Immunogenicity, effectiveness and safety of combined hepatitis A and B vaccine: a systematic literature review

ABSTRACT

Background: Hepatitis A and B are two of the most common vaccine-preventable diseases and vaccination for Hepatitis A virus (HAV) and hepatitis B virus (HBV) is recommended for those at risk of contracting HAV and/or HBV through their occupation, travel or lifestyle.

Objective: To describe the vaccine efficacy, immunogenicity, effectiveness and safety of the combined vaccine against hepatitis A and hepatitis B.

Methods: A systematic review of the literature published between 1990 and 2015.

Results: Anti-HAV seropositivity rates ranged from 96.2% to 100% and anti-HBs seroprotection rates from 82% to 100%. Antibodies persisted up to 15 years and geometric mean concentration (GMC) remained above the seropositivity cut-off value for both.

Anti-HAV and anti-HBs immune responses were lower in less immunocompetent individuals one month after completion of the immunization schedule.

The safety profiles of Twinrix^TM and monovalent hepatitis A and B vaccines were similar.

Conclusion: The vaccine offers satisfactory long-term immunogenicity rates, expected duration of protection and safety profile similar to the monovalent hepatitis A or B vaccines.

KEYWORDS:

• Hepatitis A
• Hepatitis B
• Combined vaccine
• Twinrix^TM
• Ambirix^TM
• Immunogenicity
• Safety
• Effectiveness
• Efficacy

Introduction

Hepatitis A and B are two of the most common vaccine-preventable diseases and are a serious global public health problem [1–3].

Hepatitis A virus (HAV) is primarily spread through ingestion of food or water that is contaminated with the faces of an infected individual. In 2005, 126 million cases of acute hepatitis A occurred, according to the World Health Organization (WHO) [4]. HAV infections do not cause chronic liver disease and are often asymptomatic, especially in young children. However, the disease can be severe in adolescents and adults, and can cause debilitating symptoms and acute liver failure, which is associated with high mortality [1,4]. Currently, no treatment for acute hepatitis A is available, except for supportive care or liver transplantation in the rare cases with liver failure [5].

Most children (90%) from areas with high endemicity levels of hepatitis A infections are infected before the age of 10 [1]. In countries with intermediate endemicity, higher susceptibility and disease rates occur in older age groups. In low endemicity areas, disease may occur mainly among adolescents and adults in high-risk groups, such as injecting-drug users, men who have sex with men (MSM) and people traveling to areas of high endemicity [1,4].

In these countries, a relatively large part of the disease burden is attributable to infection acquired due to importation of contaminated food and infected returning travelers. In these settings, catch-up strategies targeted at adolescents and targeted vaccination for hepatitis A of high-risk groups should be considered in low and very low endemic countries to provide individual health benefits according to the WHO [4].

Hepatitis B is a potentially life-threatening liver infection caused by the hepatitis B virus (HBV). HBV spreads through perinatal transmission, horizontal transmission (exposure to infected blood), percutaneous or mucosal exposure to infected blood, saliva, menstrual, vaginal, or seminal fluids, and sexual transmission [2,6]. Most acute infections are asymptomatic. Chronic HBV infections occur in 80–90% of infants infected during the first year of life and in about 30% of children infected before the age of 6 years. In adults, infection leads to chronic hepatitis in <5% of the cases [6]. Cirrhosis and/or liver cancer will develop in 20–30% of chronically infected adults. Worldwide, 240 million people are thought to be chronically infected with hepatitis B and approximately 780,000 persons die each year from hepatitis B infection. Prevalence of hepatitis B is highest in sub-Saharan Africa and East Asia, where 5–10% of the adult population is chronically infected. High rates of chronic infections are also found in the southern parts of eastern and central Europe [2].

Effective and safe vaccines are available for both hepatitis A and B [3,7,8]. There is no recognized correlate of protection for hepatitis A. However, anti-HAV antibody levels ranging from 10 to 33 mIU/ml, depending on the assay, are usually considered a marker of protection against HAV infection [4]. However, clinical experience suggests that protection following vaccination may be present even if there are no detectable HAV antibodies [3,4]. Anti-Hepatitis B surface antigen (HBs) antibody levels of 10 mIU/ml are the threshold for protection against HBV infection [6].

Hepatitis B vaccination is recommended by the WHO for all infants as soon as possible after birth, preferably within 24 h. As of 2014, 184 Member States vaccinate infants against hepatitis B as part of their vaccination schedules and 81% of children received the completed hepatitis B vaccine course [9]. This is a major increase compared with 31 countries in 1992.

Catch-up vaccination for hepatitis B includes people at increased risk for HBV infections, including injecting drug users, health-care workers and travelers who have not completed their hepatitis B vaccination series [6].

There is considerable overlap in groups at risk of HAV and HBV infections, including persons traveling to areas of high or intermediate endemicity, patients with chronic liver disease, patients coinfected with HCV, and persons at increased risk of disease due to sexual contact (especially MSM) [3,10,11]. Since the estimated case fatality rate for hepatitis A was 23-fold higher in patients with chronic liver disease, as compared to patients without chronic liver disease [12], hepatitis A and B vaccination has been recommended for this group [13].

Three presentations of the combined vaccine against hepatitis A and B (Twinrix^TM, Twinrix Pediatric^TM, and Ambirix^TM; GSK Vaccines, Belgium) are available [3,14], see Table 1. These vaccines are able to confer concurrent protection against the two infections while reducing the number of injections, associated costs, and other logistical issues. This has offered greater convenience to the vaccinee and has facilitated higher compliance to the schedule [15,16].

Table 1. Three presentations of combined vaccine against hepatitis A and B.

Vaccine	Target population	Formulation	Schedule
Twinrix^TM	Adults	1.0 ml containing 720 EL.U inactivated HAV and 20 µg recombinant HBsAg protein	3 doses
Twinrix Pediatric^TM	Children (1–11 years)	0.5 ml containing 360 EL.U inactivated HAV and 10 µg recombinant HBsAg protein	3 doses
Ambirix^TM	Children and adolescents (1–15 years)	1.0 ml containing 720 EL.U inactivated HAV and 20 µg recombinant HBsAg protein	2 doses

EL.U: Enzyme-linked immunoassay units; HAV: Hepatitis A virus; HBsAg: Hepatitis B surface antigen.

The objective of this systematic review was to summarize data on the efficacy, immunogenicity, effectiveness, and safety of Twinrix^TM, Twinrix Pediatric^TM, and Ambirix^TM.

Methods

Search strategy

The PubMed and Cochrane databases were searched for literature published between 1 January 1990 and 13 October 2015 (date of search). A search string combining terms on hepatitis A, hepatitis B, and vaccines was built (see Appendix 1).

Inclusion and exclusion criteria

Only primary studies, such as trials and cohort studies, relevant for the research objective were included. Review articles were excluded in this review; however, the reference lists of systematic reviews were checked to identify additional relevant primary studies. No additional relevant articles were retrieved from systematic reviews. Articles comparing Twinrix^TM, Twinrix Pediatric^TM, or Ambirix^TM with other hepatitis A or B vaccines were also included; articles that focused only on other vaccines were excluded. Other exclusion criteria were studies with a sample size less than 50, studies on treatment, vaccination coverage, case reports, economic evaluations, biochemistry or molecular studies, and animal studies.

Selection process

Relevant articles were selected by a three-step selection procedure. First, titles and abstracts identified through the search strategy were assessed on relevancy for the objectives. Forty percent of the titles and abstracts were screened in duplicate by two independent researchers. Second, the full text articles, selected in the first selection step, were screened. It was determined whether the paper indeed answered the review question. Only in that case critical appraisal of full text articles was done. A checklist was used to avoid selection of references with poor reporting or critical quality issues. During these two selection steps the above mentioned in- and exclusion criteria were kept in mind. Third, further scrutiny of articles during the data-extraction phase took place. Articles presenting data for the same study population were combined in one data-extraction table.

Definitions

Cut-off levels for HAV- and HBs antibodies were established as a reliable marker of immediate and long-term protection against infection. The lowest limit of anti-HAV antibodies required to prevent HAV infection has not been accurately defined, but levels of 10, 20, or 33 mIU/ml are used in clinical studies [3,4]. Seroconversion for HAV infection was defined as anti-HAV antibody levels above these cut-off levels (depending on assay used) after full vaccination. Seroconversion following full hepatitis B immunization was defined as an anti-HBs antibody concentration ≥1 mIU/ml, an anti-HBs antibody concentration ≥10 mIU/ml after full vaccination was considered protective against HBV infection [3,6]. Throughout this publication, seroprotection against hepatitis B is therefore defined as titer higher than 10 mIU/ml. Recent publications, however, show that individuals with non-zero anti-HBs levels show an anamnestic response to a challenge dose and indicate the persistence of immune memory to HBsAg even when anti-HBs are well below the estimated protective threshold [17].

Results

The search resulted in 1450 unique hits, of which 47 were included in this systematic review (Figure 1). In total, none of the articles described the efficacy of Twinrix^TM, Twinrix Pediatric^TM, or Ambirix^TM, 42 reported data on immunogenicity, one on effectiveness, and 38 on safety. Two articles reported data retrieved from the Vaccine Adverse Event Reporting System (VAERS) database. The majority of the articles presented data for more than one outcome. Study characteristics of all included articles are presented in Appendix 2.

Figure 1. Flow chart of the selection procedure.

Immunogenicity

Twinrix^TM

Twenty-four articles provided data on the immunogenicity of Twinrix^TM (Tables 2–4).

Table 2. Immunogenicity of Twinrix^TM in adults.

