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Effect of a partially hydrolyzed whey infant formula at weaning on risk of allergic disease in high-risk children: A randomized controlled trial

      Background

      Partially hydrolyzed whey formula (pHWF) has been recommended for infants with a family history of allergic disease at the cessation of exclusive breast-feeding to promote oral tolerance and prevent allergic diseases.

      Objective

      To determine whether feeding infants pHWF reduces their risk of allergic disease.

      Methods

      A single-blind (participant) randomized controlled trial was conducted to compare allergic outcomes between infants fed a conventional cow’s milk formula, a pHWF, or a soy formula. Before birth, 620 infants with a family history of allergic disease were recruited and randomized to receive the allocated formula at cessation of breast-feeding. Skin prick tests to 6 common allergens (milk, egg, peanut, dust mite, rye grass, and cat dander) were performed at 6, 12, and 24 months. The primary outcome was development of allergic manifestations (eczema and food reactions) measured 18 times in the first 2 years of life.

      Results

      Follow-up was complete for 93% (575/620) at 2 years and 80% (495/620) at 6 or 7 years of age. There was no evidence that infants allocated to the pHWF (odds ratio, 1.21; 95% CI, 0.81-1.80) or the soy formula (odds ratio, 1.26; 95% CI, 0.84-1.88) were at a lower risk of allergic manifestations in infancy compared with conventional formula. There was also no evidence of reduced risk of skin prick test reactivity or childhood allergic disease.

      Conclusion

      Despite current dietary guidelines, we found no evidence to support recommending the use of pHWF at weaning for the prevention of allergic disease in high-risk infants.

      Key words

      Abbreviations used:

      CMF (Cow’s milk formula), GINI (German Infant Nutritional Intervention Study), ITT (Intention to treat), MACS (Melbourne Atopy Cohort Study), OR (Odds ratio), pHWF (Partially hydrolyzed whey formula), SPT (Skin prick test)
      Partially hydrolyzed whey formulas (pHWFs) have been widely recommended to prevent the development of allergic diseases in early childhood.
      • Host A.
      • Halken S.
      Primary prevention of food allergy in infants who are at risk.
      • Prescott S.L.
      • Tang M.L.K.
      Australasian Society of Clinical Immunology and Allergy position statement: summary of allergy prevention in children.
      • Miniello V.L.
      • Francavilla R.
      • Brunetti L.
      • Franco C.
      • Lauria B.
      • Lieggi M.S.
      • et al.
      Primary allergy prevention: partially or extensively hydrolyzed infant formulas?.
      • Hays T.
      • Wood R.A.
      A systematic review of the role of hydrolyzed infant formulas in allergy prevention.
      • Heine R.G.
      • Tang M.L.
      Dietary approaches to the prevention of food allergy.
      • Pali-Scholl I.
      • Renz H.
      • Jensen-Jarolim E.
      Update on allergies in pregnancy, lactation, and early childhood.
      If beneficial, the use of pHWF is an attractive preventive strategy, because pHWFs are relatively inexpensive to manufacture. These formulas contain smaller, less immunogenic milk protein–derived peptides
      • Makinen-Kiljunen S.
      • Sorva R.
      Bovine beta-lactoglobulin levels in hydrolysed protein formulas for infant feeding.
      of reduced allergenicity that potentially enhance induction of tolerance to cow’s milk protein.
      • Fritsche R.
      • Pahud J.J.
      • Pecquet S.
      • Pfeifer A.
      Induction of systemic immunologic tolerance to beta-lactoglobulin by oral administration of a whey protein hydrolysate.
      • Pecquet S.
      • Bovetto L.
      • Maynard F.
      • Fritsche R.
      Peptides obtained by tryptic hydrolysis of bovine beta-lactoglobulin induce specific oral tolerance in mice.
      The widespread support for the use of pHWF appears to be based on the results of a Cochrane review that found “a significant reduction in infant allergy” (p 11) to be associated with prolonged feeding with pHWF compared with feeding with conventional cow’s milk formula (CMF).
      • Osborn D.A.
      • Sinn J.
      Formulas containing hydrolysed protein for prevention of allergy and food intolerance in infants.
      Despite the authors’ caution that further studies were required, this meta-analysis has been widely used to underpin many clinical guidelines in Europe, the United States, and Australia.
      • Host A.
      • Halken S.
      Primary prevention of food allergy in infants who are at risk.
      • Prescott S.L.
      • Tang M.L.K.
      Australasian Society of Clinical Immunology and Allergy position statement: summary of allergy prevention in children.
      • Miniello V.L.
      • Francavilla R.
      • Brunetti L.
      • Franco C.
      • Lauria B.
      • Lieggi M.S.
      • et al.
      Primary allergy prevention: partially or extensively hydrolyzed infant formulas?.
      • Hays T.
      • Wood R.A.
      A systematic review of the role of hydrolyzed infant formulas in allergy prevention.
      • Heine R.G.
      • Tang M.L.
      Dietary approaches to the prevention of food allergy.
      • Pali-Scholl I.
      • Renz H.
      • Jensen-Jarolim E.
      Update on allergies in pregnancy, lactation, and early childhood.
      • Grimshaw K.E.
      • Allen K.
      • Edwards C.A.
      • Beyer K.
      • Boulay A.
      • van der Aa L.B.
      • et al.
      Infant feeding and allergy prevention: a review of current knowledge and recommendations: a EuroPrevall state of the art paper.
      A major problem with meta-analyses is that often only published reports are analyzed.
      • Nieto A.
      • Mazon A.
      • Pamies R.
      • Bruno L.
      • Navarro M.
      • Montanes A.
      Sublingual immunotherapy for allergic respiratory diseases: an evaluation of meta-analyses.
      These are more likely to be positive studies because of publication bias, leading the review to overestimate the effectiveness of a treatment.
      • Turner E.H.
      • Matthews A.M.
      • Linardatos E.
      • Tell R.A.
      • Rosenthal R.
      Selective publication of antidepressant trials and its influence on apparent efficacy.
      Publication bias may have affected the results of the Cochrane review on the value of pHWF in preventing allergic disease.
      • Osborn D.A.
      • Sinn J.
      Formulas containing hydrolysed protein for prevention of allergy and food intolerance in infants.
      There is some evidence of asymmetry in the funnel plot
      • Egger M.
      • Davey Smith G.
      • Schneider M.
      • Minder C.
      Bias in meta-analysis detected by a simple, graphical test.
      generated for the meta-analysis reported within the Cochrane review
      • Osborn D.A.
      • Sinn J.
      Formulas containing hydrolysed protein for prevention of allergy and food intolerance in infants.
      (Harbord P = .0614), with the smaller studies tending to report stronger protective effects of the pHWF than the larger studies. The German Infant Nutritional Intervention Study (GINI),
      • von Berg A.
      • Koletzko S.
      • Grubl A.
      • Filipiak-Pittroff B.
      • Wichmann H.E.
      • Bauer C.P.
      • et al.
      The effect of hydrolyzed cow’s milk formula for allergy prevention in the first year of life: the German Infant Nutritional Intervention Study, a randomized double-blind trial.
      the largest in this field, reported that pHWF reduced the incidence of eczema in early childhood in a per-protocol analysis that excluded children exclusively breast-fed to 4 months of age. However, an intention-to-treat (ITT) analysis failed to show any benefit of pHWF compared with conventional CMF.
      • von Berg A.
      • Koletzko S.
      • Filipiak-Pittroff B.
      • Laubereau B.
      • Grubl A.
      • Wichmann H.E.
      • et al.
      Certain hydrolyzed formulas reduce the incidence of atopic dermatitis but not that of asthma: three-year results of the German Infant Nutritional Intervention Study.
      The primary aim of the current study was to determine whether the use of a pHWF reduced the incidence of allergic manifestations (eczema and food reactions) up to 2 years of age in high-risk infants compared with a conventional CMF. We also report results from a third comparison group in which infants received a soy formula.

