Pharmacokinetic Characterization of the Novel Pulmonary Delivery Excipient Fumaryl Diketopiperazine

Transcription

1 Journl of Dibetes Science nd Technology Volume 4, Issue 5, September 2010 Dibetes Technology Society ORIGINAL ARTICLES Phrmcokinetic Chrcteriztion of the Novel Pulmonry Delivery Excipient Fumryl Diketopiperzine Elizbeth, Ph.D., 1 Jmes P. Cssidy, M.S., 1 Pmel Hworth, B.S., 2 Dougls Heumn, M.D., 3 Sjoerd vn Mrle, M.D., 4 nd Robert A. Bughmn, Jr., Phrm.D., Ph.D. 1 Abstrct Bckground: Technosphere [Bis-3,6(4-fumrylminobutyl)-2,5-diketopiperzine (FDKP)] microprticles, the integrl component of the Technosphere inhltion system, deliver drugs to the deep lung nd hve been used to dminister insulin nd glucgon-like peptide-1 vi inhltion in clinicl studies. Three studies were conducted to chrcterize FDKP phrmcokinetics, including ssessments in subjects with dibetic nephropthy (DNP), in subjects with chronic liver disese (CLD), nd in helthy subjects. Methods: An open-lbel, nonrndomized, two-period, fixed-sequence crossover bsorption, distribution, metbolism, nd excretion (ADME) study ws conducted in six helthy nonsmoking men who received single intrvenous nd orl doses of [ 14 C]FDKP solution, with seril smpling of blood, urine, feces, nd expired ir. Additionlly, two single-dose, open-lbel, prllel-design studies with 20 mg of inhled FDKP were conducted in (1) 12 dibetic subjects with norml renl function nd 24 DNP subjects nd (2) 12 helthy subjects nd 21 CLD subjects. Results: In the ADME study, >95% of the intrvenous dose nd <3% of the orl dose were recovered in urine, with no evidence of metbolism. No significnt phrmcokinetic differences were observed between helthy subjects nd CLD subjects [geometric men (% coefficient of vrition) re under the curve from time 0 to 480 minutes (AUC ): 26,710 (34.8) nd 31,477 (28.8) ng/ml min, respectively]. Mximum observed drug concentrtion (C mx ) nd AUC were higher in DNP subjects thn in subjects with norml renl function [C mx : (59.4) ng/ml versus (44.3) ng/ml; AUC : 36,869 (47.2) ng/ml min versus 30,474 (31.8) ng/ml min]. None of the differences observed were considered cliniclly significnt. Conclusions: Fumryl diketopiperzine is predominntly clered unchnged by the kidney with essentilly no orl biovilbility. Technosphere is sfe delivery vehicle for medictions dministered vi inhltion. J Dibetes Sci Technol 2010;4(5): Author Affilitions: 1 Experimentl Phrmcology, MnnKind Corportion, Prmus, New Jersey; 2 MnnKind Corportion, Vlenci, Cliforni; 3 McGuire VA Medicl Center, Richmond, Virgini; nd 4 PRA-Interntionl, Institute for Clinicl Phrmcology, Groningen, The Netherlnds Abbrevitions: (ADME) bsorption, distribution, metbolism, nd excretion, (AUC ) re under the curve from time 0 to 480 minutes, (AUC 0 ) re under the curve from time 0 to infinity, (AUC 0 t ) re under the curve from time 0 to t, (BMI) body mss index, (C lst ) lst observed insulin concentrtion, (CLD) chronic liver disese, (C mx ) mximum observed drug concentrtion, (DNP) dibetic nephropthy, (FDKP) Bis-3,6(4-fumrylminobutyl)-2, 5-diketopiperzine, (GFR) glomerulr filtrtion rte, (HPLC) high-performnce liquid chromtogrphy, (LSC) liquid scintilltion counting, (t 1/2 ) terminl elimintion hlf-life, (t lst ) time corresponding to lst observed drug concentrtion, (t mx ) time to mximum observed drug concentrtion Keywords: chronic liver disese, dibetic nephropthy, fumryl diketopiperzine, inhltion, Technosphere Corresponding Author: Elizbeth, Ph.D., MnnKind Corportion, 61 South Prmus Rod, Prmus, NJ 07652; emil ddress epotock@mnnkindcorp.com 1164