Author	Immunized subjects	Schedule	Time point, months	Anti-HAV response				Anti-HBs response
	Study population; country; age			Seropositivity		GMC		Seroprotection*				GMC
Ambrosch [18]	Young adults, mostly medical students (N = 54); Austria; NR	3 doses at 0, 1, and 6 months	1	98		–		56				–
			2	100		–		98				–
			6	100		–		100				–
			7	100		–		100				–
Bruguera [19]	Students of medicine or nursery (N = 150); Spain; Mean (range): 20 years (18–31)	3 doses at 0, 1, and 6 months						≥1 mIU/ml		≥10 mIU/ml
			1	98.7		394		79.9		–		16
			2	100.0		1311		98.0		–		104
			6	100.0		557		100.0		–		297
			7	100.0		8895		100.0		99.3		7097
Czeschinski [20]	Job starters (N = 300); Germany; Mean (range): 21.8 years (18–30)	3 doses at 0, 1, and 6 months	7	100		5864		99.5				4889
De Schrijver [21]	Adult workers (N = 179); Belgium; Mean: 41.1 years	3 doses at 0, 1, and 6 months						≥1 mIU/ml		≥10 mIU/ml
			6	–		–		91.6		79.9		41.7
			7	–		–		99.4		97.1		749.0
Höhler [22]	Adult twins (N = 98); Germany; Mean: 54 years	3 doses at 0, 1, and 6 months and booster at 33 months	7	100		3019.0		100				1459.1
			Y2.5	100		475.0		70.7				128.6
Joines [23]	Adults (N = 829); US; Mean (range): 39.9 years (19–70)	3 doses at 0, 1, and 6 months	1	91.6		335		17.9				8
			2	97.7		636		61.2				23
			6	98.1		367		86.3				118
			7	99.6		4756		95.1				2099
Kallinowski [12]	Adults (N = 49); Germany; 18–50 years	3 doses at 0, 1, and 6 months	2	100		928		80				30
			6	100		487		88				157
			7	100		6125		100				3709
	HCV-positive adults (N = 34); Germany; 18–58 years	3 doses at 0, 1, 6, months	2	29		17		12				49
			6	71		173		65				72
			7	88		2455		82				1472
Kallinowski [24]	Anti-HAV and anti-HBs negative adults (N = 50); Germany; Mean (range): 30.5 years (18–49)	3 doses at 0, 1, and 6 months	1	90		471		28				58
			2	98		–		50				–
			7	100		5123		94				7314
Knoll [25]	Young medical students and staff members (N = 282); Germany; 17–39 years	3 doses at 0, 1, and 6 months	Anti-HAV status pre-immunization			Negative	Positive	≥1 mIU/ml		≥10 mIU/ml		Negative	Positive
								Negative	Positive	Negative	Positive
			7	100		6238	35481	100	100	100	100	9454	18288
Rendi-Wagner [26]	Adolescents and adults seeking pre-travel immunization advice (N = 990); Austria; Mean (range): 43.4 years (6–80)	3 doses at 0, 1, and 6–12 months	NR (GMC <6 months after 3rd dose)	–		–		92.3				1008
Rieger [27]	Adults (N = 304); Germany; Mean (range): 28.5 years (18–45)	3 doses at 0, 1, and 6 months		<40 years	≥40	<40 years	≥40	≥1 IU/l		≥10 IU/l
			1	91.2	82.4	91.3	68.8	34.6		23.8		1.72
			2	98.2	100.0	281.6	209.1	85.2		61.2		14.2
			6	99.1	100.0	184.0	151.4	96.9		94.7		106.8
			7	100.0	100.0	2940.5	1617.1	98.4		97.6		1684.0
Thoelen [11]	Adults (N = 843); Belgium, Lithuania, the Netherlands, Spain, Switzerland; Mean (range): 20.8 years (17–60)	3 doses at 0, 1, and 6 months						≥1 mIU/ml		≥10 mIU/ml
			1	94.3		306		70.6		33.7		10
			2	99.5		748		97.4		83.9		62
			6	99.3		434		99.2		96.7		236
			7	100.0		5404		99.7		99.3		4814
			24	100.0		1044		–		97.7		328
			36	100.0		948		–		97.9		240
			48	100.0		752		–		95.3		211
Van der Wielen [28]/Chilbek [29]	Seronegative adults (N = 596); Belgium, Czech Republic, Germany; Mean (range): 55.2 years (41–81)	3 doses at 0, 1, and 6 months						≥3.3 mIU/ml		≥10 mIU/ml
			7	97.2		2747		92.8		91.7		1154
			Y4	97.3		212.9		76.9		57.1		42.3
Van Herck [30]/Van Damme, 2012 [14]	Adults (N = 223); Belgium; 17–43 years	3 doses at 0, 1, and 6 months	Center	1	2	1	2	1		2		1	2
			7	100.0	100.0	4610.0	6591	100.0		100.0		2637.3	6601
			Y1.5	100.0	100.0	1071.2	1824	95.7		98.8		288.4	710
			Y2	100.0	100.0	854.3	1330	97.7		97.3		236.8	476
			Y3	100.0	100.0	733.7	1383	98.8		96.3		173.6	383
			Y4	100.0	100.0	571.7	1164	95.8		93.9		138.6	410
			Y5	100.0	100.0	580.5	905	96.9		93.0		115.8	330
			Y6	100.0	100.0	541.7	977	89.4		95.0		89.9	322
			Y6*	100.0	100.0	457.0	769.7	96.1		95.0		143.3	416.7
			Y7	100.0	100.0	400.5	833.0	97.8		91.4		140.9	391.2
			Y8	100.0	100.0	343.7	597.5	100.0		91.4		138.3	362.8
			Y9	100.0	100.0	343.6	573.0	97.5		89.3		125.6	451.5
			Y10	100.0	100.0	373.9	674.6	94.1		86.2		103.8	320.4
			Y11	100.0	100.0	–	–	100.0		92.0		–	–
			Y12	100.0	100.0	–	–	97.0		89.3		–	–
			Y13	100.0	100.0	–	–	100.0		87.0		–	–
			Y14	100.0	100.0	–	–	96.7		87.5		–	–
			Y15	100.0	100.0	–	–	92.9		89.3		–	–
Williams [31]	Residents of two nursing facilities (N = 225); US; 45–101 years	3 doses at 0, 1, and 6 months	30–60 days after complete vaccination	–		–		34				5.79
Wolters [32]	Adults (N = 248); Germany; Mean (range): 39.9 years (16–75)	3 doses at 0, 1, and 6 months	Following vaccination	96.2		–		88.7**				–
Wolters [33]	Adults (N = 104); Germany; Mean (range): 54 years (17–84)	3 doses at 0, 1, and 6 months	Following vaccination	71.2		–		37.5				–

GMC: Geometric mean concentration; HAV: Hepatitis A virus; HBs: Hepatitis B surface protein; HCV: Hepatitis C virus; NR: Not reported; Y: Years; -: Not reported.

Data from multiple articles regarding the same study were combined in the table. All relevant references are stated under ‘Author’.

*All rates are seroprotection, unless clearly stated as seropositivity (i.e. ≥1 mIU/ml).

**Seroprotection was defined as at least 100 IU/l anti-HBs detected [32].

Table 3. Immunogenicity of Twinrix^TM in children.

Author	Immunized subjects	Schedule	Time point, months	Anti-HAV response				Anti-HBs response
	Study population; country; age			Seropositivity		GMC		Seropositivity/protection		GMC
Flynn [16]	HIV-infected youth (N = 118); Bahamas, Brazil, South Africa, US; Mean (range): 20.4 years (12–25)	3 doses at 0, 1, and 6 months	7	–		–		75.4*		97.7
Thompson [34]	Adolescents (N = 160); Australia; Mean: 16.4 years	3 doses at 0, 1, and 6 months (2 lots)		Lot 1	Lot 2	Lot 1	Lot 2	Lot 1	Lot 2	Lot 1	Lot 2
			1	92.3	89.5	254	221	55.1	55.3	13	11
			2	100.0	98.7	574	665	89.7	97.3	29	25
			6	100.0	97.4	307	377	97.4	100.0	238	214
			7	100.0	100.0	5694	6260	98.7	100.0	4316	4436

GMC: Geometric mean concentration; HAV: Hepatitis A virus; HBs: Hepatitis B surface protein; HIV: Human immunodeficiency virus; -: Not reported.

*Outcomes and vaccine response were reported as IU/ml. However, we assume that the outcome measure should be mIU/ml. For the purpose of this review, data are handled as mIU/ml.

Table 4. Immunogenicity of Twinrix^TM following an alternative schedule.

Author	Immunized subjects	Schedule	Time point, months	Anti-HAV response				Anti-HBs response
	Study population; country; age			Seropositivity		GMC		Seroprotection*		GMC
Connor [35]	Adults (N = 250); US, Norway, Belgium, Czech Republic; ≥18 years	4 doses at 0, 7, 21–30, and 12 months	Day 37	98.5		454.6		63.2		15.7
			3	100.0		480.6		83.2		53.4
			12	96.9		–		82.1		–
			13	100.0		7110.5		96.4		7648.1
De Schrijver [21]	Adult workers (N = 194); Belgium; Mean: 42.5 years	3 doses at 0, 1, and 12 months						≥1 mIU/ml	≥10 mIU/ml
			6	–		–		89.2	70.6	43.7
			12	–		–		81.2	55.9	30.2
			13	–		–		98.9	95.6	1900.6
Frey [36]	Individuals (N = 1251); NR; Median (range): 23 years (10–80)	2 doses at 0 and 6 months	1	93.0		36		5.4		0.5
			7	100		5104		84.2		169
			12	99.7		1637		75.9		33
Kallinowski [12]	HCV-positive adults and healthy volunteers (N = 95); Germany; 18–58 years	3 doses at 0, 7, and 21 days		Healthy	HCV	Healthy	HCV	Healthy	HCV	Healthy	HCV
			Day 21	87	39	58	47	39	10	46	53
			2	100	89	424	158	98	77	835	302
			6	100	90	1416	561	98	73	808	297
Nothdurft [37]	Seronegative adults (N = 211); Germany; 18–45 years	4 doses at 0, 7, and 21 days and 12 months	1	100		845		82.0		65
			2	99.5		780		85.0		53
			3	100		628		95.2		183
			12	96.2		374		94.0		209
			13	100		9571		100		26002

GMC: Geometric mean concentration; HAV: Hepatitis A virus; HBs: Hepatitis B surface protein; HCV: Hepatitis C virus; -: Not reported.

*All rates are seroprotection, unless clearly stated as seroconversion (i.e. ≥1 mIU/ml).

Anti-HAV seropositivity 1 month after full vaccination

After full vaccination, the rate of seropositivity for anti-HAV ranged from 96.2% to 100%, regardless of the schedule used or whether the study was done in adults or children (Tables 2–4) [11,12,14,18–20,22–25,27–30,32,34–37].

Two articles reported lower seropositivity rates at month 7. In hepatitis C virus (HCV)-positive adults, the anti-HAV seropositivity was 88% after full vaccination [12]. The other study reported a seropositivity rate of anti-HAV of 71.2% following vaccination in a general population in which the frequency of seropositivity significantly decreased with increasing age; in subjects younger than 40 years, the seropositivity rate was 92%, which decreased to 63% in persons over 60 years [27,33].

The anti-HAV antibody geometric mean concentration (GMC) increased after the first two doses and decreased in all studies before the third dose. After the third dose, the GMC ranged from 2747 to 8895 mIU/ml [11,12,14,19,20,22–25,27–30,34,35], which is well above the seropositivity cut-off levels, inferring seroprotection. However, GMC decreased with increasing age and body mass index (BMI) [23,27,33] was lower in HCV-positive adults [12], and it was lower in smokers compared to nonsmokers [24,27]. One study did not find any factors significantly influencing the anti-HAV seropositivity rates [32]. Since almost all individuals become anti-HAV seropositive after vaccination, the number of nonresponders is very low, making it difficult finding significant risk factors for seropositivity. However, risk factors for GMC can be found due to the large range of GMC after vaccination.

Long term anti-HAV seropositivity

Long-term data showed that 97.3–100% of the adults were seropositive for anti-HAV up to year 4 or 15. The GMC showed a 10-fold decrease (to 373.9 and 674.6 mIU/ml) at year 10, but remained above the seropositivity cut-off values [30].

Anti-HBs seropositivity/seroprotection 1 month after full vaccination

After full vaccination, the rate of seropositivity (i.e. ≥1 mIU/ml) for anti-HBs ranged from 98.4% to 100% [11,19,21,25,27]. In general, the seroprotection rate for anti-HBs ranged from 82% to 100% [11,12,18–28,30,32,34–37]. Three articles reported, however, lower seroprotection rates at month 7. The seroprotection 30–60 days after full vaccination in residents of nursing facilities was only 34% [31]. In another population of relatively old participants (mean age 54 years), the seroprotection rate was 37.5% [33]. Seroprotection for anti-HBs was 75.4% in human immunodeficiency virus (HIV)-infected youth at month 7 [16]. Anti-HBs antibody GMC ranged from 749 to 9454 mIU/ml after the third dose [11,12,14,19–30,34,35].

Risk factors anti-HBs seroprotection

Seroprotection and GMC for anti-HBs decreased with increasing age [23,26,27,32,33,36] or BMI [27,32]. One article found that smokers had significantly lower GMC for anti-HBs antibodies compared to nonsmokers [24], but, two other articles did not find such an effect [27,32]. In residents of nursing homes, no statistically significant differences in anti-HBs response by demographic or clinic factors (i.e. age, gender, diabetes, BMI, or smoking) were found. However, the study was limited by a sample size of 86 participants, which likely decreased the ability to detect statistically significant differences [31]. One article reported on the immunogenicity of Twinrix^TM in HIV-infected youth. Predictors of vaccine response in univariate analyses were female gender, higher nadir cluster of differentiation 4 (CD4), higher baseline CD4 count or percentage, lower baseline viral load, enrollment at a US site, and perinatally infected participants (compared to those infected by high risk behavior) [16]. The effect of underlying chronic diseases on anti-HBs seroprotection rates was inconclusive [32,33]. Interestingly, GMC following full vaccination was significantly lower if the third dose was performed in the summer or autumn, compared to the winter of spring. Seroprotection rates were not influenced by seasonality [26].

