Pharmacology and Therapeutics
`
`Comparing 2.5%, 5%, and 10% BenzoyS Peroxide on
`Inflammatory Acne Vulgaris
`
`OTTO H. MILLS, JR., PH.D., ALBERT M. KLIGMAN, M.D., PH.D., PETER POCHb M.D.,
`AND HARRIET COMITE, M.[).
`
`From the Departments of Dermatology, University of
`Pennsylvania School of Medicine, Phi(adelphia,
`Pennsylvania, and Boston University School of Medicine,
`Boston, Massachusetts
`
`ABSTRACT: A 2.5% formulation of benzoyl peroxide was
`compared with it5 vehicle, andwitha5%anda tO%proprietary
`benzoyl peroxide get preparation in three double-blind studies
`involving !53 patients with mild to moderately severe ache
`vulgaris. The 2.5% benzoyl peroxide formulation was more
`effective than its vehicle and equivalent to tee 5% and tO%
`concentrations in reducing the number of inflammatory lesions
`(papules and pustules). Desquamation, enythema, and syrup-
`toms of burning with the 2.5% go!~ were less trequent than
`with the 70% preparation but equivalent to the 5% gel. The
`2.5% "formulation also Significantly reduced Propionibactetium
`acnes and the percentage of free fatty acids in the surface
`lipids after 2 weeks ot topica! application,
`
`B
`
`enzoy] peroxide, in concentrations of 5%, 10%,
`and 20%, has been used effectively in the treat-
`ment ofacne for more than 20 years.1-~ This compound
`has been shown to suppress Propionibacterium aches
`in vivo, the pr.obable basis for its therapeutic effect)
`With such concentrations, side effects such as ery-
`theme, desquamation, and b urnin$ itching, or stinging
`are fairly common. This paper describes clinical trials
`of a 2.5% benzoyl peroxide get, which was compared
`with 5% and 10% benzoyL peroxide gels in groups of
`patients with inflammatory acoe vuigaris. Antibacterial
`and lipid studies were aiso performed on the 2.5%
`benzoyJ peroxide formulation,
`
`Address for correspondence: Otto H. Mills, Jr., Ph.D., UMDNI-
`Robert Wood Johnson Medical School, One Robert Wood Johnson
`Piece, New Brunswick, NJ 08903.
`
`Subjects, Malerials, and Methods
`
`Clinical Studies
`
`The same methods were used to conduct three double-
`blind studies. After g~ving informed consent, subjects with
`mild to moderately severe inflammatory acne vulgaris of the
`face (minimum of 10 inflammatory lesions) were assigned to
`one of the three treatment groups. A total of ~ 53 s~bi~cts,
`74 men and 79 women (average age of 20 years), participated
`in the three studies. In the first, 2.5 subjects used 2.5% ben-
`zoyi peroxide gel a!ld 25, the gel vehicle for this formulation.
`In the second, 26 used the 2.5 ge] and 27, a 5% gel. The
`third study consisted of 25 subjects who used the 2.5% 8el
`and 25, a 10% benzoy~ peroxide gel The subiects received
`no medications for any reasoP, s dur;,P,g the ~-week period
`prior to the start of the study.
`The study participants were instructed to wash daily with
`a non-medicated soap, rinse, and dry with a c~ean towel
`~eiore applying the study medication to the face twice daily
`(morning a nd even ing) (or 8 weeks. Subiects were examined
`before treatment for baseline determinations and at weeks
`2, 4, 6, and B afte~ the start of treatment. A~. each visit, the
`number of fadal inflammatory lesions (papules and pustules)
`was recorded. In addition, the frequency and severity of side
`effects such as erythema, peeling, and burning were noted,
`A 8lobar assessment of improvement was also made by the
`investigator at each visit, according to the following criteria:
`"excellent," greater than 75% improvement; "good," about
`50% improvement; "~aiT," about 25% ~mprovement; and
`"poor," little or no improvement.
`
`Antibacteria~ and Lipid Studies
`
`Ten subjects who had a P. aches count of 100,000 co[onies
`per cm2 or greater on the face were selected l;or this study.
`k score of 3 or greater on the fo)licuJar porphyrin fluorescence
`scale was also required for admission. The density of P. aches,
`the degree of porphyrin ffuoresc~ence, and the ratio of [ree
`fatty acids to triglycerides found ~n |~,p~,d samples w~Te 6e-
`termined before and after 7 and 14 days of twice-daily ap-
`plications of the 2.5% benzoyt peroxide formulation to the
`face. All applications were made by laboratory technicians.
