CCL7 and TGF-ß secreted by MSCs play opposite roles in regulating CRC metastasis in a KLF5/CXCL5-dependent manner.

CXCL5 is overexpressed in colorectal cancer (CRC) and promotes distant metastasis and angiogenesis of tumors; however, the underlying mechanism that mediates CXCL5 overexpression in CRC remains unclear. Here, we successfully extracted and identified primary mesenchymal stromal cells (MSCs) and verified the promoting effects of tumor-associated MSCs on CRC proliferation and metastasis in vivo and in vitro. We found that MSCs not only promoted the expression of CXCL5 by secreting CCL7 but also secreted TGF-ß to inhibit this process. After secretion, CCL7/CCR1 activated downstream CBP/P300 to acetylate KLF5 to promote CXCL5 transcription, while TGF-ß reversed the effect of KLF5 on transcription activation by regulating SMAD4. Taken together, our results indicate that MSCs in the tumor microenvironment promoted the progression and metastasis of CRC and regulated the expression of CXCL5 in CRC cells by secreting CCL7 and TGF-ß. KLF5 is the key site of these processes and plays a dual role in CXCL5 regulation. MSCs and their secreted factors may serve as potential therapeutic targets in the tumor environment.

High ligation versus low ligation of the inferior mesenteric artery in laparoscopic rectal cancer surgery: a retrospective study on surgical and long-term outcome.

BACKGROUND: In laparoscopic low anterior resection for rectal cancer surgery, there has been controversy to whether the inferior mesenteric artery (IMA) should be ligated at the origin of its aorta (high ligation (HL)) or below the branches of the left colonic artery (LCA) (low ligation (LL)). This study was intended to clarify oncological outcome and long-term prognosis of retrospective analysis. METHODS: Analyzed the cases who underwent laparoscopic low anterior resection (LAR) in Shanghai Ruijin Hospital from January 2015 to December 2016, 357patients scheduled into 2 groups according to the level of IMA ligation: HL (n = 247) versus LL (n = 110). RESULTS: The primary endpoint is long-term outcomes, and the secondary endpoint is the incidence rate of major postoperative complications. There were no significant differences in 5-year overall survival (P = 0.92) and 5-year disease-free survival (P = 0.41). There were no differences between the clinical baseline levels in each group. The incidence of low anterior resection syndrome (LARS) in the two groups was statistically significant (P = 0.037). No significant differences were observed in operative time (P = 0.092) and intraoperative blood loss (P = 0.118). In the HL group, 6 cases (2.4%) had additional colonic excision due to poor anastomotic blood supply; none of the colonic anastomosis in the low ligation group had ischemic manifestations, and length from the proximal margin (P = 0.076), length from the distal margin (P = 0.184), the total number of lymph nodes excised (P = 0.065), and anastomotic leakage incidence (P = 0.33). CONCLUSION: Low ligation of the IMA which reserved LCA with vascular root lymph node dissection in laparoscopic low anterior resection for rectal cancer surgery may help protect the blood supply of the anastomosis, and will not increase postoperative complications while enhance recovery, without compromising radical excision and long-term prognosis.

Use of biexponential and stretched exponential models of intravoxel incoherent motion and dynamic contrast-enhanced magnetic resonance imaging to assess the proliferation of endometrial carcinoma.