Long-term persistence of circulating anti-HBs antibodies

One article on follow-up up to 10 years showed approximately a 20-fold decrease in GMC from 2637.3 or 6601 mIU/ml (depending on study center) at month 7 to 103.8 and 320.4 mIU/ml at year 10, respectively [30]. However, these levels are still well above the seroprotection cut-off values. After 15 years, this study population showed a decrease from 100% seroprotection at month 7 to 89.3% or 92.9% (depending on study center) [14]. One article reported a larger decrease from 92.8% at month 7 to 76.9% at year 4 for seropositivity and from 91.7% to 57.1% at year 4 for seroprotection. The study population was relatively old (>40 years old), which may explain the low seropositivity and seroprotection rates [29]. Subgroup analysis showed lower seropositivity rates at year 4 in subjects aged ≥61 years old, with a BMI ≥30 kg/m², receiving concomitant medication or with a current concomitant medical condition [29].

Twinrix pediatric^TM

Ten articles reported the immunogenicity of Twinrix pediatric^TM in children and/or adolescents (Tables 5 and 6).

Table 5. Immunogenicity of Twinrix Pediatric^TM in children and adolescents.

Author	Immunized subjects	Schedule	Time point/age cohort	Anti-HAV response				Anti-HBs response
	Study population; country; age			Seropositivity		GMC		Seroprotection*		GMC
Levie [38]/Beran [15]	Adolescents (N = 148); Belgium, Czech Republic; Mean (range): 13.4 years (12–15)	3 doses at 0, 1, and 6 months	1	93.2		227		29.1		10
			2	99.3		549		85.6		42
			6	99.3		299		98.0		305
			7	100		4174		100		5054
			7**	100		4516		100		4865
			24**	100		725		96.2		337
			120**	100		335.5		85.1		60.1
Ostergaard [39]	Adolescents (N = 122); Denmark, Sweden; Mean (range): 14.3 years (11–17)	3 doses at 0, 1, and 6 months		HAB	MTT + HAB	HAB	MTT + HAB	HAB	MTT + HAB	HAB	MTT + HAB
			0	2.0	3.0	7.6	7.9	0.0	0.3	1.7	1.7
			7	100	100	6739.0	5876.7	100	99.1	7654.7	6088.2
Pedersen [40]	Girls (N = 520); Canada, Denmark, Hungary, Sweden; 11.2 years	3 doses at 0, 1, and 6 months		HAB	HPV + HAB	HAB	HPV + HAB	HAB	HPV + HAB	HAB	HPV + HAB
			7	100	100	5288.4	4504.2	100	98.3	5646.5	3136.5
Roberton [41] /Marshall [10]	Children (N = 191); Australia, Belgium, Spain, Sweden; 1–11 years	3 doses at 0, 1, and 6 months	7	100		9257		100		13,683
			60	100		698.4		97.0		283.7
Van Damme [42]/Diaz Mitoma [43]	Children (N = 166); Canada; 1–15 years	3 doses at 0, 1, and 6 months		1–6 years	6–15 years	1–6 years	6–15 years	1–6 years	6–15 years	1–6 years	6–15 years
			7	100	100	6518	8499	100	100	7255	10513
			36	100	100	761	1250	97.5	98.9	339	518
			48	100	100	653	937	97.5	98.5	297	421
			60	100	100	479	881	97.4	95.2	195	263
			90	100	100	233	702	86.5	92.9	147	297
			120	–	100	–	680	–	95.5	–	162

GMC: Geometric mean concentration; HAV: Hepatitis A virus; HAB: Hepatitis A and B vaccine; HBs: Hepatitis B surface protein; HPV: Human papillomavirus; MTT: Tetravalent meningococcal conjugate vaccine having each of the polysaccharide coupled to tetanus toxoid; -: Not reported.

*All rates are seroprotection, unless clearly stated as seroconversion (i.e. ≥1 mIU/ml).

**Tests performed with new kits or equivalent validated in-house enzyme-linked immunosorbent assay (ELISA).

Data from multiple articles regarding the same study were combined in the table. All relevant references are stated under ‘Author’.

Table 6. Immunogenicity of Twinrix Pediatric^TM in children following an alternative schedule.

Author	Immunized subjects	Schedule	Time point/age cohort	Anti-HAV response		Anti-HBs response
	Study population; country; age			Seropositivity	GMC	Seropositivity/protection		GMC
Duval [44]/Gilca [45]	Children (N = 347); Canada; 8–10 years	2 doses at 0 and 6 months and challenge dose at Y7				≥1 mIU/ml	≥10 mIU/ml
			Post-second dose	100	–	97.1	96.5	3248
			Pre-challenge dose	97.8	275	92	75.1	60
			Post-challenge dose	100	8203	–	100	39573

GMC: Geometric mean concentration; HAV: Hepatitis A virus; HBs: Hepatitis B surface protein; -: Not reported.

Data from multiple articles regarding the same study were combined in the table. All relevant references are stated under ‘Author’.

Seropositivity/seroprotection after full vaccination

From month 7 to year 10 after the first vaccine dose, all subjects (100%) remained seropositive for anti-HAV antibodies [10,15,38-45]. After the third dose (month 7), the anti-HAV antibody GMC ranged from 4174 to 9257 mIU/ml. After 10 years, GMC decreased to 335.5 and 680 mIU/ml [15,43]. However, these levels are still well above the seropositivity cut-off values, inferring seroprotection.

At month 7, all subjects (100%) were seroprotected for anti-HBs antibodies [38-44]. Follow-up data were available up to 10 years after the primary vaccination; the percentage of seroprotected subjects decreased slightly, but remained above 85% [10,15,43]. After the third dose (month 7), the anti-HBs antibody GMC ranged from 4865 to 13683 mIU/ml. GMC decreased over time after vaccination. After 10 years, GMC was reported to be 60.1 and 162 mIU/ml, which is above the cut-off value for seroprotection [15,43].

Risk factors

Anti-HAV and anti-HBs antibody GMCs tended to be higher in children vaccinated when they were 6–15 years old compared to children vaccinated when they were 1–6 years old [42,43]. However, the sample size was too small to make any definite conclusion [43].

Alternative schedule

One study reported immunogenicity after 2 doses of Twinrix pediatric^TM at 0 and 6 months and a challenge dose after 7 years (Table 6). After the second dose, the seropositivity rates were 100% and 97.1% for anti-HAV and anti-HBs, respectively [44]. Seven years after primary vaccination seropositivity rates slightly decreased to 97.8% and 92%, respectively. The anti-HBs seroprotection rate decreased from 96.5% after the second dose to 75.1% after 7 years. After the challenge dose, 100% of the children seroconverted for anti-HAV and anti-HBs [44,45].

Ambirix^TM

Thirteen articles reported the immunogenicity of Ambirix^TM in children and/or adolescents (Tables 7 and 8).

Table 7. Immunogenicity of Ambirix^TM in children and adolescents.

Author	Immunized subjects	Schedule	Time point, months	Anti-HAV response				Anti-HBs response
	Study population; country; age			Seropositivity		GMC		Seroprotection*		GMC
Belderok [46]	Children with HIV and children receiving IM for rheumatic diseases (N = 160); the Netherlands; 1–16 years	2 doses at 0 and 26-30 weeks		HIV	IM	HIV	IM	HIV	IM	HIV	IM
			Dose 1	71	55	40	31	27	17	5	3
			Dose 2	99	100	327	288	97	93	483	321
Burgess [47]/Burgess [48]	Adolescents (N = 106); Australia; 12–15 years	2 doses at 0 and 6 months	1	98.1		365		43.4		17
			6	95.2		140		59.0		17.3
			7	100.0		5991.7		98.1		2791.4
			24	100.0		743.5		91.0		248.2
			36	98.8		620.3		81.9		149.3
			60	98.1		536.3		81.5		81.5
			72	100.0		529.0		84.8		108.7
Cunningham [49]	HIV-negative urban youth (N = 63); US; 15.2 years	2 doses at 0 and 6 months	7	–		–		94.6		–
Lee [50]	Youngsters (N = 51); China; 10–19 years	2 doses at 0,6 months						≥1 mIU/ml	≥10 mIU/ml
			1	98.0		379		61.2	51.0	55
			2	95.9		210		73.5	40.8	19
			6	85.7		122		79.6	51.0	17
			7	100		3701		93.9	91.8	1524
Levie [38]/Beran [15]	Adolescents (N = 142); Belgium, Czech Republic; Mean (range): 13.4 years (12–15)	2 doses at 0 and 6 months	1	99.3		349		43.0		14
			2	100		245		38.0		10
			6	100		178		68.3		20
			7	100		5487		97.9		4949
			7*	100		5242		99.3		4922
			24*	100		943		93.3		359
			120*	100		429.3		85.9		50.6
Ramonet [51]	Children and adolescents (N = 168); Argentina, Brazil, Mexico; 4–20 years	2 doses at 0 and 6 months	1	96.9		502		43.7		16
			2	98.1		284		55.6		18
			6	93.0		185		67.5		32
			7	100		6635		98.7		3362
Roberton [41] /Marshall [10]	Children (N = 204); Australia, Belgium, Spain, Sweden; 1–11 years	2 doses at 0 and 6 months	7	100		8412		98.5		7894
			60	100		576.8		94.1		150.2
Usonis [52]**	Infants (N = 90); Estonia, Lithuania; 11–18 months	2 doses at 0 and 6 months		Group 1	Group 2	Group 1	Group 2	Group 1	Group 2	Group 1	Group 2
			0	0.0	2.7	17	17	0.0	0.0	0.5	0.5
			7	100.0	100.0	8736	11016	100.0	100.0	23092	32540
Van Damme 2011 [53]	Children (N = 232); Belgium; 1–11 years	2 doses at 0 and 6 months	7	100		14790.1		98.5		9389.1
			36	100		1281.6		95.0		338.8
			48	100		874.1		89.9		264.5
			60	100		721.1		85.8		187.0
			72	100		692.3		81.7		206.2
			84	100		753.6		84.6		157.5
			96	100		544.4		81.1		102.7
			108	100		479.5		87.6		89.1
			120	100		601.6		81.7		80.7
Van der Wielen [54]	Children (N = 236); Belgium; 1–11 years	2 doses at 0 and 6 months		<6 y	≥6 y	<6 y	≥6 y	<6 y	≥6 y	<6 y	≥6 y
			1	100	99.2	339	493	22.1	34.6	5	10
			2	95.7	100	264	308	59.9	40.5	14	9.5
			6	100	96.9	202	189	84.8	74.8	48	28
			7	100	100	12050	11293	98.5	98.5	8985	7622

GMC: Geometric mean concentration; HAV: Hepatitis A virus; HBs: Hepatitis B surface protein; HIV: Human immunodeficiency virus; IM: Immunosuppressive medication; -: Not reported.

*All rates are seroprotection, unless clearly stated as seroconversion (i.e. ≥1 mIU/ml).

**In group 1, Ambirix^TM was administered concomitantly with DTPa-IPV/Hib vaccine. In group 2, Ambirix^TM was administered concomitantly with MMR vaccine.

Data from multiple articles regarding the same study were combined in the table. All relevant references are stated under ‘Author’.

Table 8. Immunogenicity of Ambirix^TM in adolescents following an alternative schedule.