`At each sample day (0, 7, 14), the skin was prepared by
`wiping the surface o[ the forehead for 30 seconds with a
`piece o(gauze saturated with 0.1% Triton X-IO0 (Rohm and
`Haas, Philadelphia, PA!, fo~,~owed by a dis~.i~ed water rinse
`
`664
`
`1 of 5
`
`Almirall EXHIBIT 2045
`Amneal v. Almirall
`IPR20I 8-00608
`
`

`

`No. I0
`
`BENZOYL PEROXIDE IN ACNE
`
`MiXsetal.
`
`665
`
`TABLE 1. Comparison of Effects of 2.5% Benzoyl Peroxide and Vehicle
`
`2.5% Benzoyi Peroxide
`
`VeMc~e
`
`week
`
`Number o|
`Subjects
`
`Mean Number
`Of Lesions"
`
`Mean %
`Reduction
`
`Number oi
`Subjects
`
`Mean Number
`of Lesions*
`
`Mean %
`Reduction
`
`0
`2
`4
`6
`8
`
`25
`25
`;t4
`
`25
`25
`
`" Papules and pUslules.
`t By ~-Iest.
`
`13.6
`10.O
`9.3
`7.5
`6.8
`
`--
`20.4%
`31.496
`44.t%
`50.9O/o
`
`25
`25
`25
`22
`25
`
`13.7
`13.8
`I3.0
`13.1
`11.2
`
`--
`-2.7%
`2.8%
`3.4%
`17.6%
`
`~ Difference in baseline counts.
`
`P Valuer tar
`Average
`T~eatmenl
`Difference
`
`0.91 :~
`<0.01
`0.02
`<0.01
`0.01
`
`and a 30-second wipe with hexane. This procedure removed
`environmental contaminants, desquamaling cells, and surface
`bacteria and lipids. Two areas were protected by plastic
`weighing boats w|tfl several per(orations to atlow evaporation
`ot sweat. After 1 hour, the site was sampled by the detergent
`scrub technique of Williamson and Kligman.9 A sterile glass
`cylinder with an internal area ot 3.8 cm~ was placed over the
`site. One ml of 0.1% Triton X-IO0 in 0.075% phosphate
`buffer (pH 7.9)was added and the surface scrubbed with a
`blunt Tel]on (Arthur H. Thomas, Philadelphia, PAl spatula
`for one minute. This procedure was repeated, and the two
`samples were pooled. Subsequently, tenfold dilutions were
`made in 0.5% buffered Triton X-100; the samples were drop-
`plated on brain heart agar with O. 1% Tween 80 ~Atias Chem-
`!cat, Wilmington, DE) and incubated anaerobically for 7 days
`in a Gas Pak (Arthur H. Thomas, Philadelphia, PA) jar system.
`P. aches was identified by colony morphology, by suscep-
`tibility to P. aches bacteriophage, and, when indicated, by
`biochemical testing?°n’
`At the other site on the forehead, lipid samples were col-
`tected by putting 2 m? ot’ hexane-containin8 methyl nervon-
`ate, as an internal standard, in the glass cup, as above. The
`site was scrubbed for 30 seconds with a blunted Teflon po-
`I)ceman, The solution was taken up on a Pasteur pipette and
`passed through a 0.45-mill!pore filter to remove skin debris
`and bacteria. It was then placed in Teflon capped glass
`screwtop vials. The vials were uncapped, dried overnight in
`a vacuum at 40 C, then ¢-~apped and stored at 40 C unti! lipid
`thin-layer chromatography was done by the method of
`Downing.t2
`Each subject was e:~amined under Wood’s light for par-
`phyrin fluorescence prior to washing and obtaining samples.~
`
`The intensity of fluorescence was graded on a 0 to 6 scale
`wilh 0 = none, 1 to 2 = mild fluorescence, 3 to 4 = moderate,
`and 5 to 6 = heavy fluorescence.
`
`Statistical Methods
`
`Fisher’s exact test1~ was used to compare treatment groups
`with respect to side effects at each visit duringthe treamlent
`period. A group t-testi" was used to compare treatment
`groups with regard to the reduction in number of papules
`and pustules from baseline. A paired I-test1~ was used to
`analyze changes from baseline counts within t~eatment
`groups. The Wilcoxon rank-sum tesP~ was used to analyze
`changes estimated by global ratings.