      Methods

       Inclusion and exclusion criteria

      Between 1990 and 1994 expectant mothers attending the Mercy Maternity Hospital, Melbourne, Australia, were invited to participate in a study of the effect of modification of the infant diet on the risk of infant allergy. Mother-baby pairs were enrolled if the unborn child had a first-degree relative with a history of eczema, asthma, allergic rhinitis, or food allergy. Information leaflets and posters outlined the project’s aim. Nurse research staff assessed eligibility and enrolled participants. This study was approved by the Mercy Maternity Hospital Ethics Committee, and all mothers provided written informed consent.

       Intervention

      There were 2 intervention formulas: a soy-based formula (ProSobee; Mead Johnson Nutrition/Bristol Myers, Melbourne, Australia) and a pHWF (NAN HA; Nestlé, Biessenhoffen, Germany). The control formula was a CMF (NAN; Nestlé, Tongala, Australia). In accordance with World Health Organization guidelines,
      WHO
      Complementary feeding: report of the global consultation: summary of guiding principles.
      mothers were encouraged to initiate and maintain breast-feeding for at least 6 months. Study formulas were introduced only at cessation, or partial cessation, of breast-feeding or as a breast milk substitute if breast-feeding was not intended.

       Trial design

      The trial was registered (retrospectively) with the Australian and New Zealand Clinical Trials Registry (ACTRN12609000734268). The trial commenced before the pHWF was available. The first 97 infants were randomized to either the CMF or soy study groups. When the pHWF became available, a new random allocation series was generated with a higher proportion allocated to the pHWF to obtain equal numbers in each formula group. An independent statistician created each of the computer generated allocation schedules. The random allocation list, containing the coded allocations, was available to research staff. Staff were blind to these allocation codes and to the group of allocation at the time of outcome assessment. Mother-baby pairs were allocated to the next sequential number as they were enrolled in the study and were assigned to the formula code allocated to that number. The cans of formula were labeled at an independent location. Parents of participants were informed of the identity of the assigned formula only after the child’s second birthday.
      Introduction of rice cereal, pureed apple, and pear was recommended from 4 months of age, and vegetables and other fruit from 6 months. Meats were introduced from 8 months, and nonrice cereals from 9 months. Dairy products, egg, fish, peanut, and nuts were avoided until 12 months of age.
      Skin prick tests (SPTs) were performed at 6, 12, and 24 months according to a standard technique
      • Aas K.
      • Belin L.
      Standardization of diagnostic work in allergy.
      by 1 of 3 allergy-trained research nurses. Allergen extracts used were cow’s milk, egg white, peanut, house dust mite, rye grass, and cat dander (Bayer, Spokane, Wash), and SPTs were read at 15 to 20 minutes.

