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Purpose: Preclinical and clinical studies have demonstrated that inhibition of prenylation can radiosensitize cell lines with activation of Ras and produce clinical response in patients with cancer. The aim of this study was to determine the maximally tolerated dose of the dual farnesyltransferase and geranylgeranyltransferase I inhibitor L-778,123 in combination with radiotherapy for patients with locally advanced pancreatic cancer. Experimental Design: L-778,123 was given by continuous intravenous infusion with concomitant radiotherapy to 59.4 Gy in standard fractions. Two L-778,123 dose levels were tested: 280 mg/m2/day over weeks 1, 2, 4, and 5 for dose level 1; and 560 mg/m2/day over weeks 1, 2, 4, 5, and 7 for dose level 2. Results: There were no dose-limiting toxicities observed in the eight patients treated on dose level 1. Two of the four patients on dose level 2 experienced dose-limiting toxicities consisting of grade 3 diarrhea in one case and grade 3 gastrointestinal hemorrhage associated with grade 3 thrombocytopenia and neutropenia in the other case. Other common toxicities were mild neutropenia, dehydration, hyperglycemia, and nausea/vomiting. One patient on dose level 1 showed a partial response of 6 months in duration. Both reversible inhibition of HDJ2 farnesylation and radiosensitization of a study patient-derived cell line were demonstrated in the presence of L-778,123. K-RAS mutations were found in three of the four patients evaluated. Conclusions: The combination of L-778,123 and radiotherapy at dose level 1 showed acceptable toxicity in patients with locally advanced pancreatic cancer. Radiosensitization of a patient-derived pancreatic cancer cell line was observed.

Caspofungin was the first in a new class of antifungal agents (echinocandins) indicated for the treatment of primary and refractory fungal infections. Higher doses of caspofungin may provide another option for patients who have failed caspofungin or other antifungal therapy. This study evaluated the safety, tolerability, and pharmacokinetics of single 150- and 210-mg doses of caspofungin in 16 healthy participants and 100 mg/d for 21 days in 20 healthy participants. Other than infusion site reactions and 1 reversible elevation in alanine aminotransferase (≥2× and

Nasal congestion in allergic rhinitis results from tissue edema and vasodilatation in the nasal mucosa. Of the mediators released by mast cells in response to allergens, prostaglandin (PG) D(2) is regarded as the most potent inducer of nasal congestion. Intranasal administration of PGD(2) reproduces the nasal blockade experienced by patients with seasonal allergic rhinitis (SAR) via its action on the PGD(2) (DP) receptor to induce nasal vasodilatation. Intranasal challenge with PGD(2) can be a useful tool for evaluating DP-receptor antagonists.The main purpose of this study was to examine the ability of MK-0524, a DP receptor antagonist in development for the treatment of SAR, to block PGD(2) induced nasal congestion in healthy volunteers.To this end, a double-blind, placebo-controlled, randomized, 3-period study was performed in 15 healthy subjects. During each period, subjects received MK-0524 25 mg, MK-0524 100 mg or placebo qd for 3 days. Twenty-four hours following the last dose, nasal provocations with PGD(2) were performed to determine the PD(75), which is the intranasal dose of PGD(2) that provokes a 75% increase in baseline total nasal airway resistance as performed by active anterior rhinomanometry.Following treatment with MK-0524, the PD(75) (mean+/-SD) was significantly shifted from 15.8 +/- 18.3 mug/nostril during the placebo period to more than 512 mug/nostril both following the 25- and 100-mg (maximum challenge dose tested) dose regimen.Whether this45 fold increase in PD(75) will induce a clinically meaningful effect of MK-0524 will require clinical study in participants with SAR.

