Metabolic targeting with recombinant methioninase combined with palbociclib regresses a doxorubicin-resistant dedifferentiated liposarcoma

https://doi.org/10.1016/j.bbrc.2018.10.119Get rights and content

Highlights

  • Ddedifferentiated liposarcoma (DDLPS) is recalcitrant and has the lowest survival rate.

  • Determine the efficacy of rMETase combined with palbociclib (PAL) against a DOX-resistant DDLPS in a PDOX mouse model.

  • The combination of PAL and rMETase significantly regressed tumor volume.

  • The combination of rMETase and PAL could be developed clinically.

Abstract

Liposarcoma is the most common type of soft tissue sarcoma. Among the subtypes of liposarcoma, dedifferentiated liposarcoma (DDLPS) is recalcitrant and has the lowest survival rate. The aim of the present study is to determine the efficacy of metabolic targeting with recombinant methioninase (rMETase) combined with palbociclib (PAL) against a doxorubicin (DOX)-resistant DDLPS in a patient-derived orthotopic xenograft (PDOX) model. A resected tumor from a patient with recurrent high-grade DDLPS in the right retroperitoneum was grown orthotopically in the right retroperitoneum of nude mice to establish a PDOX model. The PDOX models were randomized into the following groups when tumor volume reached 100 mm3: G1, control without treatment; G2, DOX; G3, PAL; G4, recombinant methioninase (rMETase); G5, PAL combined with rMETase. Tumor length and width were measured both pre- and post-treatment. On day 14 after initiation, all treatments significantly inhibited tumor growth compared to the untreated control except DOX. PAL combined with rMETase was significantly more effective than both DOX, rMETase alone, and PAL alone. Combining PAL and rMETase significantly regressed tumor volume on day 14 after initiation of treatment and was the only treatment to do so. The relative body weight on day 14 compared with day 0 did not significantly differ between each treatment group. The results of the present study indicate the powerful combination of rMETase and PAL should be tested clinically against DDLPS in the near future.

Introduction

Liposarcoma (LPS) is the most common type of soft-tissue sarcoma (STS). It consists of about 15%–25% of all STS. Histologically, LPS is classified as either well-differentiated/atypical lipomatous tumors, pleomorphic, myxoid/round cell, or de-differentiated. About 40% of the LPS are well-differentiated and 10% develop into de-differentiated liposarcoma (DDLPS) [1]. DDLPS is most common in the retroperitoneum. DDLPS contains the genomic amplification of the 12q13-15 chromosomal region of the mouse double minute 2 homolog (MDM2) gene [2,3]. Recently, Asano et al. [4] found that receptor tyrosine kinase (RTK) genes were amplified in approximately one-third of DDLPS. DDLPS has the lowest survival rate among all LPS [5] and often recurs or metastasizes due to incomplete resection and resistance to radiation, or first-line chemotherapy with doxorubicin (DOX) [[5], [6], [7]]. Small-molecule inhibitors of MDM2 and cyclin-dependent kinase 4 (CDK4) were shown to be a treatment option for DDLPS in Phase I and II clinical trials [[8], [9], [10]] with limited efficacy alone on unresectable DDLPS. Therefore, transformative individualized therapy is necessary against DDLPS.

We have developed the patient-derived orthotopic xenograft (PDOX) mouse model of cancer for discovery of transformative individualized therapy for recalcitrant cancer [11]. Our laboratory pioneered the PDOX nude mouse model with the technique of surgical orthotopic implantation (SOI) for all major cancers [[12], [13], [14], [15], [16], [17], [18], [19], [20], [21], [22], [23], [24], [25]]. The PDOX model, developed by us, has many advantages over subcutaneous-transplant models [11,26].

Cancer cells have an elevated requirement for methionine (MET) compared to normal cells. This phenomenon is termed MET dependence [27]. MET restriction arrests tumor growth and induces a selective S/G2-phase cell-cycle block of cancer cells in vitro and in vivo [[28], [29], [30], [31], [32]].

Recent studies suggest that MET dependence is duet to excessive use of MET for aberrant transmethylation reactions, termed the Hoffman effect [[33], [34], [35], [36], [37], [38]], analogous to the Warburg effect for glucose in cancer [39]. The excessive and aberrant use of MET in cancer is shown in the clinic in [11C]MET PET imaging, where high uptake of [11C]MET results in a very strong and selective tumor signal compared to normal tissue background. [11C]MET is better than [18C]fluorodeoxyglucose (FDG) for PET imaging [40], suggesting MET dependence is highly cancer-specific compared to glucose dependence [[41], [42], [43]]. Lien et al. [44] have shown that oncogenic PIK3CA (phosphoinositide-3-kinase, catalytic, alpha polypeptide) promotes MET and cysteine utilization in breast cancer cells by inhibiting the cystine transporter. Further, dietary modulation of MET can alter the levels of histone methylation [45,46]. Recently, Dai et al. [47] found no change in the location of histone H3 lysine 4 trimethylation (H3K4me3) peaks under MET restriction but found that MET restriction altered the response of H3K4me3 peak width and its biology [47].

Previous studies have demonstrated that MET the cleaving enzyme, methioninase (METase), purified from Pseudomonas putida (P. putida), is an effective antitumor agent [[48], [49], [50], [51]]. For the large-scale production of METase, the gene from P. putida was cloned in Escherichia coli (E. coli) and a purification protocol for recombinant METase (rMETase) has been established with high purity and low endotoxin [[52], [53], [54], [55], [56], [57]]. Recently, we found that oral-METase (o-METase) is superior to injectable rMETase against acquired GEM resistance in pancreatic cancer (59).

Palbociclib (PAL), a CDK 4/6 inhibitor [[59], [60], [61]], had clinical efficacy for several tumor types [10,[62], [63], [64], [65], [66]]. PAL in combination with letrozole or fulvestran was recently approved by the US Food and Drug Administration (FDA) for breast cancer [59,66].

The present report demonstrates the efficacy of rMETase combined with palbociclib on the PDOX model of DOX-resistant DDLPS.

Section snippets

Mice

Athymic nu/nu nude mice (AntiCancer Inc., San Diego, CA), 4–6 weeks old, were used in this study. Animal housing, their diet, surgical procedures, and imaging were performed as previously described [[23], [24], [25]]. The response of animals during surgery was monitored to ensure adequate depth of anesthesia. The animals were observed daily and humanely sacrificed by CO2 inhalation if they met the humane endpoint criteria as described in our previous publication [[23], [24], [25]]. All animal

Effect of treatment on tumor growth

On day 14 after treatment initiation, all treatments significantly inhibited tumor growth compared to untreated control except DOX: (DOX: p = 0.76; rMETase: p = 0.05; PAL: p < 0.05; PAL combined with rMETase p<0.001) on day 14 after initiation. PAL combined with rMETase was significantly more effective than either DOX (p < 0.001), rMETase alone (p < 0.01) or PAL alone (p < 0.01). The combination of PAL and rMETase significantly regressed tumor volume on day 14 after initiation of treatment

Discussion

DDLPS is one of the most lethal malignancies with lowest survival rate among all types of LPS [5] and often recurs or metastasizes because of lack of curative treatments. Therefore, transformative individualized therapy is needed for DDLPS. In the present study, we show that a combination of rMETase and PAL is effective against DDLPS. The strong efficacy of the PAL-rMETase combination is important and the mechanism will be further studied in the future.

Bollard et al. [65] found that PAL

Funding

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

Conflicts of interest

The authors declare that there are no potential conflicts of interest.

Acknowledgement

This paper is dedicated to the memory of A. R. Moossa, M.D., and Sun Lee, M.D.

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