Due to lack of full vascularization, the meniscus relies on diffusion through the extracellular matrix to deliver small (e.g., nutrients) and large (e.g., proteins) to resident cells. Under normal physiological conditions, the meniscus undergoes up to 20% compressive strains. While previous studies characterized solute diffusivity in the uncompressed meniscus, to date, little is known about the diffusive transport under physiological strain levels. This information is crucial to fully understand the pathophysiology of meniscus. The objective of this study was to investigate strain-dependent diffusive properties of the meniscus fibrocartilage. Tissue samples were harvested from the central portion of porcine medial menisci and tested via fluorescence recovery after photobleaching to measure diffusivity of fluorescein (332 Da) and 40K Da dextran (D40K) under 0%, 10%, and 20% compressive strain. Specifically, average diffusion coefficient and anisotropic ratio, defined as ratio of the diffusion coefficient in the direction of the tissue collagen fibers to that orthogonal, were determined. For all the experimental conditions investigated, fluorescein diffusivity was statistically faster than that of D40K. Also, for both molecules, diffusion coefficients significantly decreased, up to ~45%, as strain increased. In contrast, the anisotropic ratios of both molecules were similar and not affected by the strain applied to the tissue. This suggests that compressive strains used in this study did not alter the diffusive pathways in the meniscus. Our findings provide new knowledge on the transport properties of the meniscus fibrocartilage that can be leveraged to further understand tissue pathophysiology and approaches to tissue restoration.

Damage to the meniscus has been associated with excessive shear loads. Aimed at elucidating meniscus pathophysiology, previous studies have investigated the shear properties of the meniscus fibrocartilaginous core. However, the meniscus is structurally inhomogeneous, with an external cartilaginous envelope (tibial and femoral surface layers) wrapping the tissue core. To date, little is known about the shear behavior of the surface layers. The objective of this study was to measure the dynamic shear properties of the surface layers and derive empirical relations with their composition. Specimens were harvested from tibial and femoral surface layers and core of porcine menisci (medial and lateral, n = 10 each). Frequency sweep tests yielded complex shear modulus (G*) and phase shifts (δ). Mechanical behavior of regions was described by a generalized Maxwell model. Correlations between shear moduli with water and glycosaminoglycans content of the tissue regions were investigated. The femoral surface had the lowest shear modulus, when compared to core and tibial regions. A 3-relaxation times Maxwell model satisfactorily interpreted the shear behavior of all tissue regions. Inhomogeneous tissue composition was also observed, with water content in the surface layers being higher when compared with tissue core. Water content negatively correlated with shear properties in all regions. The lower measured shear properties in the femoral layer may explain the higher prevalence of meniscal tears on the superior surface of the tissue. The heterogenous behavior of the tissue in shear provides insight into meniscus pathology and has important implications for efforts to tissue engineer replacement tissues.

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The tropomycin kinase (Trk) receptor family contain Trk A, B and C that are encoded by the NTRK1, NTRK2, and NTRK3 genes respectively. Trk family kinases have been shown to play a very important role in the development and physiological functions of the nervous system. Recently, chimeric Trk kinases resulted from gene fusions of NTRK genes are involved in initiation and progression of various cancers such as sarcoma, thyroid, salivary gland cancers, etc. Targeting such alteration of Trk family kinases becomes a new avenue for the therapeutic intervention. ARQ 531 has biochemical IC50 values of 1.3 nM, 1.8 nM and 1.8 nM for Trk A, B and C, respectively with targeting a unique set of other kinases. In a cell based assay, ARQ 531 inhibits kinase activity of Trk A, B and C in UOS2 cells overexpressing Trk family kinases with IC50 values of 50.5 nM, 28.0 nM and 4.1 nM respectively. Pathway analysis shows that ARQ 531 abolished autophosphorylation of TMP3-Trk A in colon cancer KM-12 cells after 2-hour treatment, concomitant with marked inhibition of pAKT, pERK and pPLCγ1. Moreover, in Trk A-overexpressing K562 cells, ARQ 531 suppressed phosphorylation of Trk A and its downstream targets AKT and MAPK. Anti-proliferative activity of ARQ 531 has been demonstrated in Ba/F3 system over-expressing fusion Tel-Trk family kinases (ETV6-NTRK1, 2 or 3), exhibiting IC50 value of 300, 481, and 341 nM for Tel-Trk A, B or C respectively. In TMP3-TrkA(TMP3-NTRK1) fusion-driven KM12 cells driven by ARQ 531 strongly suppressed cell proliferation with an IC50 value of 130 nM. Subsequently, apoptotic response was observed in KM-12 cells after treatment of ARQ 531 for 24 hours, which was demonstrated by the increase in cleaved PARP1 and cleaved caspase 3. Significant anti-tumor activity of ARQ 531 was observed in KM-12 xenograft mouse model. In conclusion, ARQ 531 is a potent Trk family kinase inhibitor and the results from preclinical study provides rationale for testing ARQ 531 in a clinical setting, particularly in cancers driven by Trk fusion kinases. Citation Format: Yi Yu, Gabi Schwartz, Pascal Schamber, Ron E. Savage, Sudharshan Eathiraj, Terence Hall, Brian Schwartz. In vitro and in vivo effect of ARQ 531 on Trk family kinases [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4797.