Oral cancer nano vaccine and structuring method thereof

Abstract

The invention provides an oral cancer nano vaccine and a structuring method thereof. The vaccine comprises a water-in-oil type nano-emulsion cancer vaccine and tomato lectin which is modified on the surface of the water-in-oil type nano-emulsion cancer vaccine through a cross-linking agent of phosphatidyl ethanolamine. The structuring method comprises the steps of: 1, preparing the water-in-oil type nano-emulsion cancer vaccine; 2, synthesizing tomato lectin-phosphatidyl ethanolamine; and 3, modifying the surface of the nano-emulsion cancer vaccine. In the oral cancer nano vaccine provided by the invention, the tomato lectin is combined with intestinal tract cells M in a specific targeting manner, which is favorable for effectively improving vaccine absorptivity and activating mucous membrane and whole body dual immune; a nano emulsion is used as a vector for the vaccine so as to ensure that the vaccine effectively passes through an enzymatic barrier and a membrane barrier, and improve the bioavailability of the oral vaccine; and a superantigen SEA component is added into a cancer antigen, which is favorable for effectively activating cells CD4+T at the same time of activating cells CD8+T, and the superantigen SEA component and the cancer antigen take effect synergistically to enhance the immune effect of the cancer antigen.

Description

technical field

[0001] The invention belongs to the technical field of tumor immunotherapy, and in particular relates to an oral tumor nano vaccine and a construction method thereof. Background technique

[0002] The development of tumor vaccines has a history of more than 100 years, and a variety of vaccines have been clinically tested and achieved encouraging results. However, we should also note that the results of some vaccines that are very effective in animal experiments are disappointing in human trials. An important reason is that after inoculation using conventional routes (muscle injection, intradermal inoculation), lymphoid tissue uptake The amount of antigen is limited, so that the immune efficacy of the vaccine cannot be fully exerted. In contrast, the intestinal mucosa, which is twice the body surface area, contains extremely rich lymphoid tissue, and the vaccine can activate both mucosal and systemic immunity after oral administration, and has the characteris...

Examples

Embodiment 1

[0031] (1) Preparation of water-in-oil nanoemulsion tumor vaccine

[0032] Mix 1.0 mg of composite antigen (MAGE1 / HSP70 / MAGE3 and SEA at a molar ratio of 100:1), Span-20, Pluronic188 and 0.6 mL of soybean oil, adjust the total volume to 2.5 mL with double distilled water, and mix well to obtain oil Then, under 3000rpm magnetic stirring, the oil phase was added dropwise to 7.5mL of double-distilled water and water phase, and then moved to a vacuum high-speed shear emulsifier, under 0.7KPa vacuum, 23000 rpm under high-speed shear for 40min , the temperature was controlled below 80°C; then moved to an ice bath and ultrasonicated at 40KHz for 5 minutes, repeated ultrasonication three times, dialyzed to remove free protein, and sterilized by filtration with a 0.22μm filter membrane, and stored at 4°C for later use to obtain water-in-oil nano Emulsion tumor vaccine; the mass percentage concentration of Span-20 in the oil phase is 8%, and the mass percentage concentration of Pluronic...

Embodiment 2

[0038] (1) Preparation of water-in-oil nanoemulsion tumor vaccine

[0039]Mix 0.5 mg of composite antigen (MAGE1 / HSP70 / MAGE3 and SEA at a molar ratio of 50:1), Span-20, Pluronic188 and 0.6 mL of soybean oil, adjust the total volume to 1 mL with double distilled water, and mix well to obtain an oil phase ; then under 3000rpm magnetic stirring, the oil phase was added dropwise to the 3mL double-distilled water phase, and then moved to the vacuum high-speed shear emulsifier, under 1KPa vacuum, high-speed shear 50min under the condition of 20000 rpm, temperature control Below 80°C; then move to an ice bath for 30KHz ultrasound for 8 minutes, repeat the ultrasound three times, dialyze to remove free protein, filter and sterilize with a 0.22μm filter membrane, store at 4°C for later use, and obtain a water-in-oil nanoemulsion tumor vaccine ; The mass percentage concentration of Span-20 in the oil phase is 5%, and the mass percentage concentration of Pluronic188 is 20%;

[0040] (2)...