Author	Immunized subjects	Schedule	Time point, months	Anti-HAV response		Anti-HBs response
	Study population; country;aAge			Seropositivity	GMC	Seroprotection*	GMC
Burgess [47] /Burgess [48]	Adolescents (N = 102); Australia; 12–15 years	2 doses at 0 and 12 months	1	99.0	293.9	34.3	15.0
			12	84.2	146.5	50.5	16.1
			13	99.0	8471.8	97.0	4339.5
			24	98.8	1704.7	93.8	569.6
			36	98.8	1273.6	87.2	400.5
			60	100.0	880.6	93.0	213.5
			72	100.0	932.6	92.9	231.9

GMC: Geometric mean concentration; HAV: Hepatitis A virus; HBs: Hepatitis B surface protein.

*All rates are seroprotection, unless clearly stated as seroconversion (i.e. ≥1 mIU/ml).

Data from multiple articles regarding the same study were combined in the table. All relevant references are stated under ‘Author’.

Seropositivity/seroprotection after full vaccination

After full vaccination following the regular immunization schedule, all children were seropositive for anti-HAV [38,41,48,50–54]. Five to 10 years after primary vaccination, all children (100%) remained seropositive for anti-HAV [10,15,47,53]. After the second dose (month 7), GMC was 3701–14790.1 mIU/ml [10,15,38,41,47,48,50–54]. Long-term follow-up data showed that GMC decreased over the years, but remained above the seropositivity cut-off up to 10 years follow-up [10,15,47,53].

In one study, anti-HBs seropositivity was measured and was 93.9% after full vaccination [50], and seroprotection rates ranged from 91.8% to 100% [38,41,48–54]. Seroprotection rates decreased to at least 81.7% 10 years after primary vaccination [10,15,47,53]. GMC ranged from 1524 to 32540 mIU/ml 7 months after primary vaccination [38,41,48,50–54]. After 10 years GMC decreased to 50.6–80.7 mIU/ml [15,53].

Risk factors

One article reported the immunogenicity of Ambirix^TM in children with HIV or children receiving immunosuppressive medication (IM) for rheumatic diseases [46]. Seroprotection rates after full vaccination were 99% and 100% for anti-HAV and 97% and 93% for anti-HBs in children with HIV or receiving IM, respectively. The GMCs found after full vaccination in HIV-infected children and children on IM were 327 and 288 mIU/ml for anti-HAV antibodies and 483 and 321 mIU/ml for anti-HBs antibodies, respectively. In children infected with HIV, no statistical differences in anti-HAV antibody GMC were found regarding CD4 count, viral load, or use of combination antiretroviral therapy (cART). For HBV, a significantly higher anti-HBs antibody GMC was found in children with lower viral load and children on cART [46].

Alternative schedule

One study described the immunogenicity following two doses of Ambirix^TM at 0 and 12 months (Table 8). Seropositivity/protection rates at month 13 were 99.0% and 97.0% for anti-HAV and anti-HBs, respectively. Six years after primary vaccination the seropositivity/protection rates were 100% and 92.9%, respectively. The study demonstrated long-term persistence of immunity to hepatitis A and B, following two doses of Ambirix^TM, when given either at 0 and 6 months or 0 and 12 months [47].

Twinrix^TM vs. monovalent hepatitis A and B vaccines

Seventeen articles described a direct comparison of the immunogenicity of Twinrix^TM and monovalent hepatitis A and/or B vaccines (Tables 9–11). The anti-HAV seropositivity rates were equal for both vaccines in all studies. Anti-HAV antibody GMC was equal or higher following full vaccination with Twinrix^TM or Ambirix^TM compared to full vaccination with Havrix^TM or unspecified hepatitis A vaccines [35,50,51]. Twinrix^TM was equal or higher than other monovalent hepatitis B vaccines with regard to the anti-HBs response (Tables 9–11).

Table 9. Comparison between Twinrix^TM and concomitant administration of monovalent hepatitis A and B vaccines.

Author	Population	Vaccine and schedule		Time point	Anti-HAV response							Anti-HBs response
					Seropositivity			GMC				Seroprotection*							GMC
Ambrosch [18]	Young adults, mostly medical students; N = 162 (3 x 54); Age: NR	[1] Twinrix^TM: 3 doses at 0, 1, and 6 months [2] Havrix^TM + Engerix-B^TM (Simultaneous): 3 doses at 0, 1, and 6 months [3] Havrix^TM + Engerix-B^TM (Mixed): 3 doses at 0, 1, and 6 months		Vaccine	[1]	[2]	[3]					[1]		[2]			[3]
				1	98	98	100	–				56		78			65		–
				2	100	100	100	–				98		96			98		–
				6	100	98	100	–				100		100			98		–
				7	100	100	100	–				100		100			100		–
Connor [35]	Adults; N = 496; Age ≥18 years	[1] Twinrix^TM: 4 doses at 0, 7, and 21–30 days and 12 months [2] Havrix^TM + Engerix-B^TM: 2 and 4 doses at 0 (1, 2), and 12 months		Vaccine	[1]	[2]		[1]	[2]			[1]		[2]					[1]		[2]
				Day 37	98.5	98.6		454.6	241.7			63.2		43.5					15.7		7.5
				3	100.0	95.6		480.6	120.7			83.2		76.7					53.4		52.2
				12	96.9	86.9		–	–			82.1		77.8					–		–
				13	100.0	100.0		7110.5	3235.7			96.4		93.4					7648.1		4380.9
Czeschinski [20]	Job starters; N = 300; Mean age (range): 21.8 years (18–30)	[1] Twinrix^TM: 3 doses at 0, 1, and 6 months [2] Havrix^TM + Engerix-B^TM: 3 doses at 0, 1, and 6 months [3] Havrix^TM or Engerix-B^TM: 3 doses at 0, 1, and 6 months		Vaccine				[1]	[2]		[3]								[1]		[2]	[3]
				7	–			5997	5864		2707	–							4889		8212	4931
Flynn [16]	HIV-infected youth; N = 336; Mean age (range): 20.4 years (12–25)	[1] Twinrix^TM: 3 doses at 0, 1, and 6 months [2] Engerix-B^TM 20 mcg: 3 doses at 0, 1, and 6 months [3] Engerix-B^TM 40 mcg: 3 doses at 0, 1,and 6 months		Vaccine								[1]		[2]			[3]		[1]		[2]	[3]
				7	–			–				75.4		60			73.2		97700		52500	77600
Frey [36]	Individuals; N = 1385; Median age (range): 23 years (10–80)	[1] Twinrix^TM: 2 doses at 0,6 months [2] Vaqta + Recombivax-HB: 2 doses at 0 and 6 months		Vaccine	[1]	[2]		[1]		[2]		[1]		[2]					[1]		[2]
				1	93.0	89.1		36		31		5.4		7.9					0.5		0.7
				7	100	99.2		5104		3555		84.2		91.8					169		588
				12	99.7	99.2		1637		1263		75.9		83.6					33		110
Joines [23]	Adults; N = 829; Mean age (range): Twinrix^TM: 39.9 years (19–70)/Havrix^TM + Engerix-B^TM: 40.4 years (19–83)	[1] Twinrix^TM: 3 doses at 0, 1, and 6 months [2] Havrix^TM + Engerix-B^TM: 3 doses at 0, 1, and 6 months**		Vaccine	[1]	[2]		[1]		[2]		[1]		[2]					[1]		[2]
				1	91.6	98.1		335		444		17.9		7.5					8		6
				2	97.7	98.9		636		257		61.2		50.4					23		18
				6	98.1	98.9		367		196		86.3		75.7					118		45
				7	99.6	99.3		4756		2948		95.1		92.2					2099		1871
Knöll [25]	Young medical students and staff members; N = 282; Age range: 17–39 years	[1] Twinrix^TM: 3 doses at 0, 1, and 6 months [2] Engerix-B^TM: 3 doses at 0, 1, and 6 months	Pre-immunization anti-HAV status	Vaccine	[1]	[2]		[1]		[2]		≥1 mIU/ml		≥10 mIU/ml					[1]		[2]
												[1]	[2]	[1]			[2]
			Negative	7	100	–		6238		–		100	100	100			99		9454		7826
			Positive	7				35481				100	100	100			100		18288		8813
Nothdurft [37]	Seronegative adults; N = 479; Age range: 18–45 years	[1] Twinrix^TM: 4 doses at 0, 7, and 21 days and 12 months [2] Havrix^TM: 2 doses at 0 and 6 months Engerix-B^TM: 4 doses at 0, 7, and 21 days and 12 months		Vaccine	[1]	[2]		[1]		[2]		[1]		[2]					[1]		[2]
				1	100	99.0		845		512		82.0		83.9					65		98
				2	99.5	98.9		780		286		85.0		84.9					53		58
				3	100	97.8		628		219		95.2		90.8					183		131
				12	96.2	95.0		374		170		94.0		91.6					209		106
				13	100	100		9571		5205		100		100					26002		29196
Rendi-Wagner [26]	Adolescents and adults seeking pre-travel immunization advice; N = 1874; Mean age (range): 43.4 years (6–80)	[1] Twinrix^TM: 3 doses at 0, 1, and 6-12 months [2] Engerix-B^TM: 3 doses at 0, 1, and 6–12 months [3] GEN H-B-VAX: 3 doses at 0, 1, and 6–12 months		Vaccine								[1]	[2]			[3]			[1]	[2]	[3]
				<6 months after 3^rd dose**	–			–				92.3	87.7			79.3			1008	523	142
Rieger [27]	Adults; N = 304; Mean age (range): 28.5 years (18–45)	[1] Twinrix^TM: 3 doses at 0, 1, and 6 months [2] Vaqta + GEN H-B-VAX: 3 doses at 0, 1, and 6 months***		Vaccine	[1]	[2]		[1]		[2]			≥1 IU/l			≥10 IU/l			[1]		[2]
													[1]		[2]	[1]		[2]
			<40y	1	91.2	98.1		91.3		173.9		1	34.6		20.0	23.8		8.0	1.72		0.8
				2	98.2	99.0		281.6		157.4
				6	99.1	97.1		184.0		152.3		2	85.2		70.1	61.2		53.2	14.2		6.6
				7	100.0	100.0		2940.5		2500.3
			≥40y	1	82.4	94.7		68.8		108.2		6	96.9		90.3	94.7		79.0	106.8		27.2
				2	100.0	89.5		209.1		83.2
				6	100.0	89.5		151.4		74.5		7	98.4		99.1	97.6		98.4	1684.0		528.2
				7	100.0	100.0		1617.1		1275.6
Van der Wielen [28]/ Chlibek [29]	Seronegative adults; N = 596; Mean age (range): 55.2 years (41–81)	[1] Twinrix^TM: 3 doses at 0, 1, and 6 months [2] Havrix^TM + Engerix-B^TM: 3 doses at 0,1, and 6 months*** [3] Vaqta + HB VAX PRO: 3 doses at 0, 1, and 6 months***		Vaccine	[1]	[2]	[3]	[1]		[2]	[3]	≥3.3 mIU/ml			≥10 mIU/ml				[1]	[2]	[3]
												[1]	[2]	[3]	[1]	[2]		[3]
				7	97.2	98.9	98.9	2747		1394	3707	92.8	83.5	77.8	91.7	79.7		71.0	1154	491	179
				48	97.3	93.9	96.0	212.9		165.7	277.4	76.9	61.9	51.6	57.1	40.1		26.6	42.3	23.6	13.7

GMC: Geometric mean concentration; HAV: Hepatitis A virus; HBs: Hepatitis B surface protein; HIV: Human immunodeficiency virus; NR: Not reported

*All rates are seroprotection, unless clearly stated as seroconversion (i.e. ≥1 mIU/ml).

