Kit of tabletted alive recombinant oral bivaccine against smallpox and hepatitis b and method for vaccination at applying the kit mentioned

FIELD: medicine, immunology.

SUBSTANCE: the present innovation deals with specific prophylaxis of smallpox and viral hepatitis B. The kit contains two tablets each contains stabilizing additives, a filler and lyophilized alive viral material worked out based upon recombinant VOV strain at typical VOV properties expressing proteins preS2-S and HBs virus of hepatitis B virus, the first immunizing dosage corresponds to minimal quantity of viral material being sufficient to obtain weak immune response in the body in case of insignificant at insignificant reactogenicity, and immunizing dosage of the second - maximal quantity of viral material that causes pronounced and prolong immune response in the body at no negative side action. The technique of applying the kit of bivaccine tablets, first, one should use the 1st tablet at minimal dosage of bivaccine, as for the 2nd tablet - with maximal dosage of bivaccine it should be taken till the moment of developing humoral answer (in 7-14 d) after injecting the 1st tablet at minimal immunizing dosage of bivaccine. The innovation enables to create stable immunity.

EFFECT: higher efficiency.

4 cl, 5 ex, 6 tbl

 

The invention relates to medicine, in particular to immunology, and can be used for specific prophylaxis of smallpox and viral hepatitis C.

Known live recombinant vaccine for the prevention of hepatitis b and smallpox and vaccination, including cutaneous application of specified vaccines (RF patent No. 2073524, IPC AC 39/29; 12N 15/86, publ. 20.02.97 year). The vaccine contains a strain of recombinant vaccinia virus (BOB) GKV No. 2131 with a title (3-5)×108ooobasic units (AOE) per ml, is able to synthesize preS2 and HbsAg hepatitis b virus, and peptone as a stabilizer in the following ratio, wt.%:

The recombinant strain of WWII STB No. 2131
with the specified title90-95
Peptone5-10

The disadvantages of the known vaccine and method of vaccination is: poor performance when cutaneous vaccination with this drug, the need for qualified medical personnel for vaccination, threat syringe infection" HIV, hepatitis, etc. When vaccination scratch vaccine is also possible selection of WWII in the environment, which could lead to uncontrolled transmission to unvaccinated people with Risco the development of severe complications.

The closest analogue prototype vaccine is preformed live recombinant bisaccia against smallpox and hepatitis b based on BOB for oral use, containing in each tablet: 1. lyophilized viral material developed on the basis of a recombinant strain of WWII STB No. 2131 typical properties of WWII, expressing preS 2-Ag and HbsAg of hepatitis b virus activity of 6.5 to 8.0 lg, OOE on the tablet 2. lactose, 3. sucrose, 4. calcium stearate, 5. vanilla (RF patent No. 2076735, IPC AC 39/00; 12N 15/00, publ. 10.04.97 year).

The closest prototype of the method of oral vaccination is the way a single injection tablets biwekly. Immunizing dose of each tablet is the same (patent RF №2076735, IPC AC 39/00; 12N 15/00, publ. 10.04.97 year).

Known vaccine and its applications have 2 major drawbacks: in a single pill use with small immunizing dose not achieved a sufficient level of immune response, and in a single pill use with large immunizing dose cause severe inflammation in the upper respiratory tract, although it achieves an acceptable level of seroconversion to BOB: immune interlayer among vaccinated reaches 90%. In addition, a single application of the vaccine, even in large doses causes the formation of only short term is on immunity to BOB (within 1-3 months).

The technical result of the proposed invention is to create a set of biwekly against smallpox and hepatitis b and method of vaccination using a specified set, which would provide sufficient immune response in a long time (≥6 months) without showing side effects of the vaccine.

This technical result is achieved by the fact that the set of preformed live recombinant biwekly against smallpox and hepatitis b for oral administration according to the invention includes two tablets, each containing stabilizing additives, filler and freeze-dried live viral material developed on the basis of a recombinant strain of the war on typical properties of WWII, expressing the protein preS2-S and HBs hepatitis b virus, immunizing dose the first of which is the minimum number of viral material sufficient to obtain a weak immune response in the body with a low reactogenicity, and immunizing dose of a second - the maximum number of viral material, which causes a pronounced and long-lasting immune response in the body without the negative side onto him after administration of the first tablet biwekly. Immunizing dose of the first tablets (0,5-7,0)×106OO is, and immunizing dose of the second tablets (1,0-5,0)×107AOE. Viral material obtained on the basis of recombinant strain of WWII STB No. 2131 in chicken embryos (CE) or transplantable culture of animal cells.

The first and second tablets vaccine have set different color on the surface and inside.

This technical result is also achieved by the fact that in the method of vaccination preformed live recombinant beakley against smallpox and hepatitis b, including oral pills of biwekly, according to the invention for vaccination using a set of tablets biwekly according to claims 1 to 3 of this claims, and initially take the first pill with a minimal dose of biwekly, and the second tablet with a maximum dose of biwekly take before the time of formation of the humoral immune response (by 7-14 days) after ingestion of the first tablets with the minimum immunizing dose of biwekly.

The recombinant strain of WWII STB No. 2131 (b7,5S2-S) deposited in the state collection of viruses of the Institute of Virology. Dijanoveckog (RF patent No. 1575576, 12N 15/00, publ. 1994).