2 Introduction Technosphere technology is novel, verstile drug delivery pltform tht enbles pulmonry dministrtion of therpeutic gents. 1 The resulting drug product is composed of the ctive gent dsorbed onto the Technosphere microprticle [Bis 3,6(4 fumrylminobutyl)- 2,5-diketopiperzine (FDKP) nd polysorbte 80], which utilizes proprietry, breth-ctuted inhler for pulmonry delivery of the prticles. Fumryl diketopiperzine is phrmceuticl excipient tht, in solution nd under slightly cidic conditions, forms n rry of microcrystlline pltes. The pltes then self-ssemble into microprticles with lrge surfce re onto which vrious peptides, proteins, or other gents cn be dsorbed (Figure 1). When dried, the resulting dry powder is filled into unit-dose crtridges, which re then used in conjunction with n inhler. Technosphere prticles hve n verge dimeter of 2.5 μm, with 90% of the prticles in the respirble rnge for pulmonry delivery ( μm), s confirmed by erodynmic testing. 1 These chrcteristics enble orl inhltion of therpeutics for wide rnge of disese sttes, 1 providing n lterntive delivery route for gents currently dosed only by injection, nd by mking it possible to bypss heptic first-pss metbolism by bsorption directly into the rteril circultion. 2,3 This technology cn ccommodte therpeutics over wide moleculr size rnge 1 nd hs been used to dminister insulin nd glucgon-like peptide-1 by the pulmonry route in clinicl studies. 3,4 A number of other gents, including oxyntomodulin nd peptide YY, hve been dministered in nonclinicl studies. 5 Therpeutics dministered vi the Technosphere technology demonstrte rpid bsorption, believed to be ssocited with rpid dissolution of the microprticles rther thn ny FDKP-medited bsorption enhncement. In vitro studies using the Clu-3 lung cell line 6 nd in vivo rt studies (unpublished dt) show tht FDKP does not enhnce drug bsorption. Neither in vitro nor preclinicl in vivo investigtions hve found ny evidence of phrmcologicl effect of FDKP. Fumryl diketopiperzine ws evluted in 63 in vitro receptor-binding ssys (neurotrnsmitters, steroids, ion chnnels, secondry messengers, prostglndin, growth fctors/hormones, brin nd gut peptides, nd vrious enzymes) to evlute potentil effects. At concentrtions Figure 1. Scnning electron microgrph of Technosphere prticle. up to 100 μm, FDKP did not inhibit binding of ny substrte to these receptors. The biologicl ctivity of FDKP hs been evluted in comprehensive toxicology studies nd in rt nd dog phrmcology studies, with no observed effects on crdic, centrl nervous system, pulmonry, or renl functions (unpublished dt). To chrcterize FDKP phrmcokinetics, n bsorption, distribution, metbolism, nd excretion (ADME) study ws conducted in helthy humn subjects utilizing 14 C-rdiolbeled FDKP dministered by both intrvenous nd orl routes (study 1). Subsequently, two phse 1 studies were conducted to ssess FDKP phrmcokinetics in: (1) subjects with dibetic nephropthy (DNP) nd dibetic subjects with norml kidney function (study 2) nd (2) subjects with chronic liver disese (CLD) compred with subjects with norml heptic function (study 3). The results of these clinicl studies re reported here. Methods Study Conduct nd Study Subject Consent Ech study protocol nd the ccompnying ptient informtion were pproved by locl ethics committees nd institutionl review bords, nd ech subject gve written informed consent before prticiption. Studies were conducted in complince with the Declrtion of 1165

3 Helsinki nd under the current Interntionl Conference on Hrmoniztion Guidelines for Good Clinicl Prctice. For studies 2 nd 3, subjects were trined by site personnel on use of the MedTone inhler (MnnKind Corportion, Dnbury, CT). Study Subject nd Disese Chrcteristics In ll studies, subjects were nonsmokers willing to use dequte birth control. Exclusion criteri in ll three studies included, but were not limited to, previous exposure to n inhled insulin product, prticiption in nother clinicl study, or presence of cliniclly significnt disese. In study 1, use of ny medictions or herbl remedies within 14 dys before the first dosing (except for prcetmol) ws lso precluded. Complete physicl exmintions were performed t the screening nd follow-up visits. Any cliniclly significnt chnge ws documented. Twelve-led electrocrdiogrms