**Time point meting seroconversion: NR.

***Only two doses of Havrix^TM and/or Vaqta were administered at months 0 and 6.

Data from multiple articles regarding the same study were combined in the table. All relevant references are stated under ‘Author’.

Table 10. Comparison between Twinrix pediatric^TM and concomitant administration of monovalent hepatitis A and B vaccines.

Author	Population	Vaccine and schedule	Time point	Anti-HAV response				Anti-HBs response
				Seropositivity		GMC		Seroprotection		GMC
Duval [44] /Gilca [45]	Children; N = 704; Age 8–10 years	Twinrix pediatric^TM [TX]: 2 doses at 0 and 6 months and challenge dose at Y7 [2] Recombivax-HB [RB]: 2 doses at 0 and 6 months and challenge dose at Y7	Vaccine	[TX]	[RB]	[TX]	[RB]	[TX]	[RB]	[TX]	[RB]
			Post-second dose	100	–	–	–	–	–	3248	742
			Pre-challenge dose	–	–	275	–	75	55	60	12
			Post-challenge dose	–	–	8203	–	100	98	39573	5114

GMC: Geometric mean concentration; HAV: Hepatitis A virus; HBs: Hepatitis B surface protein; RB: Recombivax-HB; TX: Twinrix pediatric^TM; -: Not reported.

Data from multiple articles regarding the same study were combined in the table. All relevant references are stated under ‘Author’.

Table 11. Comparison between Ambirix^TM and concomitant administration of monovalent hepatitis A and B vaccines.

Author	Population	Vaccine and schedule	Time point	Anti-HAV response				Anti-HBs response
				Seropositivity		GMC		Seroprotection*				GMC
Cunningham [49]	HIV-negative urban youth; N = 123; Mean age: 15.2 years	[1] Ambirix^TM: 2 doses at 0 and 6 months	Vaccine					[1]		[2]
		[2] Recombivax-HB: 2 doses at 0and 6 months	1	–		–		94.6		87.2		–
			Week 76	–		–		88.0		81.1		–
Lee [50]	Chinese youngsters; N = 103; Mean age (range): Ambirix^TM: 14.1 y (10-19)/Simultaneous: 12.4 years (10–19)	[1] Ambirix^TM 2 doses at 0 and 6 months	Vaccine	[1]	[2]	[1]	[2]	≥1 mIU/ml		≥10 mIU/ml		[1]	[2]
								[1]	[2]	[1]	[2]
		[2] Unspecified simultaneous hepatitis A and B vaccines: 2 doses at 0 and 6 months	1	98.0	97.9	379	219	61.2	70.2	51.0	48.9	55	36
			2	95.9	91.5	210	136	73.5	70.2	40.8	42.6	19	23
			6	85.7	95.1	122	74	79.6	72.3	51.0	42.6	17	15
			7	100	100	3701	1705	93.9	95.7	91.8	95.7	1524	720
Ramonet [51]	Children and adolescents; N = 248; Age range: 4–20 years	[1] Ambirix^TM: 2 doses at 0 and 6 months [2] Havrix^TM + Engerix-B^TM: 2 doses at 0 and 6 months	Vaccine	[1]	[2]	[1]	[2]	[1]		[2]		[1]	[2]
			1	96.9	97.3	502	342	43.7		28.8		16	13
			2	98.1	94.8	284	188	55.6		48.7		18	22
			6	93.0	91.9	185	134	67.5		68.9		32	26
			7	100	98.6	6635	2728	98.7		95.9		3362	1724

GMC: Geometric mean concentration; HAV: Hepatitis A virus; HBs: Hepatitis B surface protein; HIV: Human immunodeficiency virus; -: Not reported.

*All rates are seroprotection, unless clearly stated as seroconversion (i.e. ≥1 mIU/ml).

Twinrix pediatric^TM or Ambirix^TM co-administered with other vaccines

Co-administration of Twinrix pediatric^TM or Ambirix^TM with other vaccines (i.e. quadrivalent conjugate meningococcal vaccine [MenACWY-TT], human papilloma virus [HPV], diphtheria, tetanus, acellular pertussis – inactivated poliovirus/Haemophilus influenzae type b vaccine [DTPa-IPV/Hib] or measles, mumps, rubella vaccine [MMR]) was immunologically noninferior to administration of the vaccine alone [39,40,52].

Effectiveness

One article reported data on the effectiveness of Twinrix^TM. The incidence rate of hepatitis A in Catalonia decreased from 5.44 per 100,000 person-years before universal adolescent vaccination (1992–1998) to 3.02 per 100,000 person-years after universal adolescent vaccination (2001–2006) (p < 0.001). The decrease in incidence rates of hepatitis A after introduction of universal adolescent vaccination was statistically significant in male (6.85 vs. 3.89 per 100,000 person-years, p < 0.001) and female participants (4.10 vs. 2.18 per 100,000 person-years, p < 0.001) [55].

Safety

Twinrix^TM

The most reported local solicited reaction was soreness/pain at injection site (10.5–65.6%; Table 12). The most reported systemic solicited reaction differed between articles (i.e. fatigue, headache, and malaise). Subjects receiving Twinrix^TM alone at a travel clinic had significantly lower relative risk for experiencing any general symptoms compared to subjects receiving one or more vaccines concomitantly (p > 0.05) [56]. In HCV-infected patients, the most reported local reaction was soreness at injection site and the most reported systemic reaction was headache [12].

Table 12. Solicited and unsolicited adverse events following immunization with Twinrix^TM, Twinrix pediatric^TM, or Ambirix^TM.

Reaction	Twinrix^TM, range	Twinrix pediatric^TM, range	Ambirix^TM, range
Local reactions, %
Any	41.6–65.6 [24,34,35,37]	–	50.4–54.3 [48]
Soreness/pain	10.5–65.6 [12,19–24,27,34,35,37,56]	39.1–52.2 [38–41]	13.8–54.0 [38,41,48,50–52,54]
Redness	0.0–26.0 [12,19–23,27,34,35,37,56]	11.9–30.8 [38,40,41]	1.0–29.4 [38,41,48,50–52,54]
Swelling	0–20.6 [12,19–23,27,34,35,37,56]	4.9–21.1 [38,40,41]	0.8–15.7 [38,41,48,50–52,54]
Systemic reactions, %
Any	2–58.0 [24,34,35,37]	–	40.9 [48]
Fever	0.5–7.8 [12,20,21,23,34,35,37,56]	1.3–15.6 [38,40,41]	1.7–13.3 [38,41,48,50–52,54]
Headache	2.1–38.7 [12,19–23,34,35,37,56]	13.9–42.6 [38,40,41]	7.0–31.5 [38,41,48,50–52,54]
Malaise	0.5–29.0 [12,20,21,34,37,56]	–	7.5 [51]
Fatigue	2.6–42.2 [19,21–23,34,35,37,56]	19.3–32.0 [38,40,41]	5.0–29.2 [38,41,48,50–52,54]
Nausea	1.3–11.9 [12,20,23,24,34,37]	–	4.4 [51]
Vomiting	0–7.0 [12,23,34,37,56]	–	1.2 [51]
Gastrointestinal	1.7–7.5 [21–23,35]	3.4–30.3 [38,40,41]	4.3–22.9 [38,41,48,51,52,54]
SAE, n	0–21 [12,22,23,27,34,35,37]	1–5 [38–41]	0–6 [38,41,48,50–52,54]
Unsolicited events, n	0–109 [19,22,35,37,56]	18–89 [39,41]	0–132 [41,48,50,54]

SAE: Serious adverse event; -: Not reported.

The percentage of individuals experiencing unsolicited adverse events (AEs) ranged between 0% and 20.8% [19,22,35,37,56]. Reported unsolicited events included tonsillitis, pharyngitis, cold, influenza, otitis, diarrhea, dizziness, upper respiratory tract infection, and local reactions or bruising at the injection site.

The percentage of individuals experiencing serious adverse events (SAEs) ranged between 0% and 9.5% [11,12,14,16,18,21,23,27,28,30,32,34,35,37]. Only one article reported a SAE related to the vaccine: a participant developed simultaneous headache, numbness of the face, and vomiting, each of moderate intensity [37].

Between May 2001 and September 2003, VAERS received 305 reports of possible AEs after vaccination with Twinrix^TM. Of these reports, 67.9% listed Twinrix^TM as the sole vaccine. After vaccination with only Twinrix^TM, the reported AEs were related to neurologic conditions (34.7%), liver and gastrointestinal tract (18.2%), integument (9.1%), flu-like syndrome (7.4%), musculoskeletal disorders (7.4%), allergy or anaphylaxis (6.6%), infections (4.1%), or other (12.4%) [57]. Pregnant women filed 35 reports following Twinrix^TM vaccination between 1996 and 2013, 40% of the reports did not describe an AE, 16 were maternal events, and 5 were infant/neonatal events. Reported AE included pregnancy specific AE (spontaneous abortion, elective termination, premature delivery, oligohydramnios, abruption placentae, uterine, and abdominal cramps), nonpregnancy specific AE (pelvic pain, sinusitis, nausea and dizziness, paleness, and injection site pain), and infant AE (jaundice, fetal distress, and growth under lower lip). Two serious reports after Twinrix^TM in pregnant women included oligohydramnios and neonatal jaundice [58].

Twinrix pediatric^TM

The most reported local solicited reaction was soreness/pain at injection site (Table 12). The most reported systemic reactions were fatigue and headache. Two articles reported the number of unsolicited AEs; in one study, 14.8% of the adolescents experienced an unsolicited AE [39]. In the other study, 17.4% of the children reported at least one unsolicited AE related to vaccination [41]. The nature of these AEs was not reported.

SAEs were reported in 0.4% and 0.8% of the participants following immunization with Twinrix pediatric^TM [39,41]. Only one SAE was considered to be associated with vaccination: vomiting, fever with symptoms of rhinorrhea, and cough occurring four days after receiving the first vaccine dose. The subject was hospitalized and diagnosed with viral upper respiratory tract infection and exacerbation of eczema [41].

Ambirix^TM

The most reported local solicited reactions were soreness/pain at injection site (Table 12) and redness. The most reported solicited systemic reactions were fever, fatigue, and headache (Table 12). The percentage of participants experiencing unsolicited AEs ranged between 0% and 30.2% [41,48,50,54]. The kind of unsolicited event was not reported.

The percentage of SAEs ranged from 0% to 1.7% [38,41,48,50,54]. When Ambirix^TM was co-administered with DTPa-IPV/Hib vaccine, 5% of the participants experienced a SAE, and 4.6% of the participants after had a SAE following co-administration with MMR vaccine [52]. One SAE was reported unlikely to be associated with vaccination (acute diabetes mellitus) [48] and one was reported to be associated with vaccination (febrile convulsions) [52]. All other SAEs were not associated with vaccination.

Combined vs. separate hepatitis A and B vaccines

The safety profiles of the combined vaccines and unspecified concomitant hepatitis A and B vaccines, Engerix-B^TM, simultaneous Engerix-B^TM and Havrix^TM, simultaneous Vaqta and Gen H-B-Vax and Recombivax-HB were similar [16,18,23,27,35,37,49–51]. However, the group receiving the mixed administration of Havrix^TM and Engerix-B^TM had a total local symptom score twice as high as the other two groups, probably due to the twofold volume of the vaccine and the twofold amount of aluminum hydroxide in the vaccine [18]. Furthermore, while the percentage of recorded pain reactions, indurations, or swellings was similar after Engerix-B^TM and Twinrix^TM, it was lower after Havrix^TM [20]. The percentage of reported AEs was higher following Twinrix^TM than in the group receiving concurrent Vaqta and Recombivax-HB/H-B-VaxII [36].

