Exploring the effects of extracranial injections of botulinum toxin type A on activation and sensitization of central trigeminovascular neurons by cortical spreading depression in male and female rats.

BACKGROUND: OnabotulinumtoxinA (onabotA), is assumed to achieve its therapeutic effect in migraine through blocking activation of unmyelinated meningeal nociceptors and their downstream communications with central dura-sensitive trigeminovascular neurons in the spinal trigeminal nucleus (SPV). The present study investigated the mechanism of action of onabotA by assessing its effect on activation and sensitization of dura-sensitive neurons in the SPV by cortical spreading depression (CSD). It is a follow up to our recent study on onabotA effects on activation and sensitization of peripheral trigeminovascular neurons. METHODS: In anesthetized male and female rats, single-unit recordings were used to assess effects of extracranial injections of onabotA (five injections, one unit each, diluted in 5 µl of saline were made along the lambdoid (two injection sites) and sagittal (two injection sites) suture) vs. vehicle on CSD-induced activation and sensitization of high-threshold (HT) and wide-dynamic range (WDR) dura-sensitive neurons in the SPV. RESULTS: Single cell analysis of onabotA pretreatment effects on CSD-induced activation and sensitization of central trigeminovascular neurons in the SPV revealed the ability of this neurotoxin to prevent activation and sensitization of WDR neurons (13/20 (65%) vs. 4/16 (25%) activated neurons in the control vs. treated groups, p = 0.022, Fisher's exact). By contrast, onabotA pretreatment effects on CSD-induced activation and sensitization of HT neurons had no effect on their activation (12/18 (67%) vs. 4/7 (36%) activated neurons in the control vs. treated groups, p = 0.14, Fisher's exact). Regarding sensitization, we found that onabotA pretreatment prevented the enhanced responses to mechanical stimulation of the skin (i.e. responses reflecting central sensitization) in both WDR and HT neurons. In control but not treated WDR neurons, responses to brush (p = 0.004 vs. p = 0.007), pressure (p = 0.002 vs. p = 0.79) and pinch (p = 0.007 vs. 0.79) increased significantly two hours after CSD. Similarly, in control but not treated HT neurons, responses to brush (p = 0.002 vs. p = 0.79), pressure (p = 0.002 vs. p = 0.72) and pinch (p = 0.0006 vs. p = 0.28) increased significantly two hours after CSD. Unexpectedly, onabotA pretreatment prevented the enhanced responses of both WDR and HT neurons to mechanical stimulation of the dura (commonly reflecting peripheral sensitization). In control vs. treated WDR and HT neurons, responses to dural stimulation were enhanced in 70 vs. 25% (p = 0.017) and 78 vs. 27% (p = 0.017), respectively. CONCLUSIONS: The ability of onabotA to prevent activation and sensitization of WDR neurons is attributed to its preferential inhibitory effects on unmyelinated C-fibers. The inability of onabotA to prevent activation of HT neurons is attributed to its less extensive inhibitory effects on the thinly myelinated Aδ-fibers. These findings provide further pre-clinical evidence about differences and potentially complementary mechanisms of action of onabotA and calcitonin gene-related peptide-signaling neutralizing drugs.

Tirbanibulin (KX2-391) analog KX2-361 inhibits botulinum neurotoxin serotype A mediated SNAP-25 cleavage in pre- and post-intoxication models in cells.

Botulinum neurotoxins (BoNT) inhibit neuroexocytosis, leading to the potentially lethal disease botulism. BoNT serotype A is responsible for most human botulism cases, and there are no approved therapeutics to treat already intoxicated patients. A growing body of research has demonstrated that BoNT/A can escape into the central nervous system, and therefore, identification of BoNT/A inhibitors that can penetrate BBB and neutralize the toxin within intoxicated neurons would be important. We previously identified an FDA-approved, orally bioavailable compound, KX2-391 (Tirbanibulin) that inhibits BoNT/A in motor neuron assays. Recently, a structural analog of KX2-391, KX2-361, has been shown to exhibit good oral bioavailability and cross BBB with high efficiency in mouse experiments. Therefore, in this work, we evaluated the inhibitory effects of KX2-361 against BoNT/A. Toward this goal, we first evaluated the compound for its effects on cell viability in PC12 cells, via MTT assay, and in mouse embryonic stem cell (mESC)-derived motor neurons, with imaging-based assays. Following, we tested KX2-361 in mESC-derived motor neurons intoxicated with BoNT/A holotoxin, and the compound exhibited activity against the toxin in both pre- and post-intoxication conditions. Excitingly, KX2-361 also inhibited BoNT/A enzymatic component (light chain; LC) in PC12 cells transfected with BoNT/A LC. Furthermore, our molecular docking analyses suggested that KX2-361 can directly bind to BoNT/A LC. Medicinal chemistry approaches to develop structural analogs of KX2-361 to increase its efficacy against BoNT/A may provide a critical lead compound with BBB penetration capacity for drug development efforts against BoNT/A intoxication.