       Definitions

      Outcomes up to 2 years of life, as assessed during 18 telephone interviews with parents (every 4 weeks until 64 weeks, then at 78 and 104 weeks), were defined as follows:
      • Eczema: Doctor-diagnosed eczema or any rash that was treated with topical steroid preparation (excluding rash that only affected the scalp or nappy region).
        • Lowe A.J.
        • Abramson M.J.
        • Hosking C.S.
        • Carlin J.B.
        • Bennett C.M.
        • Dharmage S.C.
        • et al.
        The temporal sequence of allergic sensitization and onset of infantile eczema.
      • Food reaction: Within 2 hours of ingesting that food, the child developed an acute skin rash (urticaria, angioedema, erythematous, or morbilliform), a flare of pre-existing eczema, signs of anaphylaxis, or vomiting.
        • Hill D.J.
        • Firer M.A.
        • Shelton M.J.
        • Hosking C.S.
        Manifestations of milk allergy in infancy: clinical and immunologic findings.
      • Any allergic manifestation: Presence of eczema or food reaction within the first 2 years of life.
      • Positive SPT: A wheal of at least 3 mm (mean) diameter with a positive (histamine) control.
      Childhood outcomes, based on parent report during telephone interviews conducted when children were age 6 or 7 years, were defined as follows:
      • Current childhood eczema: Eczema diagnosed by the family physician in the previous 12 months.
      • Current childhood asthma: Asthma diagnosed by the family physician in the previous 12 months.
      • Persistent childhood asthma: Asthma diagnosed by the family physician in the previous 12 months on at least 2 occasions at the follow-up at 5, 6, or 7 years.
      • Current childhood allergic rhinitis: One or more episodes of nasal discharge and/or congestion in the absence of an upper respiratory tract infection in the previous 12 months that either the family physician or parent attributed to allergic rhinitis (hay fever) and that was treated with an antihistamine and/or nasal steroid.
        • Lowe A.J.
        • Hosking C.S.
        • Bennett C.M.
        • Carlin J.B.
        • Abramson M.J.
        • Hill D.J.
        • et al.
        Skin prick test can identify eczematous infants at risk of asthma and allergic rhinitis.

       Outcomes

      The primary outcome was any allergic manifestation (cumulative incidence) up to 2 years of age. Secondary outcomes were the individual incidence of eczema and food reactions, reported in the first 2 years of life, and SPT reactivity at 6, 12, and 24 months. Additional secondary outcomes were the 2-year period prevalence of eczema, asthma, and allergic rhinitis at ages 6 and 7 years.

       Sample size

      A total of 176 infants per group were required to have 80% power to detect a 15% absolute difference in risk of allergic manifestation between the formula groups, assuming an α level of 0.05 and a 45% baseline risk of allergic disease within the first 2 years of life. Allowing for approximately a 15% dropout rate over the first 2 years of life, a total of 206 children per group were required.

       Statistical methods

      The primary analysis followed the ITT principle and compared the risk of any allergic manifestation between the allocated formula groups by using simple proportions and χ2 tests. The estimated associations are presented as odds ratios (OR) with 95% CIs, with the CMF as the reference group.
      Secondary analyses were also performed for the outcomes of sensitization to cow’s milk and any allergen (assessed separately at 6, 12, and 24 months, and also combined by using logistic regression models, estimated by the generalized estimating equations approach) and childhood asthma, allergic rhinitis, and eczema at ages 6 and 7 years (again by using the generalized estimating equations approach).
      A number of per-protocol analyses were performed. First, infants were excluded if they were exclusively breast-fed beyond 4 months of life. The 4-month cut-off period was selected to allow direct comparison of results with the GINI study.
      • von Berg A.
      • Koletzko S.
      • Grubl A.
      • Filipiak-Pittroff B.
      • Wichmann H.E.
      • Bauer C.P.
      • et al.
      The effect of hydrolyzed cow’s milk formula for allergy prevention in the first year of life: the German Infant Nutritional Intervention Study, a randomized double-blind trial.
      • von Berg A.
      • Koletzko S.
      • Filipiak-Pittroff B.
      • Laubereau B.
      • Grubl A.
      • Wichmann H.E.
      • et al.
      Certain hydrolyzed formulas reduce the incidence of atopic dermatitis but not that of asthma: three-year results of the German Infant Nutritional Intervention Study.
      Second, a  per-protocol analysis was performed including only those infants who had received some of the allocated formula by 4 months of age.
      To determine whether the effect of the formula on risk of allergic disease varied between those with a family history of eczema (either the mother or father) and those without (neither the mother nor father),
      • von Berg A.
      • Koletzko S.
      • Grubl A.
      • Filipiak-Pittroff B.
      • Wichmann H.E.
      • Bauer C.P.
      • et al.
      The effect of hydrolyzed cow’s milk formula for allergy prevention in the first year of life: the German Infant Nutritional Intervention Study, a randomized double-blind trial.
      a stratified analysis was performed. Interaction effects were assessed by using Wald tests.
      Adjusted associations were also estimated, to allow for any confounding due to chance imbalances at baseline. Adjustment was made for infant sex, parental smoking during pregnancy, and family history of allergic disease in all models. All statistical analysis was performed by A.J.L. using Stata statistical software (release 9.2; Stata Corp, College Station, Tex).