Set of tablets of live recombinant biwekly against smallpox and hepatitis b for oral administration contains two flat tablets # 1 and # 2, located in a blister pack. Tabla is key # 1 and # 2 have different color inside and outside, for example, yellow and grey-brown (to the best of their differences). Immunizing dose tablets # 1 is (0,5-7,0)×106AOE, and immunizing dose tablets№2 - (1,0-5,0)×107AOE. Tableted form disintegrates in the oral cavity under the influence of moisture and temperature of the body.

Example 1. The method of receiving live viral material

A. Obtaining viral material on chorioallantoic membranes of chick embryos

Way of live recombinant viral material comprises culturing recombinant BOB at chorioallantoic membranes (HAO) 12-13-daily EC then get vaccinated material in the form of homogenate HAO 20% lactose, which is subjected to freeze-drying. Dry vaccinated biological product is crushed, mixed with the components of the filler and stabilizer for the preparation of tablet mass. From the latter by pressing get tablets vaccine.

B. Obtaining viral material on transplantable animal cells

First test was conducted homogeneity of viral materials, obtained by 10-fold passage of the Museum of strain STB No. 2131 (b7,5S2-S) on a transplantable cell cultures Vero and 4647. For this studies passage material (5 and 10 passages) by analyzing the expression of HBs-antigen immune method is enzymatic assay (ELISA) and the availability of building-pre-S2-S gene into the viral DNA by polymerase chain reaction (PCR). The need for these studies is dictated by production requirements of the drug, when the number of passages of the production strain in cell culture-producenta can reach 10. As a result of these studies it was noted that passage of the material (5 and 10 passages in Vero cell cultures and 4647) does not contain impurities BOB wild-type and expresses HBs-antigen at the level of 100-200 ng/106cells. This indicates a high stability of the created genetic structure and opens up prospects for the use of any of the tested cell cultures for the production of biwekly against smallpox and hepatitis C.

For getting vaccinated material for subsequent brewing culture recombinant biwekly against smallpox and hepatitis b by the roller culture bottles with a volume of, for example, 2 liters grown monolayers transplantable cell cultures of green monkey kidney No. 4647 or VERO on growth nutrient medium, DMEM with 10%serum bovine (cattle) or 10%fetal calf serum. Used medium is drained and in roller bottles poured supporting nutrient medium, DMEM or Needle MEM with 2%cattle serum or 2%fetal calf serum in an amount of 200 ml per bottle. Cells infect strain recombinante what about BOB (STB No. 2131 (b7,5S2-S) in the dose of infection - 1,0-100,0 the FIGHT on the cage. The cultivation is carried out at a rotation speed of the bottles 3-12 rpm up to 50-70% of the cytopathic effect (when this condition is achieved the highest yield of virus). Data on the accumulation of viral material in the cell cultures are shown in table 1.

Table 1

Data on the accumulation of strain b7,5S2-S of vaccinia virus in cell cultures 4647 and Vero roller when the cultivation
The number of experimentVirus concentration in lg PFU/ml in cell culture (I95=±0,5 lg PFU/ml)
4647Vero
17,07,0
27,57,3
37,57,5
47,36,8*
Note. (*) - low density cell monolayer before infection.

Further supporting the growth medium (200 ml one bottle) is replaced by a buffer solution with a volume (20 ml), that is 10 times smaller. As the buffer solution using 0.01 M Tris to maintain a pH of 8.8 and 9.0. Replacement of the culture medium on the smaller volume of buffer (0.01 M Tris pH 8,8-9,0) before extraction Viru is and from animal cells is to reduce the amount of viral biomass (while keeping the total number of viable viral particles), reducing the financial and time costs during the process subsequent lyophilization of viral material. Getting vaccinated material is carried out after the replacement of the culture medium on the buffer solution by extraction of the virus at three times the freezing at minus 20°With at least 5 hours and thawing the cell monolayer at a temperature of + 20°C for at least 1 hour. Next, the resulting vaccinated suspension defend and separate target supernatant fraction with a titer of not less than 7.0 lg PFU/ml, which is injected stabilizing additives: peptone, sucrose and gelatos in their final concentration in the mixture (wt.%): 2,0-4,0; 2,0-3,0; 1,0-3,0 respectively, or urea, peptone, sucrose and gelatos in their final concentration in the mixture (wt.%): 1,0-2,0; 2,0-4,0; 2,0-3,0; 1,0-3,0 respectively. Received preparative liquid form is poured into the vessel placed in the lyophilic installation type SUPER-4. After conducting the standard procedure of freeze drying in the preparation is practically not reduced infectious viral activity, indicating the high stability of the strain b7,5S2-S BOB to lyophilization in the presence of the inventive stabilizing additives. Next, the dry material is ground and mixed with a filler.

Example 2. Technology of preparation of tablets and their composition

The extrusion is in the tablets of the drug produced on dvenadtsatiraundovom the Tabletpress model K-VIII (Dresden, Germany). The number of vaccinated material in each tablet lay so that the concentration of viable virus in them according to the calculated data based on the activity of the lyophilized materials were within the estimated range.