Multiple sclerosis and spasticity: the role of anaesthetic nerve blocks on rectus femoris muscle. When should stiff knee be treated with botulinum toxin?

OBJECTIVE: To compare the effect of rectus femoris diagnostic motor nerve blocks (DNB) with anaesthetics and rectus femoris muscle botulinum toxin (BoNT-A) injection in multiple sclerosis patients with unilateral stiff-knee gait. DESIGN: Prospective observational study Subjects/Patients: Multiple sclerosis patients in stable condition. METHODS: Patients underwent evaluation before and 1 hour after the anaesthetic block, and 1 month after the botulinum injection. Assessment included a 10-m walking test, a 6-minute walking test, a timed-up-and-go (TUG) test, and a Baseline Expanded Disability Status Scale (EDSS). Post-DNB and post-BoNT-A satisfaction was measured with the global assessment of efficacy scale. RESULTS: Fourteen patients with unilateral stiff-knee gait due to multiple sclerosis underwent a DNB, among whom 13 received botulinum injections in the rectus femoris muscle after a satisfying test result. Positive post-DNB results correlated with significant functional improvements after BoNT-A. Higher EDSS and longer time from diagnosis correlated with poorer post-DNB and post-BoNT-A absolute outcomes. CONCLUSION: DNB showed predictive value for BoNT-A outcomes, especially in the case of worse functional status. It effectively predicted endurance and walking speed improvement, while TUG showed greater improvement after botulinum. In cases of uncertain therapeutic benefit, nerve blocks may provide a valuable diagnostic support, particularly in patients with lower functional status.

Clinical Conditions Targeted by OnabotulinumtoxinA in Different Ways in Medicine.

OnabotulinumtoxinA (BT-A) is used in different medical fields for its beneficial effects. BT-A, a toxin originally produced by the bacterium Clostridium botulinum, is widely known for its ability to temporarily paralyze muscles by blocking the release of acetylcholine, a neurotransmitter involved in muscle contraction. The literature continually reports new hypotheses regarding potential applications that do not consider blockade of acetylcholine release at the neuromuscular junction as a common pathway. In this opinion article, it is our aim to investigate the different pathway targets of BT-A in different medical applications. First of all, the acetylcholine effect of BT-A is used to reduce wrinkles for cosmetic purposes, in the treatment of urological problems, excessive sweating, temporomandibular joint disorders, obesity, migraine, spasticity in neurological diseases, and in various cases of muscle overactivity such as cervical dystonia, blepharospasm, and essential head tremor. In another potential pathway, glutamate A, CGRP, and substance P are targeted for pain inhibition with BT-A application in conditions such as migraine, trigeminal neuralgia, neuropathic pain, and myofascial pain syndrome. On the other hand, as a mechanism different from acetylcholine and pain mediators, BT-A is used in the treatment of hair loss by increasing oxygenation and targeting transforming growth factor-beta 1 cells. In addition, the effect of BT-A on the apoptosis of cancer cells is also known and is being developed. The benefits of BT-A applied in different doses to different regions for different medical purposes are shown in literature studies, and it is also emphasized in those studies that repeating the applications increases the benefits in the long term. The use of BT-A continues to expand as researchers discover new potential therapeutic uses for this versatile toxin.

The functional lumen imaging probe contractile response pattern is the best predictor of botulinum toxin response in esophagogastric junction outflow obstruction.

BACKGROUND: Esophagogastric junction outflow obstruction (EGJOO) is a heterogenous disorder in which the correct management strategy is unclear. We assessed whether functional lumen imaging probe (FLIP) topography data could select EGJOO, which would benefit from lower esophageal sphincter Botulinum toxin (Botox) injection. METHODS: This was a single-center prospective study of adult patients meeting Chicago Classification (CC) v3.0 criteria for EGJOO. We assessed differences in pretreatment physiologic measurements on high-resolution manometry (HRM) and FLIP and other relevant clinical variables in predicting Botox response (>50% in BEDQ at 2 months). KEY RESULTS: Sixty-nine patients were included (ages 33-90, 73.9% female). Of these, 42 (61%) were Botox responders. Majority of physiologic measures on HRM and FLIP and esophageal emptying were not different based on Botox response. However, a spastic-reactive (SR) FLIP contractile response (CR) pattern predicted a Botox response with OR 25.6 (CI 2.9-229.6) when compared to antegrade FLIP CR; and OR for impaired-disordered/absent CR was 22.5 (CI 2.5-206.7). Logistic regression model using backward elimination (p value = 0.0001, AUC 0.79) showed that a SRCR or IDCR/absent response and the upright IRP predicted Botox response. Response rates in tiered diagnostic groups were: (i) CCv3.0 EGJOO (60.9%), (ii) CCv4.0 EGJOO (73.1%), (iii) CCv4.0 + FLIP REO (80%), (iv) CCv4.0, FLIP REO, and abnormal FLIP CR (84.2%), and (v) CCv4.0, FLIP REO, and SR FLIP CR (90%). CONCLUSIONS AND INFERENCES: FLIP helps identify patients with EGJOO who are likely to response to LES Botox therapy. An abnormal FLIP contractile response pattern is the single-most important predictor of a Botox response.

