Biology subjects

Beier, F.

Publications and source records attributed to Beier, F..

12 recordsLinked to original sources

bioRxiv2026

Systematic De-Risking of TCR-Mimic Therapeutics Through Proteome-Wide Off-Target Landscaping and a Generalizable Design Rule Framework

TCR-mimic (TCRm) antibodies targeting peptide-human leukocyte antigen (pHLA) complexes enable precision immunotherapy against intracellular antigens, including cancer-testis antigens (CTAs). Achieving high specificity, however, remains challenging because of the vast diversity of the human immunopeptidome and the associated risk of off-target recognition. Here, we introduce ValidaTe, a unified framework for the proteome-scale prediction, validation, and mitigation of off-target liabilities in pHLA-directed therapeutics. ValidaTe integrates rational target prioritization, peptide-centric binder selection, proteome-wide off-target prediction, and therapeutic engineering into a hierarchical de-risking workflow. Using the CTA MAGE-A4 as a proof-of-concept, we identify the TCRm antibodies VR-4 and VR-6 with superior specificity and demonstrate how this workflow enables the discovery of safer pHLA-targeted binders. Furthermore, ValidaTe establishes the basis for the WiFi (Widened Fingerprint) engineering principle, which rationally combines TCRms with complementary off-target fingerprints in trivalent T-cell engagers to minimize unintended interactions while preserving potent target-specific activity. Together, these findings establish a generalizable framework for the rational development of safer and more selective pHLA-targeted therapeutics. We further discuss how orthogonal proteomic characterization may complement this workflow as a final layer of translational safety assessment prior to clinical development. TeaserValidaTe accelerates safe pHLA-targeted immunotherapy through proteome-wide off-target mapping and WiFi design

immunology↗

bioRxiv2023

Aged male and female Panx3 KO mice develop severe osteoarthritis independent of forced mechanical use.

Osteoarthritis (OA) is a multi-factorial disease associated with aging. As the molecular mechanisms underpinning the pathogenesis of this disease are unclear, there are no disease-modifying drugs to combat OA. Pannexin 3 (PANX3) has been shown to promote cartilage loss during posttraumatic OA. In contrast, the ablation of Panx3 in male mice results in spontaneous full-thickness cartilage lesions at 24 months of age. While protected from traumatic intervertebral disc (IVD) degeneration, Panx3 knockout (KO) mice show signs of IVD disease with altered disc mechanics. Whether the deleterious effects of ablating Panx3 in aging is the result from accumulated mechanical damage is unknown. We used male and female wildtype (WT) and global Panx3 KO C57Bl6 mice aged to 18 months of age. Mice were then randomized to sedentary (SED) or forced treadmill running (FEX) for 6 weeks. Knee joint tissues including patellar tendon, quadriceps and distal patellar enthesis, and synovium were analyzed histologically and through micro-CT, along with lumbar spine IVDs. Half of male and female sedentary Panx3 KO mice developed full-thickness cartilage lesions, severe synovitis, and ectopic fibrocartilage deposition and calcification of the knee joints in comparison to all other conditions. Panx3 KO mice with severe OA show signs of quadriceps and patellar enthesitis, characterized by bone and marrow formation. Forced treadmill running did not seem to exacerbate these phenotypes in male or female Panx3 KO mice; however, it may have contributed to the development of lateral compartment OA. The IVDs of aged Panx3 KO mice displayed no apparent differences to control mice, and forced treadmill running had no further effects in either genotype. We conclude that aged Panx3 KO mice show features of late-stage primary OA including full-thickness cartilage erosion, severe synovitis, and enthesitis. These data suggest that the deletion of Panx3 is deleterious to synovial joint health in aging. LAY SUMMARYOsteoarthritis is a common joint disease, and its incidence can increase with aging. As such, no drugs are available to treat its progression. Recently, we discovered that cellular channels composed of Pannexin 3 (PANX3) proteins may contribute to the health of joint tissues. In this study, we propose a new model for age-related primary OA, using histology, microCT, and pathology assessments under sedentary and forced exercise conditions. We found that aged mice lacking the channel protein PANX3 developed late-stage OA, including severe cartilage loss, joint inflammation, and tendon attachment damage. Half of the sedentary PANX3-deficient mice showed these changes, along with abnormal bone growth. Treadmill running did not make the condition worse but may have caused additional joint damage in some areas. The spine, however, remained unaffected. These findings suggest that deleting Panx3 harms joint health with aging. Importantly, this mouse model mimics human primary OA, even without added factors like diet changes or forced exercise.

physiology↗

bioRxiv2023

Pannexin 3 deletion in mice results in knee osteoarthritis and intervertebral disc degeneration after forced treadmill running.