Below are examples of (a-d) formulations tablets №1 and №2 sets of live recombinant biwekly against smallpox and hepatitis b oral administration (in the mouth).

a) Tablet No. 1 (yellow)

Dried material
recombinant strain with BOB
activity (7,6×106OOE) tablet2,04
peptone0,12
gelatos0,06
sucrose10,10
the buffer0,001
calcium stearate2,0
dye E 1040,02
lactosethe rest is up to 100%

b) Tablet No. 2 (gray-brown)

Freeze-dried material with recombinant
the strain of world war II activity (3,9×107OOE)
pill7,20
peptone0,38
gelatos0,19
sucrose10,29
urea0,01
the buffer0,003
calcium stearate1,99
vanilla0,20
lactosethe rest is up to 100%

in) Tablet No. 1 (yellow)

Freeze-dried material with recombinant
the strain of world war II activity (1,6×106OOE)
pill1,04
peptone1,20
gelatos0,60
sucrose10,85
the buffer0,009
calcium stearate1,97
dye E 1040,02
lactosethe rest is up to 100%

g) Tablet No. 2 (gray-brown)

Freeze-dried material with recombinant
the strain of world war II activity (1,5×107OOE)
pill 10,13
peptone3,80
gelatos1,90
sucrose11,55
the buffer0,027
calcium stearate1,94
vanilla0,17
lactosethe rest is up to 100%

Example 3. Method of vaccination using the proposed set

When oral immunization tablet must be chewed and kept in the mouth to dissolve at least 2-3 minutes, then for 30 minutes is not recommended to drink, smoke and eat. It is forbidden to swallow a pill. The immunization procedure is performed under the control of the listed. The reception of the first tablet of the proposed set causes the formation of a minimal protective level of specific immunity to BOB and marker of hepatitis C. After 7-14 days (before the time of formation of the humoral immune response after ingestion of the first tablets with the minimum immunizing dose) into the body orally administered second tablet of the claimed set with a maximum dose of biwekly, then formed a stable immunity to viruses BOB and hepatitis C.

Example 4. Experimental data on the volunteers of the proposed set of tablets biwekly and method of vaccination, as well as on the individual tablets # 1 and # 2 to the induction of humoral immunity to smallpox

A vaccine. Used set of two tablets biwekly smallpox and hepatitis b with the immunizing dose (0,5-7,0)×106OOE (tablet 1) and (1,0-5,0)×107OOE (tablet No. 2)prepared in accordance with the previously described examples 1 and 2 and separate tablet No. 1 and tablet No. 2.

Samples. From each volunteer daily for 14 days produced a selection of saliva and urine, and once a week for 4 weeks, samples were taken of plasma and serum for virological and serological studies.

Serological studies. The blood serum of people investigated with the aim of determining the level of antibodies to the virus WWII (strain LIVP), HBs Ag. The titers of antibodies to BOB was evaluated in the reaction of neutralization (PH) according to the standard method [Reference, 1967]. The titer of the serum took the maximum dilution that caused a decrease in the number of AOE on HAO more than two times compared with the control. For the working dilution of the virus took its dilution in the calculation of OEE on HAO gave education from 40 to 60 winced. Similarly evaluated the antibody titers to the second world war in PH in the cell culture 4647 method plaques, which gave similar results to those obtained using EC. It is generally accepted that protective antibody titers to BOB in the serum of a person are ≥1:25.

Statistical processing of the results is the ATA. Statistical processing of the obtained results was performed by standard methods [Ashmarin IV, A. Vorobyev, 1962].

For experimental studies conducted recruiting volunteers previously vaccinated against smallpox.

Criteria for selection of candidates for volunteers were: the presence or absence of smallpox vaccination marks on visual inspection and anamnestic data (age applicants, the absence of disease viral hepatitis and vaccination against hepatitis b).

On the basis of a preliminary medical examination of the applicants for volunteer Committee composed of: head of research, physician, dentist, cardiologist, ENT specialist, neurologist, conducted the selection of the volunteers who have not been identified somatic and other diseases, which is a contraindication to vaccination.

According to the results of serological tests for the presence of markers of hepatitis b and antibodies to WWII were formed 8 groups of volunteers for clinical trials biwekly "Relax-W".

Group 1 consisted of 6 volunteers, age from 30 to 40 years, with pox marks on his shoulder. These volunteers were immunized in the conditions of a remote booster low dose (tablet No. 1) biwekly "Relax-W".

Group 2 consisted of 10 volunteer is in, the average age group of 22 to 24 years, with pox marks on his shoulder. Volunteers were immunized in the conditions of remote revaccination large dose (tablet No. 2) biwekly "Relax-W".

Group 3 is represented by 8 volunteers, average age in the group is from 18 to 19 years old, with shoulder pox marks. They were immunized in primary vaccination with a low dose (tablet No. 1) biwekly.

Group 4 consists of 12 volunteers, average age from 18 to 20 years old with no shoulder pox marks. Volunteers were immunized in primary vaccination of a large dose (tablet No. 2) biwekly.

Group 5 consists of 10 volunteers, age group: 34-40 years with pox marks on his shoulder. These volunteers were immunized with a low dose of biwekly "Relax-W" (tablet 1) and after 1 week large dose (tablet No. 2).

Group 6 consists of 10 volunteers, age group: 31-38 years with pox marks on his shoulder. These volunteers were immunized with a low dose of biwekly "Relax-W" (tablet 1) and after 2 weeks of high dose (tablet No. 2).

Group 7 consists of 11 volunteers, age group: 28-34 years with pox marks on his shoulder. These volunteers were immunized with a low dose of biwekly "Relax-W"(tablet 1) and after 1 month of high dose (tablet No. 2).