Background: It is important to assess the proliferation of endometrial carcinoma (EC) noninvasively using imaging methods. This prospective diagnostic study investigated the value of biexponential and stretched exponential models of intravoxel incoherent motion (IVIM) and dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) in predicting the Ki-67 status of EC. Methods: In all, 70 patients with EC underwent pelvic MRI. The diffusion coefficient (D), pseudo diffusion coefficient (D*), perfusion fraction (f), distributed diffusion coefficient (DDC), water molecular diffusion heterogeneity index (α), volume transfer constant (Ktrans), rate transfer constant (Kep), and volume of extravascular extracellular space per unit volume of tissue (Ve) were compared. The area under the receiver operating characteristic (ROC) curve (AUC) was used to quantify diagnostic efficacy. Multivariate logistic regression and bootstrap (1,000 samples) analyses were used to establish and evaluate, respectively, the optimal model to predict Ki-67 status. Results: D, Ktrans, and Kep were lower while α was higher in the high-proliferation group as compared with low-proliferation group (all P values<0.05). D and Kep were independent predictors of Ki-67 status in EC, and the combination of these parameters had optimal diagnostic efficacy (AUC =0.920; sensitivity 85.71%; specificity 89.29%), which was significantly better than that of D (AUC =0.753; Z=2.874; P=0.004), α (AUC =0.715; Z=3.505; P=0.001), Ktrans (AUC =0.808; Z=2.741; P=0.006), and Kep (AUC =0.832; Z=2.147; P=0.032) alone. The validation model showed good accuracy (AUC =0.882; 95% confidence interval 0.861-0.897) and consistency (C-statistic =0.902). D, Kep, Ktrans, and α showed a slightly negative (r=-0.271), moderately negative (r=-0.534), slightly negative (r=-0.409), and slightly positive (r=0.488) correlation with the Ki-67 index, respectively (all P values <0.05). Conclusions: IVIM- and DCE-MRI-derived parameters, including D, α, Ktrans, and Kep, were associated with Ki-67 status in EC, and the combination of D and Kep may serve as a superior imaging marker for the identification of low- and high-proliferation EC.

Quantitative dynamic contrast-enhanced parameters and intravoxel incoherent motion facilitate the prediction of TP53 status and risk stratification of early-stage endometrial carcinoma.

BACKGROUND: The aim of the study was to investigate the value of dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) and intravoxel incoherent motion (IVIM) in differentiating TP53-mutant from wild type, low-risk from non-low-risk early-stage endometrial carcinoma (EC). PATIENTS AND METHODS: A total of 74 EC patients underwent pelvic MRI. Parameters volume transfer constant (Ktrans), rate transfer constant (Kep), the volume of extravascular extracellular space per unit volume of tissue (Ve), true diffusion coefficient (D), pseudo-diffusion coefficient (D*), and microvascular volume fraction (f) were compared. The combination of parameters was investigated by logistic regression and evaluated by bootstrap (1000 samples), receiver operating characteristic (ROC) curves, calibration curves, and decision curve analysis (DCA). RESULTS: In the TP53-mutant group, Ktrans and Kep were higher and D was lower than in the TP53-wild group; Ktrans, Ve, f, and D were lower in the non-low-risk group than in the low-risk group (all P < 0.05). In the identification of TP53-mutant and TP53-wild early-stage EC, Ktrans and D were independent predictors, and the combination of them had an optimal diagnostic efficacy (AUC, 0.867; sensitivity, 92.00%; specificity, 80.95%), which was significantly better than D (Z = 2.169, P = 0.030) and Ktrans (Z = 2.572, P = 0.010). In the identification of low-risk and non-low-risk early-stage EC, Ktrans, Ve, and f were independent predictors, and the combination of them had an optimal diagnostic efficacy (AUC, 0.947; sensitivity, 83.33%; specificity, 93.18%), which was significantly better than D (Z = 3.113, P = 0.002), f (Z = 4.317, P < 0.001), Ktrans (Z = 2.713, P = 0.007), and Ve (Z = 3.175, P = 0.002). The calibration curves showed that the above two combinations of independent predictors, both have good consistency, and DCA showed that these combinations were reliable clinical prediction tools. CONCLUSIONS: Both DCE-MRI and IVIM facilitate the prediction of TP53 status and risk stratification in early-stage EC. Compare with each single parameter, the combination of independent predictors provided better predictive power and may serve as a superior imaging marker.

