International Efficacy Survey
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1 International Efficacy Survey on Wrinkle Treatment Products for Narhex Australia Pty Ltd By Dr Vyt Garnys Ph.D., A.R.A.C.I., A.I.M.M., Managing Director Cetec Pty Ltd Consulting - Enterprises in Technology
2 International Efficacy Survey 1. Introduction on Wrinkle Treatment Products for Narhex Australia Pty Ltd On contract to Narhex Australia Pty Ltd, Cetec Pty Ltd has conducted and managed an international efficacy trial for the Narhex 3% K-Elastin based wrinkle treatment eye cream following consultation and involvement of the following groups: Australia Cetec Pty Ltd (Inc Vic), Dr Vyt Gamys, Ph.D (UNSW), Managing Director and Project Leader and Dr David Nurse, Consultant Dermatologist, Royal Melbourne Hospital and private practitioner at 71 Mayston Road, East Hawthorn, Victoria. United Kingdom Professor Ronald Marks, Professor of Dermatology, Department of Medicine, University of Wales College of Medicine, Cardiff, UK. and France Dr J-P Cesarini, Senior Research Dermatologist, Groupe De Recherche Sur L'Oncgenese les Ultraviolets et La Pigmentation Cutanee Humaine, Fondation A de Rothschild, Cetec Pty Ltd Page 2 of 12
3 and the above overseas researchers were involved in critically reviewing the study protocol submitted by Cetec, conducting the surveys in their respective countries, submitting the silicone casts to Cetec for measurement, travelling to Australia to review the techniques used by Cetec and preliminary review of the results. In all instances, Professor Marks and Dr Cesarini expressed satisfaction with the survey and the measurement techniques. Dr Cesarini, who has seen the completed results has expressed satisfaction with them and would be willing to co-author a publication. The surveys were based on the design constraints described in the following sections. This report summarises the results of the actual measurement of the surface profile of the skin across wrinkles in the undereye region expressed as standard roughness indices. Page 3 of 12
4 2. Products The following products were trialled: Commercially available Narhex Eye Cream containing 3% K-Elastin and a Moisturiser of the same compositional base as the N arhex Eye Cream but not containing Elastin. These products were supplied by N arhex and were accepted by Cetec as described. The containers were labelled with an identifier number and the eye to be treated. 3. Subjects Middle class males and females, caucasions born in the country of study and having lived in metropolitan areas for most of life, currently living in a similar locality, aged targeted to be between years, normally distributed for age, healthy and having indoor employment with regular outdoors recreation were chosen. No connections with Narhex or Cetec and no previous use of wrinkle treatment products were allowed. To normalise for diet, living conditions, environment, socio-economic and logistic factors, husband/wife groups were chosen as a matter of preference. 4. Controls A split face technique was chosen by which Moisturiser was used on one side of the face and the Narhex Eye Cream Product on the other. Page 4 of 12
5 5. Methodology To validate the main study, a preliminary Protocol Study was carried out in The Protocol or Pilot Study was designed to test all of the procedures under full survey conditions and refine the methodology as required. This study was on 18 subjects all living within the City of Camberwell locality of Melbourne. In all cases but one, the subjects were married couples Physical Measurements All subjects were screened for suitability against survey criteria. Each subject was examined by an independent specialist dermatologist at the conclusion of the study. Twice daily (morning and prior to retiring in the evening) application for four weeks of all treatments to the under and side eye areas were requested with the general instructions supplied to be noted. Prior to the study and weekly for four weeks thereafter, Cetec applied "Silflo" silicone casts to the treated areas. These casts were aligned and mounted on a card for general examination and topographical measurement. Controlled triple light angle still photographs were taken prior to treatment and at the end of the study in a specially prepared local photographic studio and using a set protocol previously established by Narhex. The photographs were designed to show the subjects in a natural, uniformly lit condition. At the end of the study research was conducted on alternate lighting angles and conditions to highlight wrinkle development. The silicone casts of both under-eye and side-eye "crows-feet" areas of the left and right eyes were measured as closely spaced transects at 8,000 points for precise surface shape at a resolution of approximately 1:12,000 with a field of view of 72 square millimetres and an accuracy of 1.0 micrometre. Page 5 of 12