      Results

      A total of 620 infants were recruited (Fig 1). Infants allocated to the CMF and pHWF groups were similar on baseline risk factors (see this article’s Table E1 in the Online Repository at www.jacionline.org). Infants allocated to the soy formula had a higher proportion of parents with food allergy and siblings with allergic disease (Table E1). There were no differences between the groups in terms of duration of exclusive breast-feeding or age of introduction of solids (Table E1).
      Figure thumbnail gr1
      Fig 1Flow chart of participation in the MACS. CM, Cow’s milk formula.
      Approximately 50% of infants received some of the allocated formula by 4 months of age; 16.5% of infants never received their allocated formula because of either continuing breast-feeding (13.6%; n = 78/575) or using a nonallocated formula (2.9%; n = 17/575). There were no differences in rates of exposure to the allocated formula between the groups (Fig 2). The majority of mothers fully adhered to the study formula feeding protocol (breast-feeding and then weaning onto allocated formula with no other formula exposures) during the first 6 months of the child’s life (91.2%, 86.9%, and 87.4% for the CMF, pHWF and soy groups, respectively) despite only 63% of children having been exposed to the allocated formula by this age. The rates of adherence declined by 12 months of age (75.7%, 69.1%, and 76.4%, respectively).
      Figure thumbnail gr2
      Fig 2Proportion of infants exposed to the allocated formula from the time of birth (0 weeks) until 52 weeks of age.
      There were 575 (92.7%) infants followed until 24 months of age (Fig 1); 25 children were lost to follow-up (shifted residence without informing the study), 14 refused ongoing participation, and 6 children did not complete the 2-year follow-up but subsequently rejoined the study (Fig 1).

       Primary outcome

      Neither the pHWF nor the soy formula reduced the risk of allergic manifestations in the first 2 years of life (Table I).
      Table IUnadjusted associations between allocated formula and risk of allergic disease
      OutcomeConventional formula (CMF)Hydrolyzed formula (pHWF)Soy formula
      % (n/N)% (n/N)Crude OR (95% CI)% (n/N)Crude OR (95% CI)
      Any allergic manifestation: 0-1 y (228/575)37.3 (72/193)37.7 (72/191)1.02 (0.67-1.54)44.0 (84/191)1.32 (0.88-1.98)
      Any allergic manifestation: 0-2 y (300/575)48.7 (94/193)53.4 (102/191)1.21 (0.81-1.80)54.5 (104/191)1.26 (0.84-1.88)
      Secondary outcomes
       Eczema within first 2 y43.0 (83/193)48.7 (93/191)1.26 (0.84-1.88)46.1 (88/191)1.13 (0.76-1.69)
      Food reactions within first 2 y
       Any food (92/575)13.5 (26/193)15.2 (29/191)1.15 (0.65-2.04)19.4 (37/191)1.54 (0.89-2.67)
       Cow’s milk protein (17/575)3.1 (6/193)1.6 (3/191)0.50 (0.12-2.02)4.2 (8/191)1.36 (0.46-4.00)
       Cow’s milk with + SPT to cow’s milk (3/575)0 (0/193)0.5 (1/191)NE1 (2/191)NE
       Peanut with + SPT to peanut (1/575)0.5 (1/193)0 (0/191)NE0 (0/191)NE
       Egg with + SPT to egg (8/575)1.0 (2/193)0.5 (1/191)0.50 (0.04-5.59)2.6 (5/191)2.57 (0.49-13.40)
      Childhood outcomes (period prevalence at 6-7 y)
       Eczema (157/493)31.5 (51/162)33.5 (56/167)1.08 (0.69-1.68)30.5 (50/164)0.95 (0.60-1.48)
       Asthma (148/495)32.1 (52/162)28.0 (47/168)0.91 (0.57-1.45)29.7 (49/165)0.97 (0.61-1.54)
       Rhinitis (117/495)22.2 (36/162)22.0 (37/168)0.94 (0.56-1.58)26.7 (44/165)1.27 (0.77-2.10)
       Persistent asthma (120/494)25.5 (41/161)24.2 (40/165)0.88 (0.53-1.46)24.2 (40/165)0.94 (0.57-1.55)
      NE, OR not estimable.

       Secondary outcomes in the first 2 years

      There was no evidence of differences between the groups on any secondary clinical outcome (Table I) or SPT reactivity (Table II). Using a 2-mm mean wheal diameter to define a positive SPT did not change the overall pattern of results or the conclusion that there was no evidence of a difference between the groups. There were 19 (3.2%) children with large SPT (≥6 mm) wheals to cow’s milk, consistent with IgE-mediated cow’s milk allergy,
      • Sporik R.
      • Hill D.J.
      • Hosking C.S.
      Specificity of allergen skin testing in predicting positive open food challenges to milk, egg and peanut in children.
      and these children were evenly distributed between the groups (CMF, 6/197; pHWF, 4/196; and soy, 9/201).
      Table IIUnadjusted associations between allocated formula and risk of positive SPT
      OutcomeConventional formula (CMF)Hydrolyzed formula (pHWF)Soy formula
      % (n/N)% (n/N)Crude OR (95% CI)% (n/N)Crude OR (95% CI)
      Positive SPT (any allergen) at
       6 mo (95/552)16.9 (30/177)18.3 (35/191)1.10 (0.64-1.88)16.3 (30/184)0.95 (0.55-1.66)
       12 mo (146/544)29.2 (52/178)25.0 (47/188)0.81 (0.51-1.28)26.4 (47/178)0.87 (0.55-1.38)
       2 y (136/449)31.6 (50/158)26.0 (38/146)0.76 (0.46-1.25)33.1 (48/145)1.07 (0.66-1.73)
       Repeated measures
      These estimates are based on repeated measures (combining results from the 6, 12, and 24 month SPT using the generalized estimating equations approach), meaning it is not possible to report simple proportions.
      0.90 (0.61-1.33)0.98 (0.67-1.44)
      Positive SPT to cow’s milk at
       6 mo (23/552)5.1 (9/177)4.2 (8/191)0.82 (0.31-2.16)3.3 (6/184)0.63 (0.22-1.81)
       12 mo (32/544)5.1 (9/178)5.9 (11/188)1.17 (0.47-2.89)6.7 (12/178)1.36 (0.56-3.31)
       2 y (16/449)3.8 (6/158)1.4 (2/146)0.35 (0.07-1.77)5.5 (8/145)1.48 (0.50-4.37)
       Repeated measures
      These estimates are based on repeated measures (combining results from the 6, 12, and 24 month SPT using the generalized estimating equations approach), meaning it is not possible to report simple proportions.
      0.89 (0.40-1.99)1.01 (0.44-2.30)
      These estimates are based on repeated measures (combining results from the 6, 12, and 24 month SPT using the generalized estimating equations approach), meaning it is not possible to report simple proportions.