Colorectal cancer tissue-originated spheroids reveal tumor intrinsic signaling pathways and mimic patient clinical chemotherapeutic response as a rapid and valid model.

Locally advanced colorectal cancer requires preoperative chemotherapy to reduce local recurrence and metastasis rates, but it remains difficult to predict the tumor will be sensitive to which treatments. The patient-derived organoids (PDOs) are considered an effective platform for predicting tumor drug responses in precision oncology. However, it has the limitation of being time-consuming in practical applications, especially in neoadjuvant treatment. Here we used cancer tissue-originated spheroids (CTOS) method to establish organoids from a heterogeneous population of colorectal cancer specimens, and evaluated the capacity of CTOS to predict clinical drug responses. By analyzing the relationship of the activities of drug-treated CTOS, drug targets and target-related pathways, tumor intrinsic effective-target-related pathways can be identified. These pathways were highly matched to the abnormal pathways indicated by whole-exome sequencing. Based on this, we used half effective concentration gradients to classify CTOS as sensitive or resistant to chemotherapy regimens within a week, for predicting neoadjuvant treatment outcomes for colorectal cancer patients. The drug sensitivity test results are highly matched to the clinical responses to treatment in individual patients. Thus, our data suggested that CTOS models can be effectively screened ex vivo to identify pathways sensitive to chemotherapies. These data also supported organoid research for personalized clinical medication guidance immediately after diagnosis in patients with advanced colorectal cancer.

Dysbiosis of human tumor microbiome and aberrant residence of Actinomyces in tumor-associated fibroblasts in young-onset colorectal cancer.

Colorectal cancer (CRC) is the third most common form of cancer, and the incidence of sporadic young-onset colorectal cancer (yCRC) has been increasing. Microbiota residing in the tumor microenvironment are emerging tumor components. The colonic microbiome differs between patients with CRC and healthy controls; however, few studies have investigated the role of the tumor microbiota in disease diagnosis and tumorigenesis of yCRC. We performed 16S rRNA sequencing analysis to identify the microbiome in CRC and found that tumor microbial diversity decreased in yCRC. Proteobacteria and Firmicutes were the most abundant phyla in all CRC samples, and Actinomyces and Schaalia cardiffensis were the key microbiota in the yCRC group. Correlation analysis revealed that Actinomyces co-occurred with various pro-tumor microbial taxa, including Bacteroidia, Gammaproteobacteria, and Pseudomonas. An independent cohort was used to validate the results. The Actinomyces in CRC was co-localized with cancer-associated fibroblasts and activated the TLR2/NF-κB pathway and reduces CD8+ T lymphocyte infiltration in CRC microenvironment. This study suggests that tumoral microbiota plays an important role in promoting tumorigenesis and therefore has potential as a promising non-invasive tool and intervention target for anti-tumor therapy.

Traditional Chinese Medicine as an adjunct therapy in the treatment of idiopathic membranous nephropathy: A systematic review and meta-analysis.

