Most
-positive lung cancers will develop ALK-independent resistance after treatment with next-generation ALK inhibitors.
amplification has been described in patients progressing on ALK inhibitors, ...but frequency of this event has not been comprehensively assessed.
We performed FISH and/or next-generation sequencing on 207 posttreatment tissue (
= 101) or plasma (
= 106) specimens from patients with ALK-positive lung cancer to detect
genetic alterations. We evaluated ALK inhibitor sensitivity in cell lines with
alterations and assessed antitumor activity of ALK/MET blockade in ALK-positive cell lines and 2 patients with MET-driven resistance.
amplification was detected in 15% of tumor biopsies from patients relapsing on next-generation ALK inhibitors, including 12% and 22% of biopsies from patients progressing on second-generation inhibitors or lorlatinib, respectively. Patients treated with a second-generation ALK inhibitor in the first-line setting were more likely to develop
amplification than those who had received next-generation ALK inhibitors after crizotinib (
= 0.019). Two tumor specimens harbored an identical
rearrangement, one of which had concurrent
amplification. Expressing
in the sensitive H3122 ALK-positive cell line induced resistance to ALK inhibitors that was reversed with dual ALK/MET inhibition. MET inhibition resensitized a patient-derived cell line harboring both
and
amplification to ALK inhibitors. Two patients with ALK-positive lung cancer and acquired
alterations achieved rapid responses to ALK/MET combination therapy.
Treatment with next-generation ALK inhibitors, particularly in the first-line setting, may lead to MET-driven resistance. Patients with acquired
alterations may derive clinical benefit from therapies that target both ALK and MET.
Orthoester-containing copolymers are degradable under mild acidic conditions, making them attractive candidates for applications in drug delivery, photolithography, or disposable plastics with a ...built-in mechanism for environmental remediation. Orthoester linkages may be incorporated into polyether backbones by a chain-growth ring-opening and subsequent ring-closing polymerization of glycidyl acetate (GA) and methyl glycidyl acetate (MGA) monomers, where both epoxy and carbonyl groups participate in orthoester formation using a mono(μ-alkoxo)bis(alkylaluminum) (MOB) initiator. MGA exists as a mixture of (S,S)/(R,R)- and (S,R)/(R,S)-diastereomers, and MOB-initiated polymerization exhibited a strong preference for the (S,R)/(R,S)-diastereomer. Copolymerization between GA and a conventional epoxide monomer resulted in materials combining the facile degradation of polyorthoesters with the structural diversity of polyethers. We demonstrated the synthesis of degradable linear chains with methyl, allyl ether, propenyl ether, and methylene chloride pendant groups with weight average molecular weights ranging from 6.1 to 18.4 kg/mol. Additionally, we demonstrated the synthesis of degradable, polarity-switching cross-linked networks with acid-cleavable orthoester linkages and propenyl ether pendant groups that slowly degrade under mild acidic conditions without swelling.
Following anterior cruciate ligament (ACL) reconstruction surgery, a staged repair response occurs where cells from outside the tendon graft participate in tunnel integration. The mechanisms that ...regulate this process, including the specific cellular origin, are poorly understood. Embryonic cells expressing growth and differentiation factor 5 (GDF5) give rise to several mesenchymal tissues in the joint and epiphyses. We hypothesized that cells from a GDF5 origin, even in the adult tissue, would give rise to cells that contribute to the stages of repair. ACLs were reconstructed in Gdf5‐Cre;R26R‐tdTomato lineage tracing mice to monitor the contribution of Gdf5‐Cre;tdTom+ cells to the tunnel integration process. Anterior−posterior drawer tests demonstrated 58% restoration in anterior−posterior stability. Gdf5‐Cre;tdTom+ cells within the epiphyseal bone marrow adjacent to tunnels expanded in response to the injury by 135‐fold compared with intact controls to initiate tendon‐to‐bone attachments. They continued to mature the attachments yielding zonal insertion sites at 4 weeks with collagen fibers spanning across unmineralized and mineralized fibrocartilage and anchored to the adjacent bone. The zonal attachments possessed tidemarks with concentrated alkaline phosphatase activity similar to native entheses. This study established that mesenchymal cells from a GDF5 origin can contribute to zonal tendon‐to‐bone attachments within bone tunnels following ACL reconstruction.
The objective of our study was to use Gdf5‐Cre transgenic mice to trace the origin of cells that revitalize the tendon graft following ACL reconstruction in a mouse model and to examine the extent to which these cells can create mineralized attachments within bone tunnels.
There is an increasing need for efficient phenotyping and histopathology of a variety of tissues. This phenotyping need is evident with the ambitious projects to disrupt every gene in the mouse ...genome. The research community needs rapid and inexpensive means to phenotype tissues via histology. Histological analyses of skeletal tissues are often time consuming and semi-quantitative at best, regularly requiring subjective interpretation of slides from trained individuals. Here, we present a cryohistological paradigm for efficient and inexpensive phenotyping of mineralized tissues. First, we present a novel method of tape-stabilized cryosectioning that preserves the morphology of mineralized tissues. These sections are then adhered rigidly to glass slides and imaged repeatedly over several rounds of staining. The resultant images are then aligned either manually or via computer software to yield composite stacks of several layered images. The protocol allows for co-localization of numerous molecular signals to specific cells within a given section. In addition, these fluorescent signals can be quantified objectively via computer software. This protocol overcomes many of the shortcomings associated with histology of mineralized tissues and can serve as a platform for high-throughput, high-content phenotyping of musculoskeletal tissues moving forward.