       Secondary outcomes at ages 6 and 7 years

      Between 6 and 7 years of age, 80% (495/620) of children had a telephone interview. There were no differences between the groups in the rates of childhood eczema, asthma, or allergic rhinitis (Table I).

       Adjusted analysis

      Adjustment for sex, parental smoking, and family history of allergic disease did not alter the associations between the allocated group and the risk of any allergic manifestation in the first 2 years of life or any of the secondary outcomes and did not change the interpretation of the results (see this article’s Table E2 in the Online Repository at www.jacionline.org).

       Interactions with family history of eczema

      There was no evidence that pHWF protected against the development of allergic manifestation in those children with or without a family history of eczema (see this article’s Table E3 in the Online Repository at www.jacionline.org; all P values for all interaction terms >.15).

       Per-protocol analysis

      None of the per-protocol analyses produced substantially different findings from the ITT analysis. Limiting the analysis to children whose parents were compliant with the study feeding protocol did not alter the study conclusions (primary outcome OR, 1.20; 95% CI, 0.75-1.93, for pHWF [n = 132 and 146]). Excluding infants exclusively breast-fed for more than 4 months did not alter the results (OR, 1.22; 95% CI, 0.72-2.04, for pHWF [n = 110 and 121]). Similarly, including only infants who had consumed some of the allocated formula (OR, 1.16; 95% CI, 0.66-2.02 [n = 97 and 102]) or consumed the allocated formula for at least 2 weeks during the first 4 months of life (OR, 1.10; 95% CI, 0.59-2.04 [n = 82 and 80]) did not alter the results. Similar results were obtained when the analysis was limited to children who consumed the allocated formula for at least 4 (OR, 1.06; 95% CI, 0.55-2.03 [n = 73 and 73]) and 8 weeks (OR, 1.00; 95% CI, 0.48-2.09 [n = 62 and 55]) in the first 4 months of life. Limiting the analysis to children who consumed at least 100 mL per day for each of these durations produced similar results. Finally, limiting the analysis to children who were exposed to the allocated formula within the first 2 weeks of life again did not produce any evidence of benefit (OR, 0.91; 95% CI, 0.41-2.01 [n = 43 and 58]), although the reduced numbers limited the precision of this comparison. Similarly, interpretation for all secondary outcomes did not change in any of these analyses (data not shown).