6 After mathematical surface fitting, two levels of surface filtration using Fourier Transform Techniques were used to compute three indices of surface roughness (R), namely, the Relative Mean Square (R RMS), Centre Line Average (R CLA) and Peak to Valley Height (R P-V). Roughness is the standard technical term used to mathematically describe the micro undulation of a surface and has been universaly used by industry for a long time throughout the world. Cetec has conducted preliminary validation of the accuracy of measuring wrinkle imprints on silicone casts by three independent techniques namely: Active optical triangulation, Confocal Microscopy using the laser light and Profilometry using a stylus based instrument. Accurately produced, engineered test pieces were measured by optical microscopy and active optical triangulation to confirm the accuracy of the technique in being able to measure width and depth at micrometre levels. Standard roughness test pieces were used to confirm the ability of the measurement technique and the calculations to produce accurate and precise roughness data. The Cardiff group have independently measured some of the casts using profilometry and image analysis. All product tubes were weighed at the completion of the study Questionnaire The key question being sought in the final study was: That the Narhex Anti-wrinkle Cream visually and actually reduces depth and number of wrinkles after a month of application. To assist in better understanding the context of the results, a questionnaire was completed by Australian subjects and the dermatologist from each subject in all three countries. The above were supplemented by interview at each of the weekly silicone casting visits and dermatological examinations as necessary. Page 6 of 12
7 5.3. Statistical Analysis The statistical analysis of the measurement data arising from the optical scanning of the silicone casts was done through a newly available computer statistics package, SYSTAT from SYSTAT Inc, USA. Computations, plotting and data preparation was with the Lotus and Dbase4 spreadsheets. There were only a few subjects who did not proceed beyond the first week due to eye sensitivity. Narhex funded Dr Garnys of Cetec to travel to Europe to personally present the report of the preliminary study and to demonstrate the silicone casting technique to Professor Marks, Dr Cesarini and to a major international health care company. In all cases the response was extremely positive and required no modification of the methodology. 6. Analysis 6.1. Skin Surface Measurements The surface shape measurement yielded three sets of Surface Roughness measurements at two levels of filtering or sensitivity to yield information on the effects of the treatments on coarse and fine wrinkles. This data was combined with the measured subject data and subject profile data into one computer file. The four, weekly measurements on the side of the face using the product moisturiser (casts 2 to 5) and the one measurement before commencing treatment (cast 1) were considered to be an indication of the skin condition using the Narhex Eye Cream. Subtractions and percentage calculations were made for this side of the face. The wrinkle measurement results were then tabulated and plotted. Computer surface topographical plots were made of the first cast for each subject. This report shows results of Narhex Eye Cream for the three countries surveyed. The report could be easily separated to exclude any country or countries. Page 7 of 12
8 6.2. Results and Discussion At this stage, there is no prior knowledge of the status of skin wrinkles or their effects from products similar to the Narhex products since there does not seem to be much readily available publication of similar studies. It is likely that this and subsequent planned studies will be unique and pioneering. For this reason, no existing population descriptors or variances of facial under eye skin measurement are known by which comparison can be made Treatment Acceptability All subjects found the issued cream to be pleasant to use and without any side effects provided it was applied away from the immediate vicinity of the eye, as per instructions supplied with the products. Several subjects initially complained of slight irritation but this was corrected when they were counselled to apply the products in the vicinity of the cheek bone and below. The dermatological examinations found no evidence of lasting irritation or sensitisation. The silicone casting procedure did not cause any adverse reactions when properly used Photographs The results from the photographs were affected by skin oils and small changes in angle of the head and printing tones and thus could not supply the detail comparable to the surface profile measurement technique. Various changes can be seen in other subjects which could be the subject of a specific report and will be reported on fully in subsequent reports. The low angle lighting techniques developed for the study were of benefit since they highlight the facial wrinkles by oblique lighting rather than hide the features when uniform direct lighting is used. Page 8 of 12
9 7. Australian Subjects Calculations were made of the difference in skin roughness measurements of the side of the face using the active cream for the 75 Australian subjects. The results and typical examples of bar graph plots are shown in Appendix 1. Of the 45 females improvement in their coarse under-eye skin condition e.g. coarse wrinkles and expression lines, was measured, using the relative mean square method, for 67% of the subjects whilst 76% had improvements in their fine skin condition e.g. fine lines, pores and fine wrinkles. The range of both coarse and fine line improvement was between a few percent to over 80% with most of the improved females showing 10 to 30% nett reductions on an individual basis. Of the 30 males, improvement in their coarse under-eye skin condition was measured for 57 to 63% (depending on the method of calculation) of the subjects whilst 80% had improvements in their fine skin condition. The range of both coarse and fine line improvement was between a few percent to over 80% with most of the improved males showing 20 to 40% nett reductions on an individual basis. Over all of the 75 Australian subjects, 47 or 63% had nett improvements in their coarse wrinkles and 58 or 77% in their fine wrinkles after four weeks of treatment. The above reduced results have been statistically tested (t-test and F-test by analysis of variance) to be very certainly (probability 99.9%) a different group of results to the non-reduced. In other words, we are certain that the decreases are real and not subtle variations which may just be achieved by chance. No data was excluded from the analysis. Page 9 of 12
