Laboratory and clinical trials of cocamide diethanolamine lotion against head lice

Context. During the late 1990s, insecticide resistance had rendered a number of treatment products ineffective; some companies saw this as an opportunity to develop alternative types of treatment. We investigated the possibility that a surfactant-based lotion containing 10% cocamide diethanolamine (cocamide DEA) was effective to eliminate head louse infestation. Settings and Design. Initial in vitro testing of the lotion formulation versus laboratory reared body/clothing lice, followed by two randomised, controlled, community-based, assessor blinded, clinical studies. Materials and Methods. Preliminary laboratory tests were performed by exposing lice or louse eggs to the product using a method that mimicked the intended use. Clinical Study 1: Children and adults with confirmed head louse infestation were treated by investigators using a single application of aqueous 10% cocamide DEA lotion applied for 60 min followed by shampooing or a single 1% permethrin creme rinse treatment applied to pre-washed hair for 10 min. Clinical Study 2: Compared two treatment regimens using 10% cocamide DEA lotion that was concentrated by hair drying. A single application left on for 8 h/overnight was compared with two applications 7 days apart of 2 h duration, followed by a shampoo wash. Results. The initial laboratory tests showed a pediculicidal effect for a 60 min application but limited ovicidal effect. A longer application time of 8 h or overnight was found capable of killing all eggs but this differed between batches of test material. Clinical Study 1: Both treatments performed badly with only 3/23 (13%) successful treatments using cocamide DEA and 5/25 (23.8%) using permethrin. Clinical Study 2: The single overnight application of cocamide DEA concentrated by hair drying gave 10/56 (17.9%) successes compared with 19/56 (33.9%) for the 2 h application regimen repeated after 1 week. Intention to treat analysis showed no significant difference (p = 0.0523) between the treatments. Over the two studies, there were 18 adverse events possibly or probably associated with treatment, most of which were increased pruritus after treatment. Conclusions. Cocamide DEA 10% lotion, even when concentrated by hair drying, showed limited activity to eliminate head louse infestation.

113 Both "Immobile" and "Killed" were included in mortality figures. For louse eggs, those that had 114 developed fully and from which a living louse nymph had emerged completely were recorded as 115 "Hatched". In some cases the nymph had developed and started the emergence process but had 116 died before escaping from the eggshell ("Half-hatched"). Eggs in which there was no obvious 117 sign of embryonic development, specifically no appearance of the eyespot, which would normally 118 become apparent between the third and fourth day of incubation, were designated "Undeveloped", 119 whereas eggs that contained an embryo with eyespots but that failed to hatch were recorded as 120 having "Died". In the case of louse eggs, both "Died" and "Undeveloped" were included in 121 mortality figures. 122 123 In preparation for the second clinical study we ran a series of tests against louse eggs, using 124 essentially the same methods, to compare different treatment regimens of exposure time (1 hour, 2 125 hours, or overnight); washing with shampoo followed by rinsing or washing with water only; and 126 the influence of hair drying over a period of approximately 5 minutes, using an electric hair dryer 127 on "Cool" setting held at a variable distance of approximately 30cm from the test specimens, 128 compared with humidification after application of the product. The temperature during hair drying 129 was modulated by placing the fingers of the operator's hand between the air jet and the test insects 130 and by movement of the device nearer of further away during the drying process so that a 131 temperature of no greater than 40 o Celsius occurred in the dish where the insects were held, 132 measured using a thermocouple probe. Also we compared the sensitivity of louse eggs of different 133 ages to the product by collecting eggs on a single day, storing them under the same conditions and 134 then treating randomly sampled gauze squares, each bearing approximately 100 eggs, on the 135 subsequent test days. As with the first series of laboratory tests the control groups were treated in 136 the same way but using tap water for the initial exposure to fluid. 137 138 After completion of the second clinical study, the low level of efficacy achieved, despite using 139 longer application times and increasing the dose concentration of the cocamide DEA by 140 evaporation, caused us to consider the possibility that the two batches of cocamide DEA lotion 141 were different in some way. We were able to do this by performing further in vitro tests in which 142 different groups of louse eggs from the same batch were treated with samples the two batches of 143 lotion, using both 2 hour and overnight applications followed by a water rinse. 158 10 minutes to pre-washed and towel dried hair, followed by rinsing. Both products were applied 159 on a single occasion. Shampoo was supplied to both groups by the investigators applying the 160 treatment. No guidance on routine hair washing was given for this study.

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162 The second study compared two treatment regimens using cocamide DEA 10% (batch number NH 163 35101). In one group we applied the product to dry hair until it was thoroughly soaked then 164 evaporated the excess water using a hair dryer leaving the hair sticky, with the appearance of 165 having been heavily oiled. The length of the "drying" process varied according to the length and 166 thickness of the hair and ranged from approximately 3-20 minutes, which was determined by the 167 appearance and feel of the treated hair. This was then left overnight then washed off by the 168 participant or parent with plain warm water in the morning. There was no second treatment.

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170 For the other group, we also applied the product to dry hair and dried off excess water with the 171 hair dryer to the same sticky stage. This was then left for a timed 2 hour period before being rinsed 172 off with plain warm water by the investigator. A repeat treatment was given 7 days later.
174 For both groups, participants were given bottles of the same non-medicated shampoo to use for 175 normal hair washing on the third and tenth days after treatment.