Reversion of chronic to episodic migraine in working age and botulinum toxin-resistant patients treated with fremanezumab: A real-life study.

OBJECTIVES: The objectives of this real-life study were to analyze the reversion of chronic migraine (CM) to episodic migraine (EM) with fremanezumab, evaluate its benefit on the symptomatology, and determine the influence of possible clinical features on the reversion. BACKGROUND: The clinical manifestations of CM have a high impact on the quality of life of patients, and monoclonal antibodies such as fremanezumab are used as prophylactic treatment. METHODS: Diagnosed CM patients treated for at least 3 months with monthly fremanezumab were interviewed. The data to assess efficacy were before treatment and at the time of the interview: monthly headache days (MHDs), daily headache hours (DHHs), monthly symptomatic medication days (MSMDs), percentage of patients with symptomatic medication overuse (SMO), and pain intensity with the numerical rating scale (NRS) score. Possible predictors of reversion were analyzed: percentage of patients treated for at least 12 months, hypertension, diabetes mellitus, depression, anxiety, symptomatic control with non-steroidal anti-inflammatory drugs (NSAIDs), triptans or both, and amitriptyline prophylaxis. RESULTS: A total of 54 patients were included, of whom 40 (74.1%) were converters to EM. There were significant improvements in converters compared to pre-treatment in MHDs (28.0 vs. 5.0 days), as well as on the variables DHHs, MSMDs, and SMO. The percentage of erenumab failures was significantly higher in non-converters than in converters, as was the percentage of patients with anxiety. CONCLUSIONS: High reversion from CM to EM was achieved with fremanezumab and notable symptomatological improvement, establishing previous failure to erenumab and anxiety as possible detrimental factors for reversion.

The Effect of Botulinum Toxin A on the NADPH Oxidase System and Ischemia-Reperfusion Injury.

BACKGROUND: Despite the increasing popularity of various materials for ischemia-reperfusion (I/R) injury mitigation, research on botulinum toxin type A (BoNTA) remains limited. This study assesses BoNTA's efficacy in protecting flaps from I/R injury by inhibiting the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase system and reducing reactive oxygen species (ROS) production. METHODS: Seventy-six Sprague-Dawley rats were studied. We examined the effects of BoNTA on superoxide production in four rats using a lucigenin-enhanced chemiluminescence assay (LECL). Another group of 60 rats had their superficial inferior epigastric artery (SIEA) flaps treated with either BoNTA or saline and clamped for 0, 1, and 4 hours before reperfusion. Flap survival and histological outcomes were assessed five days post-operation. ROS production in SIEA flaps and femoral vessels was analyzed in 12 additional rats, post-I/R injury. RESULTS: The LECL results showed that the BoNTA group had significantly lower superoxide production compared to controls, with notable reductions at 4 hours. While no significant differences were noted at the 0 and 1-hour marks, the 4-hour mark showed significant protective effects in BoNTA-treated groups. The survival rate was 90% for BoNTA-treated rats versus 60% for controls ( P = 0.028). Significant reductions in ROS were also observed in the 4-hour I/R group. CONCLUSIONS: BoNTA effectively protects against I/R injury by inhibiting the NADPH oxidase system and reducing ROS levels. These results support further investigation into the specific mechanisms of NADPH oxidase inhibition by BoNTA and its potential clinical applications, given its safety profile. CLINICAL RELEVANCE STATEMENT: The findings from the present study are expected to provide a basis for clinical studies regarding this use of BoNTA.

Systematic review of the histological and functional effects of botulinum toxin A on masticatory muscles: Consideration in dentofacial orthopedics and orthognathic surgery.

INTRODUCTION: Botulinum toxin type A causes muscle paralysis and is widely used in the masticatory muscle for stomatognathic diseases, such as temporomandibular disorder, bruxism, or masseteric hypertrophy. Nonetheless, its muscular effect remains unclear. Better understanding could aid improved use and perhaps new indications, particularly in dentofacial orthopaedics and orthognathic surgery. METHODS: This systematic review explored the histologic and functional effects of botulinum toxin in animal and human masticatory muscles and was conducted in accordance with the recommendations of the Preferred Reporting Items for Systematic Reviews and Meta-Analyses statement. The MEDLINE, Web of Science, and Cochrane Library electronic databases were searched for relevant articles. The inclusion criteria were human or animal masticatory muscle analysis after botulinum toxin injection(s) AND histological structural/ultrastructural analysis by optical or electronic microscopy OR functional effect analysis by bite force evaluation (occlusal force analyzer) and muscle activity (electromyography). RESULTS: Of an initial 1578 articles, 44 studies were eventually included. Botulinum toxin injection in the masticatory muscle altered its histological structure and functional properties. The human and animal studies revealed ultrastructural change, atrophy, and fiber type modifications of the masticatory muscles after one injection. Botulinum toxin decreased bite force and muscle activity, but recovery was uncertain. CONCLUSIONS: Muscle forces applied on the skeleton is a key feature of facial growth. Masticatory muscle paralysis changes mechanical stress on bones, which rebalances the force applied on facial bones. This new balance could benefit dental deformity or surgical relapse. Therefore, botulinum toxin could limit the orthognathic effect of the masticatory muscles in such patients. Given the uncertain recovery, multiple injections should be avoided, and usage should not deviate from established consensus.

