Table I.
Study design and outcomes (n=37) according to treatment, animal model, tumor cell line and main outcomes.
| Author/year | Treatments | Animal model | Tumor cell lines | Outcomes | (Refs.) |
|---|---|---|---|---|---|
| Han et al, 2015 | Endragit: Enteric-coated lectin from KM grown on poplar. Toxicity test: 500, 1,000, 2,000 mg/kg, single dose, p.o. Antitumor effect: Added to feed at 1, 2 or 4% concentration. | Toxicity test: ICR mice, both sexes; Antitumor activity: Male GDFI mice | Murine B16.F10 melanoma | POSITIVE Significant inhibition of tumor growth at 4% concentration. Increased survival rate (at 2 or 4%). Significant dose-and time-dependent reduction in tumor cell viability. Mechanisms: Cycle-cell deregulation, apoptosis. | (53) |
| Delebinski et al, 2015 | VA.M; TT (OA 4.0 mg/ml + BA 0.38 mg/ml); ML-I (3.7 µg/ml); TT/ML-I (qualitatively positive); 6 groups: (1) dose escalation 20/40/60 mg/kg TT; (2) 0.3/0.4/0.5 µg/kg ML-I; (3) TT/ML-I combination; VAE given twice every 3rd day, i.v., from day 3 after tumor inoculation; (4) ARA-C 100 mg, i.v., every 3rd day, 4 doses; (5) TT/ML-I + ARA-C; and (6) control, received equal amounts of cyclodextrins. | Female NOD/SCID/II-2rg mice in pathogen-free conditions | Human HL-60 cells | POSITIVE TT/ML-I combination significantly reduced tumor weight, comparable to ARA-C. The greatest effect was observed with TT/ML-I + ARA-C combination. | (25) |
| Facina et al, 2014 | VA-Qu AE (Iscador Qu; 5 mg with 0.375 µg/ml of lectins), 0.1 ml, s.c. peritumoral, daily for 10 days | Female C57BL/6N mice | B16F10 melanoma | MIXED No effect on tumor volume. Upregulation of apoptosis-related gene expression, particularly the extrinsic apoptotic pathway. Immunomodulatory Th-1-like action, with IL recruitment and cell death activation through caspase 7, associated upstream with Card-10 and downstream with CAD. | (44) |
| Stan et al, 2013 | VA-M, ethanolic extract, 50 mg/kg, i.p., days 1, 3, 6. DOXO 2.5 mg/kg, i.p., days 1 and 6; 7 groups (VA-M yes/no, DOXO, yes/no, EAC yes/no) | Swiss female mice | EAC | POSITIVE VA-M enhanced the antitumor effect of DOXO reflected in tumor development (body weight, tumor volume and cell concentration) and oxidative stress-related enzymes. | (58) |
| Strüh et al, 2013 | VA-M AE; STT; CD as control (2.2 g/kg). Treatments and controls injected s.c. peritumorally, every 2nd day starting at day 3 after tumor injection up to 10 cycles. Experiment 1: AE 12 µg/kg of ML-I; STT 12 µg/kg of ML-I and 93 mg/kg OA. Experiment 2: AE 3.5 µg/kg of ML-I; STT 3.5 µg/kg of ML-I and 71 mg/kg OA; TT 71 mg/kg OA. | Male C57BL/6NrL mice | Murine B6F10 melanoma cells | POSITIVE Both AE and STT significantly slowed down tumor growth and increased median survival. The effect of STT was greater, but highly toxic. Both AE and STT significantly increased tumor necrosis and reduced tumor angiogenesis. These effects remained with the lower dose, but were reduced; in turn, they were not associated with toxicity. TT alone had no significant effect on tumor growth. STT and TT alone induced greater tumor necrosis compared with AE. Active caspase-3 expression was weak in all the groups. No treatment had any ef fect on proliferation of viable melanoma cells (Ki-67 positive). | (26) |