were performed t protocol-specified time points with subjects in supine position. Smoking nd pulmonry sttus (studies 2 nd 3) were reviewed t the screening, tretment, nd follow-up visits. Study 1: Humn ADME Study This phse 1, single-dose, open-lbel, nonrndomized, two-period, fixed-sequence crossover study ws conducted in six helthy volunteers. Prticipnts were men 18 nd 60 yers of ge deemed to be helthy bsed on screening physicl exmintion, medicl history, norml clinicl chemistry nd urinlysis, fsting blood glucose 110 mg/dl, nd body mss index (BMI) 30 kg/m 2 (nonobese). The study consisted of four clinic visits, with study tretment dministrtion of 14 C-lbeled FDKP (see Figure 2 for FDKP structure) on visits 2 nd 3. Subjects underwent seril smpling of blood, urine, feces, nd expired ir t both tretment visits. During visit 2, ll eligibility criteri were confirmed nd subjects received single infusion of 10 ml FDKP (3.7 MBq of [ 14 C]FDKP) intrvenously over 5 minutes t 2 mg/min for totl dose of 10 mg FDKP. Urine nd feces from ll subjects were ssessed dily by liquid scintilltion counting Figure 2. Structure of FDKP. The symbol indictes the position of the 14 C lbel. ( quick counts ). If quick counts did not meet dischrge criteri (<50 dpm/ml urine, <75 dpm/100 mg feces in 24-hour collection) on dy 4, 24-hour urine nd feces specimens were collected until criteri were met. After minimum 2-week wshout period, t visit 3, ech subject received single 100-ml orl dose of FDKP solution (0.2 mg/ml, contining 2.5 MBq of [ 14 C]FDKP). The finl 20-mg dose is n pproximte equivlent of the FDKP mount in the 60-unit Technosphere Insulin dose, cliniclly relevnt dose studied frequently in the Technosphere Insulin development progrm. 7 9 All subjects remined in the clinic until dy 4, nd quick counts were conducted s per visit 2. Visit 4 ws follow-up tht took plce 7 14 dys fter visit 3. Study 2: Dibetic Subjects with Mild or Moderte Dibetic Nephropthy or Norml Renl Function This single-dose, open-lbel, prllel, controlled study of FDKP ws conducted in dibetic subjects with norml renl function nd mild or moderte DNP. The study consisted of three clinic visits: screening, tretment, nd follow-up. Ech subject received one 20-mg (crtridge fill weight) dose of Technosphere inhltion powder t the tretment visit. Blood nd urine smples were collected serilly over 24 hours postdose. Inclusion criteri consisted of men nd women 18 nd 80 yers of ge with type 1 or type 2 dibetes confirmed by medicl records nd BMI 37 kg/m 2. Stging of subjects with DNP ws bsed on recommendtions of the Ntionl Kidney Foundtion Disese Outcomes Qulity Inititive clinicl prctice guidelines. 10 Mild DNP ws defined s glomerulr filtrtion rte (GFR) of ml/min/1.73 m 2 nd microlbuminuri ( 17 mg/g in men, 25 mg/g in women). Moderte DNP ws defined s GFR of ml/min/1.73 m 2 nd microlbuminuri ( 17 mg/g in men, 25 mg/g in women). Norml renl function required GFR 90 ml/min/1.73 m 2 without microlbuminuri (<17 mg/g in men, <25 mg/g in women). Study 3: Subjects with Mild or Moderte Chronic Liver Disese or Norml Liver Function This single-dose, open-lbel, prllel, controlled study ws conducted in nondibetic subjects with norml liver function nd mild or moderte CLD. This study consisted of screening, tretment, nd follow-up visits nd 20 mg of Technosphere inhltion powder (crtridge fill weight) dministered t the tretment visit. Blood nd urine smples were collected serilly over 24 hours postdose. 1166