The Role of Botulinum Neurotoxin A in the Conservative Treatment of Fractures: An Experimental Study on Rats.

This paper explores the role of botulinum neurotoxin in aiding fracture recovery through temporary muscle paralysis. Specifically, it investigates the effects of botulinum neurotoxin-induced paralysis of the sternocleidomastoid muscle on clavicle fractures in rats. The research aims to assess safety, effectiveness, and the impact on fracture healing. Healthy male Albino Wistar rats were divided into four groups: clavicle fracture, botulinum neurotoxin injection, both, and control. Surgeries were conducted under anaesthesia, and postoperatively, animals were monitored for 28 days. Euthanasia and radiological assessment followed, examining fracture healing and muscle changes, while tissues were histopathologically evaluated. The modified Lane-Sandhu scoring system was used for the radiographic evaluation of clavicle fractures, and the results varied from complete healing to nonunion. Histopathological examination at 28 days postfracture showed fibrous tissue, mesenchymal cells, and primary callus formation in all groups. Despite varied callus compositions, botulinum neurotoxin administration did not affect clavicle healing, as evidenced by similar scores to the control group. Several studies have explored botulinum neurotoxin applications in fracture recovery. Research suggests its potential to enhance functional recovery in certain types of fractures. Theoretical benefits include managing muscle spasticity, aiding reduction techniques, and preventing nonunion. However, botulinum neurotoxin's transient effect and nonuniversal applications should be considered. The present study found that botulinum toxin had no clear superiority in healing compared to controls, while histological evaluation showed potential adverse effects on muscle tissue. Further research is essential to understand its risk-benefit balance and long-term effects.

Botulinum toxin type A is a potential therapeutic drug for chronic orofacial pain.

BACKGROUND: Botulinum toxin type A (BTX-A), produced by the gram-positive anaerobic bacterium Clostridium botulinum, acts by cleaving synaptosome-associated protein-25 (SNAP-25), an essential component of the presynaptic neuronal membrane that is necessary for fusion with the membrane proteins of neurotransmitter-containing vesicles. Recent studies have highlighted the efficacy of BTX-A in treating chronic pain conditions, including lower back pain, chronic neck pain, neuropathic pain, and trigeminal neuralgia, particularly when patients are unresponsive to traditional painkillers. This review focuses on the analgesic effects of BTX-A in various chronic pain conditions, with a particular emphasis on the orofacial region. HIGHLIGHT: This review focuses on the mechanisms by which BTX-A induces analgesia in patients with inflammatory and temporomandibular joint pain. This review also highlights the fact that BTX-A can effectively manage neuropathic pain and trigeminal neuralgia, which are difficult-to-treat chronic pain conditions. Herein, we present a comprehensive assessment of the central analgesic effects of BTX-A and a discussion of its various applications in clinical dental practice. CONCLUSION: BTX-A is an approved treatment option for various chronic pain conditions. Although there is evidence of axonal transport of BTX-A from peripheral to central endings in motor neurons, the precise mechanism underlying its pain-modulating effects remains unclear. This review discusses the evidence supporting the effectiveness of BTX-A in controlling chronic pain conditions in the orofacial region. BTX-A is a promising therapeutic agent for treating pain conditions that do not respond to conventional analgesics.

Mobile digital gait analysis captures effects of botulinum toxin in hereditary spastic paraplegia.

BACKGROUND AND PURPOSE: Hereditary spastic paraplegias (HSPs) comprise a group of inherited neurodegenerative disorders characterized by progressive spasticity and weakness. Botulinum toxin has been approved for lower limb spasticity following stroke and cerebral palsy, but its effects in HSPs remain underexplored. We aimed to characterize the effects of botulinum toxin on clinical, gait, and patient-reported outcomes in HSP patients and explore the potential of mobile digital gait analysis to monitor treatment effects and predict treatment response. METHODS: We conducted a prospective, observational, multicenter study involving ambulatory HSP patients treated with botulinum toxin tailored to individual goals. Comparing data at baseline, after 1 month, and after 3 months, treatment response was assessed using clinical parameters, goal attainment scaling, and mobile digital gait analysis. Machine learning algorithms were used for predicting individual goal attainment based on baseline parameters. RESULTS: A total of 56 patients were enrolled. Despite the heterogeneity of treatment goals and targeted muscles, botulinum toxin led to a significant improvement in specific clinical parameters and an improvement in specific gait characteristics, peaking at the 1-month and declining by the 3-month follow-up. Significant correlations were identified between gait parameters and clinical scores. With a mean balanced accuracy of 66%, machine learning algorithms identified important denominators to predict treatment response. CONCLUSIONS: Our study provides evidence supporting the beneficial effects of botulinum toxin in HSP when applied according to individual treatment goals. The use of mobile digital gait analysis and machine learning represents a novel approach for monitoring treatment effects and predicting treatment response.