Twinrix pediatric^TM or Ambirix^TM co-administered with other vaccines

Co-administration of Twinrix pediatric^TM or Ambirix^TM with other vaccines (i.e. MenACWY-TT, HPV, DTPa-IPV/Hib, or MMR) did not significantly alter the safety profile [39,40,52]. However, the incidence of grade 3 pain was slightly higher when Twinrix pediatric^TM was co-administered with HPV vaccine than when administered alone [40]. Both local and general symptoms were more frequently reported after co-administered hepatitis A/B and MMR vaccination (48% and 46%, respectively) compared to co-administered hepatitis A/B and DTPa-IPV/Hib vaccination (18% and 32%, respectively) [52].

Discussion

Summary of results

After full vaccination with Twinrix^TM (regular and accelerated schedule), the rate of seropositivity for anti-HAV ranged from 96.2% to 100%. The rate of seroprotection for anti-HBs after full vaccination with Twinrix^TM ranged from 82% to 100%. Increasing age was associated with decreasing immunogenicity response. Immunogenicity results were equal or higher for both anti-HAV and anti-HBs following Twinrix^TM vaccination compared to monovalent hepatitis A or B vaccination. Co-administration of Twinrix pediatric^TM or Ambirix^TM with other routine childhood vaccines was immunologically noninferior to administration of the combined hepatitis A and B vaccine alone and did not significantly alter the safety profile. The safety profiles of the combined vaccines and monovalent hepatitis A and B vaccines were similar.

Risk groups for hepatitis A and/or B infection

Patients with chronic hepatitis C are at increased risk of complication once exposed to hepatitis A or B. Therefore, vaccination against these vaccine preventable diseases has been recommended for patients with chronic liver disease caused by hepatitis [12]. The immunogenicity of both monovalent and combined hepatitis A and B vaccines was lower in patients with chronic hepatitis C infection [12]. Unfortunately, the adherence to hepatitis vaccination guidelines for patients with chronic liver disease is low and there are significant variations among providers [59].

Vaccination for both HAV and HBV is also recommended in HIV-infected subjects [16]. Protection for both infections can be achieved by vaccination with Twinrix^TM, which potentially improves the patient acceptance and reduces the costs [16]. The immune response in children with HIV after full vaccination is satisfactory and comparable to the response in healthy children. However, a substantial proportion of these children was not protected after the first dose, which should be taken into account in travel health when full vaccination may not be possible due to time restrictions [46].

Although there is a lot of attention on immunization of high risk groups, only three studies describing the immunogenicity of Twinrix^TM, Twinrix pediatric^TM, or Ambirix^TM in high risk populations (i.e. HCV- or HIV-infected patients or patients on IM for rheumatic diseases) were found [12,16,46]. In children infected with HIV or on IM, the immunogenicity of Ambirix^TM was similar to healthy children. The GMC was lower but remained above the cut-off levels [46]. In HIV-infected youth, the anti-HBs seroprotection rate was lower than in healthy participants (75.4%) [16]. In HCV-infected adults, the anti-HAV seroprotection was slightly lower than in the general population (88%) [12].

Another high risk group for hepatitis A and B infection is travelers. Hepatitis A vaccination is recommended for all travelers to areas of intermediate or high prevalence. Hepatitis B vaccination is recommended for travelers likely to be at risk for hepatitis B infection, traveling to areas of intermediate or high HBV prevalence. Elderly travelers are at particular risk for morbidity and mortality related to HAV and HBV, which underscores the need for vaccination of this group. For travelers to countries with intermediate or high prevalence of both HAV and HBV, Twinrix^TM could be used to protect the traveler for both infections. An accelerated schedule, vaccination given at day 0, 7, and 21, and a booster at 12 months, could be considered for travelers [60].

Strengths and limitations

The main strengths of the methodology of this review are the systematic approach and the long search period used (1990–2015). To our knowledge there is no up-to-date systematic review available covering the immunogenicity, effectiveness, and safety of Twinrix^TM, Twinrix Pediatric^TM, and Ambirix^TM. However, only articles with a direct comparison between Twinrix^TM and separate hepatitis A or B vaccines were included, which could have limited the comparison of the immunogenicity and safety of the separate vaccines.

A few publications with poor description of the methodology were excluded (see Figure 1 for more details). However, a number of the included publications lack some details describing, for example, the exact setting, study period, randomization process or whether or not intention-to-treat analyses or according to protocol analyses were done. Furthermore, there is a broad variety of study designs used in the included publications, each with strengths and limitations (see Table A1 for more details). Comparison of the studies with respect to their quality can be challenging, but all studies show consistent immunogenicity and safety data.

Another limitation with regard to the included studies was related to the heterogeneity of the assays used. In some studies, the assay used to determine serum concentrations changed during the study period. However, the equivalence of the used assays was validated and the change of kits had no significant effect on the outcomes.

Limited to no data were found on the effectiveness and efficacy of Twinrix^TM, Twinrix pediatric^TM, or Ambirix^TM. The limited information found for the effectiveness can at least partially be explained by the fact that Twinrix^TM is not included in any universal mass vaccination programs, as is it the case for Hepatitis B and to a lesser extend Hepatitis A monovalent vaccines, which makes it difficult to perform adequate effectiveness studies in a general population. Some articles reporting on the incidence of hepatitis A or B before and after implementation of vaccination were found; however, the vaccine used was not specified. The lack of efficacy data can be explained because immunologic correlates of efficacy were well established (or at least accepted) for each component of the combination vaccine. Therefore, data from immunogenicity studies and immune noninferiority studies versus the monovalent licensed vaccines could provide a basis for license approval without the need for additional evidence studies [61]. Furthermore, there were only a few articles on the immunogenicity of safety of the vaccines in risk populations.

Conclusion

Twinrix^TM is the only licensed combined hepatitis A and B vaccine largely available, outside of China, where another combined vaccine (Bilive) is available. A combined vaccine provides benefits over separate vaccine such as fewer injections, fewer office visits, lower costs, more convenience for vaccinees, simplify logistics and administration, and potentially increase compliance [23,27,35–37,43,50,51]. This review shows that Twinrix^TM, Twinrix pediatric^TM, and Ambirix^TM (conventional and accelerated schedules) offer satisfactory immunogenicity, sustained up to 15 years after vaccination, and show an expected duration of protection and safety profile similar to the monovalent vaccines.

Expert commentary

This review covers clinical trials and observational studies that have been published between 1990 and 2015 covering the immunogenicity, effectiveness, and safety of the combined Hepatitis A&B vaccines Twinrix^TM, Twinrix Pediatric^TM, and Ambirix^TM. To our knowledge, this is the first review with a systematic approach and the first summarizing more than two decades experience with the combined vaccine. Unlike monovalent Hepatitis A and Hepatitis B vaccines, A&B combination vaccines are usually not universally recommended at national level and not used in national immunization programs. Monovalent Hepatitis A vaccines are recommended and used in national childhood vaccination programs in countries with intermediate endemicity as recommended by WHO. Hepatitis B childhood vaccination is universally recommended and implemented in at least 183 WHO Member States (the Hepatitis B antigen being often used in combination with other childhood vaccines). On the other hand, the A&B combination vaccine is mostly recommended and used in individuals at risk of exposure, providing the opportunity to protect against Hepatitis A and B individuals that would suffer from higher morbidity and mortality, if infected. The impact of vaccination and long-term safety, immunogenicity, and effectiveness with this vaccine cannot be not measured in the context of national programs as it is the case for Hepatitis A [62]or B [63]. It is therefore key to continue monitoring how Hepatitis A&B combination vaccination performs from individual studies and to do so with and using a systematic and unbiased methodology seems to be the most appropriate approach. A combined vaccine provides benefits over separate vaccine such as fewer injections, fewer office visits, lower costs, more convenience for vaccinees, simplify logistics and administration, and potentially increase compliance. This review shows that Twinrix^TM, Twinrix pediatric^TM, and Ambirix^TM (conventional and accelerated schedules) offer satisfactory immunogenicity, sustained up to 15 years after vaccination, and show a safety profile similar to the monovalent vaccines.

Five-year view

Systematic and unbiased reviews of published data are key to support regulatory authorities in their assessment of the benefit/risk of licensed vaccines over time. Additional long-term immunogenicity and safety data with Twinrix^TM, Twinrix pediatric^TM, and Ambirix^TM will become available and should be systematically gathered and reviewed in order to illustrate the benefit/risk and also inform on the need to provide a booster vaccination for Hepatitis A, Hepatitis B, or both more specifically for individuals at increased risk of exposure to these viral hepatitis.

Key issues

• Twinrix^TM, Twinrix pediatric^TM, and Ambirix^TM offer satisfactory immunogenicity, sustained up to minimum 15 years after vaccination, and show a safety profile similar to the monovalent vaccines.

• Immunogenicity results were equal or higher for both anti-HAV and anti-HBs following Twinrix^TM vaccination compared to monovalent hepatitis A or B vaccination.

• Co-administration of Twinrix pediatric^TM or Ambirix^TM with other routine childhood vaccines was immunologically noninferior to administration of the combined hepatitis A and B vaccine alone and did not significantly alter the safety profile

• Combined vaccines provide benefits such as fewer injections, fewer office visits, lower costs, more convenience for vaccinees, simplify logistics and administration, and potentially increase compliance compared to separate vaccination.

• Limited data were found describing the immunogenicity of the combined vaccines in high risk populations, including patients with chronic liver disease or HIV-infected patients. Overall, the immunogenicity was lower in high risk patients compared to healthy subjects.

Trademarks

Twinrix^TM, Twinrix pediatric^TM, Ambirix^TM, Havrix^TM and Engerix-B^TM are trademarks of the GSK group of companies.

Vaqta, Recombivax-HB and Gen H-B-Vax are trademarks of Merck & Co.

HB VAX PRO is trademark of Sanofi-Pasteur MSD.

Bilive is a trademark of SINOVAC Biotech Ltd.

Financial and competing interests disclosure

C Marano, M De Ridder and L De Moerlooze are employees of GSK group of companies and hold stock options/restricted shares from the sponsoring company. M Bakker and E Bunge report grants from GSK group of companies during the conduct of the study and outside the submitted work. This systematic review was sponsored and funded by GlaxoSmithKline Biologicals S.A., Belgium. GlaxoSmithKline Biologicals S.A. was involved in all stages of the study conduct and analysis; and also took charge of all costs associated with the development and the publishing of the manuscript. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

Acknowledgements

The authors would like to thank Angeles Ceregido (from XPE Pharma & Science on behalf of GSK Vaccines) for editing and coordinating the publication development.

Appendix 1. Search string

‘twinrix’ [Supplementary Concept] OR Twinrix[tiab] OR Ambirix[tiab] OR combined hepatitis A and B vaccin*[tiab] OR hepatitis A/B vaccin*[tiab] OR hepatitis A/hepatitis B vaccin*[tiab] OR (((‘Hepatitis A’[Mesh] OR hepatitis A[tiab]) AND (‘Hepatitis B’[Mesh] OR hepatitis B[tiab])) AND (‘Vaccination’[Mesh] OR vaccin*[tiab])) OR ((‘Hepatitis A’[Mesh] OR hepatitis A[tiab]) AND (‘Hepatitis B’[Mesh] OR hepatitis B[tiab]) AND (‘Immunization’[Mesh] OR immun*[tiab]))

Limit: Publication date from January 1990 to present (October 13th, 2015)

Appendix 2. Study characteristics

Table A1. Study characteristics.