Background Aprepitant is a neurokinin1 receptor antagonist that, in combination with a corticosteroid and a 5-hydroxytryptamine3 receptor antagonist, has been shown to be very effective in the prevention of chemotherapy-induced nausea and vomiting. At doses used for the management of chemotherapy-induced nausea and vomiting, aprepitant is a moderate inhibitor of cytochrome P4503A4 and may be used in conjunction with corticosteroids such as dexamethasone and methylprednisolone, which are substrates of cytochrome P4503A4. The effects of aprepitant on the these 2 corticosteroids were evaluated. Methods Study 1 was an open-label, randomized, incomplete-block, 3-period crossover study with 20 subjects. Treatment A consisted of a standard oral dexamethasone regimen for chemotherapy-induced nausea and vomiting (20 mg dexamethasone on day 1, 8 mg dexamethasone on days 2 to 5). Treatment B was used to examine the effects of oral aprepitant (125 mg aprepitant on day 1, 80 mg aprepitant on days 2 to 5) on the standard dexamethasone regimen. Treatment C was used to examine the effects of aprepitant on a modified dexamethasone regimen (12 mg dexamethasone on day 1, 4 mg dexamethasone on days 2 to 5). All subjects also received 32 mg ondansetron intravenously on day 1 only. Study 2 was a double-blind, randomized, placebo-controlled, 2-period crossover study with 10 subjects. Subjects in one group received a regimen consisting of 125 mg methylprednisolone intravenously on day 1 and 40 mg methylprednisolone orally on days 2 to 3. Subjects in the other group received oral aprepitant (125 mg aprepitant on day 1, 80 mg aprepitant on days 2 to 3) in addition to the methylprednisolone regimen. Results In study 1, the area under the concentration-time curve from 0 to 24 hours (AUC0-24) of oral dexamethasone on days 1 and 5 after the standard dexamethasone plus ondansetron regimen (treatment A) was increased 2.2-fold (P < .010) with coadministration of aprepitant (treatment B). Coadministration of aprepitant with the modified dexamethasone plus ondansetron regimen (treatment C) resulted in an AUC0-24 for dexamethasone similar to that observed after the standard dexamethasone plus ondansetron regimen (treatment A). In study 2, aprepitant increased the AUC0-24 of intravenous methylprednisolone 1.3-fold on day 1 (P < .010) and increased the AUC0-24 of oral methylprednisolone 2.5-fold on day 3 (P < .010). Conclusions Coadministration of aprepitant with dexamethasone or methylprednisolone resulted in increased plasma concentrations of the corticosteroids. These findings suggest that the dose of these corticosteroids should be adjusted when given with aprepitant. Clinical Pharmacology & Therapeutics (2003) 74, 17–24; doi: 10.1016/S0009-9236(03)00066-3

The penetration of 1 g of intravenous ertapenem once daily for 3 days in suction-induced skin blisters was evaluated. Ten forearm blisters were formed ( n = 12) 12 h prior to the last dose. Concentrations of ertapenem in blister fluid exceeded 4 μg/ml (the MIC at which 90% of the isolates tested are eliminated) for the entire dosing interval. The area under the concentration-time curve for 0 to 24 h ratio of blister fluid to plasma was 61% (90% confidence interval, 56, 65%) suggesting good blister penetration.

Study Objectives To evaluate the safety and potential pharmacokinetic interaction between indinavir and clarithromycin. Study Methods In a randomized, three-period, crossover fashion, 12 healthy adults received the following for 1 week: 800 mg oral indinavir sulfate every 8 hours with placebo, 500 mg oral clarithromycin every 12 hours with placebo, and indinavir sulfate with clarithromycin. Plasma indinavir, clarithromycin, and 14-hydroxyclarithromycin concentrations were determined after the last dose in each treatment period. Results Administration of indinavir sulfate with clarithromycin caused a statistically significant increase in four pharmacokinetic parameters: a 58% increase in plasma indinavir concentrations at 8 hours (P = .029), a 47% increase in values for clarithromycin area under the plasma concentration versus time curve from time zero to the last measured concentration [AUC(0-12h); P = .0002], and 49% and 48% decreases in 14-hydroxyclarithromycin AUC(0-12h) and maximum plasma concentration (Cmax) values, respectively (P = .0001 and P = .0001). These effects are not considered to be clinically significant in view of the insignificant effects on the values for indinavir area under the plasma concentration versus time curve from time zero to the last measured concentration [AUC(0-8h)] and Cmax, as well as the safety profile of clarithromycin. Conclusions The combination of indinavir sulfate and clarithromycin is generally well tolerated and can be coadministered without dose adjustment. (Clin Pharmacol Ther 2000;67:351-9.) Clinical Pharmacology & Therapeutics (2000) 67, 351–359; doi: 10.1067/mcp.2000.105151