4 Inclusion criteri consisted of men nd women 18 nd 80 yers of ge with BMI 35 kg/m 2. Chronic liver disese stging ws bsed on the Child Pugh score with recommendtions of the Americn Assocition for the Study of Liver Diseses prctice guidelines. 11 Mild CLD ws defined s Child Pugh clss A (5 6 points), nd moderte CLD ws defined s Child Pugh clss B (7 9 points). Disese history included histopthologicl confirmtion or clinicl imging nd lbortory findings. Subjects without CLD hd n bsence of clinicl evidence of liver disese, negtive heptitis B nd heptitis C serology, nd norml liver function tests. Bionlyticl Methods Seril blood smples were collected into tubes on ice. In study 1, plsm ws nlyzed for rdioctivity by liquid scintilltion counting (LSC). Urine nd feces were collected nd pooled for specified timed intervls. Urine smples were nlyzed for rdioctivity (LSC), wheres the rdioctivity in feces ws mesured vi smple combustion (Pckrd 307 smple oxidizer). Expired ir ws ssessed for 14 CO 2 with hymine hydroxide s trpping solution nd phenolphthlein in ethnol s n indictor. Subjects exhled vi tube plced into vil contining the solution until the solution becme colorless ( 2 minutes), indicting neutrliztion by CO 2. Vils were stored t 4 C until ssy on site for 14 C rdioctivity by LSC. For studies 2 nd 3, smples for the FDKP ssy were centrifuged t 4 C within 1 hour of collection nd frozen until FDKP quntifiction by liquid chromtogrphy with tndem mss spectrometry. Metbolites in Urine nd Feces Urine nd plsm smples with level of rdioctivity >4000 DPM/ml were nlyzed by high-performnce liquid chromtogrphy (HPLC) with rdiochemicl detection (Tble 1), method sensitive enough to seprte the cis nd trns FDKP isomers. The injection volume ws 75 μl to prevent overloding of the column. Phrmcokinetic Prmeters Phrmcokinetic prmeters included mximum plsm drug concentrtion (C mx ), time to C mx (t mx ), nd re under the curve from time 0 to t (AUC 0 t, clculted by the liner-trpezoidl method). Terminl elimintion hlf-life (t 1/2 ) ws clculted s ln(2)/λ z, where λ z is the terminl elimintion rte constnt estimted from log liner regression nlysis of the terminl elimintion phse of the concentrtion time profile, nd re under the curve from time 0 to infinity (AUC 0 ) ws Tble 1. High-Performnce Liquid Chromtogrphy: Conditions for Profiling Serum nd Urine Smples Column Gurd crtridge Mobile phses Mobile phse grdient Flow rte Lun phenyl-hexyl 3-µm pcking, mm ID Phenyl, 3 µm, 4 2 mm ID A 1000:1 (v/v) wter:tfa B 900:100:1 (v/v) MeOH:THF:TFA Time (min) %A %B ml/min ID, inside dimeter; TFA, trifluorocetic cid; MeOH, methnol; THF, tetrhydrofurn. clculted s AUC 0 tlst + C lst /λ z, where C lst is the lst observed drug concentrtion nd t lst is the time corresponding to C lst. In study 1, dditionl prmeters included mss blnce, clculted s the sum of the mount of 14 C rdioctivity excreted in urine, feces, nd expired ir divided by the mount of rdioctivity dosed. Phrmcokinetic nlyses were performed using WinNonlin Professionl, version or higher (Phrsight, Inc., Mountin View, CA), nd summry sttistics nd sttisticl nlyses were performed using SAS, version 8.2 or higher (SAS Institute, Inc., Cry, NC). Results Study 1: Humn ADME Four of the six enrolled subjects completed the study. Two subjects were withdrwn fter the intrvenous dose but before orl dosing (prior to strting visit 3) due to elevted levels of urinry cotinine (>100 ng/ml), in violtion of inclusion criteri (i.e., nonsmoking sttus). Hence, the phrmcokinetic nlysis included six helthy subjects following intrvenous dosing nd four subjects following orl dosing (Tble 2). Following intrvenous dministrtion, >90% of totl rdioctivity elimintion occurred within 8 hours of dosing (Tble 3, Figure 3A). An verge of 97.2% of totl rdioctivity ws recovered in the urine nd 1.65% ws recovered in the feces. Anlyses of urine nd dosing mterils by HPLC demonstrted tht ll rdioctivity peks detected in urine were present in the dosing mteril in the sme proportions. There ws no evidence 1167

