Amytrapper Catheter: A Prototype Extracorporeal Device That Traps Blood Amyloid-β in a Rat Model of Alzheimer’s Disease

Abstract

Alzheimer’s disease (AD) presents a pressing global health challenge, with amyloid-β (Aβ) accumulation in the brain being a hallmark feature. While monoclonal antibodies targeting Aβ have shown cognitive benefits, safety concerns remain. Here, we introduce the Amytrapper catheter, a novel extracorporeal device developed by Recombinant Technologies to trap circulating Aβ using a retro-inverso peptide conjugated to polyethylene glycol. Through in vivo experiments using a rat model of AD, we demonstrate significant reductions in blood Aβ levels and behavioral improvements following Amytrapper catheter treatment. This innovative approach holds promise as a disease-modifying therapy for AD, offering a complementary strategy to existing treatments and advocating for further clinical development.

Keywords

amyloid Alzheimer’s disease Alzheimer’s therapy plasmapheresis amyloid β-peptides Amytrapper catheter extracorporeal device amyloid removal

Introduction

Alzheimer’s disease (AD) represents a profound global health challenge and is the most prevalent cause of dementia in older adults, affecting about 6.7 million people in the US with a projected increase to 13.8 million by 2060 in the absence of timely development of medical breakthroughs to prevent, slow or cure AD.^1,2 According to the Centers for Disease Control and Prevention, USA, AD is the seventh most common cause of death claiming even more people than diabetes.³ Owing to the increased life expectancy of people and overall aging population, the incidence of AD is expected to double over the next 20 years leading to increased treatment costs.⁴

Amyloid-β (Aβ) is widely recognized as a pivotal factor in both the initiation and progression of AD.⁵ Recent FDA approvals of several monoclonal antibodies such as aducanumab (Aduhelm, Biogen), lecanemab (Eisai), and donanemab (Eli Lilly), which demonstrated cognitive benefits alongside reductions in brain Aβ burden, signal a promising shift towards disease-modifying therapies targeting Aβ.⁶ However, several experts and new meta-analysis of the clinical trials conducted for these antibodies agree that the risks of these monoclonal antibody treatments for people with AD might outweigh the benefits.⁷ For example, aducanumab and donanemab treatment was associated with the increase of amyloid-related imaging abnormalities with microhemorrhages in patients, which are characterized by microhemorrhages and superficial siderosis.⁷ Amyloid-related imaging abnormalities characterized by edema which displays symptoms such as headache, vomiting, and confusion, were also significantly increased in patients who received lacenumab and donanemab.⁷

Moreover, the intricate regulation of Aβ concentration within the brain involves a dynamic interplay between influx/efflux mechanisms across the blood-brain barrier, mediated by receptors such as the receptor for advanced glycation end products and the low-density lipoprotein receptor-related protein-1.^8,9 In AD patients, dysregulation of these mechanisms impedes Aβ clearance from the brain, exacerbating its accumulation. Emerging evidence supports the peripheral Aβ clearance hypothesis, which posits that modulating Aβ levels in the bloodstream can influence brain Aβ levels, thereby presenting a potential avenue for therapeutic intervention.^10-12 Indeed, studies in animal models and clinical trials have demonstrated cognitive improvements following peripheral Aβ clearance strategies, underscoring the importance of investigating novel amyloid-binding compounds and procedures.^10-12

Recombinant Technologies has been a pioneer in the development of non-immune, peptide-based therapeutics for AD.^13-16 Our innovative approach, exemplified by the Amytrapper catheter, offers a novel strategy for peripheral Aβ clearance by utilizing a retro-inverso peptide conjugated to polyethylene glycol (PEG) (the MW of PEG conjugate is 5120) to trap circulating Aβ. This treatment modality holds promise for improving the lives of AD patients and complements existing therapeutic regimens. In this study, we assessed the efficacy of the Amytrapper catheter in trapping Aβ in vivo through a rat model of Alzheimer’s disease and showed blood amyloid reduction coupled with behavioral improvement. Through the experimental analysis presented here, we aim to advance the development of an effective treatment for AD, projecting Amytrapper catheter as a beneficial extracorporeal device to deplete circulating amyloid with the ultimate goal of using this clinically viable design as a disease-modifying agent to provide clinical benefits to patients.