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177 Participants 178 Participants were recruited from respondents to an invitation letter and information sheet 179 distributed through schools or via general practitioners. Parents of children with lice telephoned 180 the study co-ordinator and made an appointment for a home visit. We visited within 24 hours to 181 check prospective participants for living lice using a plastic detection comb (Albyn of Stonehaven, 182 Stonehaven, Scotland). If an infestation was found, consisting of one or more live lice, the person 183 was invited to join the study and the parent of guardian guided through a written consent 184 procedure followed by child assent. All household members were offered examination and, if 185 found to be infested, the opportunity to join the study if fitting the enrolment criteria. In this 186 context the intensity of an infestation was partially subjective: Heavy = more than one louse found 187 with the first stroke of the comb; Medium = one louse found with the first stroke; and Light = lice 188 found only after several strokes of the comb over different parts of the head. Lice were returned to 189 the head because treatment followed. The number of participants with each level of infestation is 190 shown in Tables 1 and 2.   191 192 Consenting participants provided baseline demographic data including age, gender, hair 193 characteristics, concurrent medications, and medical history. Some demographic characteristics, 194 such as hair dryness and thickness, were subjective assessments made by the investigators on the 195 day and were intended to serve only as a guideline. In both studies the lower age limit was 4 196 years. People who were sensitive to paraben preservatives; who had persistent disorders of the 197 scalp such as eczema or psoriasis; who had received a head louse treatment within the 4 weeks 198 prior to entry; or had undergone antibiotic treatment or had their hair bleached, colour treated, or 199 permanently waved within the previous 4 weeks, were excluded. All treatments and assessments 200 were performed in the participant's home. No payment was offered for participation.

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202 For the first study, between July 1998 and January 1999, we recruited 44 participants (35 children 203 and 9 adults), out of the 120 planned, divided equally between cocamide DEA 10% and 1% 204 permethrin creme rinse treatments (Figure 1). Of these one person treated with permethrin was 205 lost to follow up before any post-treatment checks could be made. Six others (four treated with 206 cocamide DEA and two with permethrin) were withdrawn at the participant's request due to lack 207 of efficacy. This left 19 participants treated with cocamide DEA and 18 with permethrin with 208 complete data sets before the sponsor requested an early termination on grounds of lack of 209 efficacy for both products.
210 211 In the second study we recruited 112 out of 120 planned enrolments between October and 212 December 1999. All participants were treated with cocamide DEA 10% divided between two 213 treatment regimens: either a single overnight application or two 2 hour applications a week apart.
214 There were seven participants withdrawn (Figure 2). Three people, one from the single 215 application group and two from the double application group dropped out because a baby from the 216 family was hospitalised and the others were unable to keep appointments. Four participants were 217 withdrawn by investigators due non-compliance, one from the single application group and three 218 from the double application group. This left 54 people treated with one overnight application and 219 51 who received two 2 hour applications. Recruitment for this study was also terminated early at 220 the sponsor's request on grounds of lack of efficacy. 254 Sample sizes were estimated on the basis that it would be possible to detect equivalence with 95% 255 confidence, assuming that the underlying rates of efficacy would be 90%. At this level we 256 estimated that a group size of 50 would provide 80% power and 61 would give 90% power.
257 Therefore, a sample size of 60 per group was selected on the basis that it would provide at least 258 85% power, even allowing for possible post-randomisation protocol violations. The estimate also 259 showed that if the underlying rates of the treatments averaged 90%, this sample size would also 260 have a power of 90% to detect a difference of around 18% with 95% confidence. In the first series of in vitro tests, comparing the 3.5% and 10% lotions that we performed prior to 290 clinical studies, we found the 10% mixture was the more effective using an exposure for 60 291 minutes followed by washing with shampoo that resulted in the death of all but one laboratory 292 reared clothing/body lice when reviewed the following day ( Table 1). The effect on louse eggs 293 was also encouraging indicating that a high proportion of eggs could be inhibited from hatching, 294 with a significant (p < 0.01, 95% CI 0.014 to 0.085) increase in the proportion of eggs that showed 295 no sign of development post-treatment compared with the control group, suggesting penetration of 296 the egg structure by the active component of the lotion (Table 2). Manuscript to be reviewed 298 Because it was clear from the first clinical study that inability to kill the louse eggs contributed 299 significantly to the failure to cure we conducted a second in vitro study to investigate whether 300 older eggs were harder to kill. In the original in vitro test louse eggs 24-48 hours old were used 301 whereas on the head there would be eggs at different development stages from newly laid through 302 to hatching. We found that as the eggs aged they became less susceptible to the treatment, with 5 303 day and older eggs requiring exposures longer than 2 hours to stop louse nymphs from emerging 304 and by the seventh day not all eggs were prevented from hatching using an overnight exposure. In 305 addition the proportion of embryos that failed to develop eyespots (the first definitive indication of 306 embryonic development) was lower when exposed to a 2 hour treatment than overnight and by the 307 6 th day there was no difference from the control group because all embryos that would develop had 308 reached the "eyespot" stage between days 5 and 6 ( Table 3).  Table 4). Use of shampoo to remove the product rather than simple water rinsing was 316 potentially the most important factor for reduction of activity (p = 0.0004, 95% CI 0.078 to 0.304), 317 especially against older louse eggs, which showed no significant difference from the control group 318 on overall failure to hatch (Table 4). Manuscript to be reviewed 328 proportion of 10-14 year old participants ( Table 6) and a non-significant trend (p = 0.073) in the 329 single application group for more people to have "Average" thickness hair.  Table 7). The differences were highly significant (p < 0.00001) for both a 2 hour 357 treatment and an overnight treatment suggesting that if we had used the earlier material in the 358 second trial it could have proved more effective. 387 apart from two that were attributed to viral infections (1 head cold, 1 sore throat with fever), one 388 of continued itch due to lice from before treatment, and one in which a child was sprayed in the 389 face with an unrelated detergent product by a sibling; all events were some form of application site 390 reaction to the treatment. All these reactions were of short duration and all resolved spontaneously 391 (