The superior cervical ganglion (SCG) is a well-characterized model of neural development, in which several regulatory signals have been identified. Vasoactive intestinal peptide (VIP) has been found to regulate diverse ontogenetic processes in sympathetics, though functional requirements for high peptide concentrations suggest that other ligands are involved. We now describe expression and functions of pituitary adenylate cyclase-activating polypeptide (PACAP) during SCG ontogeny, suggesting that the peptide plays critical roles in neurogenesis. PACAP and PACAP receptor (PAC1) mRNA's were detected at embryonic days 14.5 (E14.5) through E17.5 in vivo and virtually all precursors exhibited ligand and receptor, indicating that the system is expressed as neuroblasts proliferate. Exposure of cultured precursors to PACAP peptides, containing 27 or 38 residues, increased mitogenic activity 4-fold. Significantly, PACAP was 1000-fold more potent than VIP and a highly potent and selective antagonist entirely blocked effects of micromolar VIP, consistent with both peptides acting via PAC1 receptors. Moreover, PACAP potently enhanced precursor survival more than 2-fold, suggesting that previously defined VIP effects were mediated via PAC1 receptors and that PACAP is the more significant developmental signal. In addition to neurogenesis, PACAP promoted neuronal differentiation, increasing neurite outgrowth 4-fold and enhancing expression of neurotrophin receptors trkC and trkA. Since PACAP potently activated cAMP and PI pathways and increased intracellular Ca2+, the peptide may interact with other developmental signals. PACAP stimulation of precursor mitosis, survival, and trk receptor expression suggests that the signaling system plays a critical autocrine role during sympathetic neurogenesis.

Background. Indinavir, an inhibitor of the human immunodeficiency virus type 1 (HIV-1) protease, is approved for the treatment of HIV infection in adults when antiretroviral therapy is indicated. We evaluated the safety and pharmacokinetic profile of the indinavir free-base liquid suspension and the sulfate salt dry-filled capsules in HIV-infected children, and studied its preliminary antiviral and clinical activity in this patient population. In addition, we evaluated the pharmacokinetic profile of a jet-milled suspension after a single dose. Methods. Previously untreated children or patients with progressive HIV disease despite antiretroviral therapy or with treatment-associated toxicity were eligible for this phase I/II study. Three dose levels (250 mg/m2, 350 mg/m2, and 500 mg/m2 per dose given orally every 8 h) were evaluated in 2 age groups ( Results. Fifty-four HIV-infected children (ages 3.1 to 18.9 years) were enrolled. The indinavir free-base suspension was less bioavailable than the dry-filled capsule formulation, and therapy was changed to capsules in all children. Hematuria was the most common side effect, occurring in 7 (13%) children, and associated with nephrolithiasis in 1 patient. The combination of indinavir, lamivudine, and zidovudine was well tolerated. The median CD4 cell count increased after 2 weeks of indinavir monotherapy by 64 cells/mm3, and this was sustained at all dose levels. Plasma ribonucleic acid levels decreased rapidly in a dose-dependent way, but increased toward baseline after a few weeks of indinavir monotherapy. Conclusions. Indinavir dry-filled capsules are relatively well tolerated by children with HIV infection, although hematuria occurs at higher doses. Future studies need to evaluate the efficacy of indinavir when combined de novo with zidovudine and lamivudine.

This study explored cooperation between the retinoic acid (RA) and protein kinase C (PKC) pathways during differentiation of the multipotential human teratocarcinoma (TC) cell line NTERA-2 clone D1 (abbreviated NT2/D1). We report here that, compared to RA treatment alone, RA combined with the PKC stimulator 12-O-tetradecanoylphorbol-13-acetate (TPA) enhanced the regulated expression of the immunophenotypic differentiation markers SSEA-3, a globo-series carbohydrate, and the ganglio-series carbohydrate antigens GD2 and GD3. Northern analysis and transient transfection assays revealed that TPA co-treatment augmented the RA-induced expression and activation of the RA nuclear receptor-β (RAR-β), one early marker of RA response in NT2/D1 cells. This finding was extended with transient co-transfection experiments using a PKC-α expression vector which revealed that the PKC pathway can augment the activation of RAR-β by RA. These experiments establish PKC as a modulator of RAR-β expression in NT2/D1 cells. Similarly, experiments showed that RA can modulate activation of the PKC-responsive AP- 1 complex, a transcription factor rapidly activated by TPA. Northern analysis and transient transfection assays revealed that, compared to TPA treatment alone, RA and TPA augmented the expression and transcriptional activity of AP-1 in NT2/D1 cells. In contrast, transient transfection assays revealed no cooperative effect between RA and TPA in HeLa cells, indicating that this effect in NT2/D1 cells is cell type-specific. In summary, these studies show that stimulation of the PKC second messenger pathway can modulate tumor differentiation and transcriptional activation of a retinoid receptor associated with RA response.