In group 8 included 4 volunteer, age GRU is PE: 32-35 years, with pox marks on his shoulder. They were immunized with a low dose of biwekly "Relax-W" (tablet 1) and 3-6 months large dose (tablet No. 2).

Summary of the research results presented in tables 2-4.

Analysis of the data given in table 2, showed that a single immunization of volunteers with a small dose of biwekly "Relax-W" (tablet No. 1) does not generate the desired level of seroconversion to BOB. Once the immunization large dose of biwekly (tablet No. 2), creating a suitable level of seroconversion to BOB - 90%, gives a strong manifestation of reactogenicity - up to 50% among vaccinated volunteers.

Vaccination of volunteers previously vaccinated against smallpox, the first small dose of biwekly (tablet 1) and after 7-14 days high dose (tablet No. 2) allows you to create the level of seroconversion to BOB up to 100% with minimal manifestations of reactogenicity (table 3). Another important advantage offered by us set and schemes of vaccination is the duration of the immune response to BOB: if a single vaccination immune interlayer among vaccinated volunteers in 3 months is reduced to 20%, while two-time vaccination within 6 months of the immune layer is 80% (table 4).

We were not able to achieve an acceptable level of seroconversion to HBsAg hepatitis b virus in vaccinated ox is narow. However, two additional immunization commercial hepatitis b vaccine allows you to create protective levels of antibodies to HBsAg in people vaccinated according to our proposed scheme vaccination with the use of the proposed set as twice the immunization of people with the help of this set is already creating basic immunity to HBsAg (table 3).

Table 2

The results of clinical trials of oral biwekly after a single vaccination low dose (tablet No. 1) or large (tablet No. 2) volunteer groups 1 and 2, previously vaccinated against smallpox, and groups 3 and 4 were not previously vaccinated against smallpox
The estimated rateA group of volunteers
1234
The number of subjects1610812
Immunizing dosetablet # 1tablet # 2tablet # 1tablet # 2
1Local reactions in the maximum version:number2326
severity2 4High4High4High
1General reactions in the maximum version:number3426
severity3Weak5Average5Average5Average
6The presence of virus in samples during 14 days of observation after vaccination:salivaNoNoNoNo
plasmaNoNoNoNo
urineNoNoNoNo
The percentage of volunteers who have significant humoral immune response to the virus (1 month after vaccination):ospowiki31906383
Notes:

1 - General and local reactions occurred, as a rule, in the period from the 7th till the 13th day after vaccination;

2 - mild redness of the throat, increasing the submandibular lymph nodes to 0 cm; < / br>
3 - increase of body temperature up to 37.2°;

4 - purulent angina, increased submandibular lymph nodes up to 1.5 cm;

5 - weakness, increased body temperature to 39.0°;

6 - conducted a study of WWII in samples of saliva, plasma and urine samples taken from volunteers daily in the period from the 1st to the 14th day after vaccination;

td align="center"> plasma
Table 3

The results of clinical trials of oral biwekly in a double vaccination of volunteers previously vaccinated against smallpox low dose (tablet 1) and then a large (tablet No. 2)
The estimated rateA group of volunteers
5678
The number of subjects1010114
Immunizing dosetablet # 1 and after 1 week the tablet No. 2tablet No. 1 and 2 weeks tablet No. 2tablet No. 1 and through 1 month tablet No. 2tablet # 1 and 3-6 months tablet No. 2
The introduction of small doses (primary immunization)Local reactions in the maximum variant1 number2210
severityweak2weak2weak2weak2
General reactions in the maximum variant1number2121
severityweak3weak3weak3weak3
The introduction of a large dose (secondary immunization)Local reactions in the maximum version:number0000
Severity----
General reactions in the maximum version:number0000
severity
The presence of virus in samples during 14 days of observation after vaccination:salivaNoNoNoNo
NoNoNoNo
urineNoNoNoNo
The percentage of volunteers who have significant humoral immune response to the virus:ospowiki10090,055,650
Note:

1 - General and local reactions occurred, as a rule, in the period from the 7th till the 13th day after vaccination;

2 - mild redness of the throat, increasing the submandibular lymph nodes up to 0.8 cm;

3 - increase of body temperature up to 37.2°C.

Table 4

Comparative data on the dynamics of titers of antibodies to vaccinia virus in volunteers orally vaccinated once a small dose of biwekly (tablet No. 1) or high dose (tablet No. 2) and twice with an interval of 1-2 weeks a set of biwekly
The number of volunteerTitle smallpox neutralizing antibodies through different periods of observation:
0 days30 days90 days180 days270 d is to
A single vaccination with beakley in small (tablet No. 1) or high dose (tablet No. 2) volunteers, previously immunoserology and not immunoserologic against smallpox*
761:51:1251:251:25
771:51:251:51:5
80<1:5>1:1251:1251:25
811:5>1:251:51:5
311:51:1251:5<1:5
35<1:5>1:25<1:5<1:5
14<1:51:25<1:5<1:5
15<1:51:125<1:5<1:5
17<1:51:1251:51:5
82<1:51:1251:5<1:5
DV is a multiple of vaccination dial biwekly with an interval of 1-2 weeks *
11:51:1251:1251:125
21:51:51:251:5
3<1:51:1251:251:125
4<1:5>1:1251:125>1:125
51:51:251:251:25
61:51:251:251:125
7<1:51:251:25>1:125
8<1:51:251:251:125
91:51:125>1:1251:5
10<1:51:125>1:125<1:5
11<1:51:125>1:125
12<1:51:1251:125
13<1:51:1251:125
14<1:51:1251:125
Note:

*- shows the titers of antibodies to vaccinia virus only volunteers observed within 6-9 months.

