(W1130-09-51) Development of Crystalline Linsitinib Formulations for Local Reduction of Adipose Tissue

Purpose: Linsitinib is a small molecule inhibitor of insulin growth factor type 1 receptor (IGF-1R), which transmits insulin’s actions to metabolic responses in systemic tissues, including adipose tissue. IGF1-R and its role in thyroid eye disease (TED) has also been highlighted by the recent approval of teprotumumab, a systemically dosed biologic that blocks IGF-1R. The potential for a sustained delivery system that can provide similar efficacy with reduced systemic side effects would be of use in TED and other conditions where excessive white fat accumulation is implicated. Here, we describe the formulation of linsitinib nano- and microcrystals that inhibit adipogenesis in orbital fibroblasts in vitro and provide progressive, sustained loss of abdominal white fat in mice in vivo.

Methods: Linsitinib crystal size was reduced using wet-bead milling in the presence of 1.0 mm Zr beads using a TissueLyser. Various stabilizers were screened to facilitate stability in solution, ease in redispersion, and ability to inject through a small-gauge needle. Cell survival and adipogenesis assay was performed in orbital fibroblasts. Female CD1 mice,10-12 weeks old, were used to characterize the efficacy of linsitinib microcrystals (Lin-MCs) in reducing white fat. An incision was made in the abdomen to allow for injecting 40 µL of crystals or free linsitinib dissolved in polyethylene glycol 400 (Lin-PEG-400) at 4 different sites (1 mg linsitinib) in the peritoneal muscle on Day 0. Another control group received 150 µL of Lin-PEG-400 (7.5 mg/mL) peroral daily at a dose previously described to reduce visceral body fat in adult mice (45 mg/kg) [1]. The skin was then closed up using sutures and surgical staples, and dosed with Buprenex SR analgesic. Mice were weighed daily through day 7. Mice were sacrificed on day 7 to collect the abdominal fat pad and liver.

Results: Various sized linsitinib nanocrystals (196.1 ± 3.1 nm, 0.29 ± 0.11 PDI and 476.1 ± 45.3 nm, 0.31 ± 0.13) were formulated for testing in human-derived orbital fibroblasts in vitro. Linsitinib and linsitinib nanocrystals similarly inhibited orbital fibroblast adipogenesis in adipogenic media in vitro. To test the effect of sustained release of linsitinib on white fat in vivo, larger microcrystals were formulated. Lin-MCs milled in the presence of cellulose derivatives and polyvinyl alcohol (PVA) resulted in the formation of sediment within 24 hrs that was not easily dispersible and not easily passed through a needle. Lin-MCs milled in the presence of 0.25% hyaluronic acid (HA) (500 kDa) resulted in stable formulation (6.1 ± 2.5 µm) and easily passed through a small gauge needle for administration. Lin-MCs milled in the presence of 0.25% HA (500 kDa) were then tested in mice. All mice lost weight initially, which was consistent across the three groups on Day 2. Mice dosed with the Lin-MCs injection showed continued weight loss (-7.3 ± 0.8%) that was significant (**p < 0.01) on days 6 and 7 compared to the Lin-PEG-400 injection (-2.7 ± 0.8%). Similarly, mice dosed with Lin-MCs showed significant reduction (*p < 0.05) in the weight of the abdominal fat pad at day 7. As a measure of systemic toxicity, we performed qPCR analysis on gluconeogenic genes in the liver. While the weight of the livers relative to body weight was consistent across treatment groups, there was a trend toward increase in expression of some genes in the oral dosed group despite the short duration of dosing due to the toxicity.

Conclusion: A sustained delivery crystalline linsitinib suspension provided effective fat reduction with reduced systemic side effects to oral drug dosing. Such a formulation may be useful in the local treatment of TED and other conditions where excessive white fat accumulation is implicated.

References: Tajima, Kazuki, et al. "Metabolic recovery of lipodystrophy, liver steatosis, and pancreatic β cell proliferation after the withdrawal of OSI-906." Scientific Reports 7.1 (2017): 4119.

Acknowledgements:

Funding: This study was supported by Research to Prevent Blindness and the Helmsley Foundation.
Conflict of Interest: LME, YRN, and FR are co-inventors on patent applications related to methods of use and formulations for therapies to treat adipose tissue and diseases involving IGF-1R. The terms of these arrangements are being managed by Johns Hopkins University in accordance with its conflict-of-interest policies. All other authors have no conflicts of interest to declare.