Pannexin 3 (Panx3) is a glycoprotein that forms mechanosensitive channels expressed in chondrocytes and annulus fibrosus cells of the intervertebral disc (IVD). Evidence suggests Panx3 plays contrasting roles in traumatic versus aging osteoarthritis (OA) and intervertebral disc degeneration (IDD). However, whether its deletion influences the response of joint tissue to mechanical stress is unknown. The purpose of this study was to determine if Panx3 deletion in mice causes increased knee joint OA and IDD after forced treadmill running. Male and female wildtype (WT) and Panx3 knockout (KO) mice were randomized to either a no exercise group (sedentary; SED) or daily forced treadmill running (forced exercise; FEX) from 24 to 30 weeks of age. Knee cartilage, tibial secondary ossification center and IVD histopathology were evaluated by histology. Both male and female Panx3 KO mice developed larger superficial defects of the tibial cartilage after forced treadmill running compared to SED WT mice. Additionally, both male and female Panx3 KO mice developed greater bone area of the tibial secondary ossification center with running. In the lower lumbar spine, both male and female Panx3 KO mice developed histopathological features of IDD after running compared to SED WT mice. These findings suggest that the combination of deleting Panx3 and forced treadmill running induces OA and causes histopathological changes associated with degeneration of the IVDs in mice.

physiology↗

bioRxiv2023

Anabolic phenotype in cartilage-specific Mitogen-inducible gene-6 knockout mice is independent of Transforming growth factor-α

Background/ObjectiveOsteoarthritis (OA) is a whole joint disorder with no disease modifying treatment currently available. The Epidermal Growth Factor Receptor (EGFR) signaling pathway plays an important role in cartilage/bone development and its ligand transforming growth factor- (TGl{middle dot}) is upregulated in OA. In contrast, Mitogen-inducible gene 6 (Mig6) is a negative regulator of EGFR, and our studies demonstrate that cartilage-specific Mig-6 deletion results in anabolic effects on cartilage and formation of chondro-osseus nodules (CON). We aimed to attenuate EGFR signaling by inhibiting TGF production in cartilage-specific Mig6 deficient mice, to test whether this would prevent the formation of CONs. MethodsWe generated double knockout mice by crossing cartilage-specific Mig-6fl/flCol2a1-Cre+/- and whole-body Tgfa+/- mice to generate experimental and control wild-type mice. Knee and elbow sections were stained with toluidine blue to examine articular cartilage thickness and cell density, and tartrate-resistant acid phosphatase for osteoclast activity. Additionally, immunohistochemistry was completed to analyze phospho-EGFR and SOX9. ResultsMig-6 deficient mice display cartilage thickening and CONs at 12 weeks in both the elbow and knee joints, which is independent of TGF ligand presence. Similarly, articular cartilage cell density is increased in Mig-6fl/flCol2a1-Cre+/-Tgfa-/- and Mig-6fl/flCol2al-Cre+/- mice, but not Tgfa-/- mice, and displays increased SOX9 and phospho-EGFR staining. ConclusionThe articular cartilage displays increased thickness, increased cell density, and CON formation independent of the presence of TGF, suggesting the anabolic phenotype in the Mig6-deficient mice is independent of TGF/EGFR binding. The anabolic phenotype may be due to an alternative EGFR ligand activation, or perhaps due to other non-EGFR specific mechanism. More research is required to elucidate the exact pathway responsible for the anabolic effects.

physiology↗

bioRxiv2021

Pannexin 3 deletion reduces fat accumulation and inflammation in a sex-specific manner