Example 5. Comparative experimental studies on volunteers of the proposed set of tablets biwekly and method of vaccination for the induction of humoral immunity to hepatitis b

The vaccines. In this work, we used two series of "Biwekly smallpox and hepatitis In embryonic live recombinant, tablets (Relax-W)No. 10 and No. 11 with a concentration CENTURIES 3,9×107 and 7.6×106 OOE/tablet, respectively, prepared according to the EPR project in accordance to the previously described method (Patent RF №2076735, 1997), the past quality control in accordance with the requirements of the project Fund in the OPF CC scientifically defense Ministry, SRC VB "Vector" and certified in gisk named after. Laurasia. To enhance the immune response to HBsAg used commercial recombinant yeast liquid hepatitis b vaccine "Combitech".

Samples. From each volunteer once a month for 6 months was selected samples of plasma and serum serological studies.

Serologic is the cue studies. Detection of antibodies to HBsAg, the presence of HBsAg and antibodies to HBCor Ag in the blood serum of people registered with certified MOH Russia enzyme immunoassay systems "Vecto HBs Ag-antibody-strip"Ag "Recomthe In strip and Vecto HBCor-antibody-strip" (the production company "Vector-best, Novosibirsk region, palzoo). The sensitivity of the last test systems for CCA risk - 0.2 ng/ml is generally Accepted that protective concentration of antibodies to HBsAg in the serum of human blood are ≥10 mIU/ml

Processing of test results. Statistical processing of the obtained results was performed by standard methods [Ashmarin and sparrows, 1962; Sachs, 1976].

For experimental studies conducted recruiting volunteers previously vaccinated against smallpox.

Criteria for selection of candidates for volunteers were: the presence or absence of smallpox vaccination marks on visual inspection and anamnestic data (age applicants, the absence of disease viral hepatitis and vaccination against hepatitis b).

On the basis of a preliminary medical examination of the applicants for volunteer Committee composed of: head of research, physician, dentist, cardiologist, ENT specialist, neurologist, conducted the selection of the volunteers who have not been identified somatic and the other is their diseases, which is a contraindication to vaccination.

According to the results of serological tests for the presence of markers of hepatitis b and antibodies to WWII were formed 2 groups of volunteers for clinical trials set biwekly "Relax-W".

Group 1 represented by 7 volunteers (aged 22 to 44 years). These volunteers were twice immunized with recombinant monovalent "Combitech".

Group 2 represented by 4 volunteers (age from 37 to 45 years of age, previously vaccinated against smallpox and vaccination with signs. These volunteers were immunized in the conditions of remote revaccination first recombinant monovalent "Combitech", month dose of biwekly 6,9 lg OOE) and after 1 month is very dose of 7.6 lg OOE.

The results of these investigations are given in tables 5 and 6.

As can be seen from the data protective titers of antibodies to HBsAg was observed in 40% of the volunteers.

From table 6 it is seen that the protective titers of antibodies to BB - from 75,0% volunteers, and protective titers of antibodies to HBsAg - 100.0% of volunteers, and it should be noted that the increase in titers of HBsAg was observed after revaccination with beakley.

Comparing the results presented in tables 5 and 6, it can be concluded that application of a set of tablets biwekly after immunization vaccine PR is against hepatitis b can significantly enhance the immune response to HBsAg. So if the second immunization vaccine "Combitech" induces the immune response to HBsAg 40% of vaccinated volunteers (3 of 7), immunization with a set of tablets biwekly - in 100% of vaccinated volunteers (4 of 4). It should be noted that in this experiment was used is not the most successful scheme double oral immunization beakley: the interval between doses doses biwekly amounted to 1 month.

Table 5

Dynamics of humoral immunity to HBsAg in volunteers group 1 immunized twice vaccine "Combitech in doses of 20 mcg
The number of volunteerAge, yearThe day after vaccinationThe concentration of antibodies to HBsAg, mIU/ml
60320<10
30<10
30Re-introduction of the vaccine "Combitech"
60<10
180<10
71420<10
30<10
Re-introduction of the vaccine "Combitech"
603200
180500
90440<10
30<10
30Re-introduction of the vaccine "Combitech"
60<10
62310<10
28100
60<10
61320<10
30<10
30Re-introduction of the vaccine "Combitech"
60<10
180<10
91240<10
30<10
30Re-introduction of the vaccine "Combitech"
60<10
180<10
4220<10
30<10
30Re-introduction of the vaccine "Combitech"
60500
180<10