`
`Results
`
`Results on efficacy and side effect data are presented
`separately for each study. No subject was obliged to
`drop out of any study because of adverse effects.
`
`Study 1: 2.5% Benzoyt Peroxide Versus Vehicle
`
`The 2.5% ibenzoyF peroxide was more effective than
`the vehicle in reducing the number of inflammatory
`lesions (papules and pustules) at alt follow-up visits
`(Table 1 ).
`The 2.5% benzoyl peroxide was also significantly
`more effective than the vehicle in global ratings at all
`evaluations. Mild peeling, burning, and itching were
`
`TA8L~ 2. Comparison of Effect 0[2.5% and $.0% Benzoyl Peroxide Gel
`
`2.5°5 Benzoyl Peroxide
`
`5% genzoyl Peroxide
`
`week
`
`Number of
`Subjecls
`
`Mean Number
`of Lesions*
`
`Mean %
`Red~clion
`
`Number of
`5ubjecis
`
`Mean Number
`of Lesions
`
`Mean %
`Reduction
`
`0
`2
`4
`6
`8
`
`26
`26
`25
`26
`26
`
`¯ Papules and pustules.
`f 8y t.test.
`
`21.3
`14.8
`13.3
`10.0
`9.6
`
`~
`32.2%
`40.3°/0
`543%
`553%
`
`27
`27
`27
`25
`25
`
`?9.4
`13.5
`12.9
`t 0.6
`7÷8
`
`~
`30.6%
`35,1%
`47.2%
`57.7%
`
`~ Difference in baseline counts.
`
`P Valuer/or
`Average
`Treatment
`Difference
`
`0.47:~
`0.75
`0,5t
`0.41
`0,94
`
`2 of 5
`
`

`

`666
`
`INTERNATIONAl_ IOURNAL OF DERMATOLOGY De~ember 1986
`
`Vol, 25
`
`TAaLE 3. Comparison of Effec~ of 2.5% and 10% Benzoyl Peroxide Gel
`
`2.5% BenzoyI Peroxide
`
`10%% Senzoy1, Peroxide
`
`Week
`
`Number of
`Subjecls
`
`Mean Number
`of Lesions*
`
`Mean %
`Reduction
`
`Number of
`Subjects
`
`Mean Number
`of Lesions
`
`Mean %
`Reduction
`
`0
`2
`4
`6
`B
`
`25
`25
`24
`24
`24
`
`* Papules and pustules.
`By t-test.
`
`T9.7
`16. I
`12.8.
`10.9
`10.5
`
`--
`18.3%
`35.0%
`44.7%
`46.79/o
`
`24
`25
`24
`23
`24
`
`23.7
`19.0
`"t4.9
`14.5
`t 3.2
`
`--
`19.8%
`37.to/o
`38.8%
`44.7°/Q
`
`~ Difference in baseline counts.
`
`P Valvet Jor
`Average
`Treatmenl
`Difference
`
`.1 7~
`.66
`.47
`.75
`,57
`
`more frequent in the benzoyt peroxide group than in
`the vehicle group, but only statistically significantly so
`for peeling at week 8. There was no significant differ-
`ence in the incidence of erythema, although 2.5%
`benzoyi peroxide more often induced erythema at
`week 2.
`
`Study 2: 2.3% Versus 5.0% Benzoyl Peroxide
`
`No significant difference in efficacy between the
`2.5% and the 5.0% benzoyl peroxide formulations was
`noted. In both groups, a significant reduction in pa-
`pules and pustules was observed at 2, 4, 6, and 8 weeks
`{Table 2). The global ratings confirmed the lack of sig-
`nificant difference in efficacy between the 2.5% and
`5.0% gel formulations. There was no significant dif
`ference between the two preparations in regard to
`burning, peeling, or erythema.
`
`Study 3: 2.5% Versus 10% Benzoyl Peroxide
`
`Both 2.5% and 10% benzoyl peroxide gels reduced
`the number of papules and pustules from baseline
`counts, but there was no statistically significant differ-
`ence between the two groups (Table 3). Statistical
`evaluation of the investigator’s global response ratings
`also indicated that there was no significant difference
`between the efficacy of 2.5% and 10% benzoyl per
`oxide,
`There was a statistically significant difference in the
`frequency and severity of burning, erythema, and
`peeling among subjects who used 10% benzoyl per-
`oxide than among those who used the 2.5% concen-
`tration at all follow-up visits (Table 4).