      Discussion

      This randomized controlled trial failed to show any beneficial effect of the pHWF for the prevention of any allergic disease outcome up to 7 years of age in high-risk children compared with a conventional cow’s milk–based formula.
      This is the second largest trial to randomize individual infants to receive either pHWF or a conventional cow’s milk formula. An ITT analysis of the largest study, the GINI study,
      • von Berg A.
      • Koletzko S.
      • Grubl A.
      • Filipiak-Pittroff B.
      • Wichmann H.E.
      • Bauer C.P.
      • et al.
      The effect of hydrolyzed cow’s milk formula for allergy prevention in the first year of life: the German Infant Nutritional Intervention Study, a randomized double-blind trial.
      • von Berg A.
      • Koletzko S.
      • Filipiak-Pittroff B.
      • Laubereau B.
      • Grubl A.
      • Wichmann H.E.
      • et al.
      Certain hydrolyzed formulas reduce the incidence of atopic dermatitis but not that of asthma: three-year results of the German Infant Nutritional Intervention Study.
      • von Berg A.
      • Filipiak-Pittroff B.
      • Kramer U.
      • Link E.
      • Bollrath C.
      • Brockow I.
      • et al.
      Preventive effect of hydrolyzed infant formulas persists until age 6 years: long-term results from the German Infant Nutritional Intervention Study (GINI).
      also failed to demonstrate a clear benefit of pHWF over conventional formula for the outcomes of allergic manifestations and eczema up to 12 months, and childhood eczema, asthma, or allergic rhinitis (2-year period prevalence at 6 years).
      • von Berg A.
      • Filipiak-Pittroff B.
      • Kramer U.
      • Link E.
      • Bollrath C.
      • Brockow I.
      • et al.
      Preventive effect of hydrolyzed infant formulas persists until age 6 years: long-term results from the German Infant Nutritional Intervention Study (GINI).
      Although 1 report from GINI showed some benefit for the cumulative prevalence of allergic manifestations up to 3 years of age (relative risk, 0.77; 95% CI, 0.61-0.98),
      • von Berg A.
      • Filipiak-Pittroff B.
      • Kramer U.
      • Link E.
      • Bollrath C.
      • Brockow I.
      • et al.
      Preventive effect of hydrolyzed infant formulas persists until age 6 years: long-term results from the German Infant Nutritional Intervention Study (GINI).
      a previous analysis of the same outcome within GINI did not (population odds ratio, 0.94; 95% CI, 0.73-1.20).
      • von Berg A.
      • Koletzko S.
      • Filipiak-Pittroff B.
      • Laubereau B.
      • Grubl A.
      • Wichmann H.E.
      • et al.
      Certain hydrolyzed formulas reduce the incidence of atopic dermatitis but not that of asthma: three-year results of the German Infant Nutritional Intervention Study.
      Most of the reported benefits of pHWF in GINI are from a per-protocol analysis in which children were excluded if they had not received the allocated formula within the first 4 months of life. It is well accepted that the main conclusions of a randomized controlled trial should be based on an ITT analysis because per-protocol analysis can bias the findings.
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      • Osborn D.A.
      • Sinn J.
      Formulas containing hydrolysed protein for prevention of allergy and food intolerance in infants.
      suggested a benefit of pHWF for the prevention of “any allergic manifestation in infancy” compared with conventional CMF (pooled OR, 0.73; 95% CI, 0.59-0.90). However, when our results are added to this pooled estimate, there is no longer evidence of a protective effect of pHWF for “any allergic disease in infancy” (pooled OR, 0.91; 95% CI, 0.79-1.05). The ITT analysis of the 2 largest studies in this area (GINI
      • von Berg A.
      • Filipiak-Pittroff B.
      • Kramer U.
      • Link E.
      • Bollrath C.
      • Brockow I.
      • et al.
      Preventive effect of hydrolyzed infant formulas persists until age 6 years: long-term results from the German Infant Nutritional Intervention Study (GINI).
      and Melbourne Atopy Cohort Study [MACS]) failed to show any benefit of pHWF compared with conventional CMF, whereas studies with far fewer participants showed stronger protective effects of the pHWF.
      • Marini A.
      • Agosti M.
      • Motta G.
      • Mosca F.
      Effects of a dietary and environmental prevention programme on the incidence of allergic symptoms in high atopic risk infants: three years’ follow-up.
      • Vandenplas Y.
      • Hauser B.
      • Van den Borre C.
      • Clybouw C.
      • Mahler T.
      • Hachimi-Idrissi S.
      • et al.
      The long-term effect of a partial whey hydrolysate formula on the prophylaxis of atopic disease.

      Lam BCC, Yeung CY. The effect of breast milk, infant formula and hypoallergenic formula on incidence of atopic manifestation in high risk infants. 6th Annual Scientific Meeting of Hong Kong College of Physicians and Hong Kong College of Paediatricians, Hong Kong 1992 (October).

      These conflicting results suggest that publication bias may have had an impact on this Cochrane review.
      • Osborn D.A.
      • Sinn J.
      Formulas containing hydrolysed protein for prevention of allergy and food intolerance in infants.
      Alternatively, the effect of pHWF may be influenced by previous breast-feeding. The studies that demonstrated a strong protective effect of pHWF were of a small number of children who received pHWF without receiving any breastfeeding.
      • Marini A.
      • Agosti M.
      • Motta G.
      • Mosca F.
      Effects of a dietary and environmental prevention programme on the incidence of allergic symptoms in high atopic risk infants: three years’ follow-up.
      • Vandenplas Y.
      • Hauser B.
      • Van den Borre C.
      • Clybouw C.
      • Mahler T.
      • Hachimi-Idrissi S.
      • et al.
      The long-term effect of a partial whey hydrolysate formula on the prophylaxis of atopic disease.

      Lam BCC, Yeung CY. The effect of breast milk, infant formula and hypoallergenic formula on incidence of atopic manifestation in high risk infants. 6th Annual Scientific Meeting of Hong Kong College of Physicians and Hong Kong College of Paediatricians, Hong Kong 1992 (October).