30
Table 6

Dynamics of humoral immunity to CC and HBsAg in volunteers group 9, first immunized by vaccine "Combitech", month dose of biwekly 6,9 lg, OOE and after another month dose of biwekly 7,6 lg OOE in terms of remote booster
The number of volunteerAge, yearThe presence of pox marksThe day after vaccinationThe titer of antibody in PH to BBThe concentration of antibodies to HBsAg, mIU/ml
64370<1:5<10
0Immunization vaccine "Combitech"
30--10
30Immunization dose of biwekly 6,9 lg OOE
601:5˜10
60Immunization dose of biwekly 7,6 lg OOE
901:25150
65370<1:5˜10
Immunization vaccine "Combitech"
30-<10
Immunization dose of biwekly 6,9 lg OOE
60<1:5<10
60Immunization dose of biwekly 7,6 lg OOE
901:2540
99430<1:5<10
Immunization vaccine "Combitech"
30-<10
30Immunization dose of biwekly 6,9 lg OOE
60<1:5<10
60Immunization dose of biwekly 7,6 lg OOE
901:12530
103450<1:5<10
Immunization vaccine "Combitech"
30-<10
30Immunization dose of biwekly 6,9 lg OOE
60<1:5500
60 Immunization dose of biwekly 7,6 lg OOE
901:5200

1. A set of preformed live recombinant biwekly against smallpox and hepatitis b for oral administration, comprising two tablets containing stabilizing additives, filler and freeze-dried live viral material developed on the basis of recombinant virus strain of the war on typical properties of vaccinia virus expressing preS 2-Ag and HbsAg hepatitis b virus, immunizing dose the first of which is (0,5-7,0)·106ooobasic units (AOE) - the minimum number of viral material, sufficient to receive the base of the immune response in the body, and immunizing dose of the second is (1,0-5,0)·107OOE, is the maximum amount of viral material, which causes an immune response in the body without the negative side onto him after administration of the first tablet biwekly, and viral material obtained on the basis of recombinant strain of vaccinia virus STB No. 2131 in chicken embryos or human culture of animal cells.

2. The kit according to claim 1, characterized in that the first and second tablets of the set have a different color.

3. Method of vaccination preformed live recombinant beakley against smallpox, hepati the while, including oral pills of biwekly, characterized in that for vaccination using a set of tablets biwekly according to claims 1 and 2 of the present claims, and initially take the first pill with a minimal dose of biwekly, and the second tablet with a maximum dose of biwekly take before the time of formation of the humoral immune response after ingestion of the first tablets with the minimum immunizing dose.

4. The method according to claim 3, characterized in that the second tablet with a maximum dose of biwekly take over 7-14 days after taking the first tablet with a minimal dose of biwekly.



 

Same patents:

FIELD: gene engineering.

SUBSTANCE: invention relates to method for modification target endogenic gene or chromosomal locus in eucaryotic cells. Claimed method includes production of large cloned genomic fragment having more than 20000 n.p. and designing based on the same large targeting vector (LTVEC) by using bacterial homological recombination. Further LTVEC is introduced into eucaryotic cells to modify endogenic gene or chromosomal locus. Finally assay is carried out to determine of allele modification in such cells. Also disclosed is application said cells for generation of organisms carrying such genetic modification.

EFFECT: method for modification with large DNA sequences.

26 cl, 6 dwg, 2 tbl, 5 ex

FIELD: molecular biology, biochemistry, medicine, oncology.

SUBSTANCE: invention relates to DNA sequences found in analysis of mDNA from squamous carcinoma cellular lines of different origin wherein these DNA sequences represent transcripts from rearranged genes SCCA1 and SCCA2. Result of rearrangement is formation of fused gene consisting of exon 2-7 of gene SCCA1 and exon 8 of gene SCCA2, or exons 2-7 of gene SCCA2 and exon 8 of gene SCCA1. Prepared expressing vectors comprising above said combinations of exons of two genes provide synthesis of corresponding fused protein in host-cell. Proposed sequences of nucleic acids and genetic constructions based on thereof represent novel agents for diagnosis squamous carcinomas.

EFFECT: valuable biological and medicinal properties of transcripts.

8 cl, 9 dwg, 1 tbl, 5 ex

FIELD: biotechnology, in particular isolated DNA molecule providing plant disease resistance and method for providing of disease resistance to plants.

SUBSTANCE: DNA molecular containing N1M1 is isolated. Recombinant vector including active in plant promoter functionally bonded with said DNA is constructed and plant is transformed by this vector.

EFFECT: decreased technological charges and increased land productivity.

9 cl, 37 dwg, 18 tbl, 10 ex

The invention relates to biotechnology and genetic engineering and can be used to create transgenic animals and plants

The invention relates to biotechnology

The invention relates to propertytaxsession systems that require cleavage product a predecessor to the new polypeptide capable of restoring dichloroindophenol and oxidized glutathione to DNA that encodes this polypetide, farmkompanijam comprising the polypeptide, monoclonal antibodies against the indicated polypeptide

The invention relates to biotechnology, in particular genetic engineering, is a recombinant plasmid pCVA designed for the transcription of the genes of the ribozymes in the composition of sequences of virus-associated PHK (VA PHK) adenovirus birds FAV1 (CELO) in eukaryotic cells

FIELD: veterinary science, virology, biotechnology.

SUBSTANCE: the suggested vaccine contains the suspension of avirulent and purified antigenic material out of the strain N 101 of cattle rotavirus (Research Institute of Animal Protection) obtained in mono-layer culture cells MDBK or SPEV at accumulation degree being 106 viral particles/ml, not less and activity in IEA being 5.0 log2, not less, and target additives: aluminum hydroxide and saponin in efficient ratios. The strain is deposited in collection of FGU VGNKI under registration number - a culture strain N 101 Research Institute of Animal Protection-DEP. The suggested vaccine is highly immunogenic, safe and areactogenic, it is capable to protect cattle stock against epizootic agent of rotaviral infection circulating in Russian territory.