`
`also a significant reduction in the ratio of free fatty
`acids to triglycerides.
`
`Discussion
`
`The 2.5% benzoyl peroxide formuPation was signif-
`icantly more effective than its vehicle in reducing the
`number of papules and pustules and was comparable
`to the 10% benzoyl peroxide by lesion counts. By the
`same measurement, there were no differences be-
`tween the 2.5% gel and the 5% benzoyl peroxide gel;
`both were clinically effective. The incidence of irrita-
`tion was lower with 2.5O/o than with ! 0O/o benzoyl per-
`oxide. It should be pointed out that in two of these
`clinical studies there would need to have been much
`larger patient groups to assure "statistical power" for
`differences between treatments. The differences be-
`tween the 2.5% benzoyl peroxide and its vehic[e is
`not a question. A clear significant difference between
`these two exists. When the 2.5O/o versus 50 and 2.5%
`versus IOOA~ studies were reviewed (or "st, a*,isU.caI,
`power," it is evident that, with the number of subjects
`involved, the power of the test was not high enough
`to assure a differer~ce that was statistica!ly significant.
`However, we feel these studies are clinically significant
`and present important information for clinicians and
`those working in dermatopharmacology.
`Also, these studies do not represent a titration of
`percent.concentration of drug in the same vehicle. The
`2.5% formulation vehicle was different from those of
`
`TABLE 4. Frequency and Severity o(Burning, Erythema, and
`Peeling* (Total Number of Reports for 8 Weeks)
`
`2.5% genzoyP Peroxide Ge!
`
`i 0% 8enzoyt Peroxide Gel
`
`Bacteriology and Free Fatty Acids
`
`Mild Moderate
`
`Mild
`
`Moderate
`
`A marked reduction in the quantity of P. aches was
`observed after 1 week (Table 5). The intensity of fol-
`licular porphyrin fluorescence was also reduced by 1
`week and markedly suppressed by 2 weeks. There was
`
`Burning
`Erythema
`Peeling
`
`20
`22
`50
`
`1
`4
`9
`
`BurninR
`Erythema
`Peeling
`
`57
`51
`36
`
`20
`30
`.55
`
`¯ Possibly o~ prohab|y rel, a~.ed ~o druB.
`
`3 of 5
`
`

`

`No. 10
`
`BENZOYL PEROXIDE IN ACNE
`
`Mil!5 et al.
`
`667
`
`TABLE 5- EffeCt Of Topical Application of 2.5% Benzoyl Peroxide Gel on Quantitative P. aches Counts,
`Follicular Po~phyrin Fluorescence and Free Fatty Acids in Skin Surface Lipids
`
`P. aches (Iog/cm=}
`
`Follicular Porphyrin Fluorescence
`(Grades 0-6)
`
`Flee Falty Acids/Trigtyceddes
`
`Week 0
`
`Week !
`
`Week 2
`
`Week 0
`
`Wee~k I
`
`Week 2
`
`Week 0
`
`Week f
`
`Week 2
`
`5,8285
`6.2775
`5.2775
`6.6455
`6.1015
`5.6243
`6.7383
`6.3647
`6.4372
`5,7035
`6,083
`--
`
`4.9254
`4.3647
`3.7547
`4.0223
`5.5538
`3.1684
`5.6657
`4.0223
`5.7383
`4.6021
`4.504
`<0.001
`
`4.4694
`3.6455
`3.6455
`4.4994
`4.0223
`3.4025
`5.6021
`3.7383
`5.6455
`3.1684
`4.108
`<0.00~
`
`6
`6
`6
`5
`4
`4
`5
`6
`5
`6
`5.30
`--
`
`4
`5
`6
`4
`3
`4
`3
`4
`3
`4
`4,00
`<0.0!