10 Various other changes were seen in the rest of this group but a significant effect in more than half of the subjects is highly significant considering this was a normal cross-section of the population with no prior selection of subjects most likely to be assisted. In this sense this study differs from a hospital related clinical study. The results are clear and discriminate well even small changes in the skin surface. The Pearson correlation statistics shown an excellent concordance of the three methods of calculation of the roughness data for fine and coarse features for males (Correlation coefficients of about 0.9 and 0.8 respectively) and adequate for females (0.9 for fine and 0.6/0.9 for coarse). The general (or population) statistics on the nett percentage changes for all (including those with increases and decreases) females and males showed nett mean and median reductions in wrinkles over the month's trial. For all males reductions of about 3% for coarse wrinkles and 14% for fine wrinkles were observed whilst for females there were nett reductions of 8% for coarse and 7 to 14% for fine wrinkles. The individual results for each subject were plotted and are presented in Appendix. Clearly there are a wide range of effects observed and hence an extremely difficult statistical analysis problem if the within-month time effects are to be studied. Considerable efforts have been made to apply various mathematical regression computations to better describe the time effects on skin wrinkles of the products but without success over the whole survey population. There may be some cluster patterns but this type of detailed analysis is beyond the fmancial resources allocated by Narhex to this project and in any event may not be conclusive despite more effort. Page 10 of 12
11 8. UK Subjects The following is a summary and comments of the UK results as measured by Cetec using the surface imaging technique on the casts taken by Professor Marks' group in Cardiff on Husband/Wife subjects currently living in Wales. The results are tabulated in Appendix 2. Over all of the 26 subjects, (69% to 77%) depending on measurement calculations of females and 62% of males showed nett smoothing of coarse wrinkles over the initial (pre-application) skin condition. For fine wrinkles, over all females, 69% again showed nett smoothing at 4 weeks whilst males were much more affected at 77 to 85% depending on the method of roughness computation. There were not strong age effects. At 4 weeks, nett coarse wrinkle smoothing was achieved in 17, or 65% of all of the 26 subjects and 19, or 73% for fine wrinkles. The range of actual individual smoothing of the active treatment over the placebo at 4 weeks ranged from 4 to 45% for the coarse wrinkles and from 11 to 42% for the fine wrinkles for females. For males coarse wrinkle improvements were seen within the range of 8 to 33% and 0 to 42% in fine wrinkles. The general (or population) statistics on the nett percentage changes for all (including those with increases and decreases) females and males showed nett mean and median reductions in wrinkles over the month's trial. For all males reductions of about 6% for coarse wrinkles and 10% for fine wrinkles were for coarse and 10% for fine wrinkles. The Pearson correlation statistics show an excellent concordance of the three methods of calculation of the roughness data for fme and coarse features for males and females (Correlation coefficients of about 0.9). All of the above comments must be conditional on the small sample of subjects examined and the particularly small cohort of the 37 to 50 year old age group. Page 11 of 12
12 However subject to statistical tests of significance, there seem to be a consistent and sensible trend for the UK results suggesting that, within the ranges quoted above, the active product does have a nett effect in further smoothing both male and female skin over and above the starting condition. 9. French Subjects The French picture is weaker than for the UK or Australia and will have to be examined further. The data is appended. The report is being prepared for further review by the collaborators but since this study was designed to be a measurement study, irrespective and transcending dermatological examination, the above comments are unlikely to be modified. This study has not been designed to answer why the product works - a biochemical and dermatological question. Instead, superior measurement techniques have been used to measure changes which do not require dermatological skills except that dermatological opinion was essential for the design of the study and to warrant that the skin conditions were normal and remained so after the application of the products. Dr Vyt. Garnyo Ph.D, A.R.A.C.I., A.I.M.M. Managing Director Page 12 of 12
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