| Podlech et al, 2012 | Iscador Qu; (1) 100 µg/ML, culture medium then 2 µg, intratumor, (2) 1 µg, s.c., periodically (NR), (3) 20 µl (100 µg/ML), intratumor, single dose. | Athymic CD1.deficient NMRI nude mice; VMDk mice | LNT-229; SMA560 glioma cells | MIXED Pretreatment reduced tumor growth, however this was not due to altered proliferation or cell death. Systemic treatment was associated with slight, non-significant inhibition of tumor growth, whereas intratumor administration had significant ef fect. | (52) |
| Li et al, 2011 | CM-1 of Chinese mistletoe grown on poplar. Treated groups: 10 µg.kg−1 CM-1 every 3rd day or 20 µg/kg−1 every 10 days, i.v. | Female athymic nude (Balb/c nu nu) mice | CRC CLY and HT-29 cells | POSITIVE CM-1-treated groups exhibited a dose-dependent reduction in tumor growth and high tumor growth inhibition rate. Downregulation of specific miRNA induced by CM-1 was not due to suppression of host gene transcription, but to direct degradation of their precursors. There was direct correlation between miR-135a&b levels and cellular sensitivity to CM-1. | (69) |
| Thies et al, 2008 | ML-I from leaves of mistletoe grown on poplar (40%), apple (30%) and red oak (30%) trees collected in the winter. ML stock solution with 963 µg/ML ML-I, ~1% ML-II and <0.5% ML-II. Treatment groups: 30, 150 and 500 ng/kg in 200 µl PBS, i.p., 19 days. | Pathogen-free male Balb/c SCID/SCID mice | Human melanoma MV3 | MIXED Primary tumor: Significant reduction of tumor growth with low dose, significant increase of tumor weight with intermediate vs. low dose. The apoptosis rate was significantly higher in all 3 treatment groups, significantly higher with low vs. intermediate and high doses. There was an increased number of infiltrating DC in all treatment groups, significant increase of DC apoptosis with intermediate and high doses vs. low dose; significant increase in the number of infiltrating DCs with low dose vs. controls. Lung metastases: Significant reduction in number with low dose vs. control. Mean number increased with intermediate and high dose vs. low dose. Vascular count increase with high dose vs. controls. | (33) |
| Seifert et al, 2008 | VA-A (Helixor), 160 µg/ml ML-II/III and 5.10% ML-I; VA-P, 725 µg/ml ML-II/III; treated groups: 1, 50, 100 mg/kg, i.p, 4 d/wk., 3 wks., 1 day after tumor inoculation. Controls: CPA 100 mg/kg on day 3. | Male CB-17 SCID/SCID mice; specific-pathogen free | Human ALL NALM.6 cells | POSITIVE Both VA extracts exhibited concentration-dependent inhibition of cell proliferation, due to apoptosis; even low concentrations of extracts induced dose. dependent induction of early apoptosis markers. All extracts at all doses increased survival; there were 3 long-term survivors, all treated with VA-A (2 from 50 mg and 1 from 100 mg groups). None of these animals exhibited leukemia or lymphoma at necropsy | (59) |
| Cebovic et al, 2008 | CO2 extract of leaves of mistletoe grown on plum collected in winter; 2.0 ml/kg/d, i.p., duration NR, several doses tested but NR | Male and female Hann: NMRI mice | EAC | POSITIVE Volatile extract includes TT. Reduction of tumor cell number and viability; reduction of antioxidative enzyme activity in tumor bearing animals. | (27) |
| Beuth et al, 2006 | VA-A (Helixor) 3.6 µg/ml ML-I, 354 µg/ml ML-III; VA-M, 230 µg/ml ML-I, 500 µg/ml ML-III. Treated groups: 5 mg/µl/injection, i.p., of VA-A or VA-M on days 5, 6, 7, 12, 13 and 14 | Male Balb/c mice | BT474 human ductal breast carcinoma | POSITIVE Both extracts induced a significant reduction in tumor weight; reduction of tumor cell proliferation rate; increased cell necrosis and apoptosis rates. | (68) |