Reference	Country	Study design	Study population, N, age	Vaccine used	Laboratory test used	Conflict of interest	Main study limitations reported by the authors
Ambrosch J Med Virol 1994 [18]	NR	Comparative study*	Healthy young adults, mostly medical studentsN = 162 (54 subjects received Twinrix^TM)Age: NR	Twinrix^TM3 doses at 0, 1, and 6 monthsComparator: Havrix^TM+ Engerix-B^TM3 doses at 0, 1,and 6 months	HAV: HAVAB (Abbott)^a ELISA inhibition assay (SmithKline Beecham Biologicals)^b HBV: AUSAB (Abbott)^c CORAB (Abbott)^a AUSRIA (Abbott)^a	Not reported
Belderok Vaccine 2013 [46]	The Netherlands	Phase IV interventional study	Children known to be infected with HIV and children receiving IM for treatment of rheumatic diseasesN = 160Age range: 1–16 years	Ambirix^TM2 doses at 0, 26–30 weeksComparator: None	HAV: Axsym HAVAB 2.0 (Abbott)^a Anti-HAV Total HAVT (Biomerieux)^a HBV: Axsym CORE (Abbott)^a Axsym AUSAB (Abbott)^a	None	None
Bruguera Vaccine 1996 [19]	Spain	Randomized trial	Healthy young students of medicine or nurseryN = 150Mean age (SD; range): 20 years (1.4; 18–31)	Twinrix^TM3 doses at 0, 1, and 6 monthsComparator: None	HAV: ETI-ABHAVK (Sorin Biomédica)^a Enzymum ELISA (Boehringer Mannheim)^d HBV: ETI-AB-COREK (Sorin Biomédica)^a ETI-ABAVK ELISA (Sorin Biomédica)^a ETI-MAK ELISA (Sorin Biomédica)^a AUSAB (Abbott)^c	GSK	None
Burgess Vaccine 2001 [48]Burgess Vaccine 2010 [47]	Australia	Open, randomized study with follow-up	Healthy subjects who had not been previously vaccinated against hepatitis A and BN = 240Age range: 12–15 years	Ambirix^TM2 doses at 0 and 6 months or 0 and 12 monthsComparator: None	HAV: Enzymum ELISA (Boehringer Mannheim)^d Enzygnost (DADE Behring)^e HBV: AUSAB (Abbott)^c,f AUSRIA (Abbott)^a CORAB (Abbott)^a	GSK	Small sample size at 6-year follow-up.
Connor J Travel Med 2007 [35]	Belgium, Czech Republic, Norway, US	Prospective, open-label, randomized, multicenter, comparative study	Healthy adults who were seronegative for anti-HAV, anti-HBs, and anti-HBc antibodies and for HBsAgN = 496 (250 subjects received Twinrix^TM)Age: ≥18 years	Twinrix^TM4 doses at 0, 7, and 21–30 days and 12 monthsComparator: Havrix^TM + Engerix-B^TM2 and 4 doses at 0 (1,2), and 12 months	HAV: Enzyme immunoassay (Behring)^eHBV: AUSAB (Abbott)^f	GSK	None
Cunningham Pediatr Infect Dis J 2010 [49]	US	Phase II, randomized, single-blinded trial	HIV-negative urban youthN = 123 (63 subjects received Twinrix^TM)Mean age (SD): 15.12 years (1.58)	Ambirix^TM2 doses at 0 and 6 monthsComparator: Recombivax HB2 doses at 0 and 6 months	HAV: NAHBV:- EIA (Abbott)^a- ECI (Ortho Vitros)^a	None	Significant loss to follow-up (due to hurricane Katrina).Study was not designed to compare both vaccines.Relatively small sample size
Czeschinski Vaccine 2000 [20]	Germany	Randomized study	Healthy job startersN = 300 (75 subjects received Twinrix^TM)Mean age (range): 21.8 years (18–30)	Twinrix^TM3 doses at 0, 1, and 6 monthsComparator: Havrix^TM+ Engerix-B^TM3 doses at 0, 1, and 6 months	HAV: ETI-AB-HAVK-3 (Sorin Biomédica)^bHBV: IMx AUSAB (Abbott)^g	Not reported	Exact antibody titers could be assessed for only 43 of 74 subjects receiving Twinrix
De Schyver J Viral Hepat 2011 [21]	Belgium	Randomized, stratified, controlled study	Healthy adult workers, who are not occupationally exposed to HBVN = 397Mean age: Study group: 42.5 years Reference group: 41.4 years	Twinrix^TM Study group: 3 doses at 0, 1, and 12 months Reference group: 3 doses at 0, 1, and 6 months Comparator: None	HAV: NRHBV:- ETI-AB-AUK-3 (DiaSorin)^c- ETI-AB-CORTEK PLUS (DiaSorin)^c- ETI-MAK-4 (DiaSorin)^c	GSK	None
Dominguez J Viral Hepat 2008 [55]	Spain	Surveillance study	Subjects with viral hepatitisN = 7536All ages	Twinrix^TM3 doses at 0, 1, and 6 monthsComparator: None	HAV: NRHBV: NR	None	Possibly underestimation of number of cases, since only reported clinical cases are included in the surveillance system (cases not reported for various reasons and asymptomatic infection will be missed)
Duval Vaccine 2005 [44] GilcaPediatr Infect Dis J2009 [45]	Canada	Open label study	Children eligible to be vaccinated according to the regular immunization program in Quebec cityN = 704 (347 children received Twinrix^TM)Mean age (95% CI): Twinrix^TM: 114 months (108–119) Recombivax-HB: 114 months (109–120)	Twinrix pediatric^TM2 doses + challenge dose at 0 and 6 months + 7 yearsComparator: Recombivax-HB2 doses + challenge dose at 0 and 6 months + 7 years	HAV: HAV Total Assay EIA (Bio-Rad)^dHBV: AUSAB EIA (Abbott)^c ELISA (ORTHO, Raritan)^a EIA 2.0 (Bio-Rad)^a	Not reported	Nonrespondents from the primary study were excluded in the follow up study, which might lead to an overestimation of the response to a challenge dose
Flynn J AIDS 2011 [16]	Bahamas, Brazil, South Africa, US	Randomized trial	HIV-infected youthN = 336 (118 receiving Twinrix^TM)Mean age: 20.44 years	Twinrix^TM3 doses at 0, 1, and 6 monthsComparator: Engerix-B^TM20 mcg (recommended) or 40 mcg (increased)3 doses at 0, 1, and 6 months	HAV: NRHBV: Abbott EIA or Ortho Vitros ECI^g	None	None
Frey J Infect Dis 1999 [36]	NR	Open, randomized trial	Healthy individualsN = 1,385 (1252 receiving Twinrix^TM)Median age (range): Twinrix^TM: 23 years (10–80) VAQTA + Recombivax-HB: 22 years (11–59)	Twinrix^TM2 doses at 0 and 24 weeksComparator: VAQTA+ Recombivax-HB2 doses at 0 and 24 weeks	HAV: HAVAB (Abbott)^gHBV: AUSAB RIA (Abbott)^g	Merck and Company Inc.	The wide age range of study participants may have contributed to less-than-optimal response to the combination vaccine
HöhlerVaccine 2007 [22]	NR	Extension of a primary open, prospective, single-site study	Healthy mono- and dizygotic twinsN = 98Mean age (SD): 54 years (7.1)	Twinrix^TM3 doses at 0, 1, and 6 months and booster dose at 33 monthsComparator: None	HAV: Enzygnost (DADE Behring)^eHBV: AUSAB EIA (Abbott)^f	GSK	None
Joines Vaccine 2001 [23]	US	Open-label, randomized, multicenter, controlled clinical trial	Healthy adultsN = 829 (414 subjects received Twinrix^TM)Mean age (range): Twinrix^TM: 39.9 years (19–70) Havrix^TM+ Engerix-B^TM: 40.4 years (19–83)	Twinrix^TM3 doses at 0, 1, and 6 monthsComparator: Havrix^TM+ Engerix-B^TM2 and 3 doses at 0 (1), and 6 months	HAV: HABAV (Abbott)^dHBV: AUSAB (Abbott)^c	Not reported	None
Kallinowski Z Gastroenterol 2003 [12]	NR	Open, multicenter study	Caucasian patients with chronic HCV-infection and healthy volunteers (i.e. oversea travelers or healthy hospital staff)N = 178Mean age (SD), years: HCV+/Accelerated: 39 (10) Healthy/Accelerated: 33 (7) HCV+/Standard: 35 (8) Healthy/Standard: 30 (8)	Twinrix^TM Standard: 3 doses at 0, 1, and 6 months Accelerated: 3 doses at 0, 7, and 21 days Comparator: None	HAV: ELISA (Abbott)^dHBV: ELISA (Abbott)^c	None	None
Kallinowski Liver 1996 [24]	Germany	Prospective cohort study*	Anti-HAV and anti-HBs negative volunteersN = 50Mean age (range): 30.5 years (18–49)	Twinrix^TM3 doses at 0, 1, and 6 monthsComparator: None	HAV: ELISA (SmithKline Beecham Biologicals)^bHBV: ELISA (SmithKline Beecham Biologicals)^g	Not reported	None
Knöll Vaccine 2000 [25]	Germany	Comparative study*	Healthy young medical students and staff membersN = 282 (134 receiving Twinrix^TM)Age range: 17–39 years	Twinrix^TM3 doses at 0, 1, and 6 monthsComparator: Engerix-B^TM3 doses at 0, 1, and 6 months	HAV: AXSYM HAVAB (Abbott)^bHBV:- AXSYM CORE (Abbott)^c- AXSYM AUSAB (Abbott)^c
Lee J Med Virol 1999 [50]	China	Open, randomized study	Healthy Chinese youngstersN = 103 (51 subjects received Twinrix^TM)Mean age (SD; range): Twinrix^TM: 14.1 years (2.68; 10–19) Concomitant: 12.4 years (2.21; 10–19)	Ambirix^TM2 doses at 0 and 6 monthsComparator: Concomitant hepatitis A and B vaccines (not specified)2 doses at 0 and 6 months	HAV: HAVAB (Abbott)^a Enzymun (Boehringer Mannheim)^a HBV: AUSRIA II (Abbott)^a CORAB (Abbott)^a AUSAB (Abbott)^a	GSK	None
Levie Vaccine 2002 [38] Beran Vaccine 2010 [15]	Belgium, Czech republic	Open, randomized, comparative, multicenter study	Subjects aged between 12 and 15 years at the time of first vaccinationN = 300Mean age (range): 13.4 years (12–15 years)	Ambirix^TM2 doses at 0 and 6 monthsTwinrix pediatric^TM3 doses at 0, 1, and 6 monthsComparator: None	HAV: Enzymun (Boehringer Mannheim)^d Enzygnost (Boehringer)^e HBV: AUSAB RIA (Abbott)^c AUSAB EIA (Abbott)^f Equivalent validated in-house ELISA^f	Not reported	None
Moro Am J Obstet Gynecol 2014 [58]	US	Review of reports	Pregnant women who had received hepatitis A or combined hepatitis A and B vaccinationN = 35Maternal age, median (range): 28 years (19–39 years)	Twinrix^TMNumber of doses and schedule: NR	NA	None	VAERS is a passive surveillance system that may be prone to biased reporting and inconsistency in the quality and completeness of reports.VAERS generally cannot determine whether a vaccine caused an AE.VAERS does not collect data on the number of individuals vaccinated; therefore, it is not possible to calculate incidence or prevalence of AE.
Nothdurft Vaccine 2002 [37]	Germany, UK	Randomized. multicenter study	Healthy subjects with normal liver function test, seronegative for anti-HAV, anti-HBs, and anti-HBc, and none had previously been vaccinated against HA or HBN = 479 (239 subjects received Twinrix^TM)Mean age (range): Twinrix^TM: 24.9 years (18–45 years) Havrix^TM+ Engerix-B^TM: 27.0 years (18–48 years)	Twinrix^TM4 doses at 0,7,21 days and 12 monthsComparator: Havrix^TM+ Engerix-B^TM2 and 4 doses at 0 (7, 21) days, and 12 months	HAV: Enzymun ELISA (Boehringer Mannheim)^dHBV: AUSAB (Abbott)^c	Not reported	None