•Type V collagen regulates the collagen fibril nanostructure and micromechanics of both the fibrocartilage and hyaline cartilage layers in temporomandibular joint condyle.•Reduction of type V ...collagen leads to decreased cell density and aberrant cell clustering in both fibrous and hyaline layers.•Loss of type V collagen leads to reduced cell proliferation and β-catenin expression in the fibrous layer, indicating its role in maintaining the progenitor cell niche in condylar cartilage.•Ablation of type V collagen at the post-weaning age results in pronounced thinning of the hyaline layer, highlighting the interplay between type V collagen and mechanoregulation of condylar cartilage growth.•The role of type V collagen in regulating cell fate is specific to the progenitor cells in condylar cartilage, and is absent in knee cartilage.
This study queried the role of type V collagen in the post-natal growth of temporomandibular joint (TMJ) condylar cartilage, a hybrid tissue with a fibrocartilage layer covering a secondary hyaline cartilage layer. Integrating outcomes from histology, immunofluorescence imaging, electron microscopy and atomic force microscopy-based nanomechanical tests, we elucidated the impact of type V collagen reduction on TMJ condylar cartilage growth in the type V collagen haploinsufficiency and inducible knockout mice. Reduction of type V collagen led to significantly thickened collagen fibrils, decreased tissue modulus, reduced cell density and aberrant cell clustering in both the fibrous and hyaline layers. Post-natal growth of condylar cartilage involves the chondrogenesis of progenitor cells residing in the fibrous layer, which gives rise to the secondary hyaline layer. Loss of type V collagen resulted in reduced proliferation of these cells, suggesting a possible role of type V collagen in mediating the progenitor cell niche. When the knockout of type V collagen was induced in post-weaning mice after the start of physiologic TMJ loading, the hyaline layer exhibited pronounced thinning, supporting an interplay between type V collagen and occlusal loading in condylar cartilage growth. The phenotype in hyaline layer can thus be attributed to the impact of type V collagen on the mechanically regulated progenitor cell activities. In contrast, knee cartilage does not contain the progenitor cell population at post-natal stages, and develops normal structure and biomechanical properties with the loss of type V collagen. Therefore, in the TMJ, in addition to its established role in regulating the assembly of collagen I fibrils, type V collagen also impacts the mechanoregulation of progenitor cell activities in the fibrous layer. We expect such knowledge to establish a foundation for understanding condylar cartilage matrix development and regeneration, and to yield new insights into the TMJ symptoms in patients with classic Ehlers-Danlos syndrome, a genetic disease due to autosomal mutation of type V collagen.
Lesions of the distal deep digital flexor tendon (DDFT) are frequently diagnosed using MRI in horses with foot pain. Intralesional injection of biologic therapeutics shows promise in tendon healing; ...however, accurate injection of distal deep digital flexor tendon lesions within the hoof is difficult. The aim of this experimental study was to evaluate accuracy of a technique for injection of the deep digital flexor tendon within the hoof using MRI‐guidance, which could be performed in standing patients. We hypothesized that injection of the distal deep digital flexor tendon within the hoof could be accurately guided using open low‐field MRI to target either the lateral or medial lobe at a specific location. Ten cadaver limbs were positioned in an open, low‐field MRI unit. Each distal deep digital flexor tendon lobe was assigned to have a proximal (adjacent to the proximal aspect of the navicular bursa) or distal (adjacent to the navicular bone) injection. A titanium needle was inserted into each tendon lobe, guided by T1‐weighted transverse images acquired simultaneously during injection. Colored dye was injected as a marker and postinjection MRI and gross sections were assessed. The success of injection as evaluated on gross section was 85% (70% proximal, 100% distal). The success of injection as evaluated by MRI was 65% (60% proximal, 70% distal). There was no significant difference between the success of injecting the medial versus lateral lobe. The major limitation of this study was the use of cadaver limbs with normal tendons. The authors conclude that injection of the distal deep digital flexor tendon within the hoof is possible using MRI guidance.
To determine the safety and short-term efficacy of intrabursal administration of botulinum toxin type B (BTXB) to alleviate lameness in horses with degenerative injury to the podotrochlear apparatus ...(PA).
10 Quarter Horses with degenerative injury to the PA.
Degenerative injury to the PA was confirmed with diagnostic analgesia and imaging. Then, BTXB (3.8 to 4.5 U/kg) was injected into the podotrochlear (navicular) bursa of each horse. Three horses were used in a safety evaluation. Subsequently, video recordings of lameness evaluations were obtained for 7 client-owned horses 5 days before (baseline) and 7 and 14 days after BTXB treatment and used to determine the effect of BTXB injection on lameness; 1 horse was removed from the study 8 days after BTXB treatment. Three investigators who were unaware of the treated forelimbs or time points separately reviewed the recordings and graded the lameness of both forelimbs of the horses.
Improvement in lameness of the treated forelimbs was detected at 1 or both time points after BTXB administration in all horses. However, all horses had some degree of lameness at the end of the study. Two horses developed transient increases in lameness 48 to 72 hours after treatment; lameness resolved uneventfully.
Intrabursal injection of BTXB temporarily alleviated chronic lameness in horses with degenerative injury to the PA, without causing serious short-term adverse effects. Further investigation into the potential use of BTXB in horses affected by degenerative injury to the PA is warranted.
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