`
`2
`3
`4
`2
`2
`2
`1
`3
`2
`2
`2.30
`<O.OOl
`
`0.47
`0.84
`0.38
`0.22
`! ,77
`0.93
`1,25
`0.22
`! .05
`0.19
`0.732
`--
`
`0.16
`0.73
`0.16
`0.19
`1.12
`0.86
`1.19
`0.16
`0.99
`0.19
`0.575
`<0.02
`
`0,14
`0.66
`0.14
`0.11
`0.86
`0.55
`0.89
`0.14
`0.60
`0.10
`0.4;9
`<0_01
`
`Subject
`Number
`
`I
`2
`3
`4
`5
`6
`7
`8
`9
`10
`Mean
`p vaJue
`
`the .5% and 10% formulations, but we think it impor-
`tant that we saw these results with percent concentra-
`lions of drug that were one-fourth to one-half less than
`the highest concentration,
`The laboratory results from the in vivo study of P.
`aches showed the 2.5% benzoyt peroxide gel reduced
`the anerobic population by 97% after twice-daily
`treatments for 1 week and by 99% after 2 weeks. This
`outcome is in agreement with previous work using a
`different 2.5% benzoyl peroxide formulation.’s
`Regarding the clinical changes of peeling and ery-
`thema and the symptoms of burning, there were no
`differences between 2.5% versus 5%, but differences
`did exist between the 2.5% and its vehicle and the
`2.5% and 10% formulations. With this in mind, the
`lower concentration of benzoyl peroxide should be
`useful for treating patients with easily irritated skin.
`Also, in combination topical therapy with comedolytic
`agents, 2.5% benzoyl peroxide might lessen the ex-
`pected degree of irritation.
`
`Drug Names
`
`2.5% benzoyl peroxide: Clear by Design
`5.0% ber~zoyl peroxide: Desquam X-5
`10.0% benzoyl peroxide: Desquam X-lO
`
`References
`
`1, Frank L. Active oxygen ache therapy--oxygenatmn vs, reduction
`of the follicular structures. Cutis. 1965;I :306-308.
`
`2. Pace WE. A henzoyl peroxide-suJfur cream for ache vulgads.
`(:an Med Assoc I. 1965;93:252-254.
`3. Oanto JL, Maddin WS, Steward WD, et al. A controlled trial of
`benzoyl peroxide and precipitated sulfur cream in ache vul-
`garis, Appl Ther, 1966;8"624-625.
`4. Belknap BS. Treatment of acne with 5 percent benzoyl peroxide
`gel or 0.05 percent of retinoic acid cream. Cuffs. 1979;23:
`856-859.
`5. SmilhEB, PadillaRS, McCaheJM, etaI. Benzoylperoxidelotion
`(20 percent) in acne. Culls. 1980;25:90-92.
`6. Kligman AM, Mills OH, McGinley KJ, et al. Ache therapy with
`tretinoin in combination wilh antibiotics, Acfa Derm Verereol.
`1975;74(Suppl):111-115.
`7. Martin RJ, Kahn G, Oooding )W, et aL Cutaneous porphyrin
`fluorescence as an indication of antibiotic absorption and ef-
`fecliveness, cuffs. 1973;12:758-764.
`8. McGinley KI, Webster CF, Leyden II. Facial follicular porphyrin
`0uorescence: correlation with age and density of Propmni-
`bacterium acnes. Br J DermatoL 1980;102:437-441.
`9. Williamson P, Kligman AM. A new method for the quantitalive
`investigation of cutaneous bacteria. I Invest Dermatol.
`1965;45:498-503.
`io Marples RR, McGinley K), Corynebacterium aches and the an-
`aerobic diphtheroids from human skin. ~1 Meal MicrobioL
`1974.7:349-357.
`1 I. McGinley KJ, Webster GF, Leyden JJ. Regional variation of cu-
`taneous propionibacterium. J Appl [nv Microbiol. 1978;35:
`62 -66.
`12. Downing DT, Pholodensitometry in the thin-layer chromato-
`graphic analysis of neutral lipids. I Chromatogr. 1968;38:91 -
`99.
`13. Ostle B. Statistical inference: testing hypotheses: statistics in re-
`search, Iowa City, IA: iowa State University Press, 1963;119-
`321.
`!4, Seigel S. The case of k independent samples: nor~pararnelri(cid:128)
`statistics for the behavioral sciences. New York: McGraw-
`Hill, 19s6:101-117.
`15. Leyden JJ, McGinley KI, Mitls OH, el al. Topical antibiotics and
`topica! antimicto~ial agents in acne therapy. Acta Derm Ve-
`nereol. 1980;89(Suppl):75-82.
`
`4 of 5
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