This report investigated safety and dosing recommendations of intravenous caspofungin in hepatic insufficiency. In the single-dose study, 8 patients each with mild and moderate hepatic insufficiency received 70 mg of caspofungin. In the multiple-dose study, 8 patients with mild hepatic insufficiency and 13 healthy matched controls received 70 mg on day 1 and 50 mg daily on days 2 through 14. Eight patients with moderate hepatic insufficiency received 70 mg on day 1 and 35 mg daily on days 2 through 14. Caspofungin was generally well tolerated with no discontinuations due to serious or nonserious adverse experiences. The area under the concentration-time profile over the interval of last quantifiable point to infinity (AUC(0-infinity)) geometric mean ratio (GMR) (90% confidence interval [CI]) for mild hepatic insufficiency/historical controls was 1.55 (1.32-1.86) in the single-dose study and for mild hepatic insufficiency/concurrent controls was 1.21 (1.04-1.39) for day 14 area under the concentration-time profile calculated over the interval 0 to 24 hours (AUC(0-24h)) following multidose. The AUC(0-infinity) GMR (90% CI) for moderate hepatic insufficiency/historical controls was 1.76 (1.51-2.06) following 70 mg; AUC(0-24h) GMR (90% CI) for moderate hepatic insufficiency/concurrent controls was 1.07 (0.90-1.28) on day 14 after 35 mg daily. No dosage adjustment is recommended for patients with mild hepatic insufficiency. A dosage reduction to 35 mg daily following the 70-mg loading dose is recommended for patients with moderate hepatic insufficiency.

Farnesyl:protein transferase (FPTase) inhibitors were developed as anti-Ras drugs, but they fail to inhibit Ki-Ras activity because Ki-Ras can be modified by geranylgeranyl:protein transferase type-I (GGPTase-I). L-778,123, an inhibitor of FPTase and GGPTase-I, was developed in part because it can completely inhibit Ki-Ras prenylation. To support the clinical development of L-778,123, we developed pharmacodynamic assays using peripheral blood mononuclear cells (PBMCs) to measure the inhibition of prenylation of HDJ2 and Rap1A, proteins that are FPTase- and GGPTase-I substrates, respectively. We validated these assays in animal models and show that inhibition of HDJ2 prenylation in mouse PBMCs correlates with the concentration of FPTase inhibitors in blood. In dogs, continuous infusion of L-778,123 inhibited both HDJ2 and Rap1A prenylation in PBMCs, but we did not detect inhibition of Ki-Ras prenylation. We reported previously results from the first L-778,123 Phase I trial that showed a dose-dependent inhibition of HDJ2 farnesylation in PBMCs. In this report, we present additional analysis of patient samples from this trial and a second Phase I trial of L-778,123, and demonstrate the inhibition of both HDJ2 and Rap1A prenylation in PBMC samples. This study represents the first demonstration of GGPTase-I inhibition in humans. However, no inhibition of Ki-Ras prenylation by L-778,123 was detected in patient samples. These results confirm the pharmacologic profile of L-778,123 in humans as a dual inhibitor of FPTase and GGPTase-I, but indicate that the intended target of the drug, Ki-Ras, was not inhibited.

This study evaluates the safety and potential pharmacokinetic interaction between indinavir and trimethoprim/sulfamethoxazole (TMP/SMZ). In a randomized, three-period crossover fashion, 12 healthy adults received 1 week of indinavir sulfate 400 mg orally every 6 hours with placebo, TMP 160 mg/SMZ 800 mg orally every 12 hours with placebo, and indinavir sulfate with TMP/SMZ. Plasma indinavir, SMZ, and TMP concentrations were determined after the last dose of each treatment period. Concomitant administration resulted in a 17% decrease in geometric mean trough plasma indinavir concentrations (p = 0.032), an 18% increase in geometric mean AUC 0-12h and C max TMP values (p = 0.031 and 0.030, respectively), and a 5% increase in geometric mean AUC 0-12h SMZ values (p = 0.039). None of these effects was considered clinically significant. The combination of indinavir sulfate and TMP/SMZ is generally well tolerated, with no clinically significant pharmacokinetic interaction being noted.