Early effect of onabotulinumtoxinA on EEG-based functional connectivity in patients with chronic migraine: A pilot study.

OBJECTIVE: In this pilot prospective cohort study, we aimed to evaluate, using high-density electroencephalography (HD-EEG), the longitudinal changes in functional connectivity (FC) in patients with chronic migraine (CM) treated with onabotulinumtoxinA (OBTA). BACKGROUND: OBTA is a treatment for CM. Several studies have shown the modulatory action of OBTA on the central nervous system; however, research on migraine is limited. METHODS: This study was conducted at the Neurology Unit of "Policlinico Tor Vergata," Rome, Italy, and included 12 adult patients with CM treated with OBTA and 15 healthy controls (HC). Patients underwent clinical scales at enrollment (T0) and 3 months (T1) from the start of treatment. HD-EEG was recorded using a 64-channel system in patients with CM at T0 and T1. A source reconstruction method was used to identify brain activity. FC in δ-θ-α-ß-low-γ bands was analyzed using the weighted phase-lag index. FC changes between HCs and CM at T0 and T1 were assessed using cross-validation methods to estimate the results' reliability. RESULTS: Compared to HCs at T0, patients with CM showed hyperconnected networks in δ (p = 0.046, area under the receiver operating characteristic curve [AUC: 0.76-0.98], Cohen's κ [0.65-0.93]) and ß (p = 0.031, AUC [0.68-0.95], Cohen's κ [0.51-0.84]), mainly involving orbitofrontal, occipital, temporal pole and orbitofrontal, superior temporal, occipital, cingulate areas, and hypoconnected networks in α band (p = 0.029, AUC [0.80-0.99], Cohen's κ [0.42-0.77]), predominantly involving cingulate, temporal pole, and precuneus. Patients with CM at T1, compared to T0, showed hypoconnected networks in δ band (p = 0.032, AUC [0.73-0.99], Cohen's κ [0.53-0.90]) and hyperconnected networks in α band (p = 0.048, AUC [0.58-0.93], Cohen's κ [0.37-0.78]), involving the sensorimotor, orbitofrontal, cingulate, and temporal cortex. CONCLUSION: These preliminary results showed that patients with CM presented disrupted EEG-FC compared to controls restored by a single session of OBTA treatment, suggesting a primary central modulatory action of OBTA.

Botulinum Toxin-A for the Treatment of Myogenous Temporomandibular Disorders: An Umbrella Review of Systematic Reviews.

OBJECTIVE: Temporomandibular disorders (TMDs) encompass several conditions that cause pain and impair function of the masticatory muscles (M-TMDs) and temporomandibular joints. There is a large interest among clinicians and researchers in the use of botulinum toxin-A (BoNT-A) as a treatment for M-TMD. However, due to the lack of consistent evidence regarding the efficacy as well as adverse events of BoNT-A, clinical decision making is challenging. Therefore, this umbrella review aimed to systematically assess systematic reviews (SRs) evaluating BoNT-A treatment effects on pain intensity, mandibular movements, and adverse events in patients with M-TMDs. METHOD: An electronic search was undertaken in the databases MEDLINE, EMBASE, CINAHL, Cochrane Central Registry of Controlled Trials (CENTRAL), Web of Science, Epistemonikos, ClinicalTrials.gov, and ICTRP to identify SRs investigating BoNT-A effects on M-TMDs, published from the inception of each database until 6 December 2023. The quality of evidence was rated according to the critical appraisal checklist developed by the umbrella review methodology working group. Only high-quality SRs were included. RESULTS: In total, 18 SRs were included. BoNT-A was shown to be more effective than placebo to reduce pain intensity, but not compared to standard treatments. Additionally, BoNT-A was not superior to placebo or standard treatments regarding improvement of mandibular movements. BoNT-A was considered to have a higher risk for adverse events on muscle and bony tissue compared with other treatments. CONCLUSION: The synthesis in this umbrella review provides the highest level of evidence present. Taken together, there are indications of effectiveness of BoNT-A for treatment of M-TMDs, supported by moderate evidence. However, considering the risk of causing serious adverse events, treatment with BoNT-A is recommended to be the last treatment alternative.

A Validation of the Equivalence of the Cell-Based Potency Assay Method with a Mouse LD50 Bioassay for the Potency Testing of OnabotulinumtoxinA.

