu/nu nude mice and transgenic RFP expressing athymic nu/nu mice (four to 6 weeks) were obtained from AntiCancer Inc. (San Diego, CA). All surgical procedures and imaging were performed in accordance with an AntiCancer Institutional Animal Care and Use Committee (IACUC)-protocol specifically approved for this study, and in accordance with the principles and procedures outlined in the National Institutes of Health Guide for the Care and Use of Animals under Assurance Number A3873-1.

Non-invasive red fluorescence imaging of colon cancer growing on the peritoneal surface of the PDOX model

Orthotopically-implanted tumors grew in the peritoneum of non-transgenic nude mice. These tumors had sufficient red fluorescent stroma from previous growth in RFP nude mice to permit non-invasive imaging through the abdominal wall (Fig. 1A-D). Confocal laser microscopy showed persistence of the RFP-expressing stroma in frozen sections (Fig. 2A, B).

Tumor histology

Histologically, the untreated control tumor mainly comprised viable carcinoma cells. (Fig. 2C) In contrast, tumor treated with the combination of

Discussion

Recently, a paper appeared with the title “The new anticancer era: tumor metabolism targeting” (Borriello and Della Ragione, 2017). However, this “new anticancer era” started in 1959 with the observation of Sugimura et al. that depriving animals of MET arrested tumor growth (Sugimura et al., 1959). The Warburg effect refers to the significantly increased uptaked glucose by cancer cells. In addition to calorie restriction, specific amino acid restriction has been studied in the past to treat

Declaration of Competing Interest

JHP, HO, KM, TH, SNY, ZZ, NS, JY and RMH are or were unsalaried associates of AntiCancer Inc. QH, MZ and YT are employee of AntiCancer Inc. AntiCancer Inc. uses PDOX models for contract research. The Authors declare that there are no potential conflicts of interest.

Acknowledgements

This paper is dedicated to the memory of A.R. Moossa, Sun Lee, MD, Professor Li Jiaxi and Masaki Kitajima, MD.

Funding

This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

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