5 of metbolism following intrvenous dosing. The rdiochromtogrm from representtive urine smple (Figure 4) shows the cis nd trns FDKP isomers, with no other rdioctivity peks evident. Following orl dosing, >90% of rdiolbel ws eliminted within 48 hours postdose, with n verge of 2.45% of totl rdioctivity recovered in the urine nd pproximtely 95.1% recovered in the feces (Tble 3, Figure 3B). A negligible frction of the dose ws recovered in expired ir (0.03% of the dose). Phrmcokinetic prmeters nd 14 C rdioctivity of FDKP in plsm re summrized in Tble 4 nd Figures 5 nd 6. chrcteristics mong the three renl function groups, with the exception of rce; the mjority of subjects in the mild DNP group were of Hispnic origin. Given the smll number of subjects, the determintion of rcil differences on FDKP phrmcokinetics ws not possible; Study 2: Dibetic Subjects with Dibetic Nephropthy nd Norml Renl Function All 36 enrolled subjects completed the study (Tble 5). No significnt differences were noted in subject Tble 2. Men (SD ) Demogrphics nd Bseline Chrcteristics of Subjects in Study 1 Intrvenous dose popultion (n = 6) Orl dose popultion (n = 4) Age, yers (SD) 29.8 (14.9) 34.0 (17.2) Weight, kg (SD) 77.7 (10.5) 77.2 (13.6) Height, cm (SD) (12.7) (15.2) BMI, kg/m 2 (SD) 23.9 (1.2) 24.5 (1.0) Rce, n (%) Cucsin 5 (83.3) 3 (75.0) Asin 1 (16.7) 1 (25.0) Stndrd devition. Tble 3. Summry of 14 C Rdioctivity Phrmcokinetic Prmeters in Urine, Feces, nd Expired Air: Study 1 Prmeter 10-mg IV dose (n = 6) Urine Feces Expired ir Urine 20-mg orl dose (n = 4 ) Feces Expired ir Figure 3. Men (±SE) cumultive excretion of totl rdioctivity in (A) urine nd (B) feces following 10-mg intrvenous nd 20-mg orl FDKP dose. Ae, b µg ,030 5 Fe, % Mss blnce, % (SD) 98.9 (2.1) 97.6 (1.5) Two subjects did not receive orl dose of study drug becuse they were withdrwn from the study. b Ae, totl mount of 14 C rdioctivity excreted in urine or feces, or estimted totl mount of 14 C rdioctivity excreted in expired ir; Fe, percentge of dose of 14 C rdioctivity excreted in urine, feces, or expired ir; IV, intrvenous; SD, stndrd devition. Figure 4. Representtive rdiochromtogrm of 0- to 4-hour urine smple following intrvenous dministrtion of 14 C-lbeled FDKP showing the cis (first pek) nd trns (second pek) isomers of FDKP. 1168

6 however, there is no expecttion tht FDKP clernce is ffected by rce, s there were no obvious differences in FDKP prmeters in the four rcil groups within ech renl function ctegory. Fumryl diketopiperzine exposure [FDKP re under the curve from time 0 to 480 minutes (AUC )] ws pproximtely 18 nd 25% higher in subjects with mild nd moderte DNP, respectively, compred with subjects with norml renl function (Tble 6, Figure 7). Fumryl diketopiperzine t 1/2 ws similr in subjects with norml renl function nd those with mild DNP nd ws 80 minutes longer in subjects with moderte DNP. This difference in t 1/2 resulted in more noticeble difference in exposure over longer smpling period in subjects with moderte DNP, with n pproximte 52% greter exposure when AUC 0 ws compred with subjects with norml renl function. The longer t mx observed in subjects with moderte DNP (30 minutes versus 12 minutes in subjects with norml renl function nd 15 minutes in those with mild DNP) is most likely lso ttributble to the observed longer t 1/2. Over 24 hours, urinry elimintion ws lmost complete, with pproximtely 20% of the FDKP dose excreted in the urine of ll subjects, with slower ccumultion in Tble 4. Summry of Phrmcokinetic Prmeters nd 14 C Rdioctivity of FDKP in Plsm: Study 1 Prmeter Intrvenous dose (10 mg) n = 6 Orl dose (20 mg) n = 4 AUC 0, ng/ml min 75,240 (5640) NC AUC 0 tlst, ng/ml min 74,791 (5640) 5,587 (3028) C mx, ng/ml (130.5) 8.5 (1.6) t mx, h NC 6 (1, 16) t 1/2, min 150 (24) NC F orl NA (0.0174) Prmeters re presented s men (stndrd devition) except for t mx, which is presented s medin (min, mx). AUC 0 tlst, re under the plsm concentrtion time curve from time 0 to t, where t is time of lst quntifible concentrtion; F orl, biovilbility following orl dose; NA, not pplicble; NC, not clculted. Figure 6. Men (±SE) totl rdioctivity in plsm following 20-mg FDKP orl dose (liner scle). Figure 5. Men (±SE) totl rdioctivity in plsm following 10-mg FDKP intrvenous dose. Figure 7. Men (±SE) FDKP concentrtion time profiles in subjects with dibetes nd dibetic nephropthy nd norml kidney function (study 2). 1169

