Viscum album : from high dilutions to phytotherapy

Background : Besides conventional anticancer therapy, cancer patients use complementary and alternative medicine like the medicinal herb mistletoe (Viscum album L.), used in oral form or injections. For this condition Viscum album (VA) is also used in high dilutions, in injections, as in Brazil. Aim: Our aim is to compare results of basic research about these two presentations: herbal medicine and high dilutions and to look for an eventual continuity of action between the molecular form (herbal medicine) and the informative one (high dilutions) Method: About VA in herbal medicine : a lot of references in immunology, in particular those published by the team of Srini Kaveri and Stephan Baumgartner.(1) (2) (3) (4) (5) About VA in high dilutions : the work in immunology of Aloiso Cunha de Carvalho and Leoni Bonamin published in a thesis. (6) And then to compare those results with our practice as veterinarian on dogs and cats . Results : VA in herbal medecine: from basic research results, we know that in low concentration, an immunostimulary action is observed while in higher doses it is a cytotoxic activity. These two properties are found in our practical use of VA in dogs or cats : stabilization of the size of tumors for the low concentrations, necrosis of tumors and abscess for the high concentrations. VA in high dilutions (from D3 to D30) : from basic research, an immunostimutating effect is observed. More exactly, on a murin model of tumor of Ehrlich, it is observed an improvement of quality of life, a reduction of the tumor immunosuppressive effect, an enlargement of local inflammations (Oedema) for all concentrations, but only 3D reduces tumor growth, without modifying local necrosis and the immunological parameters. In our practice, on two cases of osteosarcoma on dogs, we verify those properties using VA both in herbal medicine and in high dilutions. Conclusions: From these results, both in basic research and in the practice, we should estimate there is probably a continuity of action fom VA, in High Dynamised Dilutions ( no more molecule), to VA in herbal medicine (molecular activity). From immunostimation to cytotoxicity, from information to molecular activity. Does each Viscum album’s preparation touch the same level of the Immune System ? The same cells ? The same receptors ? A lot of questions remain… References 1) Duong Van Huyen JP and al. Variable sensitivity of lymphoblastoid cells to apoptosis induced by Viscum album Qu FrF, a therapeutic preparation of mistletoe lectin. Chemotherapy. (2001) 2) Duong Van Huyen JP and al. Induction of apoptosis of endothelial cells by Viscum album: a role for anti-tumoral properties of mistletoe lectins. Mol Med. (2002) 3)Duong Van Huyen JP and al Comparative study of the sensitivity of lymphoblastoid and transformed monocytic cell lines to the cytotoxic effects of Viscum album extracts of different origin.. Chemotherapy. (2003) 4) Estko M and al. Tumour cell derived effects on monocyte/macrophage polarization and function and modulatory potential of Viscum album lipophilic extract in vitro. BMC Complement Altern Med. 2015 5) Saha C, and al Differential Effects of Viscum album Preparations on the Maturation and Activation of Human Dendritic Cells and CD4⁺ T Cell Responses. Molecules. 2016 Jul 14;21(7) 6) Aloisio Cunha de Carvalho , ATIVIDADE ANTI-NEOPLÁSICA DE Viscum album (L) EM TUMORES EXPERIMENTAIS: REVISÃO CRÍTICA E ESTUDO EXPERIMENTAL EM TUMOR DE EHRLICH, Thesis Sao Paulo

Characterization of Sialic Acid Affinity of the Binding Domain of Mistletoe Lectin Isoform One

Sialic acid (Sia) is considered as one of the most important biomolecules of life since its derivatives and terminal orientations on cell membranes and macromolecules play a major role in many biological and pathological processes. To date, there is only a limited number of active molecules that can selectively bind to Sia and this limitation has made the study of this glycan challenging. The lectin superfamily is a well-known family of glycan binding proteins, which encompasses many strong glycan binding peptides with diverse glycan affinities. Mistletoe lectin (ML) is considered one of the most active members of lectin family which was initially classified in early studies as a galactose binding lectin; more recent studies have suggested that the peptide can also actively bind to Sia. However, the details with respect to Sia binding of ML and the domain responsible for this binding are left unanswered because no comprehensive studies have been instigated. In this study, we sought to identify the binding domain responsible for the sialic acid affinity of mistletoe lectin isoform I (MLI) in comparison to the binding activity of elderberry lectin isoform I (SNA), which has long been identified as a potent Sia binding lectin. In order to execute this, we performed computational carbohydrate-protein docking for MLB and SNA with Neu5Ac and β-Galactose. We further analyzed the coding sequence of both lectins and identified their glycan binding domains, which were later cloned upstream and downstream to green fluorescent protein (GFP) and expressed in Escherichia coli (E. coli). Finally, the glycan affinity of the expressed fusion proteins was assessed by using different biochemical and cell-based assays and the Sia binding domains were identified.

Therapeutic Potential of Medicinal Plant Proteins: Present Status and Future Perspectives

Biologically active molecules obtained from plant sources, mostly including secondary metabolites, have been considered to be of immense value with respect to the treatment of various human diseases. However, some inevitable limitations associated with these secondary metabolites like high cytotoxicity, low bioavailability, poor absorption, low abundance, improper metabolism, etc., have forced the scientific community to explore medicinal plants for alternate biologically active molecules. In this context, therapeutically active proteins/peptides from medicinal plants have been promoted as a promising therapeutic intervention for various human diseases. A large number of proteins isolated from the medicinal plants have been shown to exhibit anti-microbial, anti-oxidant, anti-HIV, anticancerous, ribosome-inactivating and neuro-modulatory activities. Moreover, with advanced technological developments in the medicinal plant research, medicinal plant proteins such as Bowman-Birk protease inhibitor and Mistletoe Lectin-I are presently under clinical trials against prostate cancer, oral carcinomas and malignant melanoma. Despite these developments and proteins being potential drug candidates, to date, not a single systematic review article has documented the therapeutical potential of the available biologically active medicinal plant proteome. The present article was therefore designed to describe the current status of the therapeutically active medicinal plant proteins/peptides vis-à-vis their potential as future protein-based drugs for various human diseases. Future insights in this direction have also been highlighted.

