Subject demographics.

<p>Subject demographics.</p

Mean pharmacokinetic parameters of STA-9090.

<p>*NR: not reportable due to insufficient terminal phase characterization.</p

Structure of STA-1474.

<p>Shown is the chemical structure of STA-1474, a water-soluble phosphate prodrug of ganetespib.</p

Response of cutaneous mast cell tumors to STA-1474 treatment.

<p>a) Patient #19 had recurrent grade 3 cutaneous MCTs. This patient received STA-1474 on the 1 hour infusion protocol twice per week and experienced a partial response to therapy of the cutaneous lesions after 7 treatments. b) Patient #24 had a large previously untreated cutaneous MCT. This patient received STA-1474 on the 1 hour infusion protocol twice per week and experienced a partial response to therapy of the cutaneous lesions after 4 treatments.</p

Serial CT demonstrating regression of oral malignant melanoma following treatment with STA-1474.

<p>Patient # 5 had an aggressive oral malignant melanoma that had invaded into the nasal cavity. During the 4^th treatment cycle with STA-1474, an extravasation event occurred resulting in altered drug pharmacokinetics. A marked decrease in the oral mass was observed 7 days later and a subsequent CT scan confirmed a partial response to therapy. Shown are two representative matched CT images of tumor before and after treatment.</p

Response of metastatic thyroid carcinoma and OSA to STA-1474 treatment.

<p>a) Patient #14 had a locally recurrent thyroid carcinoma with metastatic disease to the lungs. This patient received STA-1474 on the 8 hour infusion protocol and experienced a partial response to therapy of both the locally recurrent and metastatic disease. Shown are representative radiographs before and after 15 treatments with STA-1474. The yellow arrows point to the metastatic pulmonary nodules. b) Patient #18 had metastatic OSA to the lungs following amputation and chemotherapy for an appendicular OSA. This patient received STA-1474 on the 8 hour infusion protocol and experienced a partial response to therapy. Shown are representative radiographs before and after 4 treatments with STA-1474. The yellow arrows point to the metastatic pulmonary nodules.</p

Analysis of HSP90 and HSP70 expression in dogs before and after treatment with STA-1474.

<p>a) PBMCs were collected from normal control dogs (cont 1 and cont 2) and study dogs before and 24 hours after treatment with STA-1474 at 9.5 mg/kg over 8 hours. Protein lysates were generated and following SDS-PAGE, Western blotting was performed for HSP70. Blots were then stripped and reprobed for β-actin. b) Tumor biopsies and PBMCs were collected from dogs treated with 5 mg/kg STA-1474 over 1 hour before treatment and at 7 and 24 hours post treatment. Protein lysates were generated and an ELISA was performed to detect HSP70. Results are expressed as HSP70 protein (ng/ml) and percent of baseline. c) Protein lysates generated from the tumor biopsies described above were subjected to SDS-PAGE followed by Western blotting for HSP90. Blots were then stripped and re-probed for β-actin.</p

Pharmacokinetic analysis of STA-1474 following administration using three different dosing regimens.

<p>a) Mean ganetespib plasma concentration – time profile for each dosing regimen b) Representative ganetespib plasma concentration – time profiles for two patients in each dosing regimen.</p

The HSP90 Inhibitor Ganetespib Alleviates Disease Progression and Augments Intermittent Cyclophosphamide Therapy in the MRL/lpr Mouse Model of Systemic Lupus Erythematosus

<div><p>Systemic lupus erythematosus (SLE) is a complex, systemic autoimmune disease with a diverse range of immunological and clinical manifestations. The introduction of broad spectrum immunosuppressive therapies and better management of acute disease exacerbations have improved outcomes for lupus patients over recent years. However, these regimens are burdened by substantial toxicities and confer significantly higher risks of infection, thus there remains a significant and unmet medical need for alternative treatment options, particularly those with improved safety profiles. Heat shock protein 90 (HSP90) is a ubiquitously expressed molecular chaperone that acts as an important modulator of multiple innate and adaptive inflammatory processes. Of note, accumulating clinical and experimental evidence has implicated a role for HSP90 in the pathogenesis of SLE. Here we evaluated the potential of HSP90 as a therapeutic target for this disease using the selective small molecule inhibitor ganetespib in the well-characterized MRL/lpr autoimmune mouse model. In both the prophylactic and therapeutic dosing settings, ganetespib treatment promoted dramatic symptomatic improvements in multiple disease parameters, including suppression of autoantibody production and the preservation of renal tissue integrity and function. In addition, ganetespib exerted profound inhibitory effects on disease-related lymphadenopathy and splenomegaly, and reduced pathogenic T and B cell lineage populations in the spleen. Ganetespib monotherapy was found to be equally efficacious and tolerable when compared to an effective weekly dosing regimen of the standard-of-care immunosuppressive agent cyclophosphamide. Importantly, co-treatment of ganetespib with a sub-optimal, intermittent dosing schedule of cyclophosphamide resulted in superior therapeutic indices and maximal disease control. These findings highlight the potential of HSP90 inhibition as an alternative, and potentially complementary, strategy for therapeutic intervention in SLE. Such approaches may have important implications for disease management, particularly for limiting or preventing treatment-related toxicities, a major confounding factor in current SLE therapy.</p></div

Ganetespib prevents the development of renal damage in a pilot prophylactic dosing study.

<p>(A) Mean pharmacokinetic plasma concentration-time profile after i.v. administration of ganetespib to MRL/lpr mice. Ganetespib was dosed at 50 mg/kg twice weekly and samples were taken after the second dose. Data represent mean ± SD (n = 3). (B) Average proteinuria scores (±SE) are plotted for vehicle and ganetespib-treated mice over the time course of the study. Average proteinuria scores for non-lupus parental strain (MRL/mp) mice from 8–15 weeks of age are included for comparison. (C) HSP70 induction as a pharmacodynamic marker of ganetespib activity in kidneys from drug-treated animals. Kidneys were harvested at 6 and 24 hours following the second intra-weekly dose of ganetespib (or vehicle) and lysates immunoblotted for HSP70 expression. GAPDH included as a loading control. Each lane represents renal tissue from an individual animal. (D) Representative H&E stained renal histopathology showing extensive chronic inflammation and thickening of the basement membrane with crescent formation in the glomerulus of a vehicle-treated mouse (<i>left panel</i>). In contrast, the kidneys of a ganetespib-treated animal showed no evident pathologic changes (<i>right panel</i>). Original magnification, 40X; scale bar represents 50 μm. (E) To evaluate total lupus-related renal damage 7 primary characteristics of GN (glomerular capillary deposits, hypercellularity, necrosis, tubular atrophy, intratubular casts, interstitial chronic inflammation, and fibrosis) were scored for each animal and summed for a composite disease score. The average summed pathology scores for each treatment group are plotted. (E) Representative IgG immunohistochemical staining of renal tissues harvested from vehicle- and ganetespib-treated animals at 22 weeks of age. Red arrows highlight the marked reduction in glomerular IgG deposition seen following HSP90 inhibitor treatment. Original magnification 20X: scale bar represents 100 μm. (F) Quantification of IgG glomerular immunoreactivity observed in vehicle and drug treated animals.</p