7 subjects with moderte DNP compred with the other subjects (Figure 8). Study 3: Subjects with Chronic Liver Disese nd Norml Liver Function Study enrollment ws stopped t 33 subjects before the trget enrollment of 36 subjects ws chieved (Tble 5) due to the prolonged enrollment period necessry to identify CLD subjects. No significnt differences in subject chrcteristics were noted mong the three heptic function groups. Fumryl diketopiperzine exposure (AUC ) ws higher in subjects with CLD thn in those with norml liver function; however, this difference ws slight (Tble 7, Figure 9). No differences in FDKP C mx nd t mx nd t 1/2 were observed. Over 24 hours, urinry elimintion ws lmost complete with, on verge, <25% of the FDKP dose excreted in urine for ll subjects (Figure 10). Differences in men cumultive excretion mounts were skewed for the mild CLD group by one subject with high excretion vlues; this pprent difference ws negted when tht subject ws excluded. Tble 5. Men (SD ) Demogrphics nd Bseline Chrcteristics of Subjects in Studies 2 nd 3 Dibetic subjects without DNP (n = 12) Mild DNP (n = 15) Moderte DNP (n = 9) Norml (n = 12) Mild CLD (n = 15) Moderte CLD (n = 6) Age, yers (SD) 58 (7) 63 (9) 52 (13) 51 (7) 58 (8) 58 (3) BMI, kg/m 2 (SD) 31 (4) 29 (4) 33 (3) 28 (4) 30 (4) 28 (2) Sex Mle (%) 7 (58) 11 (73) 6 (67) Femle (%) 5 (42) 4 (27) 3 (33) Weight, kg (SD) 88 (16) 80 (13) 97 (11) 92 (18) 95 (15) 88 (11) Height, cm (SD) 167 (11) 165 (7) 170 (8) 181 (8) 178 (10) 176 (7) Rce, n Cucsin Blck Hispnic Other Stndrd devition. Tble 6. Geometric Men (% Coefficient of Vrition) FDKP Phrmcokinetic Prmeters in Dibetic Nephropthy: Study 2 Prmeter Dibetic without DNP (n = 12) Dibetic nephropthy Mild (n = 15) Moderte (n = 9) Mild nd moderte (n = 24) AUC 0 480, ng/ml min 30,474 (32) 36,090 (43) 38,206 (54) 36,869 (47) AUC 0, ng/ml min 37,137 (29) 44,168 (43) 56,393 (58) 48,406 (53) C mx, ng/ml (44) (55) (66) (59) t mx, min 12 (3 45) 15 (3 76) 30 (9 153) 16 (3 153) t 1/2, min (52) (46) (65) (64) Ae , mg b 3.7 (1.2) 3.7 (1.7) 2.8 (1.7) 3.3 (1.7) t mx is presented s medin (rnge). b Ae is presented s rithmetic men (stndrd devition). 1170

8 Sfety: All Studies Administrtion of FDKP ws generlly well tolerted in ll studies, nd tretment-relted dverse events were mild with no ssocited discontinutions. No severe dverse events or deths occurred during these studies. Figure 8. Men (±SE) cumultive mount of FDKP excreted in urine in subjects with dibetes nd dibetic nephropthy nd norml kidney function (study 2). In study 1, ll tretment-relted dverse events were mild in intensity except for one instnce of iridocyclitis of moderte intensity, not relted to the study drug. Cough, the most common tretment-emergent dverse event, ws reported by 19 (52.8%) subjects in study 2 nd by 17 (51.5%) subjects in study 3. In both studies, ll coughs were nonproductive, mild in intensity, nd occurred within 10 minutes of inhltion nd s single or intermittent occurrence. Figure 9. Men (±SE) FDKP concentrtion time profiles in subjects with norml nd impired heptic function (study 3). Figure 10. Men (±SE) cumultive mount of FDKP excreted in urine (Ae) in subjects with impired nd norml heptic function (study 3). Tble 7. Geometric Men (% Coefficient of Vrition) FDKP Phrmcokinetic Prmeters in Chronic Liver Disese: Study 3 Prmeter Norml (n = 12) Chronic liver disese Mild (n = 15) Moderte (n = 6) Mild nd moderte (n = 21) AUC 0 480, ng/ml min 26,710 (35) 31,001 (26) 32,700 (36) 31,477 (29) AUC 0, ng/ml min 32,320 (33) 36,505 (26) 40,085 (41) 37,494 (32) C mx, ng/ml (49) (35) (42) (36) t mx, min 7.5 (3 20) 6.0 (3 60) 6.0 (3 45) 6.0 (3 60) t 1/2, min 190 (27) 173 (30) 198 (45) 180 (36) Ae , mg b 3.2 (1.1) 4.5 (2.7) 3.4 (1.2) 4.1 (2.4) t mx is presented s medin. b Ae is presented s rithmetic men (stndrd devition). 1171