| Duong Van Huyen et al, 2006 | Iscador Qu FrF, 20 µg/dose, once/day, i.p., 7 days starting from the day of tumor inoculation | C57BL/6 and B16.129-II2b (IL-12 deficient) mice | B16F1 melanoma | POSITIVE The chosen dose was 20 µg, which was an effective dose that did not promote lethality. Reduced tumor weight and restoration of proliferative capacity of splenocytes in tumor-bearing animals, associated with increased secretion of IL-12. | (67) |
| Pryme et al, 2004 | Pure Ml-I 10 mg/day as dietary supplement in different concentrations (2.5, 5.0, 10.0 mg ML-I/d) | Female NMRI mice | Krebs II NHL cells adapted to grow as intraperitoneal ascites tumor | POSITIVE Reduction in the number of mitotic figures; intra-tumor lymphoid infiltrate doubled, with a corresponding increase in tumor necrosis; accelerated cell turnover inside tumor, and increased number of apoptotic cells. 1 1% of treated animals no longer had histological evidence of viable tumors on day 1 1. | (70) |
| Yoon et al, 2003 | Aqueous extract (KM-110) and lectin (KML-C) of KM. KM-110 100 µg/animal; KML-C 0.05, 0.02, 0.005 µg/animal, i.v.; lung metastasis: 1 and 2 days before tumor inoculation; liver and spleen metastasis 1 day after tumor inoculation | Experimental lung metastasis: Balb/c; C57BL/6 mice Experimental liver and spleen metastasis: f CDF1 mice Spontaneous lung metastasis: C57BL/6 mice | Experimental lung metastasis: colon26-M3-1 carcinoma; B16.BL6 melanoma, 2.5×104 Experimental liver and spleen metastasis: L51784.ML25 lymphoma cells, 4×104 Spontaneous lung metastasis: B16.BL6 melanoma, 5×105 | POSITIVE KML-C prophylactically and therapeutically inhibited tumor metastasis of both lung metastatic tumor lines and reduced tumor metastasis of hematogenous tumor cells. KML-C 5.0 ng/animal, 5 days after inoculation, 5 times at 3.day intervals significantly inhibited spontaneous lung metastasis. KML-C 550 ng/mouse increased NK cell activity. KML-c at various doses given 2 days before inoculation increased macrophage-mediated cytotoxicity anti B16.BL6. | (66) |
| Braun et al, 2002 | sME Iscador Spezial, 2, 20, 100 µg/animal, s.c., 3 times per wk., 24 h after tumor inoculation | Inbred male Balb/c mice | RAW-117 P (liver); L-1 (lung) lymphosarcoma | POSITIVE Dose-dependent reduction in liver and lung tumor colonies and inhibition of local tumor growth (weight and volume) in all treated groups. Thymocytes upregulation; increased number of leukocytes, lymphocytes and monocytes in peripheral blood. | (49) |
| Elsässer-Beile et al, 2001 | rML 300 or 1,500 µg/ml in solution (EDTA, NaCl, PBS, PVP); 0.1 ml per intravesical catheter. Control group: NMU on wks. 0,2,4,6 Treated groups: 30 or 150 ng on weeks 8 to 13 or 14 to 19 | Inbred female Fischer 344 rats | NMU-induced urinary bladder carcinoma | POSITIVE All 4 treated groups exhibited significant reduction of rates of atypical hyperplasia/neoplastic transformation compared to controls. No change in any of the assessed cytokines. | (22) |
| Braun et al, 2001 | Helixor ME-A 160 ng/ml, Helixor ME-P® 725 µg/ml of ML-II/III (both >95%), 5 and 50 µg/mouse, s.c., i.p., 3 times/wk, 24 h after tumor inoculation | Inbred male Balb/c mice | RAW-117 P (liver); L-1 (lung) lymphosarcoma | POSITIVE Dose-dependent reduction of liver and lung colonies; i.p. significantly greater effect. Dose-dependent increase of leukocyte, lymphocyte and monocyte count. | (48) |