BACKGROUND: Idiopathic membranous nephropathy (IMN) is one of the most common causes of nephrotic syndrome in adults involving multiple targets and factors. The effect of conservative nonimmunosuppressive or immunosuppressive therapies is unsatisfactory and with many side effects. Traditional Chinese medicine (TCM) can regulate immune function and improve kidney function. PURPOSE: To evaluate the total effective rate, curative rate, recurrence rate and adverse events of TCM alone or TCM as an adjunctive therapy for IMN. METHODS: Randomized controlled trials (RCT) comparing either TCM alone or the combination of TCM to western medicine (WM) therapies for patients with IMN were retrieved by searching English and Chinese database. Risk of bias summary was used to assess the methodological quality of eligible studies. Dichotomous data were presented using odds ratios (OR). The primary outcome measure was the total effective rate. Secondary outcomes included curative rate, recurrence rate and adverse events. RESULTS: 29 RCTs involving 1883 participants met the inclusion criteria. There was no statistically significant difference between the therapy of TCM alone and WM on the total effective rates (OR: 2.00; 95% CI: 0.80-4.98; P = 0.14) and curative rate (OR: 1.66; 95%CI: 0.66-4.22; p = 0.28). However, compared to basic treatment or immunosuppressive therapies alone, results showed that TCM as an adjunctive therapy had beneficial effects on the total effective rate (OR: 2.59; 95% CI: 1.38-4.86; P = 0.003 and OR: 3.01; 95% CI: 2.25-4.04; P < 0.00001) and curative rate (OR: 3.01; 95%CI: 1.24-7.28; p = 0.01 and OR: 1.73; 95%CI: 1.10-2.71; p = 0.02). In addition, the combination of TCM treatment could reduce the recurrence rate (OR: 0.28; 95% CI: 0.12-0.68; P = 0.004) and adverse reactions (OR: 0.38; 95% CI: 0.27-0.54; p < 0.00001). CONCLUSION: The results indicate that TCM is well-tolerated for the treatment of IMN. However, there remains a need for large-scale and high-quality trials.

Lipidomics reveals association of circulating lipids with body mass index and outcomes in IgA nephropathy patients.

IgA nephropathy (IgAN) is a leading cause of chronic kidney disease (CKD), which are commonly accompanied by dyslipidemia. Obesity is also associated with dyslipidemia and risk of CKD, but the relation of the dyslipidemia patterns with obesity and disease progression in IgAN patients remains unknown. Traditional Chinese medicine (TCM) and the combined treatment with corticosteroids and TCM have been shown to be of benefit for IgAN patients, but predictive markers for guiding these treatments are lacking. Here, we quantified 545 lipid species in the plasma from 196 participants, including 140 IgAN patients and 56 healthy volunteers, and revealed an altered plasma lipidome in IgAN patients as compared to healthy participants. Association analysis showed that a sub-group of glycerides, particularly triacylglycerols (TGs) containing docosahexaenoic acid, were positively associated with high body mass index (BMI) in under- or normal weight IgAN patients, while several free fatty acids and sphingomyelins were positively associated with high BMI in overweight or obese IgAN patients. Further, our study suggested that elevated levels of eight lipids, mainly TG species containing linolenic acid, were independent risk factors for IgAN progression and also reported the prospective association of circulating lipids with treatment outcomes in IgAN. Taken together, our findings may not only help to achieve precision medicine but also provide a knowledge base for dietary intervention in the treatment of IgAN.

Low-density lipoprotein receptor-related protein 1 is an essential receptor for trichosanthin in 2 choriocarcinoma cell lines.

Type-I ribosome-inactivating protein-trichosanthin (TCS) exhibits selective cytotoxicity toward different types of cells. It is believed that the cytotoxicity results from the inhibition of ribosomes to decrease protein synthesis, thereby indicating that there are specific mechanisms for TCS entry into target cells to reach the ribosomes. Low-density lipoprotein (LDL) receptor-related protein 1 (LRP1) is a large scavenger receptor that is responsible for the binding and endocytosis of diverse biological ligands on the cell surface. In this study, we demonstrated that 2 choriocarcinoma cell lines can significantly bind and internalize TCS. In contrast, Hela cell line displayed no obvious TCS binding and endocytosis. Furthermore LRP1 gene silencing in JAR and BeWo cell lines blocked TCS binding; TCS could also interact with LRP1.The results of our study established that LRP1 was a major receptor for phagocytosis of TCS in JAR and BeWo cell lines and might be the molecular basis of TCS abortificient and anti-choriocarcinoma activity.

The Efficacy and Mechanism of Chinese Herbal Medicine on Diabetic Kidney Disease.

