DRUG PERMEATION THROUGH SKIN Mario Grassi Department of Chemical Engineering (DICAMP) UINVERSITY OF TRIESTE

1 - SKIN STRUCTURE non-living layer of keratin-filled cells surrounded by a lipid-rich extracellular matrix viable tissue devoid of blood vessels. contains capillary loops Adapted from M. Prausnitz et al. NATURE REVIEWS, DRUG DISCOVERY, 3, (2004),115

2 – PERMEATION ROUTES Sweat pore 1 2 3 Hair Dermis Hair follicle Sebaceous gland Epidermis Stratum Corneum Sweat gland Sweat duct Adapted from B. W. Barry, Adv. Drug. Del. Review, 54, (2002), S31-40

3 – EXPERIMENTAL SET UP 1

DRUG CONCENTRATION INCREASE 4 – MODELLING 1 Cd = Cd0 M = M0 Drug DRUG DISSOLUTION Css = 0 Cde = 0 DRUG DIFFUSION Css = 0 DRUG CONCENTRATION INCREASE Cr = 0

FICK LAW S. Corneum Dermis + Epidermis stagnant layer

SOLID SURFACE VARIATION: MONODISPERSED PARTICLES SYSTEM SOLID DRUG Mass balance Particles initial surface area

SAMPLING TECNIQUE receiver drug concentration just before sampling. receiver drug concentration just after sampling.

CASE STUDY: ACYCLOVIR PERMEATION THROUGH RAT SKIN - Recordati , Milano - ANTIVIRAL, WHITE CRYSTALLINE POWDER (R = 5.7 mm) - SURFACE AREA = 3370 cm2/g (mercury porosimeter) - U.V. PEAK ABSORBANCE 251 nm - SOLUBILITY in PBS (pH = 7.4; 37°C) Cs = 2.62 mg/cm3 - ACYCLOVIR DIFF. COEFF. (PBS, 37°C) Dss = 7.8*10-6 cm2/s (IDR) - ACYCLOVIR DISS. CONST. (PBS, 37°C) KD = 5*10-5 cm/s RAT SKIN: Male hairless rats (Rnu eutimic, Charles River, MI, Italy) 5–7 weeks old Full-thickness skin removed from abdomen by incision of the outermost layer with a surgical bisturi. Stratum corneum was separated from the dermis–epidermis by placing the full-thickness skin (dermis-side down) on a filter paper saturated with a 1% trypsin) solution at the temperature of 37 ◦C for 4 h.

STAGNANT LAYER THICKNESS: hss Sh = f Rem Scn Sh = d/hss Sc = h/(r D) Re = wrd2/h r = fluid density D = drug diff. coeff. d = stirrer diameter h = fluid viscosity. Dissolution T = 25°C w = 388 rpm w = 505 rpm w = 605 rpm hss1 ==> Sh1 hss2 ==> Sh2 hss3 ==> Sh3 T = 37°C hss4 ==> Sh4 hss5 ==> Sh5 hss5 ==> Sh6

f = 3.2*10-6 m = 1.2 n = 0.95 Sc = 1147 Re = 5737 hss = 0.011 cm

PARTITION COEFFICIENTS: Kp1,2,3,4 The full skin/Acyclovir solution and the one-layer skin/Acyclovir solution partition coefficients Kskin and Kde are determined by immersion of both fragments in the Acyclovir solution (PBS pH 7.4) for 4 h at 37°C Kskin =0.547 Kde =0.95 Kp1 =0.5 (literature) Kp3 = Kp2*Kp1 Kp2 = Kskin*(1+G)-Kp1/(G*Kp1) G = hsd/hsc= 11.7 Kp1 = 0.5 Kp2 = 1.1 Kp4 = 1 Kp3 = 0.55

5 – RESULTS DAV = (1.7 ± 0.6)*10-6 cm2/s FIVE WEEKS OLD RATS: ONE LAYER (DERMIS-EPIDERMIS) Data (symbols) not corrected for dilution D = 2*10-6 cm2/s Lines: model best fitting D = 2*10-6 cm2/s D = 1*10-6 cm2/s DAV = (1.7 ± 0.6)*10-6 cm2/s

SEVEN WEEKS OLD RATS: ONE LAYER (DERMIS-EPIDERMIS) Data (symbols) not corrected for dilution D = 9.5*10-7 cm2/s Lines: model best fitting D = 6.5*10-7 cm2/s DAV = (8 ± 2)*10-7 cm2/s

DAV = (1.33 ± 1)*10-9 cm2/s FIVE WEEKS OLD RATS: FULL SKIN D = 2.5*10-9 cm2/s Data (symbols) not corrected for dilution Lines: model best fitting D = 1*10-9 cm2/s D = *10-10 cm2/s DAV = (1.33 ± 1)*10-9 cm2/s

SEVEN WEEKS OLD RAT: FULL SKIN D = 6.5*10-10 cm2/s Data (symbols) not corrected for dilution Lines: model best fitting

COMMENTS 1 Regardless animal age, stratum corneum represents the main barrier to drug permeation (Dde  1000 Dsc) 2 Skin permeability seems to decrease with age 3 A considerable inter-animal variability is observed 4 Model simulations reveal that pseudo steady state conditions are met after 2 hours for young animals (one layer skin), while they are met after 6 hours in the remaining cases

6 – COMPARISON THIS APPROACH / COMMON APPROACH 2 DATA CORRECTION FOR DILUTION = corrected drug concentration after “n” samplings = experimental drug concentration after “n” samplings = experimental drug concentration after “i” samplings = sampling volume = receiver volume

Dm = drug diff coeff through membrane h = membrane thickness ASYMPTOTIC FICK EQUATION SOLUTION 1) Pseudo steady state conditions 2) Cd = C0; Cr  0 (sink conditions) 3) Stagnant layer is neglected Dm = drug diff coeff through membrane h = membrane thickness S = membrane area Kp = partition coefficient

ASSUMING “m” AND “q” AS INDEPENDENT FITTING PARMETERS: IS FITTED ON EXPERIMENTAL DATA TO GET Dm

FROM DIFFUSION COEFFICIENT TO RESISTANCE TRADITIONAL APPROACH THIS APPROACH

Dermis + epidermis (one layer skin) Experiment number identifier N Animal age (weeks) F-value Ro (s/cm) Rm (s/cm) Dermis + epidermis (one layer skin) 1 5 732 (5.9  0.014)*104 (6.1  0.016)*104 2 261 (4.9  0.001)*104 (5.5  0.012)*104 3 136 (12.6  0.06)*104 (12.1  0.059)*104 4 7 864 (21.1  1.62)*104 (19.2  1.35)*104 5906 (15.6  0.814)*104 (15.0  0.765)*104 Full skin 6 595 (4.83  0.75)*106 (2.49  0.175)*106 45 (26.4  24.7)*106 (10.7  2.96)*106 8 99965 (13.7  6.7)*106 (7.68  0.014)*106 9 57 (12.7  5.2)*106 (5.76  0.13)*106 This approach Traditional approach

7 – REFERENCES 1) N. Coceani, I. Colombo, M. Grassi, Int. J. Pharm. 254 (2003) 197 –210. 2) Chien, Y.W. (Ed.), 1987. Transdermal Controlled Systemic Medications. Marcell Dekker, Inc., New York, Basel (Chapter 2).