EFFECT: higher efficiency.

14 cl, 9 ex, 10 tbl

FIELD: medicine, biology.

SUBSTANCE: the present innovation deals with the technology for manufacturing the series of preparations being of immunoregulating action and could be applied for manufacturing vaccines and preparations for the purposes mentioned. It is necessary to dissolve 3000 g sugar in bidistilled water up to saturated syrup to sterilize it due to boiling followed by cooling it up to about 45-50°C, then one should add as a basis 30 g protein bile fraction of two ad more even-toes animals dissolved in 200 g bidistilled water, the mixture should be thoroughly mixed to dry it up to the end in oven drier at 45-50°C, then the mixture should be ground to sterilize the powder obtained that contains a target product with quartz irradiation to be packed into gelatinous capsules NN 1-3 at 0.5-1.5 g dosage. The innovation provides the development of reliable, economically profitable technology for manufacturing native preparations of immunoregulating action that enables to shorten terms of therapy, prolong terms of remission, achieve complete recovery in case of viral diseases and intoxications and decrease lethality percentage in case of the diseases mentioned.

EFFECT: higher efficiency.

6 ex

FIELD: medicine, pharmacology, bioorganic chemistry, pharmacy.

SUBSTANCE: invention relates to the effective using amount of β-L-2'-deoxynucleoside of the formula (I) or (II) used in manufacturing a medicinal agent used in treatment of hepatitis B, pharmaceutical compositions containing thereof, and methods for treatment of hepatitis B. Proposed agent shows the enhanced effectiveness in treatment of hepatitis B.

EFFECT: enhanced and valuable medicinal properties of agent.

83 cl, 6 tbl, 11 ex

FIELD: medicine.

SUBSTANCE: method involves carrying out revaccination of children, not reaching the age of 6 years from May to September, next year within the period from May to September.

EFFECT: enhanced effectiveness of revaccination.

1 tbl

FIELD: medicinal plants, chemical technology.

SUBSTANCE: method involves extraction of milled licorice (Glycyrrhiza uralensis, Fischer) roots with 0.5% aqueous solution of NH4OH. The total triterpene acids of extract are precipitated with concentrated H2SO4 followed by extraction of total triterpene acids with 1% solution of H2SO4 in acetone, precipitation of glycyrrhizic acid triammonium salt with 25% solution of NH4OH and its conversion to monoammonium salt by re-crystallization from glacial CH3COOH and purification of the latter by re-crystallization from 85% ethanol. Invention provides enhanced yield of the end product.

EFFECT: improved preparing method, enhanced yield.

1 dwg, 1 ex

FIELD: organic chemistry, medicine, pharmacy.

SUBSTANCE: invention describes derivatives of piperazine of the general formula: or their pharmaceutically acceptable salts wherein Ra - R8a mean phenyl; R8b means pyridyl, or R8 means naphthyl; R1 means hydrogen atom; R2 - R9, R10, R11 mean substituted phenyl; R9, R10, R11 mean substituted pyridyl or pyrimidyl; R9, R10, R11 mean substituted pyridyl-N-oxide or pyrimidyl-N-oxide; R12, R13 mean substituted oxazolyl, naphthyl, fluorenyl, compounds of formulae , or ; R3 means hydrogen atom, (C1-C6)-alkyl, (C1-C6)-alkoxy-(C1-C6)-alkyl, (C3-C10)-cycloalkyl, (C3-C10)-cycloalkyl-(C1-C6)-alkyl; R8 means phenyl; R8 means phenyl-(C1-C6)-alkyl, or R8 means thienyl-(C1-C6)-alkyl; R4, R5, R7 and R13 mean independently hydrogen atom or (C1-C6)-alkyl; R6 means hydrogen atom or (C1-C6)-alkyl; R8 means 1-3 substitutes that mean independently hydrogen atom, halogen atom, (C1-C6)-alkoxyl or -CF3; R8a means 1-3 substitutes that mean independently hydrogen atom, halogen atom, -CF3, -CF3O, -CN; R14 means phenyl, -NHCOCF3 and imidazolyl; R8b means 1-3 substitutes that mean independently hydrogen atom or halogen atom; R9 and R10 mean independently (C1-C6)-alkyl, halogen atom, -NR17R18, -OH, -CF3 and -OCH3; R11 means R9, hydrogen atom, phenyl, -NO2, -CN, -CH2F, -CHF2, -CHO, -CN=NOR17, pyridyl, pyridyl-N-oxide, pyrimidinyl, pyrazinyl, -N(R17)CONR18R19, -NHCONH-(chloro-(C1-C6)-alkyl), -NHCONH-((C3-C10)-cycloalkyl-(C1-C6)-alkyl), -NHCO-(C1-C6)-alkyl, -NHCOCF3, -NHSO2N-((C1-C6)-alkyl)2, -NHSO2-(C1-C6)-alkyl, -N(SO2CF3)2, -NHCO2-(C1-C6)-alkyl, (C3-C10)-cycloalkyl, -SR20, -OSO2-(C1-C6)-alkyl, -SO2CF3, hydroxy-(C1-C6)-alkyl, -CONR17R18, -CON(CH2CH2-O-CH3)2, -OCONH-(C1-C6)-alkyl, -Si(CH3)3 or -B(OC(CH3)2)2; R12 means (C1-C)-alkyl or R14-phenyl; R14 means 1-3 substitutes that mean independently hydrogen, (C1-C6)-alkyl, -CF3, -CO2R17, -CN, (C1-C6)-alkoxyl and halogen atom; R15 and R16 mean independently hydrogen atom and (C1-C6)-alkyl, or R15 and R16 mean in common (C2-C5)-alkylene group and in common with carbon atom to which they are bound form (C3-C6)-spiran ring; R17, R18 and R19 mean independently hydrogen atom or (C1-C6)-alkyl; R20 means (C1-C6)-alkyl. Also, invention describes pharmaceutical compositions containing these compounds and using novel compounds as CCR5 antagonists in treatment of HIV infection, arthritis, asthma, cerebrospinal sclerosis and other diseases.