9 Discussion The Technosphere inhltion system cn be used to deliver vrious therpeutic gents vi the pulmonry route. Fumryl diketopiperzine, the primry component of Technosphere prticles, is novel, phrmcologiclly inert compound tht is bsorbed rpidly into the systemic circultion. 1 Becuse dministrtion of Technosphere prticles results in systemic FDKP exposure, the phrmcokinetic properties of this compound were chrcterized. Inhled drugs my enter the systemic circultion vi two routes: (1) frction of the dose enters through the lung (directly into the centrl comprtment) nd (2) frction of the dose is swllowed. Therefore, FDKP metbolism ws ssessed following both intrvenous nd orl routes of dministrtion to fully chrcterize its clernce fter inhltion. Following intrvenous dministrtion, lmost complete renl clernce of unchnged FDKP ws observed. In the sme study, FDKP demonstrted essentilly no orl biovilbility; hence, the route of elimintion of drug following pulmonry dosing is expected to be predominntly renl, s ny swllowed drug is not expected to be systemiclly biovilble t cliniclly relevnt doses. In studies 2 nd 3, the cumultive mount of FDKP excreted in the urine fter inhltion ws mesured following dosing. Approximtely 20 23% of the dose ws excreted t 24 hours in ll groups studied. Becuse the ADME study showed lmost complete urinry FDKP elimintion by 8 hours following intrvenous dosing nd becuse FDKP is bsorbed rpidly from the lung, this quntity ws ssumed to reflect FDKP biovilbility following inhltion. Absolute biovilbility ws lso clculted from men FDKP AUC 0 in studies 2 nd 3 nd FDKP AUC 0 following intrvenous dministrtion in study 1 (24.7 nd 21.5%, respectively), confirming the vlue determined from urinry excretion dt. Fumryl diketopiperzine biovilbility less thn unity cn be ttributed to the fct tht some of the drug product remins in the inhler nd some is swllowed during inhltion. Differences observed in FDKP rte nd extent of exposure between subjects with DNP nd dibetic subjects without renl disese re consistent with the predominnt renl clernce of the compound. However, differences observed were not considered cliniclly significnt, s FDKP exposure is expected to remin within the sfety mrgins with repeted dosing, bsed on the NOAEL (no observed dverse effect level) estblished in nonclinicl studies (unpublished dt) nd n ccumultion rtio (R = AUC 0 /AUC 0 480, bsed on geometric mens of AUCs) of 1.22 (norml renl function) nd 1.31 (mild nd moderte DNP) with three-times-per-dy dosing. When dt from subjects with moderte DNP were exmined, n pproximte 50% increse in FDKP exposure (bsed on AUC 0 ) ws observed compred with dibetic subjects without renl impirment. This finding ws without expected clinicl implictions due to the lck of FDKP phrmcologicl ctivity following extensive in vitro, preclinicl, nd toxicologicl studies. Interestingly, lthough inhled FDKP is clered predominntly by the kidneys, study 3 showed n increse in FDKP exposure in subjects with mild nd moderte CLD compred with helthy subjects. It is well estblished tht decresed renl function occurs in subjects with heptic disese 12,13 nd tht reduced renl excretion of number of drugs clered unchnged in urine (i.e., furosemide, bumetnide, cimetidine, nd rnitidine) in subjects with heptic impirment hs been reported. 13 Therefore, it is likely tht the slightly elevted FDKP exposure in subjects with CLD is linked to impired renl clernce ssocited with reduced heptic function. However, this smll, reltive increse in FDKP exposure ws not considered cliniclly significnt. In studies 2 nd 3, phrmcokinetic dt following inhltion were vilble from subjects without CLD or DNP. Most prmeters were comprble between these two groups, with somewht higher exposure (C mx ) nd lter t mx in study 2. This difference is most likely ttributed to the difference in popultions nd the potentil effect of the dibetic stte on renl clernce of FDKP in study 2. In studies 2 nd 3, cough ws the common tretmentemergent dverse event. The cough ws mild nd ws observed s single or intermittent occurrence. This finding ws not surprising, s cough is often ssocited with the inhltion of dry powder. 14 Conclusions In n ADME study conducted in helthy mle volunteers, >95% of the intrvenous FDKP dose nd <3% of the orl FDKP dose were recovered unchnged in urine. Orl biovilbility ws negligible, nd no evidence of 1172

