Preparation of an antitumor and antivirus agent: chemical modification of α-MMC and MAP30 from Momordica Charantia L. with covalent conjugation of polyethyelene glycol.
BACKGROUND
Alpha-momorcharin (α-MMC) and momordica anti-HIV protein (MAP30) derived from Momordica charantia L. have been confirmed to possess antitumor and antivirus activities due to their RNA-N-glycosidase activity. However, strong immunogenicity and short plasma half-life limit their clinical application. To solve this problem, the two proteins were modified with (mPEG)(2)-Lys-NHS (20 kDa).


RESULTS
In this article, a novel purification strategy for the two main type I ribosome-inactivating proteins (RIPs), α-MMC and MAP30, was successfully developed for laboratory-scale preparation. Using this dramatic method, 200 mg of α-MMC and about 120 mg of MAP30 was obtained in only one purification process from 200 g of Momordica charantia seeds. The homogeneity and some other properties of the two proteins were assessed by gradient SDS-PAGE, electrospray ionization quadruple mass spectrometry, and N-terminal sequence analysis as well as Western blot. Two polyethylene glycol (PEG)ylated proteins were synthesized and purified. Homogeneous mono-, di-, or tri-PEGylated proteins were characterized by matrix-assisted laser desorption ionization-time of flight mass spectrometry. The analysis of antitumor and antivirus activities indicated that the serial PEGylated RIPs preserved moderate activities on JAR choriocarcinoma cells and herpes simplex virus-1. Furthermore, both PEGylated proteins showed about 60%-70% antitumor and antivirus activities, and at the same time decreased 50%-70% immunogenicity when compared with their unmodified counterparts.


CONCLUSIONS
α-MMC and MAP30 obtained from this novel purification strategy can meet the requirement of a large amount of samples for research. Their chemical modification can solve the problem of strong immunogenicity and meanwhile preserve moderate activities. All these findings suggest the potential application of PEGylated α-MMC and PEGylated MAP30 as antitumor and antivirus agents. According to these results, PEGylated RIPs can be constructed with nanomaterials to be a targeting drug that can further decrease immunogenicity and side effects. Through nanotechnology we can make them low-release drugs, which can further prolong their half-life period in the human body.
School of Medical Laboratory Science, Chengdu Medical College, Chengdu, China.
Yao
Y
Meng
Shuangfeng
S
Liu
Juan
J
Li
Yanfa
Y
Meng
Xiaojun
X
Zhao
D000818:Animals; D000970:Antineoplastic Agents / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification / Q000494:pharmacology*; D000998:Antiviral Agents / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification / Q000494:pharmacology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002522:Chlorocebus aethiops; D018259:Herpesvirus 1, Human / Q000187:drug effects; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood; D029777:Momordica charantia / Q000737:chemistry; D011092:Polyethylene Glycols / Q000737:chemistry*; D051381:Rats; D017207:Rats, Sprague-Dawley; D054788:Ribosome Inactivating Proteins / Q000737:chemistry* / Q000276:immunology / Q000302:isolation & purification / Q000494:pharmacology*; D054790:Ribosome Inactivating Proteins, Type 2 / Q000737:chemistry* / Q000276:immunology / Q000302:isolation & purification / Q000494:pharmacology*; D012639:Seeds / Q000737:chemistry; D014709:Vero Cells
D000970:Antineoplastic Agents; D000998:Antiviral Agents; D007074:Immunoglobulin G; C066347:MAP30 protein, Momordica charantia; D054790:Ribosome Inactivating Proteins, Type 2; D011092:Polyethylene Glycols; C039582:MMC protein, Momordica charantia; D054788:Ribosome Inactivating Proteins