(1) Background: At present, the only potency assay approved in China for the in-country testing of botulinum toxin type A for injection products is the mouse bioassay (MBA). The Chinese market for neurotoxin products is rapidly expanding, but MBAs are subject to high variability due to individual variations in mice, as well as variations in injection sites, in addition to the limited number of batches tested for one MBA. Compared with the mLD50 method, the cell-based potency assay (CBPA) developed for the potency testing of onabotulinumtoxinA (BOTOX) by AbbVie not only does not use any experimental animals but also allows for significant time and cost savings. Due to the significant benefits conferred by the replacement of the mLD50 assay with CBPA in China, the CBPA method has been transferred, validated, and cross-validated to demonstrate the equivalence of the two potency methods. (2) Methods: The differentiated SiMa cells were treated with both BOTOX samples and the reference standard, and the cleaved SNAP25197 in the cell lysates was quantified using Chemi-ECL ELISA. A 4-PL model was used for the data fit and sample relative potency calculation. The method accuracy, linearity, repeatability, and intermediate precision were determined within the range of 50% to 200% of the labeled claim. A statistical equivalence of the two potency methods (CBPA and mLD50) was initially demonstrated by comparing the AbbVie CBPA data with NIFDC mLD50 data on a total of 167 commercial BOTOX lots (85 50U lots and 82 100U lots). In addition, six lots of onabotulinumtoxinA (three 50U and three 100U) were re-tested as cross-validation by these two methods for equivalence. (3) Results: The overall assay's accuracy and intermediate precision were determined as 104% and 9.2%, and the slope, R-square, and Y-intercept for linearity were determined as 1.071, 0.998, and 0.036, respectively. The repeatability was determined as 6.9%. The range with the acceptable criteria of accuracy, linearity, and precision was demonstrated as 50% to 200% of the labeled claim. The 95% equivalence statistic test using margins [80%, 125%] indicates that CBPA and mLD50 methods are equivalent for both BOTOX strengths (i.e., 50U and 100U). The relative potency data from cross-validation were within the range of ≥80% to ≤120%. (4) Conclusions: The CBPA meets all acceptance criteria and is equivalent to mLD50. The replacement of mLD50 with CBPA is well justified in terms of ensuring safety and efficacy, as well as for animal benefits.

Transition of endochondral bone formation at the normal and botulinum-treated mandibular condyle of growing juvenile rat.

OBJECTIVE: The aim of this study was to understand the temporal and spatial distribution of canonical endochondral ossification (CEO) and non-canonical endochondral ossification (NCEO) of the normal growing rat condyle, and to evaluate their histomorphological changes following the simultaneous hypotrophy of the unilateral masticatory closing muscles with botulinum toxin (BTX). DESIGN: 46 rats at postnatal 4 weeks were used for the experiment and euthanized at postnatal 4, 8, and 16 weeks. The right masticatory muscles of rats in experimental group were injected with BTX, the left being injected with saline as a control. The samples were evaluated using 3D morphometric, histological, and immunohistochemical analysis with three-dimensional regional mapping of endochondral ossifications. RESULTS: The results showed that condylar endochondral ossification changed from CEO to NCEO at the main articulating surface during the experimental period and that the BTX-treated condyle presented a retroclined smaller condyle with an anteriorly-shifted narrower articulating surface. This articulating region showed a thinner layer of the endochondral cells, and a compact distribution of flattened cells. These were related to the load concentration, decreased cellular proliferation with thin cellular layers, reduced extracellular matrix, increased cellular differentiation toward the osteoblastic bone formation, and accelerated transition of the ossification types from CEO to NCEO. CONCLUSION: The results suggest that endochondral ossification under loading tended to show more NCEO, and that masticatory muscular hypofunction by BTX had deleterious effects on endochondral bone formation and changed the condylar growth vector, resulting in a retroclined, smaller, asymmetrical, and deformed condyle with thin cartilage.

The impact of the migraine treatment onabotulinumtoxinA on inflammatory and pain responses: Insights from an animal model.

OBJECTIVE: Migraine, a prevalent and debilitating disease, involves complex pathophysiology possibly including inflammation and heightened pain sensitivity. The current study utilized the complete Freund's adjuvant (CFA) model of inflammation, with onabotulinumtoxinA (BoNT/A) as a treatment of interest due to its use in clinical migraine management. Using an animal model, the study sought to investigate the role of BoNT/A in modulating CFA-induced inflammation, alterations in pain sensitivity, and the regulation of calcitonin gene-related peptide (CGRP) release. Further, we aimed to assess the changes in SNAP-25 through western blot analysis to gain insights into the mechanistic action of BoNT/A. METHODS: BoNT/A or control was administered subcutaneously at the periorbital region of rats 3 days before the induction of inflammation using CFA. Periorbital mechanical sensitivity was assessed post-inflammation, and alterations in CGRP release were evaluated. Changes in SNAP-25 levels were determined using western blot analysis. RESULTS: Upon CFA-induced inflammation, there was a marked increase in periorbital mechanical sensitivity, with the inflammation side showing increased sensitivity compared to other periorbital areas. BoNT/A did decrease the withdrawal thresholds in the electronic von Frey test. Despite not being able to observe differences in pain thresholds or CGRP release, BoNT/A reduced baseline release under CFA inflamed conditions. Analysis of SNAP-25 levels in the trigeminal ganglion revealed both intact and cleaved forms that were notably elevated in BoNT/A-treated animals. These findings, derived from western blot analysis, suggest an effect on neurotransmitter release. CONCLUSION: Our investigation highlights the role of BoNT/A in reducing baseline CGRP in the context of inflammation and its involvement in SNAP-25 cleavage. In contrast, BoNT/A did not appear to alter facial pain sensitivity induced by inflammation, suggesting that mechanisms other than baseline CGRP could be implicated in the elevated thresholds in the CFA model.