Diabetic kidney disease (DKD) is the most common microvascular complication of diabetes and is one of the main causes of end-stage renal disease (ESRD) in many countries. The pathological features of DKD are the hypertrophy of mesangial cells, apoptosis of podocytes, glomerular basement membrane (GBM) thickening, accumulation of extracellular matrix (ECM), glomerular sclerosis, and tubulointerstitial fibrosis. The etiology of DKD is very complicated and many factors are involved, such as genetic factors, hyperglycemia, hypertension, hyperlipidemia, abnormalities of renal hemodynamics, and metabolism of vasoactive substances. Although some achievements have been made in the exploration of the pathogenesis of DKD, the currently available clinical treatment methods are still not completely effective in preventing the progress of DKD to ESRD. CHM composed of natural products has traditionally been used for symptom relief, which may offer new insights into therapeutic development of DKD. We will summarize the progress of Chinese herbal medicine (CHM) in the treatment of DKD from two aspects. In clinical trials, the Chinese herbal formulas were efficacy and safety confirmed by the randomized controlled trials. In terms of experimental research, studies provided evidence for the efficacy of CHM from the perspectives of balancing metabolic disorders, reducing inflammatory response and oxidative stress, antifibrosis, protecting renal innate cells, and regulating microRNA and metabolism. CHM consisting of different ingredients may play a role in synergistic interactions and multiple target points in the treatment of DKD.

Research on ribosome-inactivating proteins from angiospermae to gymnospermae and cryptogamia.

Ribosome-inactivating Proteins (RIPs) are a group of cytotoxin proteins that usually contain a RNA N-glycosidase domain, which irreversibly inactivates ribosome, thus inhibiting protein synthesis. During the past 14 years (1990-2004), the studies conducted in our laboratory had been focusing on the structure and enzymatic mechanism of several PIPs. Herein, we briefly described a summary of the studies conducted mainly in our laboratory on RIPs from angiospermae to gymnospermae and cryptogamia as follows. (1) Cinnamomin is a novel type II RIP isolated from mature seeds of camphor tree. Like ricin, it specifically removes the adenine at A4324 in rat liver 28S rRNA. We systematically studied this low-toxic RIP in term of its enzymatic mechanism, the primary and crystal structure and the nucleotide sequence of its gene, the genetic expression, and its physiological role in the seed cell and the toxicity to human cancer cells and insect larvae. The cleavage of supercoiled double-stranded DNA was its intrinsic property of cinnamomin A-chain, its N- and C-terminal regions were found to be required for deadenylation of rRNA and also necessary for deadenylation of supercoiled double-stranded circular DNA. These results strongly excluded the possibility that cleavage of supercoiled DNA was due to nuclease contamination. (2) Trichosanthin, an abortifacient protein, was purified from the Chinese medicinal herb, Tian-hua-fen, obtained from root tubers of Chinese trichosanthes plant. We proved that trichosanthin was a RNA N-glycosidase, inactivating eukaryotic ribosome by hydrolyzing the N-C glycosidic bond of the adenose at site 4324 in rat 28S rRNA, and inhibited protein synthesis in vitro. (3) A unique Biota orientalis RNase (RNase Bo) was extracted from the mature seeds of the cypress cypress tree (Oriental arborvita), which was gymnospermae plant. It cleaved only a specific phosphodiester bond between C4453 and A4454 of 28S RNA in rat ribosomes, producing a small RNA-fragment (S-fragment), thus inhibiting protein synthesis and belonging to RNase-like RIP, similar to α-sarcin, a special RIP. (4) Lamjapin, the first RIP purified from kelp, the marine cryptogamia algal plant, was shown to be the first single-chained RNA N-glycosidase from marine plant to date. It hydrolyzed rat ribosomal 28S RNA to produce meanly a rather smaller RNA, shorter than the diagnostic R-fragment under the restricted condition. The significance of existence of type I RIP in the lower marine algal plant was briefly discussed.

The three-dimensional structure and X-ray sequence reveal that trichomaglin is a novel S-like ribonuclease.