BackgroundPannexin 3 (PANX3), is a channel-forming glycoprotein that enables nutrient-induced inflammation in vitro, and genetic linkage data suggests it regulates body mass index. Here, we characterized inflammatory and metabolic parameters in global Panx3 knockout (KO) mice in the context of forced treadmill running (FEX) and high fat diet (HFD). MethodsC57BL/6N (WT) and KO mice were randomized to either a FEX running protocol or no running (SED) from 24 until 30 weeks of age. Body weight was measured biweekly, and body composition was measured at 24 and 30 weeks of age. Male WT and KO mice were fed a HFD from 12 - 28 weeks of age. Metabolic organs were analyzed for a panel of inflammatory markers and PANX3 expression. ResultsIn females there were no significant differences in body composition between genotypes, which could be due to the lack of PANX3 expression in female white adipose tissue, while male KOs fed a chow diet had lower body weight, and lower fat mass at 24 and 30 weeks of age, which was reduced to the same extent as 6 weeks of FEX in WT mice. Additionally, male KO mice exhibited significantly lower expression of multiple pro-inflammatory genes in white adipose tissue compared to WT mice. While on a HFD body weight differences were insignificant, in KO mice, multiple inflammatory genes were significantly differently expressed in quadriceps muscle and white adipose tissue resulting in a more anti-inflammatory phenotype compared to WT mice. The lower fat mass in male KO mice may be due to significantly fewer adipocytes in their subcutaneous fat compared to WT mice. Mechanistically, adipose stromal cells (ASCs) cultured from KO mice grow significantly slower than WT ASCs. ConclusionPANX3 is expressed in male adult mouse adipose tissue and may regulate adipocyte numbers, influencing fat accumulation and inflammation.

physiology↗

bioRxiv2021

Genetic deletion of interleukin-15 is not associated with major structural changes following experimental post-traumatic knee osteoarthritis in rats

Post-traumatic Osteoarthritis (PTOA) is a degenerative joint disease, leading to articular cartilage breakdown, osteophyte formation, and synovitis, caused by an initial joint trauma. Pro-inflammatory cytokines increase catabolic activity and may perpetuate inflammation following joint trauma. Interleukin-15 (IL-15), a pro-inflammatory cytokine, is increased in OA patients, although its roles in OA pathophysiology are not well characterized. IL-15 levels appear to correlate to self-reported pain levels, and polymorphisms in the IL-15 receptor alpha gene correlate to a 1.5-fold increase in OA symptoms. This could be due to IL-15 effects on the activity of proteinases, such as matrix metalloproteinases (MMP) -1, -3, and -7. Here we utilized Il15 deficient rats to examine the role of IL-15 in PTOA pathogenesis in an injury-induced model of OA. OA was surgically induced in Il15 deficient rats and control wild-type rats to compare PTOA progression. Semi-quantitative scoring of the articular cartilage, subchondral bone, osteophyte size, and synovium was performed by two blinded observers. Analyses of articular cartilage damage, subchondral bone damage, and osteophyte formation revealed no significant difference between Il15 deficient rats and wild-type rats following PTOA-induction. Similarly, synovitis scoring across 6 parameters found no significant difference between genetic variants. Overall, IL-15 does not appear to play a key role in the development of structural changes in this surgically-induced rat model of PTOA.

genetics↗

bioRxiv2020

Pancreatic cancer stem cells (CSCs) are regulated via a feedback-loop of telomerase activity and stemness factors

To date, it is still unclear how cancer stem cells (CSCs) regulate their stemness properties, and to what extent they share common features with normal stem cells. Telomerase regulation is a key factor in stem cell maintenance. In this study, we investigate how telomerase regulation affects cancer stem cell biology in pancreatic ductal adenocarcinoma (PDAC), and delineate the mechanisms by which telomerase activity and CSC properties are linked. Using primary patient-derived pancreatic cancer cells, we show that CSCs have higher telomerase activity and longer telomeres than bulk tumor cells. Inhibition of telomerase activity, using genetic TERT-knockdown or pharmacological inhibitor (BIBR1532) resulted in CSC marker depletion in vitro, and reduced tumorigenicity in vivo. Furthermore, we identify a positive feedback loop between stemness factors (KLF4, SOX2, OCT3/4, NANOG) and telomerase, which is essential for the self-renewal of pancreatic CSCs. Disruption the balance between telomerase activity and stemness factors, eliminates CSCs via induction of DNA damage and apoptosis, opening future perspectives to avoid CSC driven therapy resistance and tumor relapse in PDAC patients.

cancer biology↗

bioRxiv2020

Glycogen synthase kinase 3 alpha/beta deletion induces precocious growth plate remodeling and cell loss in mice