Sequential administration of peripheral nerve blocks and onabotulinumtoxinA for the treatment of chronic migraine and other headache disorders-A retrospective tolerability and safety study.

OBJECTIVE: To determine the tolerability and safety of concurrent peripheral nerve blocks and onabotulinumtoxinA treatment during a single outpatient clinic procedure visit. BACKGROUND: Procedural interventions are available for the treatment of headache disorders. OnabotulinumtoxinA and peripheral nerve blocks are used as alternatives or in addition to oral therapies to reduce the frequency and intensity of migraine attacks. There is currently a lack of safety data focusing on the sequential administration of local anesthetic via peripheral nerve blocks and onabotulinumtoxinA during a single clinical encounter for the treatment of headache. The primary aim of the study was to determine the safety and tolerability of concurrent peripheral nerve blockade and onabotulinumtoxinA injections during a single outpatient clinic procedure visit. We hypothesized that the dual intervention would be safe and well tolerated by patients with chronic migraine and other headache disorders. METHODS: A retrospective chart review was performed using clinical data from patients seen by multiple providers over a 16-month timeframe at one outpatient headache clinic. Patients were identified by procedure codes and those receiving peripheral nerve block(s) and onabotulinumtoxinA injections during a single encounter within the study period were eligible for inclusion. Inclusion criteria were (1) patients 18 years and older who were (2) receiving both peripheral nerve blocks and onabotulinumtoxinA injections for the treatment of chronic migraine. Patients were excluded if they were under age 18, received their procedure outside of the clinic (emergency room, inpatient ward), or were receiving sphenopalatine ganglion blocks. Age- and sex-matched patients who received one procedure, either peripheral nerve blocks or onabotulinumtoxinA, were used for control. The primary outcome of this safety study was the number of adverse events that occurred in the dual intervention group compared to the single intervention control arms. Information regarding adverse events was gathered via retrospective chart review. If an adverse event was recorded, it was then graded by the reviewer utilizing the Common Terminology Criteria for Adverse Events ranging from Grade 1 Mild Event to Grade 5 Death. Additionally, it was noted whether the adverse event led to treatment discontinuation. RESULTS: In total, 375 patients were considered eligible for inclusion in the study. After age and sex matching of controls, 131 patients receiving dual intervention were able to be compared to 131 patients receiving onabotulinumtoxinA alone and 104 patients receiving dual intervention were able to be compared to 104 patients receiving peripheral nerve block(s) alone. The primary endpoint analysis showed no significant difference in total adverse events between dual intervention compared to nerve blocks alone or onabotulinumtoxinA alone. The number of adverse events that led to treatment discontinuation approached but did not reach statistical significance for those receiving dual intervention versus onabotulinumtoxinA alone in the number of adverse events that led to treatment termination (4.6%, 6/131 vs. 0.8%, 1/131, p = 0.065); however, the number of patients who discontinued therapy was not significantly different between those groups (2.3%, 3/131 vs. 0.8%, 1/131; p = 0.314; odds ratio 0.3 [0-3.2]; p = 0.338). CONCLUSIONS: In this retrospective chart review, there was no significant difference in adverse events or therapy discontinuation between patients receiving sequential peripheral nerve block(s) and onabotulinumtoxinA injections versus those receiving either peripheral nerve block(s) or onabotulinumtoxinA injections alone. As a result, we concluded that the combination procedure is likely safe and well tolerated in routine clinical practice.

Botulinum toxin A attenuates osteoarthritis development via inhibiting chondrocyte ferroptosis through SLC7Al1/GPX4 axis.

Osteoarthritis (OA) is a prevalent joint degenerative disease, resulting in a significant societal burden. However, there is currently a lack of effective treatment option available. Previous studies have suggested that Botulinum toxin A (BONT/A), a macromolecular protein extracted from Clostridium Botulinum, may improve the pain and joint function in OA patients, but the mechanism remains elusive. This study was to investigate the impact and potential mechanism of BONT/A on OA in vivo and in vitro experiment. LPS increased the levels of ROS, Fe2+and Fe3+, as well as decreased GSH levels, the ratio of GSH / GSSH and mitochondrial membrane potential. It also enhanced the degeneration of extracellular matrix (ECM) and altered the ferroptosis-related protein expression in chondrocytes. BONT/A rescued LPS-induced decrease in collagen type II (Collagen II) expression and increase in matrix metalloproteinase 13 (MMP13), mitigated LPS-induced cytotoxicity in chondrocytes, abolished the accumulation of ROS and iron, upregulated GSH and the ratio of GSH/ GSSH, improved mitochondrial function, and promoted SLC7A11/GPX4 anti-ferroptosis system activation. Additionally, intra-articular injection of BONT/A inhibited the degradation of cartilage in OA model rats. This chondroprotective effect of BONT/A was reversed by erastin (a classical ferroptosis agonist) and enhanced by liproxstatin-1 (a classic ferroptosis inhibitor). Our research confirms that BONT/A alleviates the OA development by inhibiting the ferroptosis of chondrocytes, which revealed to be a potential therapeutic mechanism for BONT/A treating the OA.