Trichomaglin is a protein isolated from root tuber of the plant Maganlin (Trichosanthes Lepiniate, Cucurbitaceae). The crystal structure of trichomaglin has been determined by multiple-isomorphous replacement and refined at 2.2 A resolution. The X-ray sequence was established, based on electron density combined with the experimentally determined N-terminal sequence, and the sequence information derived from mass spectroscopic analysis. X-ray sequence-based homolog search and the three-dimensional structure reveal that trichomaglin is a novel S-like RNase, which was confirmed by biological assay. Trichomaglin molecule contains an additional beta sheet in the HV(b) region, compared with the known plant RNase structures. Fourteen cystein residues form seven disulfide bridges, more than those in the other known structures of S- and S-like RNases. His43 and His105 are expected to be the catalytic acid and base, respectively. Four hydrosulfate ions are bound in the active site pocket, three of them mimicking the substrate binding sites.

Both N- and C-terminal regions are essential for cinnamomin A-chain to deadenylate ribosomal RNA and supercoiled double-stranded DNA.

Cinnamomin is a type II ribosome-inactivating protein and its A-chain exhibits RNA N-glycosidase activity to remove an adenine in the conserved sarcin/ricin loop of the largest RNA in ribosome, arresting protein synthesis at the elongation step. In this report, deadenylation of both rRNA and supercoiled DNA by native and recombinant cinnamomin A-chain expressed in Escherichia coli was demonstrated. However, the mutants of cinnamomin A-chain devoid of N-terminal 52 or/and C-terminal 51 amino acid residues lost both the activity of RNA N-glycosidase and the ability to release adenines from supercoiled DNA. Additionally, supercoiled DNA could not be cleaved into nicked and linear forms by these mutants. These results indicate that both N- and C-terminal regions are essential for the cinnamomin A-chain to deadenylate rRNA and supercoiled DNA. It was suggested that phosphodiester bonds in the extensively deadenylated region of supercoiled DNA would become fragile and liable to be broken spontaneously owing to the existence of tension in the supercoiled DNA.

Fluorescence labeling of short RNA by oxidation at the 3'-end.

In RNA nanotechnology, construction of nanoparticles involves conjugation of functionalities, cross-linking of modules, labeling of RNA subunits, and chemical modification of nucleotides. Efficiency and sensitivity are important for the RNA labeling, which also can be used as probes in microarrays, Northern blotting, and gel-shift assays. Here, we describe a method for fluorescence labeling of short RNA at the 3'-end by oxidation. The 3'-terminus of in vitro-transcribed short RNA is oxidized by sodium periodate, and fluorescein-5-thiosemicarbazide is added after removal of excess oxidant. Purified short RNA with fluorescence is then applied for detection of RNA-protein interaction by gel-shift assay.

Cinnamomin: a multifunctional type II ribosome-inactivating protein.

Plant ribosome-inactivating proteins (RIPs) are a group of toxic proteins that can irreversibly inactivate ribosomes by specifically removing the conserved adenine base from the "Sarcin/Ricin domain" of the 28S RNA in ribosome. Cinnamomin is a novel type II RIP isolated in our laboratory from the mature seeds of camphor tree. Besides site-specific deadenylation of the A4324 in the Sarcin/Ricin domain of rat ribosome, this protein could also release the adenine base from DNA molecules at multiple sites and from AMP, ADP, dAMP and adenosine. Furthermore, cinnamomin displays cytotoxicity to carcinoma cells and insect larvae by modifying their ribosomal RNA. These functions possessed by cinnamomin shed a new light on the possible application of cinnamomin in the field of immunotoxin design and transgenic reagents. In this review, we introduce the major recent results on cinnamomin obtained in our laboratory, including purification of this protein, characterization of its enzymatic mechanism, structure and function, gene pattern, physiological role and its biological implications in cytotoxicity.

Simultaneous existence of cinnamomin (a type II RIP) and small amount of its free A- and B-chain in mature seeds of camphor tree.