10 metbolism ws observed. FDKP ws well tolerted in ll popultions studied. The increse in overll exposure due to mild or modertely impired renl function ws not cliniclly significnt. Fumryl diketopiperzine (Technosphere) is sfe delivery vehicle for medictions dministered vi inhltion. Funding: Studies discussed in this rticle were funded by MnnKind Corportion, Vlenci, CA. Acknowledgments: We cknowledge the following principl investigtors: Robert J. Schwb, M.D., Quli Clinicl Reserch, Omh, NE; Sherwyn L. Schwrtz, M.D., Cetero Reserch, Inc., Sn Antonio, TX; Ernesto Fuentes, M.D., Elite Reserch Institute, Mimi, FL; Krishn K. Pudi, M.D., Upstte Phrmceuticl Reserch, Greenville, SC; nd Richrd A. Preston, M.D., M.B.A., Division of Clinicl Phrmcology, University of Mimi, West Mimi, FL. 8. Rosenstock J, Lorber DL, Gnudi L, Howrd CP, Bilheimer DW, Chng PC, Petrucci RE, Boss AH, Richrdson PC. Prndil inhled insulin plus bsl insulin glrgine versus twice dily bisprt insulin for type 2 dibetes: multicentre rndomised tril. Lncet. 2010;375(9753): Peyrot M, Rubin RR. Ptient reported outcomes in dults with type 2 dibetes using meltime insulin monomer humn (rdna origin) inhltion powder (Technosphere insulin inhltion powder) or metformin + secretgogue or both [bstrct]. Dibetologi. 2009;52(Suppl 1):S396. Abstrct Ntionl Kidney Foundtion. Prt 5. Evlution of lbortory mesurements for clinicl ssessment of kidney disese. Am J Kidney Dis. 2002;39(2 Suppl 1):S Murry KF, Crithers RL Jr; AASLD. AASLD prctice guidelines: evlution of the ptient for liver trnsplnttion. Heptology. 2005;41(6): Morgn DJ, McLen AJ. Clinicl phrmcokinetic nd phrmcodynmic considertions in ptients with liver disese. An updte. Clin Phrmcokinet. 1995;29(5): Verbeeck RK. Phrmcokinetics nd dosge djustment in ptients with heptic dysfunction. Eur J Clin Phrmcol. 2008;64(12): Siekmeier R, Scheuch G. Inhled insulin--does it become relity? J Physiol Phrmcol. 2008;59 (Suppl 6): Disclosure: Dr., Mr. Cssidy, Ms. Hworth, nd Dr. Bughmn re employed by MnnKind Corportion. Drs. Heumn nd Vn Mrle were principl investigtors. Dr. Heumn hs received reserch grnt support from MnnKind Corportion. References: 1. Leone-By A, Grnt M. Technosphere/insulin: mimicking endogenous insulin relese. 2nd ed. In: Rthbone MJ, Hdgrft M, Roberts M, Lne M, editors. Modified relese drug delivery technology. Vol 2. New York: Inform Helthcre USA, Inc.; p Rve K, E, Boss AH, Mrino M, Costello D, Chen R. Phrmcokinetics nd liner exposure of AFRESA compred with the subcutneous injection of regulr humn insulin. Dibetes Obes Metb. 2009;11(7): Mrino MT, Costello D, Bughmn R, Boss A, Cssidy J, Dmico C, vn Mrle S, vn Vliet A, Richrdson PC. Phrmcokinetics nd phrmcodynmics of inhled GLP-1 (MKC253): proof-of-concept studies in helthy norml volunteers nd in ptients with type 2 dibetes. Clin Phrmcol Ther. 2010;88(2): Boss AH, Yu W, Ellermn K. Prndil insulin: is inhled enough? Drug Dev Res. 2008;69: Grnt ML, Greene S, Stowell GW, Dniels S, Smithson A, Villnuev S, Leone-By A. Mimicking endogenous peptide secretion by inhltion [poster]. Presented t: 21st Americn Peptide Symposium; June 7-12, 2009; Bloomington, IN. 6. Angelo R, Rousseu K, Grnt M, Leone-By A, Richrdson P. Technosphere insulin: defining the role of Technosphere prticles t the cellulr level. J Dibetes Sci Technol. 2009;3(3): Phillips M, Amin N, Boss AH, Richrdson P. Pulmonry functions (over 2 yers) in dibetic subjects treted with Technosphere insulin or usul ntidibetic tretment [bstrct]. Dibetologi. 2009;52(Suppl 1):S361. Abstrct