The effect of multiple doses of indinavir on the pharmacokinetics of a single dose of theophylline was investigated in 16 healthy male subjects using a randomized, double-blind, placebo-controlled, parallel-group study design. On days 1 and 7, all of the subjects received a single oral 250 mg dose of theophylline. From days 2 to 7, the subjects received orally administered 800 mg doses ofindinavir or a matched placebo every 8 hours. On day 7, theophylline and indinavir (or a placebo) were coadministered. The geometric mean AUC (0-24 h) of theophylline increased 18% when coadministered with indinavir compared to when theophylline was administered alone. This small increase in AUC (0.24 h) , although considered statistically significant, did not meet the prespecified criterion for clinical significance. The geometric mean C max of theophylline, when coadministered with indinavir, was within 8% of theophylline when administered alone. The mean t max (± SD) value for theophylline, when coadministered with indinavir (0.9 ± 0.5 h), was comparable to that observed for theophylline alone (1.0 ± 0.5 h). In conclusion, the administration of multiple doses of indinavir followed by a single dose of theophylline did not appear to result in a clinically significant pharmacokinetic interaction for theophylline.

To evaluate a potential pharmacokinetic interaction of coadministration of fluconazole, and indinavir, a human immunodeficiency virus (HIV) protease inhibitor, 13 patients were enrolled in a multiple-dose, three-period, placebo-controlled, crossover study. Patients were randomly assigned to receive indinavir at 1,000 mg every 8 h for 7 1 3 days (with fluconazole placebo), fluconazole at 400 mg once daily for 8 days (with indinavir placebo), and indinavir with fluconazole in combination. The pharmacokinetics of both drugs were measured on day 8 of each treatment period. The peak concentration in plasma ( C max ) and the time to reach C max were obtained by inspection, and the area under curve (AUC) was calculated for indinavir and fluconazole for each treatment period in which the respective drugs were administered. There was a marginally ( P = 0.08) statistically significant decrease in the AUC from 0 to 8 h (AUC 0–8 ) for indinavir when it was administered with fluconazole. However, the magnitudes of the decreases in C max and the concentration at 8 h postdosing ( C 8 ) were not as great as the decrease in AUC 0–8 . Although the 90% confidence interval for the geometric mean ratio was within the hypothesized limits, the clinical significance is not clear. Indinavir coadministration with fluconazole had no statistically ( P > 0.5) or clinically significant effect on the C max and C 8 of indinavir. Fluconazole coadministration with indinavir had no statistically or clinically significant effect on the pharmacokinetics of fluconazole. One patient was discontinued because of mild to moderate abdominal pain and diarrhea while on indinavir and fluconazole in combination. No serious adverse experience according to the results of laboratory tests was noted. Total bilirubin levels in serum were mildly increased in most patients treated with indinavir. This was not clinically significant and was not affected by the coadministration of fluconazole. Although the values of the pharmacokinetic parameters for indinavir decrease in the presence of fluconazole, indinavir and fluconazole can be administered concomitantly to HIV-infected patients without adjustment of the dose of either drug, and both drugs are generally well tolerated.

Losartan, on orally active, nonpeptide angiotensin II receptor antagonist is being developed as a therapeutic agent for the treatment of hypertension and heart failure. Many patients requiring anticoagulant therapy with warfarin also may have hypertension or heart failure, and thus, are potential candidates for losartan therapy. This study was designed to investigate whether losartan at likely dosage levels would alter the anticoagulant response to warfarin. In a two-period, placebo-controlled, randomized, crossover study, ten healthy male subjects received a single oral dose of 30 mg warfarin sodium on the seventh day of a 13-day treatment with losartan, 100 mg daily by mouth, or placebo. Multiple plasma samples were collected over a 6-day period after both warfarin doses for the measurements of R- and S-warfarin concentrations and prothrombin times. The pharmacokinetics of R- and S-warfarin were comparable in the absence and presence of losartan (no significant effects of losartan on area under the curve, Cmax, or tmax). Losartan also had no significant effect on the anticoagulant effect of warfarin, as assessed by the area under the prothrombin time versus time curve and the maximum response for prothrombin time. The lack of pharmacokinetic or pharmacodynamic interaction between warfarin and losartan observed in this investigation suggests that a clinically important interaction between these drugs is unlikely to occur in patients requiring concomitant administration of both drugs.