| Schaffrath et al, 2001 | rML, 0.3, 3, 30, 150 ng/kg, s.c., every 2nd day, starting 2 days after tumor inoculation | Inbred male Balb/c mice | RAW-117 P (liver); L-1 (lung) lymphosarcoma | POSITIVE Increased survival rate; reduced number of liver and lung tumor colonies. Increased number of leukocytes in RA W-117 P | (23) |
| Burger et al, 2001 | Lektinol, 405 µg/ml of ml, from poplar, 0.3, 3, 30, 300 ng/kg/d, i.p. or s.c., once per day, 5 days/wk. for up to 4 wk.; positive controls: DOXO, 8 mg/kg, i.v., on days 0 and 14; 5.FU, 100 mg/kg, i.p., on days 0, 7, 14 | C57BL/6, Balb/c mice, both genders | B16.F10 melanoma, C8 38 colon carcinoma, F9 testicular carcinoma, Lewis lung carcinoma or Renca renal cell carcinoma. | MIXED Dose-dependent tumor growth inhibition (Renca, colon, testicle) borderline effect; greatest inhibitory effect in Renca model with dose 300 ng, however, the sensitivity of the 3 tumor types to VA was similar. In Renca model VA as effective but less toxic than DOXO; colon carcinoma responded better to 5.FU (comparable toxicity). Lewis carcinoma and melanoma did not respond to VA. | (45) |
| Timoshenko et al, 2001 | ML-I 1 ng/kg dissolved in PBS and 50 µg/ml mouse albumin to concentration 0.22 ng/ml, 100 µl injection, s.c., days 7, 10, 13, 16, 19 after tumor inoculation; IL-2, i.p., 105 Cetus Units/ml, 100 µl injection, every 8 h, 5 days. Experiment 1: 2 rounds with 4.day interval. Experiment 2: 1 round. 5 groups: (1) healthy mice, (2) tumor inoculated (3) IL-2 alone (4) ML-I alone and (5) IL-2 + ML-I | Female C3He/Hej mice | C3L5 murine mammary adenocarcinoma, 1 × 106 | NEGATIVE ML-I alone increased primary tumor growth and development and growth of metastases. In combination, ML-I did not modify the tumor growth suppressive action of IL-2. ML-I did not cause capillary leakage neither inhibited the one caused by IL-2 and associated nitric oxide changes. | (47) |
| Park et al, 2001 | KML, 10, 30, 50 ng/animal; (1) Tumor growth: 10.50 ng/mouse, i.p., 3 times per wk. (2) Antimetastatic effect: 10.50 ng/mouse 2 days before and 1 day after tumor inoculation | Female C57BL/6 mice | Melanoma B16.BL6 | MIXED No effect on tumor growth; increased survival. Dose-dependent inhibition of tumor cell proliferation via apoptosis, dose-dependent anti-angiogenesis effect. No effect on cell cycle. | (46) |
| Zarkovic et al, 2001 | Isorel, 50 µl, every 2nd day, 3 consecutive wks., s.c. or s.c. contralateral to tumor (local vs. systemic) | Female CBA/HZgr mice | Murine mammary carcinoma | POSITIVE Inhibition of tumor growth with local administration; increased apoptosis and necrosis in both treated groups, reduction of mitosis with local treatment only, reduction of tumor invasiveness in both treated groups. Tumor infiltration without mononuclear cells in controls. | (65) |
| Mengs et al, 2000 | AE, 30 or 300 µg/ml ml, i.v., 3 days/wk., 4 consecutive wks., total volume 0.1 ml | Female C57BL/6J mice. | Urinary bladder carcinoma MB49 | POSITIVE Increased survival; disappearance of tumor-both effects statistically significant, non-dose-dependent. | (54) |
| Schumacher et al, 2000 | rML, 20, 150, 500 ng/kg, Monday to Friday until day 83 or 120% of initial body weight | SCID/SCID mice | SoTü 3 human ovarian cystoadenoma | POSITIVE Significantly longer survival with highest dose; symptom-free interval increased among animals that developed malignant ascites. | (24) |