Onabotulinum toxin A improves neurogenic detrusor overactivity following spinal cord injury: a systematic review and meta-analysis.

STUDY DESIGN: Systematic review and meta-analysis. OBJECTIVES: The current study aimed to assess the efficacy and safety of Onabotulinum toxin A (OBTX-A) treatment for neurogenic detrusor overactivity (NDO) in spinal cord injury (SCI) patients. SETTING: Iran. METHODS: All relevant articles of clinical trials and cohort studies indexed in PubMed/MEDLINE, Embase, Scopus, and Web of Science databases up to September 6, 2022, that addressed OBTX-A treatment for NDO following SCI were included. The quality of eligible studies was evaluated using Cochrane criteria. Also, the weighted mean difference (WMD) was measured with a random-effect model. RESULTS: Regarding the overall efficacy after OBTX-A treatment in the short term, volume per void (VV) (WMD = 118.8, 95% CI: 90.9-146.7, p < 0.01), incontinence-quality of life (IQoL) (WMD = 24.3, 95% CI: 15.8-32.8, p < 0.01), and maximum cystometric capacity (MCC) (WMD = 144.5, 95% CI: 132.3 to 156.7, p < 0.01) significantly increased, while maximum detrusor pressure during storage (MDP) (WMD = -30.5, 95% CI: -35.9 to -25.1, p < 0.01) showed a significant decrease. Furthermore, compared to the placebo group at the 200-unit dose, there was a significant increase in MCC (WMD = 113.5, 95% CI: 84.7 to 142.3, p < 0.01) and a significant decrease in MDP (WMD = -27.2, 95% CI: -39.2 to -15.1, p < 0.01). Urinary tract infection (UTI), hematuria, and autonomic dysreflexia were the most common side effects, occurring at rates of 29.6%, 14.8%, and 13.4%, respectively. CONCLUSION: Our findings highlighted the effectiveness and safety of OBTX-A as a promising treatment of NDO following SCI.

Synergism of Anti-CGRP Monoclonal Antibodies and OnabotulinumtoxinA in the Treatment of Chronic Migraine: A Real-World Retrospective Chart Review.

BACKGROUND: Many patients with chronic migraine do not achieve clinically meaningful improvement in their headache frequency with monotherapy. The burden associated with chronic migraine calls for a multifaceted treatment approach targeting multiple aspects of migraine pathophysiology. OBJECTIVE: The aim of this study was to evaluate the effect of concurrent anti-calcitonin gene-related peptide (CGRP) monoclonal antibody (mAb) and onabotulinumtoxinA (onabot) treatment on median monthly migraine days (MMD) in patients with chronic migraine, through a retrospective study. METHODS: The electronic medical records of Cleveland Clinic patients either concurrently (dual therapy) or consecutively (monotherapy) treated with anti-CGRP mAbs and onabot between June 2018 and November 2021 were extracted. Only adult patients (≥ 18 years of age) were included in this study. MMDs for 194 concurrently treated (86.6% female and a median [interquartile range] age of 51 [41-61] years) and 229 consecutively treated (88.2% female and median age of 47 [IQR 39-57] years) patients were examined at baseline, after first therapy of either anti-CGRP mAb or onabot, and following dual therapy for 3 consecutive months. The reduction of MMDs for each treatment group were compared. The same approach was utilized to compare consecutive monotherapy at separate times (n = 229) and dual-therapy groups. RESULTS: The initial treatment of the dual-therapy group reduced the median (IQR) MMDs from 30 (30-30) to 15 (12-30) [p < 0.0001]. After initiation of dual therapy, the median MMDs was further decreased from 15 (12-30) to 8 (3-22) [p < 0.0001]. A majority [132/194 (68.0%)] of the dual-therapy patients reported a ≥ 50% reduction in MMD and 90/194 (46.4%) reported a ≥ 75% reduction. For the consecutive monotherapy group, median MMDs changed from a baseline of 30 (25-30) to 15 (8-25) from onabot monotherapy and decreased from 25 (15-30) to 12 (4-25) after anti-CGRP mAb monotherapy. Almost half (113/229 [49.3%] from onabot, and 104/229 [45.4%] from anti-CGRP mAb) of these patients achieved a ≥ 50% reduction in MMDs and a minority (38/229 [16.6%] from onabot, and 45/229 [19.7%] from anti-CGRP mAb) achieved a reduction of ≥ 75%. Additionally, dual therapy showed significant improvement in MMDs compared with monotherapy of either treatment (p < 0.0001). CONCLUSION: Dual therapy of anti-CGRP mAbs and onabot may be more efficacious than monotherapy, possibly due to their synergistic mechanisms of action.