Embodiment 3

[0044] (1) Preparation of water-in-oil nanoemulsion tumor vaccine

[0045] Mix 1.5 mg of composite antigen (MAGE1 / HSP70 / MAGE3 and SEA at a molar ratio of 150:1), Span-20, Pluronic188 and 0.6 mL of soybean oil, adjust the total volume to 4 mL with double distilled water, and mix well to obtain an oil phase Then under 3000rpm magnetic stirring, the oil phase is added dropwise in the 12mL double-distilled water phase, and then moved to the vacuum high-speed shear emulsifier, under 0.5KPa vacuum, high-speed shear 30min under 25000 rpm conditions, temperature Control the temperature below 80°C; then move to an ice bath for 50KHz ultrasound for 3 minutes, repeat the ultrasound three times, dialyze to remove free protein, filter and sterilize with a 0.22μm filter membrane, store at 4°C for later use, and obtain a water-in-oil nanoemulsion tumor Vaccine; the mass percent concentration of Span-20 in the oil phase is 10%, and the mass percent concentration of Pluronic188 is 15%;

[004...

technical field

[0001] The invention belongs to the technical field of tumor immunotherapy, and in particular relates to an oral tumor nano vaccine and a construction method thereof. Background technique

[0002] The development of tumor vaccines has a history of more than 100 years, and a variety of vaccines have been clinically tested and achieved encouraging results. However, we should also note that the results of some vaccines that are very effective in animal experiments are disappointing in human trials. An important reason is that after inoculation using conventional routes (muscle injection, intradermal inoculation), lymphoid tissue uptake The amount of antigen is limited, so that the immune efficacy of the vaccine cannot be fully exerted. In contrast, the intestinal mucosa, which is twice the body surface area, contains extremely rich lymphoid tissue, and the vaccine can activate both mucosal and systemic immunity after oral administration, and has the characteris...

Examples

Embodiment 1

[0031] (1) Preparation of water-in-oil nanoemulsion tumor vaccine

[0032] Mix 1.0 mg of composite antigen (MAGE1 / HSP70 / MAGE3 and SEA at a molar ratio of 100:1), Span-20, Pluronic188 and 0.6 mL of soybean oil, adjust the total volume to 2.5 mL with double distilled water, and mix well to obtain oil Then, under 3000rpm magnetic stirring, the oil phase was added dropwise to 7.5mL of double-distilled water and water phase, and then moved to a vacuum high-speed shear emulsifier, under 0.7KPa vacuum, 23000 rpm under high-speed shear for 40min , the temperature was controlled below 80°C; then moved to an ice bath and ultrasonicated at 40KHz for 5 minutes, repeated ultrasonication three times, dialyzed to remove free protein, and sterilized by filtration with a 0.22μm filter membrane, and stored at 4°C for later use to obtain water-in-oil nano Emulsion tumor vaccine; the mass percentage concentration of Span-20 in the oil phase is 8%, and the mass percentage concentration of Pluronic...

Embodiment 2

[0038] (1) Preparation of water-in-oil nanoemulsion tumor vaccine

[0039]Mix 0.5 mg of composite antigen (MAGE1 / HSP70 / MAGE3 and SEA at a molar ratio of 50:1), Span-20, Pluronic188 and 0.6 mL of soybean oil, adjust the total volume to 1 mL with double distilled water, and mix well to obtain an oil phase ; then under 3000rpm magnetic stirring, the oil phase was added dropwise to the 3mL double-distilled water phase, and then moved to the vacuum high-speed shear emulsifier, under 1KPa vacuum, high-speed shear 50min under the condition of 20000 rpm, temperature control Below 80°C; then move to an ice bath for 30KHz ultrasound for 8 minutes, repeat the ultrasound three times, dialyze to remove free protein, filter and sterilize with a 0.22μm filter membrane, store at 4°C for later use, and obtain a water-in-oil nanoemulsion tumor vaccine ; The mass percentage concentration of Span-20 in the oil phase is 5%, and the mass percentage concentration of Pluronic188 is 20%;

[0040] (2)...

Embodiment 3

[0044] (1) Preparation of water-in-oil nanoemulsion tumor vaccine

[0045] Mix 1.5 mg of composite antigen (MAGE1 / HSP70 / MAGE3 and SEA at a molar ratio of 150:1), Span-20, Pluronic188 and 0.6 mL of soybean oil, adjust the total volume to 4 mL with double distilled water, and mix well to obtain an oil phase Then under 3000rpm magnetic stirring, the oil phase is added dropwise in the 12mL double-distilled water phase, and then moved to the vacuum high-speed shear emulsifier, under 0.5KPa vacuum, high-speed shear 30min under 25000 rpm conditions, temperature Control the temperature below 80°C; then move to an ice bath for 50KHz ultrasound for 3 minutes, repeat the ultrasound three times, dialyze to remove free protein, filter and sterilize with a 0.22μm filter membrane, store at 4°C for later use, and obtain a water-in-oil nanoemulsion tumor Vaccine; the mass percent concentration of Span-20 in the oil phase is 10%, and the mass percent concentration of Pluronic188 is 15%;

[004...

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