EFFECT: valuable medicinal properties of compounds and pharmaceutical compositions.

29 cl, 30 tbl, 31 ex

FIELD: chemistry of proteins, virology, pharmacy.

SUBSTANCE: invention relates to novel physiologically active protein conjugates that can be used in medicine. Proposed conjugates is formed by a protein molecule and polyethylene glycol and corresponds to the formula: wherein protein represents interferon α-2b; n and n1 have values providing an average molecular mass of links from about 7500 Da to about 35000 Da. Conjugate shows the enhanced stability as compared with the known analogs. Also, invention relates to a pharmaceutical composition possessing an antiviral activity and containing the indicated conjugate, and to a method for control of viral infection.

EFFECT: valuable medicinal and biological properties of conjugates.

4 cl, 1 dwg, 6 ex

FIELD: organic chemistry, medicine, pharmacy.

SUBSTANCE: invention relates to derivatives of pyrazine of the general formula (I): and their salts wherein R1 represents hydrogen atom or halogen atom; R2 represents hydrogen atom or protected or unprotected group of monophosphoric, diphosphoric or triphosphoric acid; R3, R4, R5 and R6 can be similar or different and represent hydrogen atom, halogen atom, substituted or unsubstituted, protected or unprotected hydroxyl group or amino-group; or R4 and R6 taken in common form a simple bond; A represents oxygen atom or methylene group; n = 0 or 1; Y represents oxygen atom pr NH-group. Compounds elicit the excellent antiviral activity and useful as a therapeutic agent in treatment of viral infections. Also, invention describes a pharmaceutical composition based on compounds of the formula (I) and derivatives of fluoropyrazine carboxamide as intermediate compounds used in synthesis of compounds of the formula (I).

EFFECT: valuable medicinal properties of compounds and pharmaceutical composition.

13 cl, 6 tbl, 57 ex

FIELD: microbiology, biotechnology, medicine, in particular production of living vaccines against viral infections.

SUBSTANCE: invention relates to living vaccines for rectal administration against viral infections based on attenuated recombinant Salmonella strains bearing protective viral antigens. Suppository for immunoprophylaxis of viral infections contains (per one 2 g suppository, mass %): cell slurry of attenuated recombinant Salmonella strains transformed with pGEX-2T-TBI, pcDNA-TCI, or pKHBc bearing gene of protective viral antigens, mixed with suppository base in amount of 106-109 living cells 1 %; fatty base 94 %; and emulsifier T2 5 %. As fatty solid cooking oil (94 %); or hydrolyzed cotton oil (94 %); or mixture of cooking oil (60 %), Cocoa fat (24 %), and solid wax (10 %).

EFFECT: agent for inducing of humoral and cell immune response to respective viral antigen.

8 tbl, 11 ex, 3 dwg

FIELD: organic chemistry, medicine, pharmacy.

SUBSTANCE: invention describes a compound of the formula (I): wherein X means alkylene group; Y means -CO-, -CS- or -SO2-group; Z represents a simple bond or -NR5-group; R1 represents unsubstituted phenyl or phenyl substituted with halogen atom. (C1-C20)-alkyl group; R2 is chosen from -alkyl, -alkyl-O-alkyl; R3 and R4 represent alkyl; R5 represents hydrogen atom or (C1-C10)-alkyl group; Also, invention describes intermediate compounds - derivatives of imidazopyridine-4-amine, 2-phenoxypyridine and 4-phenoxypyridine. Proposed compounds and pharmaceutical compositions are able to stimulate biosynthesis of different cytokines and can be used in treatment of viral and tumor diseases.

EFFECT: valuable medicinal properties of compounds and pharmaceutical compositions.

32 cl, 1 tbl, 9 ex

FIELD: microbiology, biotechnology, medicine, in particular production of living vaccines against viral infections.

SUBSTANCE: invention relates to living vaccines for rectal administration against viral infections based on attenuated recombinant Salmonella strains bearing protective viral antigens. Suppository for immunoprophylaxis of viral infections contains (per one 2 g suppository, mass %): cell slurry of attenuated recombinant Salmonella strains transformed with pGEX-2T-TBI, pcDNA-TCI, or pKHBc bearing gene of protective viral antigens, mixed with suppository base in amount of 106-109 living cells 1 %; fatty base 94 %; and emulsifier T2 5 %. As fatty solid cooking oil (94 %); or hydrolyzed cotton oil (94 %); or mixture of cooking oil (60 %), Cocoa fat (24 %), and solid wax (10 %).

EFFECT: agent for inducing of humoral and cell immune response to respective viral antigen.

8 tbl, 11 ex, 3 dwg

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