BACKGROUND/AIMS To pilot a pharmacodynamic (PD) model of short-term androgen administration that may be used to predict long-term effects of androgen administration in postmenopausal women. The study was specifically designed to identify biomarkers that evaluate early skin response to exogenous androgens. The clinical consequences of androgen administration include hirsutism and acne. Skin biopsies were performed to document morphological changes in the pilosebaceous unit (PSU) and gene expression changes (Taqman and microarray). Functional skin response was gauged by measuring sebum excretion rates (SER). METHODS A double-blind, randomized, placebo-controlled, parallel-group study was conducted in 36 healthy postmenopausal female subjects to identify early biomarkers of androgen administration. Subjects were randomized to receive either 2.5 mg of transdermal testosterone gel (TTG; AndroGel®), 300 μg of TTGtransdermal testosterone gel, or placebo gel, daily for 6 weeks in a 1:1:1 ratio. Skin biopsies were performed to document morphological changes in the pilosebaceous unit (PSU) and gene expression changes (Taqman and microarray). Five-mm skin punch biopsies were obtained from the back of subjects at baseline, after 6 weeks of treatment, and 4 weeks post treatment. Biopsies were analyzed for histological changes characterized by total sebaceous gland volume and sebocyte cell size. Skin biopsies were also analyzed for molecular skin changes using skin Taqman/microarray assays. Measurements of sebum excretion rates (SER) were obtained at baseline and after 6 weeks of treatment. RESULTS Six weeks of treatment with the 2.5-mg TTGdose of AndroGel® increased sebaceous gland volume on average by 42% (p=0.0410) and sebum excretion rateSER by 52% (p=0.0021) relative to placebo. Microarray and Taqman analyses on skin samples also demonstrated changes in the 2.5-mg TTG AndroGel® group. Histological and molecular findings were noted to be reversible 4 weeks after discontinuing therapy. Analysis of Total Sebaceous Gland Volume (106 μm3) at Week 6 (See Tables 1 and 2) Mean (SD) % Change from Baseline Treatment N Baseline Week 6 Mean (SD) LS Mean 95% CI Placebo 13 19.58 (32.51) 24.00 (32.27) 4.42 (33.53) 4.42 (−21.79, 30.63) 300 ug TTG 13 37.20 (39.16) 45.59 (47.20) 8.40 (51.97) 8.40 (−17.81, 34.61) 2.5 mg TTG 10 31.93 (36.34) 77.91 (71.76) 45.98 (53.01) 45.98 (16.10, 75.87) Analysis Results for SER at Week 6 Geometric Mean (CV) % Change from Baseline Treatment N Baseline Week 6 Geometric Mean (CV) Geometric LS Mean† 95% CI‡ Placebo 13 0.67 (0.52) 0.67 (0.51) −0.14 (0.29) −0.14 (18.22, −15.65) 300 ug TTG 12 0.81 (0.55) 0.89 (0.56) 10.80 (0.38) 10.80 (32.08, 7.05) 2.5 mg TTG 10 0.82 (0.43) 1.24 (0.42) 52.09 (0.22) 52.09 (84.37, 25.46) CONCLUSIONS TTG treatment is associated with marked increases in sebaceous gland volume and SER, consistent with side effects including hirsutism and acne. Biomarkers sufficiently robust to support using to test for androgen-mediated skin effects in postmenopausal women were identified. Clinical Pharmacology & Therapeutics (2005) 79, P84–P84; doi: 10.1016/j.clpt.2005.12.299

Cell surface and cryptic insulin receptors were solubilized from the particulate fraction of murine 3T3-L1 adipocytes with buffer containing 1% Triton X-100. Solubilized receptors were affinity crosslinked with 125I-labeled insulin and disuccinimidyl suberate and characterized by sodium dodecyl sulfate/polyacrylamide gel electrophoresis and autoradiography after specific immunoprecipitation. Two insulin-binding polypeptides were identified: the more abundant protein had a Mr of 130,000, corresponding to the size of the hormone-binding subunit of insulin receptors on the surface of target cells; the second polypeptide exhibited a Mr of 200,000 and appears to be a component of the latent pool because it was unaffected when 3T3-L1 adipocytes were exposed to trypsin under conditions that result in a 95% reduction in cell surface insulin-binding activity and the loss of the Mr 130,000 polypeptide in crosslinking experiments. Unexpectedly, the population of Mr 200,000 molecules in intact cells was accessible for limited cleavage by chymotrypsin, yielding a Mr 195,000 insulin-binding polypeptide. When 3T3-L1 adipocytes received a 15-min pulse of [35S]methionine, the predominant immunoprecipitated polypeptide had a Mr of 180,000. During a 1.5-hr chase, radioactivity in the Mr 180,000 species rapidly declined while the latent Mr 200,000 polypeptide became intensely labeled. After a 5-hr chase period, broad protein bands with Mrs of 130,000 and 90,000 were visualized as the major immunoprecipitated radioactive polypeptides. Thus, the Mr 180,000 species may be a very early biosynthetic precursor that may be subsequently processed to a Mr 200,000 form and one or both of the smaller receptor subunits at the cell surface.