      • de Seta L.
      • Sian P.
      • Cirillo G.
      • DiGruttola M.
      • Cimaduomo L.
      • Coletta S.
      The prevention of allergic diseases with a hypoallergenic formula: a follow-up at 24 months: the preliminary results.
      By contrast, studies including MACS that randomized a larger number of infants to pHWF at weaning from breast milk showed a much weaker effect.
      • von Berg A.
      • Filipiak-Pittroff B.
      • Kramer U.
      • Link E.
      • Bollrath C.
      • Brockow I.
      • et al.
      Preventive effect of hydrolyzed infant formulas persists until age 6 years: long-term results from the German Infant Nutritional Intervention Study (GINI).
      • Willems R.
      • Duchateau J.
      • Magrez P.
      • Denis R.
      • Casimir G.
      Influence of hypoallergenic milk formula on the incidence of early allergic manifestations in infants predisposed to atopic diseases.
      The negative findings of our study, and others that randomized before birth, may be a result of infants having less formula than those starting pHWF from birth. In addition, the effect of pHWF may be modified by previous breast-feeding; human breast milk contains a number of important immunologically active components
      • Field C.J.
      The immunological components of human milk and their effect on immune development in infants.
      that may both modify induction of allergen tolerance
      • Verhasselt V.
      • Milcent V.
      • Cazareth J.
      • Kanda A.
      • Fleury S.
      • Dombrowicz D.
      • et al.
      Breast milk-mediated transfer of an antigen induces tolerance and protection from allergic asthma.
      and have an effect on the impact of pHWF on the risk of allergic disease. These possibilities might explain why the per-protocol (exposure within the first 4 months) analysis of the GINI study showed stronger evidence of a protective effect than the ITT, because the per-protocol analysis would have selected out those who were predominantly breast-fed. To resolve this issue conclusively, studies that randomize large numbers of infants to specific infant formula and preclude previous breast-feeding are required. However, this may be logistically difficult and potentially unethical given the current evidence on benefits of breast-feeding.
      The importance of the first 4 months of life for dietary interventions to prevent allergic disease has been emphasized in the literature
      • Host A.
      • Halken S.
      • Muraro A.
      • Dreborg S.
      • Niggemann B.
      • Aalberse R.
      • et al.
      Dietary prevention of allergic diseases in infants and small children.
      • Greer F.R.
      • Sicherer S.H.
      • Burks A.W.
      Effects of early nutritional interventions on the development of atopic disease in infants and children: the role of maternal dietary restriction, breastfeeding, timing of introduction of complementary foods, and hydrolyzed formulas.
      because this is believed to be the critical time of oral immune tolerance development. However, we were unable to show an impact of the pHWF even when we limited our analysis to those children exposed within this time frame. Similarly, including only infants with a significant exposure to the allocated formula within the first 4 months of life did not reveal a protective effect of pHWF.
      In contrast with the GINI study,
      • von Berg A.
      • Koletzko S.
      • Grubl A.
      • Filipiak-Pittroff B.
      • Wichmann H.E.
      • Bauer C.P.
      • et al.
      The effect of hydrolyzed cow’s milk formula for allergy prevention in the first year of life: the German Infant Nutritional Intervention Study, a randomized double-blind trial.
      we did not observe any difference in the effect of a pHWF between children with or without a family history of eczema. It is highly unlikely that pHWF has a differential effect on the basis of family history of eczema.
      Our study has a number of important strengths. We have studied the effect of a pHWF on high-risk children until they were 7 years of age, when the diagnosis of asthma
      • Martinez F.D.
      • Wright A.L.
      • Taussig L.M.
      • Holberg C.J.
      • Halonen M.
      • Morgan W.J.
      Asthma and wheezing in the first six years of life.
      and allergic rhinitis is clearer. Skin prick tests were performed on 3 occasions to cow’s milk as well as 5 other common allergens. This allowed the assessment of a specific effect of pHWF on the risk of cow’s milk sensitization as well as on atopic diseases. The rate of follow-up during early life was exceptional. The sample size in this study was sufficient to detect important differences between the formulas in allergy prevention. It was not designed to demonstrate equivalence.
      The design of our study has some weaknesses. The allocation sequence was available to the research staff throughout the study. Therefore, the research staff would have known the coded group of allocation for the next participant to be enrolled. Despite this, examination of the enrollment into the study indicates that it was time-consecutive, and the staff members undertaking the distribution of formulas were blind to the formula codes. Thus selection bias and ascertainment bias were unlikely to influence the results of this study. In addition, we relied in part on parent-reported outcomes that have not been validated. However, none of the current definitions of eczema
      • Hanifin J.
      • Rajka G.
      Diagnostic features of atopic dermatitis.
      • Williams H.C.
      • Burney P.G.
      • Pembroke A.C.
      • Hay R.J.
      The U.K. Working Party’s diagnostic criteria for atopic dermatitis, III: independent hospital validation.
      have been validated in children under the age of 2 years, although a standardized assessment at the time of SPTs within this study may have improved the measurement of this outcome. We have demonstrated good agreement between the International Study of Asthma and Allergies in Childhood definitions of eczema, asthma, and hay fever and those used in this study at age 6 to 7 years (all κ values ≥.74; unpublished data A. Lowe, December 2007).
      This study tested the effect of a pHWF at weaning on the incidence of allergic manifestations. It does not provide information concerning the impact of exclusive feeding with pHWF nor other forms of partially or extensively hydrolyzed or amino acid-based formula.

       Conclusion

      There was no evidence that introducing pHWF at the cessation of breast-feeding reduced the risk of allergic manifestations, including eczema, asthma, and allergic rhinitis, in this study of high-risk infants. Our findings do not support the recommendation that pHWF should be used after breast-feeding as a preventive strategy for infants at high risk of allergic diseases.
      Clinical implications
      The authors found no evidence to support the use of pHWF at weaning for the prevention of allergic disease in infants with a family history of allergic disease.
      We thank Dr John Thorburn, FRACP, for assistance in patient recruitment and administrative assistance and the Mercy Maternity Hospital Department of Obstetrics for participant recruitment. We thank Anne Balloch for assistance with data management and all of the MACS children and parents for their participation and ongoing support for this study.