Glycogen synthase kinase (GSK) 3 acts to negatively regulate multiple signaling pathways, including canonical Wnt signaling. The two mammalian GSK3 proteins (alpha and beta) are at least partially redundant. While Gsk3a KO mice are viable and display a metabolic phenotype, abnormal neuronal development and accelerated aging, Gsk3b KO animals die late in embryogenesis or at birth. Selective Gsk3b KO in bone delayed development of some bones, whereas cartilage-specific Gsk3b KO mice are normal except for elevated levels of GSK3alpha protein. However, the collective role of these two GSK3 proteins in cartilage was not evaluated. To address this, we generated tamoxifen-inducible, cartilage-specific Gsk3a/Gsk3b KO in juvenile mice and investigated their skeletal phenotypes. We found that cartilage-specific Gsk3a/Gsk3b deletion in young, skeletally immature mice causes precocious growth plate remodeling, culminating in shorter long bones and hence, growth retardation. These mice exhibit inefficient breathing patterns at later stages and fail to survive. The disrupted growth plates in KO mice showed progressive loss of cellular and proteoglycan components and Sox9 positive cells, with increased staining for osteocalcin and type II collagen. In addition, an increase in osteoclast recruitment and cell apoptosis was observed in growth plates. Surprisingly, changes in articular cartilage of Gsk3a/Gsk3b KO mice were mild compared to growth plates, signifying differential regulation of articular cartilage vs growth plate tissues. Taken together, these findings emphasize a crucial role of two GSK3 proteins in skeletal development, in particular in the maintenance and function of growth plates. SignificanceGrowth plate cartilage dynamics determine the rate of endochondral bone growth and thus, our final height. These processes are disturbed in many genetic and acquired diseases, but the intracellular mechanisms responsible for normal growth plate function, as well as the cessation of growth plate activity in puberty, are poorly understood. Here, we demonstrate that specific removal of both GSK3 genes (Gsk3a and Gsk3b) in postnatal cartilage of mice leads to a severe reduction of endochondral bone growth, premature remodelling of the growth plate, and early death. In contrast, articular cartilage is only mildly affected by deletion of both genes. These studies identify GSK3 signaling as a key regulator of growth plate dynamics and endochondral bone growth.

developmental biology↗

bioRxiv2019

Overexpression Of Mig-6 In Limb Mesenchyme Leads To Accelerated Osteoarthritis In Mice

BackgroundMitogen-inducible gene 6 (Mig-6) is a tumour suppressor gene that is also associated with the development of osteoarthritis (OA)-like disorder. Recent evidence from our lab and others showed that cartilage-specific Mig-6 knockout (KO) mice develop chondro-osseous nodules, along with increased articular cartilage thickness and enhanced EGFR signaling in the articular cartilage. Here, we evaluate the phenotype of mice with skeletal-specific overexpression of Mig-6. MethodsSynovial joint tissues of the knee were assessed in 12 and 36 weeks-old skeleton-specific Mig-6 overexpressing (Mig-6over/over) and control animals using histological stains, immunohistochemistry, semi-quantitative OARSI scoring, and microCT for skeletal morphometry. Measurement of articular cartilage and subchondral bone thickness were also performed using histomorphometry. ResultsOur results show only subtle developmental effects of Mig-6 overexpression. However, male Mig-6over/over mice show accelerated cartilage degeneration at 36 weeks of age, in both medial and lateral compartments of the knee. Immunohistochemistry for SOX9 and PRG4 showed decreased staining in Mig-6over/over mice relative to controls, providing potential molecular mechanisms for the observed effects. ConclusionOverexpression of Mig-6 in articular cartilage causes no major developmental phenotype but results in accelerated development of OA during aging. These data demonstrate that precise regulation of the Mig-6/EGFR pathway is critical for joint homeostasis.

genetics↗

bioRxiv2019

Exposure to the RXR agonist SR11237 in early life causes disturbed skeletal morphogenesis in a rat model

Longitudinal bone growth occurs through endochondral ossification (EO), controlled by various signaling molecules. Retinoid X Receptor (RXR) is a nuclear receptor with important roles in cell death, development, and metabolism. However, little is known about its role in EO. In this study, the agonist SR11237 was used to evaluate RXR activation on EO.\n\nRats given SR11237 from post-natal day 5 to 15 were harvested for micro-computed tomography scanning and histology. In parallel, newborn CD1 mouse tibiae were cultured with increasing concentrations of SR11237 for histological and whole mount evaluation.\n\nRXR agonist-treated rats were smaller than controls, and developed dysmorphia of the growth plate. Cells invading the calcified and dysmorphic growth plate appeared pre-hypertrophic in size and shape corresponding with P57 immunostaining. Additionally, SOX9 positive cells were found surrounding the calcified tissue. The epiphysis of SR11237 treated bones showed increased TRAP staining, and additional TUNEL staining at the osteo-chondral junction. MicroCT revealed morphological disorganization in the long bones of treated animals. Isolated mouse long bones treated with SR11237 grew significantly less than their DMSO controls.\n\nThis study demonstrates that stimulation of the RXR receptor causes irregular ossification, premature closure of the growth plate, and disrupted long bone growth in rodent models.

developmental biology↗

bioRxiv2019

Overexpression of Mig-6 in Cartilage Induces An Osteoarthritis-Like Phenotype In Mice.

