Viral hepatitis type A (hepatitis A) is one of the most common infectious diseases in our country. It has occurred several times of fulminant epidemic by fecal-oral route. The most effective method to prevent hepatitis A is vaccination of hepatitis A vaccine. Now there are two kinds of hepatitis A vaccine for sale that are attenuated live vaccine and inactivated vaccine. Insufficient attenuation during process of production of attenuated live vaccine will make safety of vaccine can not be assured. But excessive attenuation may decrease immunogenicity of vaccine. Virulence can be eliminated through fecal after vaccination. The possibility of virulence reversion of virus in vivo has not been eliminated completely. In addition, storage time of attenuated live vaccine is short, activity is unstable and immune dose is big. Due to these defects of attenuated live vaccine, developed countries have stop developing live vaccine basically. Now there are at least three kinds of inactivated hepatitis A vaccines being used in many countries. Crowd trial shows that these inactivated hepatitis A vaccine have favorable safety and immunogenicity. Medicine and biological products test institute of China and Tangshan Yian bioengineering limited company developed a inactivated hepatitis A vaccine together. Approved by relative department we started phase Ⅰ clinical trial to observe the safety and immunogenicity of that vaccine.

1. Materials and methods
1.1 Vaccine  Vaccine for trial is inactivated hepatitis A vaccine produced by Tangshan Yian bioengineering Co Ltd (homemade inactivated hepatitis A vaccine for short). This vaccine was produced by adopting TZ84 strain of hepatitis A virus to inoculate human embryo lung diploid cell 2BS strain, culture and breed, harvest, purification inactivation with formaldehyde and absorption by aluminium hydroxide. It is 0.5ml per bottle that contains 1,000 unit of inactivated hepatitis A virus. Batch number is 971201. Control vaccine is Havrix inactivated hepatitis A vaccine produced by SmithKline Reecham Biologicals (SmithKline inactivated hepatitis A vaccine for short). It is 1.0ml per bottle that contains 720 ELISA units of inactivated hepatitis A virus. Batch number is VHA203C4.

1.2 Vaccinate objects

Use EIA methods to detect HBsAg during screening of susceptible and the reagent was production of Sino-foreign Shanghai industrial Kehua biotechnology Co Ltd. Undergraduates of a university are eighteen to twenty-one years old (20±0.81) with negative Anti-HAV and HBsAg, normal liver function (ALT, serum bilirubin) and nomal temperature. And there was no abnormal finding during physical examination to heart, lung liver and spleen and drug allergy history. Assigned vaccinate objects screened into two groups randomly. Intramuscular injected it in deltoid muscle of upper arm at 0 and the third month respectively. The study group which contained sixteen objects (eleven male and five female) was vaccinated with 1000 units of homemade inactivated hepatitis A vaccine every time; The control group which contained fifteen objects (eight male and seven female) was vaccinated with 720 ELISA units of SmithKline inactivated hepatitis A vaccine every time.

1.3 Indexes of safety

Observe local and systematic reaction at 8, 24, 48 and 72 hours after injection. Systematic reaction contains fever, acratia, headache, dizziness, nausea, inappetence and eryvash; Local reaction contains redness, swelling, fever, pain, bubble and maculopapule in injection part. Indexes of liver function contain ALT and serum bilirubin.

1.4 Indexes of immunogenicity

Use EIA competitive inhibition method for qualitative determination of antibody of hepatitis A virus (HAV). The reagent was production of Tangshan Yian bioengineering Co Ltd. Competitive inhibition rate that was not larger than 50% indicated that antibody of hepatitis A virus is negative (during susceptive object screening, competitive inhibition rate that was not larger than 40% indicated that antibody of hepatitis A virus is negative). Use EIA competitive inhibition method for quantitative determination of antibody of hepatitis A virus. The reagent is production of medicine and biological products test institute of China. Use WHO standard serum of antibody of hepatitis A virus as contrast to determinate serum titer of antibody. The sensitivity is 29mIU/ml. Use EIA double sandwich assay to determinate serum IgM antibody of hepatitis A. The reagent was production of Tangshan Yian bioengineering co Ltd. Took blood sample in 0,1,3,4 month respectively. Storage the serum for determination of antibody of HAV in low temperature and carry through determination after the last blood taking.

1.5 Statistical analysis

Use Fisher exact examination to compare seroconversion of serum antibody of two groups; Serum titer of antibody was shown with geometrical mean (GMT) and 95% confidence interval (95% CI). Use T test to compare two groups. Abandon the patients with negative serum antibody during calculating GMT.

2. Results

2.1 Safety

Mild reddish swelling was noted in one subject from the study group eight hours after primary vaccination, the diameter was 0.5 cm with no induration and disappeared after twenty-four hours. Nausea was complained by one subject after eight hours and fatigue was complained by one subject after twenty-four hours.  In control group, one subject complained local fever after eight hours. Another subject complained local reddish swelling whose diameter was 0.4cm and it disappeared after twenty-four hours. One subject complained nausea and dizziness after eight hours and one subject complained dizziness after forty-eight hours. No local and systematic reaction was noted in both study group and control group eight to seventy-two hours after booster vaccination. Took blood and determined ALT and serum bilirubin one month after primary vaccination and booster vaccination. And no abnormal liver function was noted in both study group and control group.

2.2 Immunogenicity

Seroconversion rates of antibody of HAV were 94%, 100%, and 100% in the study group at 1, 3 and 4 (1 month after booster) months, respectively. The corresponding figures were 73%, 80% and 100% in the control group. Compared the seroconversion of two groups at three points and there were no statistical difference. Geometric mean antibody titers were 139.2 mIU/ml, 137.7 mIU/ml and 1066.7 mIU/ml at 1, 3 and 4 months respectively in the study group, and 104.3 mIU/ml, 111.3 mIU/ml, and 760.7 mIU/ml in the control group. There was no statistical difference between GMT of two groups at 1 and 3 months after primary vaccination and 1 month after booster vaccination (Table 1). IgM antibody of HAV of all the vaccination objects was negative at 1 and 3 months after primary vaccination and 1 month after booster vaccination.  

Table 1 Geometric mean titers (mIU/ml) and 95% confidence interval (95% CI) of anti-HAV in initially HAV-seronegative subjects

3 .Discussions

It appeared temporary mild reddish swelling or fever in local in local injection place in several vaccination objects after vaccination of homemade inactivated hepatitis A vaccine and control vaccine. There were several vaccination objects complaining mild headache and dizziness or fatigue and nausea in both groups. It shows that they are not specific side effect of homemade vaccine. There was no liver function abnormal after vaccination. It shows that homemade inactivated hepatitis A vaccine is safe. Both seroconversion rate of antibody of HAV and geometric mean antibody titer of study vaccine were higher than or equal to control vaccine at 1 and 3 months after primary vaccination and 1 month after booster vaccination. It shows that the immunogenicity of homemade vaccine is good. The dose of control vaccine is the recommended dosage in specification 720 ELISA units and the dose of adult can be 1440 ELISA unit too. If increase the dose of control vaccine, it seroconversionn rate of antibody of HAV and geometric mean antibody titer may be higher.

References

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