      Appendix

      Table E1Comparison of baseline factors between the allocated formula groups
      Baseline factorHydrolyzed formula (pHWF) (n = 206)Soy formula (n = 208)Conventional formula (CMF) (n = 206)
      Male infant50.8%51.0%51.9%
      Maternal history of allergic disease
       Asthma42.9%46.6%40.3%
       Eczema38.5%36.1%42.2%
       Hay fever61.5%59.1%61.2%
       Food allergy37.4%43.8%34.5%
      Paternal history of allergic disease
       Asthma21.6%27.1%28.4%
       Eczema19.1%18.4%24.0%
       Hay fever47.5%48.3%42.6%
       Food allergy20.6%28.5%14.7%
      Demographic factors
       Median maternal age (y) (IQR)31 (29-34)32 (29-34.5)31 (28-34)
       Median paternal age (y) (IQR)33 (30-36)33 (31-36)32.5 (29.5-36.5)
       Median maternal education (y) (IQR)15 (12-15)15 (12-15)15 (11-15)
       Median paternal education (y) (IQR)15 (12-15)15 (12-15)15 (11-15)
       Median SES of father’s occupation (IQR)
      SES classified using the Australian National University (ANU)-3 system,E1 which ranges from 0 to 100, with higher values indicating higher SES.
      48.6 (34.0-61.9)43.6 (29.2-62.6)41.6 (27.2-61.9)
      Home environment
       Owner-occupied home84.0%81.7%80.1%
       Any gas cooking75.1%73.2%79.7%
       Any gas heating74.1%66.8%71.8%
       Any pet70.1%70.8%65.7%
       Maternal smoking during pregnancy7.8%4.8%10.2%
       Paternal smoking during pregnancy19.0%16.4%22.7%
      Sibling factors
       No older siblings43.2%33.7%43.7%
       Any older sibling with food allergy35.4%50.5%26.2%
       Any older sibling with eczema35.0%46.6%33.5%
       Any older sibling with asthma32.5%45.7%27.7%
       Any older sibling with hay fever19.5%26.4%19.4%
      Early diet (wk)
       Median duration of exclusive breast-feeding (IQR)14 (3-20)15 (1-21)13 (1-20)
       Median duration of any breast-feeding (IQR)
      Excludes 37 infants who were not breast-fed.
      42 (22-60)47 (17-64)44 (24-60)
       Median age of introduction to solid foods (IQR)20 (18-22)19 (16-24)20 (17-22)
      IQR, Interquartile range; SES, socioeconomic status.
      SES classified using the Australian National University (ANU)-3 system,
      • McMillan J.
      • Jones F.L.
      The ANU3_2 scale: a revised occupational status scale for Australia.
      which ranges from 0 to 100, with higher values indicating higher SES.
      Excludes 37 infants who were not breast-fed.
      Table E2Adjusted associations between allocated formula and risk of positive SPT and allergic disease
      OutcomeHydrolyzed formula (pHWF)Soy formula
      adjusted OR (95% CI)adjusted OR (95% CI)
      Any allergic manifestation
       0-1 y (228/575)†0.97 (0.63-1.48)1.23 (0.81-1.88)
       0-2 y (300/575)†1.22 (0.81-1.85)1.21 (0.80-1.84)
      Secondary outcomes
       Eczema within first 2 y (264/575)†1.24 (0.82-1.88)1.11 (0.73-1.68)
      Positive SPT within first 2 y∗
       Any allergen0.88 (0.59-1.30)0.92 (0.61-1.38)
       Cow’s milk0.79 (0.35-1.77)0.78 (0.32-1.92)
       Any food reaction within first 2 y0.95 (0.51-1.75)1.21 (0.67-2.19)
      Childhood outcomes (period prevalence) at 6-7 y
       Eczema (157/493)1.10 (0.70-1.72)0.90 (0.57-1.42)
       Asthma (148/495)‡0.82 (0.50-1.33)0.82 (0.50-1.34)
       Rhinitis (117/495)§0.91 (0.54-1.55)1.24 (0.74-2.09)
      All ORs compared to the conventional CMF group. All models adjusted for infant sex and parental smoking during pregnancy unless otherwise stated. Also adjusted for ∗parent and sibling food allergy, †parent and sibling eczema, ‡parent and sibling asthma, or §parent and sibling allergic rhinitis.
      Table E3Unadjusted associations between allocated formula and risk of allergic disease outcomes according to family history of eczema
      OutcomeNo family history of eczemaFamily history of eczema presentP value for interaction
      Conventional formula (CMF)Hydrolyzed formula (pHWF)Conventional formula (CMF)Hydrolyzed formula (pHWF)
      % (n/N)% (n/N)OR (95%CI)% (n/N)% (n/N)OR (95% CI)
      Primary outcome
       Any allergic manifestation: 0-2 y49.4 (38/77)49.4 (41/83)1.00 (0.54-1.86)47.4 (54/114)55.7 (59/106)1.39 (0.82-2.37).43
      Secondary outcomes
       Eczema in first 2 y40.3 (31/77)44.6 (37/83)1.19 (0.64-2.24)44.7 (51/114)50.9 (54/106)1.28 (0.75-2.18).86
      Positive SPT within first 2 y
       Cow’s milk (3 mm+)
      These estimates are based on repeated measures (combining results from the 6, 12, and 24 month SPT using the generalized estimating equations approach), meaning it is not possible to report simple proportions.
      0.95 (0.24-3.84)0.88 (0.34-2.30).93
       Any allergen (3 mm+)
      These estimates are based on repeated measures (combining results from the 6, 12, and 24 month SPT using the generalized estimating equations approach), meaning it is not possible to report simple proportions.
      1.18 (0.63-2.18)0.78 (0.47-1.28).31
      Childhood outcomes at age 6-7 y
       Eczema16.4 (11/67)25.4 (18/71)1.83 (0.79-4.23)40.9 (38/93)38.9 (37/95)0.89 (0.52-1.53).16
       Asthma28.4 (19/67)26.8 (19/71)1.10 (0.53-2.32)35.5 (33/93)28.4 (27/95)0.76 (0.41-1.38).44
       Allergic rhinitis17.9 (12/67)25.4 (18/71)1.54 (0.67-3.51)24.7 (23/93)20.0 (19/95)0.73 (0.37-1.44).17
      These estimates are based on repeated measures (combining results from the 6, 12, and 24 month SPT using the generalized estimating equations approach), meaning it is not possible to report simple proportions.

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