BackgroundOsteoarthritis (OA) is the most common form of arthritis and characterized by degeneration of articular cartilage. Mitogen-inducible gene 6 (Mig-6) has been identified as a negative regulator of the Epidermal Growth Factor Receptor (EGFR). Cartilage-specific Mig-6 knockout (KO) mice display increased EGFR signaling, an anabolic buildup of articular cartilage and formation of chondro-osseous nodules. Since our understanding of the EGFR/Mig-6 network in cartilage remains incomplete, we characterized mice with cartilage-specific overexpression of Mig-6 in this study.\n\nMethodsUtilizing knee joints from cartilage-specific Mig-6 overexpressing (Mig-6over/over) mice (at multiple time points), we evaluated the articular cartilage using histology, immunohistochemical staining and semi-quantitative OARSI scoring at multiple ages. MicroCT analysis was employed to examine skeletal morphometry, body composition, and bone mineral density.\n\nResultsOur data show that cartilage-specific Mig-6 overexpression did not cause any major developmental abnormalities in articular cartilage, although Mig-6over/over mice have slightly shorter long bones compared to the control group. Moreover, there was no significant difference in bone mineral density and body composition in any of the groups. However, our results indicate that Mig-6over/over male mice show accelerated cartilage degeneration at 12 and 18 months of age. Immunohistochemistry for SOX9 demonstrated that the number of positively stained cells in Mig-6over/over mice decreased relative to controls. Immunostaining for MMP13 staining is increased in areas of cartilage degeneration in Mig-6over/over mice. Moreover, staining for phospho-EGFR (Tyr-1173) and lubricin (PRG4) was decreased in the articular cartilage of Mig-6over/over mice.\n\nConclusionOverexpression of Mig-6 in articular cartilage causes no major developmental phenotype; however these mice develop earlier OA during aging. These data demonstrate that Mig-6/EGFR pathways is critical for joint homeostasis and might present a promising therapeutic target for OA.

physiology↗

bioRxiv2019

Liver X Receptor activation regulates genes involved in lipid homeostasis in developing chondrocytes

ObjectiveOsteoarthritis (OA) is the most common type of arthritis and causes debilitating symptoms and decreased quality of life. Currently available treatment options target symptoms but do not address the underlying issue of joint tissue degeneration. As such, a better understanding of the molecular mechanisms maintaining cartilage health is needed for developing novel therapeutic strategies. Liver X Receptors (LXRs) are nuclear receptors that have been previously shown to offer protection against OA. This is potentially due to suppression of chondrocyte hypertrophy in endochondral bone growth in response to LXR activation. In order to better understand the regulatory mechanisms behind this effect, we aimed to systematically examine LXRs effects on growth plate chondrocyte gene expression.\n\nMethodsPrimary chondrocytes isolated from the long bones of E15.5 mice were treated with the specific LXR agonist, GW3965, and RNA was isolated for Affymetrix microarrays followed by real time qPCR validation. Bioinformatics analyses were performed using Gene Ontology (GO) and KEGG pathway analysis. Immunohistochemistry was conducted to examine protein localization of LXR and identified targets in GW3965-treated E15.5 tibiae compared to control.\n\nResultsActivation of LXR in primary growth plate chondrocytes resulted in differential regulations of various genes involved in lipid metabolism, including several genes involved in cholesterol efflux. This pattern was compared to LXR activation in immature murine articular chondrocytes (IMACs), which revealed similar roles in lipid homeostasis. Immunohistochemical analysis of LXR and its identified targets Abca1 and Srebf1 revealed preferential protein localization to pre-hypertrophic and resting chondrocytes in GW3965-treated tibial growth plates compared to controls.\n\nConclusionOur findings show for the first time that LXR activation alters expression of lipid metabolism genes in growth plate chondrocytes, in part through activation of molecules responsible for cellular cholesterol efflux. This provides insight into potential mechanisms through which LXR regulates cellular metabolism to alter chondrocyte behavior and phenotype.

physiology↗