| Kunze et al, 2000 | ML-I, 1 ng/kg, twice/week, s.c., 6 or 15 months | Pathogen-free female Wistar rats | BBN-induced bladder carcinoma | NEGATIVE No difference in tumor growth or spectrum of histological types of carcinomas. In the groups treated for 15 months, tumor size was 2x greater. No significant stimulation of any cell population in lamina propria. | (40) |
| Antony et al, 1999 | Iscador M: (1) For spleen cell activation, 1.66 mg/dose/animal, i.p., 5 days. (2) For treatment: 1.66 mg/dose, i.p., daily, 10 days | C57BL/6 mice | B16.F10 melanoma | POSITIVE Significant inhibition of lung metastasis (in vivo activated spleen cells), increased with concomitant treatment; similar effect in metastasis-bearing mice. In vitro activation of spleen cells. Increased survival of all treated groups (best ef fect: In vivo activation + treatment). Markers: Reduction of hydroxyproline, sialic acid and GGTP (cell proliferation). | (64) |
| Lenartz et al, 1998 | ML-1. (1) Systemic: 1, 10 ng/kg, s.c., 2 wk. (2) Local: 10, 100 ng, single dose, i.p. | Adult female Fischer 344 rats | F98 glioma | POSITIVE Reduction in tumor volume in all treated groups, only significant with lowest dose, both routes. Dose-dependent increased tumor necrosis with local administration only. | (50) |
| Yoon et al, 1998 | KM AE grown on oak. Survival: 100 µg single dose i.v. Metastasis inhibition: 100 µg i.v., 2 and 4 days before tumor inoculation. Dose/route test: 1.100 µg/dose, i.v., s.c., p.o., intranasal | Specific-pathogen free female C57BL/6, Balb/c, CDF1 mice | B16.BL6 melanoma; 26.M3.1 colon carcinoma; spleen and liver metastases of L5178Y.ML25 lymphoma | POSITIVE Administration 2 days before tumor inoculation significantly inhibited melanoma and colon carcinoma metastasis, 4 days before melanoma1 metastasis only. Similar pattern in lymphoma; systemic routes had dramatic effect even in low dose. Enhanced NK cell activity. Increased survival. | (51) |
| Weber et al, 1998 | Lektinol, 30 µg/ml of bioactive mls; 4 groups: (1) Vehicle, (2.4) 3, 30, 150 ng/kg ML, respectively, i.v., 1 hour after tumor injection then 4 days/wk., 3 wk. | Male Balb/c mice | B16.F10 melanoma | POSITIVE Dose-dependent inhibition of metastasis; reduction of melanoma cell count in lungs. In vitro: stimulation of macrophages, CD4+CD8 lymphocytes; significant enhancement of anti-CD8, no ef fect on anti-CD4. | (55) |
| Zarkovic et al, 1998 | Isorel M; pretreatment of tumor cells with whole extract, ML-1 and 2 fractions-high and low molecular weight, 0.1 µg/ml | Male C57BL; GoZgr mice | Melanoma | POSITIVE All tested preparations inhibited metastasis formation. | (21) |
| Kunze et al, 1998 | Lectin, 1 ng/kg, s.c., twice per wk., 15 months | Female Wistar rats; specific pathogen-free | MNU-induced bladder carcinoma | NO EFFECT The incidence of bladder carcinoma did not differ between treated and control groups. There was no dif ference in the population of inflammatory cells infiltrating the tumor. | (41) |
| Zarkovic et al, 1997 | Isorel M, 100 mg/kg, single dose, i.p. | Male C57BLGoZgr mice | B16F10 melanoma | POSITIVE Tumor growth reduced in treated animal, with a reduction of tumor viable cells. Tumor necrosis and abundant inflammatory response. In vitro tests: Strong stimulation of lymphocytes, with increased cytotoxic activity; increased sensitivity of tumor cells to cytotoxic activity of lymphocytes; inhibition of growth of tumor cells more than normal cells. | (63) |