Transiently Expressed Mistletoe Lectin II in Nicotiana benthamiana Demonstrates Anticancer Activity In Vitro

Mistletoe (Viscum album) extracts have been used as alternative and complementary therapeutic preparations in multiple cancers for decades. Mistletoe lectins (ML-I, ML-II, and ML-III) are considered to be the main anticancer components of such preparations. In the present study, ML-II was transiently expressed in Nicotiana benthamiana using the pEAQ-HT expression system. Expression levels of up to 60 mg/kg of the infiltrated plant tissue were obtained, and a three-fold increase was achieved by adding the endoplasmic reticulum (ER) retention signal KDEL to the native ML-II sequence. The native protein containing His-tag and KDEL was purified by immobilized metal affinity chromatography (IMAC) and gel filtration. We found that the recombinant ML-II lectin was glycosylated and retained its carbohydrate-binding activity. In addition, we demonstrated that plant produced ML-II displayed anticancer activity in vitro, inhibiting non-small cell lung cancer H460 and A549 cells with EC50 values of 4 and 3.5 µg/mL, respectively. Annexin V-448A and PI double staining revealed that cell cytotoxicity occurred via apoptosis induction. These results indicate that ML-II transiently expressed in N. benthamiana plants is a promising candidate as an anticancer agent, although further optimization of production and purification methods is required to enable further in vitro testing, as well as in vivo assays.

Alterations in IL-6/STAT3 Signaling by Korean Mistletoe Lectin Regulate the Self-Renewal Activity of Placenta-Derived Mesenchymal Stem Cells

Korean mistletoe (Viscum album L. var. coloratum) lectin (VCA) is known as an anticancer drug. However, it is not clear whether VCA affects the self-renewal activity of mesenchymal stem cells (MSCs). Therefore, the objectives of this study were to analyze the effect of VCA on the proliferation of MSCs and expression of stemness markers. We also evaluated the usefulness of placenta-derived MSCs (PD-MSCs) as a screening tool. VCA was stably administered to MSCs, and analyzed self-renewal activities. The effect of IL-6 signaling on MSC proliferation was explored by quantitative methylation-specific PCR (qMSP) and western blot analysis. Compared with the control condition, low concentrations of VCA (10 pg/mL) induced an increase in the self-renewal activity of MSCs. Interestingly, a low concentration of VCA promoted IL-6 signaling in PD-MSCs through altered IL-6/STAT3 gene methylation. Furthermore, inhibition of IL-6 expression in PD-MSCs using an anti-IL-6 antibody caused a decrease in their self-renewal activity through IL-6/STAT3 signaling by altering IL-6/STAT3 gene methylation. These findings provide helpful data for understanding the mechanism of MSC self-renewal via VCA and show that VCA may be useful as a functional natural product for developing efficient therapies using placenta-derived stem cells.

Mistletoe-Based Drugs Work in Synergy with Radio-Chemotherapy in the Treatment of Glioma In Vitro and In Vivo in Glioblastoma Bearing Mice

Background. Extracts from Viscum album L. (VE) are used in the complementary cancer therapy in Europe for decades. VE contain several compounds like the mistletoe lectins (MLs) 1-3 and viscotoxins and also several minor ingredients. Since mistletoe lectin 1 (ML-1) has been described as the main component of VE harboring antitumor activity, purified native or recombinant ML-1 has been recently used in clinical trials. MLs stimulate the immune system, induce cytotoxicity, are able to modify the expression of cancer-associated genes, and influence the proliferation and motility of tumor cells. Objective. In this study our goal was to determine anticancer effects of the VE ISCADOR Qu, of recombinant ML-1 (Aviscumine), and of native ML-1 in the treatment of glioblastoma (GBM), the most common and highly malignant brain tumor in adults. Additionally we were interested whether these drugs, used in combination with a temozolomide-(TMZ)-based radio-chemotherapy, provide synergistic effects. Methods. Cell culture assays, ex vivo murine hippocampal brain slice cultures, human GBM cryosections, and a xenograft orthotopic glioblastoma mouse model were used. Results. In cells, the expression of the ML receptor CD75s, which is also expressed in GBM specimen, but not in normal brain, correlates with the drug-induced cytotoxicity. In GBM cells, the drugs induce cell death in a concentration-dependent manner and reduce cell growth by inducing cell cycle arrest in the G2/M phase. The cell cycle arrest was paralleled by modifications in the expression of cell cycle regulating genes. ML containing drugs, if combined with glioma standard therapy, provide synergistic and additive anticancer effects. Despite not reaching statistical significance, a single intratumoral application of Aviscumine prolonged the median survival of GBM mice longer than tumor irradiation. Moreover, intratumorally applied Aviscumine prolonged the survival of GBM-bearing mice if used in combination with irradiation and TMZ for further 6.5 days compared to the radio-chemotherapy. Conclusion. Our results suggest that an adjuvant treatment of glioma patients with ML-containing drugs might be beneficial.