Cinnamomin, a type II ribosome-inactivating protein (RIP), was isolated from the mature seeds of camphor tree (Cinnamomum camphora). In this paper, small amount of free A- and B-chain of cinnamomin were found to be present in the mature seed cell of C. camphora besides the intact cinnamomin. Our results demonstrated that camphorin, a type I RIP previously reported to coexist with cinnamomin in the seeds of C. camphora, actually was the A-chain of cinnamomin. The percentage of free A- and B-chain in the total cinnamomin was 2.6-2.8% in the seed extract. Of these free A- and B-chain approximate 80% already existed in the seed cell, only about 20% were produced during the purification operation. As the enzymatic activity to reduce disulfide bond of cinnamomin in the seed extract of C. camphora was detected, we proposed that the free A- and B-chain were derived from the enzymatic reduction of the interchain disulfide bond of cinnamomin. It was demonstrated that the endogenous type II RIPs of several plant species, such as Cinnamomum porrectum, Cinnamomum bodinieri and Ricinus communis, could be enzymatically reduced into the free A- and B-chain in their respective seed cells. The function of the free A-chain in the seed cell and the possibility that metabolic enzymes might be involved in the reduction of the interchain disulfide bond of type II RIPs in vivo are discussed.

In vitro interaction of eukaryotic elongation factor 2 with synthetic oligoribonucleotide that mimics GTPase domain of rat 28S ribosomal RNA.

Eukaryotic elongation factor 2 (eEF2) catalyzed the translocation of peptidyl-tRNA from the ribosomal A site to the P site. In this paper, the interaction between eEF2 and GTD RNA, a synthetic oligoribonucleotide that mimicked the GTPase domain of rat 28S ribosomal RNA, was studied in vitro. The purified eEF2 could bind to GTD RNA, forming a stable complex. Transfer RNA competed with GTD RNA in binding to eEF2, whereas poly(A), poly(U) and poly(I, C) did not interfere with the interaction between eEF2 and GTD RNA, demonstrating that the tertiary structure of RNA might be necessary for the recognition of and binding to eEF2. The complex formation of eEF2 with GTD RNA was inhibited by SRD RNA, a synthetic oligoribonucleotide mimic of Sarcin/Ricin domain RNA of rat 28S RNA. Similarly, GTD RNA inhibited the interaction between eEF2 and SRD RNA. This fact implies that these small oligoribonucleotides probably share similar recognition or binding identity elements in their tertiary structures. In addition, the binding of eEF2 to GTD RNA could be obviously weakened by the ADP-ribosylation of eEF2 with diphtheria toxin. These results indicate that eEF2 behaves differently from prokaryotic EF-G in binding to ribosomal RNA.

Bioactivities of ricin retained and its immunoreactivity to anti-ricin polyclonal antibodies alleviated through pegylation.

Ricin has long been employed to construct immunotoxins, whose efficacy was often undermined by immunogenicity. Pegylation (modification of proteins with polyethylene glycol, PEG) was one of those recently developed approaches to circumvent immunogenicity of legions of drugs. Herein, pegylation of ricin was found to have barely changed the RNA N-glycosidase activity and protein synthesis inhibiting activity of ricin, but remarkably altered the cytotoxicity of ricin on hepatoma cell line (BEL7404) or the immunoreactivity with polyclonal anti-ricin antibodies. This result suggested that the attached PEG or monomethyloxyl polyethylene glycol (mPEG) groups did not hinder ricin from hydrolyzing ribosomal RNA, but indeed covered some areas on the surface of ricin molecule, including those involved in the interaction with cellular receptors and epitopes recognized by polyclonal antibodies. Pegylation, masking certain epitopes of ricin, might contribute to alleviate the immunogenicity of the toxin. Approach in this work, if applied to thereby constructed immunotoxins, would help improve the prospective efficacy of these toxins.