| Jurin et al, 1997 | Isorel M; strength 60, 2 protocols: (1) Single dose, 1 day before, simultaneous, 1 day after sheep RBC injection; 140 or 1,400 mg/kg/animal. (2) Repeated doses: a) normal mice and tumor-bearing mice: 14 or 140 mg/kg, daily, 14 consecutive days; b) normal mice: 14, 140 or 140 mg/kg, daily, 5 wk. (25 doses); route NR | Male and female CBA/Hzgr mice | methylcholantrene-induced fibrosarcoma | IMMUNOMODULATING EFFECT 1) Isorel single dose before sheep RBC injection restored the immune response; 140 mg/kg had greatest effect. 2) Dose-dependent immunostimulating effect; prolonged application (5 wk.) had dose-dependent immunosuppressing effect on normal mice only. | (62) |
| Antony et al, 1997 | Iscador M, prophylactic, 5 doses, 1.66 mg/kg/dose, 4 groups: Control, before tumor inoculation, with tumor then 10 consecutive days, 5 days after tumor, 10 consecutive days | C57BL/6 mice | B16.F10 melanoma | POSITIVE Reduction of lung metastasis formation; increased survival rate of metastasis-bearing animals; lifespan increased in all treated groups. Markers: reduction of hydroxyproline (fibrosis), sialic acid (melanoma development). | (61) |
| Kunze et al, 1997 | Purified lectin, 50 µg/ml, 1 ng/kg, s.c., twice per wk., 7 h after tumor induction and then for 6 months | Pathogen-free adult, female Wistar rats | NMU-induced urinary bladder carcinoma | NO EFFECT No significant differences in tumor rates, characteristics or histology. | (42) |
| Kuttan et al, 1997 | 5% Iscador M, 1.66 mg/dose, route NR; 2 experiments: Carcinogenesis, twice/wk., 15 wks. Metastasis inhibition: 5 days before, simultaneous with and after tumor inoculation, 10 days in each group | Female Swiss albino and C57BL/6 mice | Methylcholantrene-induced sarcoma; B16.F10 melanoma for metastasis induction | POSITIVE Reduction of tumor size and development; reduction of metastasis formation; increased survival. Markers: Reduction of hydroxyproline and sialic acid. | (60) |
| Kuttan et al, 1996 | Iscador; (1) 1 mg/dose, i.p., twice/wk., 15 wks. (2) dilutions 0 to 1:3,000 (1.66 mg-1.66 µg), i.p., twice/wk., 15 wks. | Female Swiss albino mice | Methylcholantrene-induced sarcoma | POSITIVE Iscador fully inhibited sarcoma development; 100% of animals survived; effect was dose-dependent. | (56) |
KM, Korean mistletoe; p.o., oral route; VA-M, mistletoe grown on apple tree; TT, triterpenes; OA, oleanolic acid; BA, betulinic acid; ml-I, lectin-I; VAE, Viscum album extract; wk(s)., week(s); i.v., intravenous; ARA-C, cytarabine; NOD, non-obese diabetic; SCID, sever combined immunodeficiency; VA-Qu, mistletoe grown on oak tree; AE, aqueous extract; s.c., subcutaneous; Th, T-helper; IL, interleukin; CAD, caspase-activated DNase; i.p. intraperitoneal; DOXO, doxorubicin; EAC, Ehrlich ascites carcinoma; STT, soluble TT; CD, cyclodextrins; NR, not-reported; CM-1, Chinese mistletoe lectin >90% identical with European mistletoe ml-I; CRC, colorectal cancer; miR, microRNA; DC, dendritic cells; VA-A, mistletoe grown on fir; VA-P, mistletoe grown on pine; CPA, cyclophosphamide; ALL, acute lymphoblastic leukemia; NHL, non-Hodgkin lymphoma; FrF, extract prepared without fermentation; NK, natural killer cell; rML, recombinant lectin; 5-FU, 5-fluorouracil; BBN, N-butyl-N-(4-hydroxybutyl) nitrosamine; GGTP, gamma-glutamyl transpeptidase; PVP, polyvinylpyrrolidone; NMU; N-methyl-N-nitrosourea; RBC; red blood cells.