Ostergaard Vaccine 2012 [39]	Denmark, Sweden	Phase III, open, randomized controlled study	Healthy children who previously completed routine childhood vaccinationsN = 611Mean age (SD; range): M-TT + HAB: 14.3 years (1.89; 11–17) HAB: 14.3 years (1.94; 11–17)	Twinrix^TM (Pediatric or Adult, depending on age)3 doses at 0, 1, and 6 monthsComparator: MenACWY-TT1 dose at 0 months	HAV: Enzygnost ELISA (DADE Behring)^eHBV: In-house developed anti-HBs ELISA^a	GSK	None
Pedersen J Adolesc Health 2012 [40]	Canada, Denmark, Hungary, Sweden	Randomized, open, controlled, parallel group study	Healthy girlsN = 813Mean age (SD): HPV+HAB: 11.2 years (2.04) HAB: 11.2 years (1.99)	Twinrix pediatric^TM3 doses at 0, 1, and 6 monthsComparator: Cervarix^TM3 doses at 0, 1, and 6 months	HAV: Enzygnost ELISA (DADE Behring)^eHBV: HBs assay developed by GSK Biologicals^f Axsym Core (Abbott)^a	GSK	None
Ramonet Arch Med Res 2002 [51]	Argentina, Brazil, Mexico	Randomized study	Healthy children and adolescentsN = 248 (168 subjects received Twinrix^TM)Age range: 4–20 years	Ambirix^TM2 doses at 0 and 6 monthsComparator: Havirx + Engerix-B^TM2 doses at 0 and 6 months	HAV: Enzymun (Boehringer Mannheim)^dHBV: AUSRIAII (Abbott)^c CORAB (Abbott)^c AUSAB (Abbott)^c	GSK	None
Rendi-Wagner Vaccine 2001 [26]	Austria	Uncontrolled study	Healthy adolescents and adults who were seeking pre-travel immunization adviceN = 1874 (990 subjects received Twinrix^TM)Mean age (range): 43.4 years (6–80)	Twinrix^TM3 doses at 0, 1, and 6–12 monthsComparator: Engerix-B^TM or Gen H-B-Vax3 doses at 0, 1, and 6–12 months	HAV: NAHBV: AUSAB EIA (Abbott)^c	Not reported	None
Rieger Int J Occup Med Environ Health 2004 [27]	Germany	Open, controlled, randomized, monocentric study with two parallel groups	Healthy subjectsN = 304 (152 subjects received Twinrix^TM)Mean age (SD): 29.0 years (7.84)	Twinrix^TM3 doses at 0, 1, and 6 monthsComparator: Vaqta+ Gen H-B-Vax2 and 3 doses at 0 (1), and 6 months	HAV: ETI-AB-HAVK-3 (DiaSorin)^bHBV: ETI-AB-AUK-3 (DiaSorin)^c ETI-AB-COREK-2 (DiaSorin)^a	GSK	None
Roberton Vaccine 2005 [41] Marshall Vaccine 2010 [10]	Australia, Belgium, Spain, Sweden	Randomized, open-label, multicenter study with follow-up	Healthy childrenN = 511 (Ambirix^TM: 255, Twinrix pediatric^TM: 256)Mean age (SD; range), years: 6.0 (3.1; 1–11)	Ambirix^TM2 doses at 0 and 6 monthsTwinrix pediatric^TM3 doses at 0, 1, and 6 monthsComparator: None	HAV: Enzygnost (DADE Behring)^eHBV: Axsym HBsAg (Abbott)^f Axsym Core (Abbott)^f AUSAB EIA (Abbott)^f In-house ELISA assay^f	Not reported	None
Thoelen Vaccine 1999 [11]	Belgium, Lithuania, the Netherlands, Spain, Switzerland	Review of six clinical trials	Healthy adultsN = 843Mean age (SD; range): 20.8 years (4.2; 17–60)	Twinrix^TM3 doses at 0, 1, and 6 monthsComparator: None	HAV: Enzymum (Boehringer Mannheim)^dHBV: Radioimmunoassay (Abbott)^c
Thompson Int J Infect Dis 1998 [34]	Australia	Double-blind randomized clinical study	Healthy adolescentsN = 160Mean age: 16.4 years	Twinrix^TM3 doses at 0, 1, and 6 monthsComparator: None	HAV: Modified HAVAB (Abbott)^d Enzymun ELISA (Boehringer Mannheim)^d HBV: AUSRIA II (Abbott)^c AUSAB (Abbott)^c CORAB (Abbott)^c	Not reported	None
Usonis Vaccine 2005 [52]	Estonia, Lithuania	Comparative study	Healthy infants born to mothers not known to be seropositive for HBsAg, hepatitis B antigen or anti-HBc antibodiesN = 90Mean age (range): Lithuania: 16.5 months (15–18) Estonia: 12.5 months (11–15)	Ambirix^TM2 doses at 0 and 6 monthsComparator: None	HAV: Enzymum (Boehringer)^d HBV: AUSAB (Abbott)^c Axsym (Abbott)^c CORAB (Abbott)^c	Not reported	None
Van Damme Pediatr Infect Dis J 2011 [53]	Belgium	Open-label single group primary study	Healthy subjects without history of hepatitis or vaccination against HABN = 232Mean age 10 years after primary vaccination (SD): 17 years (2.84)	Ambirix^TM2 doses at 0 and 6 monthsComparator: None	HAV: Enzygnost (DADE Behring)^eHBV: AUSAB EIA (Abbott)^f Equivalent validated in-house ELISA^f	GSK	None
Van Damme J Med Virol 2001 [42] Diaz-Mitoma Vaccine 2008 [43]	Canada	Open study and a double-blind randomized study	Healthy childrenN = 166Age range: 1–15 years	Twinrix pediatricTM3 doses at 0, 1, and 6 monthsComparator: None	HAV: Enzymun (Boehringer Mannheim)^d Enzygnost (DADE Behring)^e HBV: AUSAB RIA (Abbott)^c AUSAB EIA (Abbott)^f	GSK	None
Van Der Wielen Vaccine 2006 [28] Chlibek J Travel Med 2011 [29]	Belgium, Czech Republic, Germany	Open, randomized, parallel group trial	Subjects seronegative for anti-HBs antibodies, anti-HBc antibodies, anti-HAV and HBsAgN = 596 (199 receiving Twinrix^TM)Mean age at first dose (SD; range): 55.2 years (9.6; 41–81)	Twinrix^TM3 doses at 0, 1, and 6 monthsComparator: Havrix^TM + Engerix-B^TM/Vaqta + HB-VAX-PRO2 and 3 doses at 0 (1), and 6 months	HAV: Enzygnost (DADE Behring)^eHBV: AUSAB (Abbott)^f	GSK	None
Van Der Wielen Pediatr Infect Dis J 2000 [54]	Belgium	Open, single center study	Healthy children/adolescentsN = 236Mean age: <6 years: 4.2 years ≥6 years: 9.1 years	Ambirix^TM2 doses at 0 and 6 monthsComparator: None	HAV: Enzymun (Boehringer Mannheim)^dHBV: AUSAB RIA (Abbott)^c CORAB (Abbott)^c AUSRIAII (Abbott)^c	Not reported	None
Van Herck Travel Med Infect Dis 2007 [30] Van Damme J Med Virol 2012 [14]	Belgium	Follow up of two double-blind, randomized trials	Healthy adultsN = 223 (Center 1: 116, Center 2: 107)Range (primary studies): 17–39 years and 17–43 years	Twinrix^TM3 doses at 0, 1, and 6 monthsComparator: None	HAV: Enzymun ELISA (Boehringer Mannheim)^d Enzygnost kit (DADE Behring)^e HBV: AUSAB RIA (Abbott)^c AUSAB EIA (Abbott)^f	GSK	None
Vilella Vaccine 2005 [56]	Spain	Prospective observational study	Subjects traveling to hepatitis A and hepatitis B highly endemic areas, who were candidates to receive HAB vaccineN = 998Mean age (SD; range): 23 years (4.5; 11–54)	Twinrix^TM3 doses at 0, 1, and 6 monthsComparator: None	HAV: NAHBV: NA	GSK	None
Williams Vaccine 2012 [31]	US	Outbreak study	Residents of nursing facilitiesN = 209Age range: 45–101 years	Twinrix^TM3 doses at 0, 1, and 6 monthsComparator: None	HAV: NAHBV: Monolisa (Bio-Rad) GS HbsAg EIA 3.0 (Bio-Rad) Vitros aHBc (Othro-Clinical diagnostics) Vitros HBcM (Othro-Clinical diagnostics) Vitros HbsAg (Othro-Clinical diagnostics) Vitros aHBs (Othro-Clinical diagnostics)	Not reported	Population not necessarily representative of all older adults, older persons with diabetes, or other long-term care facility residents -It is possible that some of the residents that were categorized as susceptible to HBV based on serologic results might have received HB vaccine in the past and either did not achieve a primary response or had antibody levels wane over time; such residents might be expected to have a more vigorous response to re-vaccination, which would have potentially increased the proportion who achieved seroprotection in this study
Wolters Hum Vaccin 2009 [32]	Germany	Multicenter trial with prospective and retrospective cohort	Prospective: Seronegative subjectsRetrospective: Subjects who had been vaccinated three times in the pastN = 422Mean age (range): Prospective: 38.9 years (18–79) Retrospective: 39.9 years (16–75)	Twinrix^TM3 doses at 0, 1, and 6 monthsComparator: None	HAV: Enzygnost (DADE Behring)^bHBV: Enzygnost (DADE Behring)^g Architect i2000SR (Abbott)^g	Not reported	Anti-HAV antibody concentration was measured only up to 80 IU/l The 4–8 week interval for antibody control was not strictly adhered to (might be especially problematic for HBV-low responders)
Wolters Vaccine 2003 [33]	Germany	Retrospective study	Patients vaccinated following manufacturer recommendationsN = 104Mean age (range): 54 years (17–84)	Twinrix^TM3 doses at 0, 1, and 6 monthsComparator: None	HAV: Axsym HAVAB 2.0 (Abbott)^bHBV: Axsym AUSAB II (Abbott)^g	Not reported	- Relatively old study population
Woo Vaccine 2006 [57]	Worldwide	Passive surveillance study	Recipients of Twinrix for whom a report of possible AE after vaccinated was received by VAERSN = 305 reportsMinimum age: 2 years	Twinrix^TMNumber of doses and schedule: NR	NA	None	Underreporting, incomplete information in many reports, inadequate data regarding the number of doses administered, and lack of direct and unbiased comparison group

CI: Confidence interval; EIA: Enzyme immunoassay; ELISA: Enzyme-linked immunosorbent assay; HAB: Hepatitis A and B; HAV: Hepatitis A virus; HBc: Hepatitis B core antigen; HBs: Hepatitis B surface antigen; HBV: Hepatitis B virus; HCV: Hepatitis C virus; HIV: Human immunodeficiency virus; HPV: Human papillomavirus; M-TT: Tetravalent meningococcal conjugate vaccine coupled to tetanus toxoid; NA: Not applicable; NR: Not reported; RIA:Radioimmunoassay; SD: Standard deviation.

*Study design not reported in the article. Design defined by Pallas.

^aCut-off: NR.

^bCut-off: 20 (m)IU/(m)l.

^cCut-off: 1 (m)IU/(m)l.

^dCut-off: 33 (m)IU/(m)l.

^eCut-off: 15 (m)IU/(m)l.

^fCut-off: 3.3 (m)IU/(m)l.

^gCut-off: 10 (m)IU/(m)l.