Materials and Methods

Rat AD Model

Transgenic rats used for the study belonged to the McGill-R-Thy1-APP line with the genetic background of HsdBrl:WH Wistar strain rats. These rats contain a transgenic construct with the cDNA coding for the human amyloid precursor protein (hAPP751) under the control of the murine Thy1.2 promoter. This transgenic construct carries both the Swedish and Indiana FAD mutations in the cDNA coding for hAPP751. The homozygous McGill-R-Thy1-APP rats display an extended phase of intracellular Aβ accumulation and a progressive extracellular Aβ deposition in the form of amyloid plaques. Transgenic rats homozygous for the mutated hAPP751 transgene and their wild-type littermates were used in this study. Behavioral studies show that spatial cognitive impairments appear as early as 3 months of age in these transgenic rats, coinciding with the pathological early AD intracellular accumulation of Aβ oligomers.¹⁷ The use of these transgenic rats was conducted in accordance with the original Material Transfer Agreement established between Recombinant Technologies and McGill University, located in Montreal, Canada, which holds a patent on this model. In total, there were 24 transgenic rats equally distributed between genders and randomized for two study groups (Amytrapper treatment group and the control catheter treatment group).

Testing of the Amytrapper Catheter in Vivo

Generation of catheter coated with RI-Mal-PEG4 conjugated peptide has been described in our previous publication.¹³ Uncoated catheters (without conjugated peptide) served as the control catheter. The procedure involved the utilization of these transgenic rats equipped with Vascular Access Buttons (VAB, Instech Lab, PA). These VABs served as blood passing ports essential for the extracorporeal procedure. At least 2 weeks prior to treatment rats underwent surgery for VAB implantation. The surgical implantation of the VABs was conducted at the breeding facility (Taconic Biosciences, Germantown, NY). Subsequent device testing on the rats was carried out at the Oklahoma City VA Health Care System, Oklahoma City, Oklahoma. Briefly, anaesthetized rats were connected to the device (Amytrapper catheter or control catheter) (Figure 1) and blood was pumped through the catheter with a peristaltic pump along with an injection of heparin (1200 I/U per kg) to prevent clotting.¹⁸ The procedure was run for 45 min every 10 days for 2 months. Rat handling and care procedures were followed according to recommendations and approval of Oklahoma City VA Health Care System Institutional Animal Care and Use Committee.

Figure 1.

In vivo Experimental Setup for Testing the Amytrapper Catheter in a McGill-R-Thy1-APP Alzheimer’s Disease Rat Model. A Vascular Access Button (VAB) is Surgically Implanted in the Rat to Facilitate Blood Flow During the Extracorporeal Procedure. Blood is Drawn From the Animal via a Peristaltic Pump, Passed Through the Amytrapper Catheter for Aβ Capture, and Returned to the Rat. The Rat’s Temperature and Other Vital Signs are Closely Monitored Throughout the Process

Estimation of Aβ42 Bound to Catheters

After extracorporeal process on AD rats, Amytrapper catheters (Retro-inverso peptide coated catheter) and uncoated control catheters were prepared for estimation of catheter bound Aβ. To minimize nonspecific binding, all catheters were blocked overnight at 4°C with a 1% BSA in PBS solution (pH 7.4, Sigma). Following this, catheters were washed 5 times with TBST and then incubated with a mouse monoclonal anti-Aβ 6E10 antibody (1:2000, Bio Legend, San Diego CA) for 1 h at 37°C. Another TBST washing step (5 times) was performed before incubating the catheters with a secondary goat anti-mouse HRP antibody (1:5000, Invitrogen, MA) for 1 hour at 37°C. A final TBST wash (5 times) ensured the removal of any unbound secondary antibody. Catheters were incubated with 350 μL of SureBlue TMB substrate (SeraCare Life Science, Milford, MA) for 5-10 min at room temperature. The substrate solution was then collected into separate microcentrifuge tubes, and 350 μL of 1N HCl was added to stop the reaction. Finally, 100 μL from each tube was transferred to a 96-well clear plate and read at 450 nm on the Elx800 plate reader (Agilent, Santa Clara, CA). The absorbance reading provides a measure of the amount of bound Aβ42 on each catheter.

Estimation of Aβ42 in Plasma and CSF by ELISA

SensoLyte anti-human Aβ42 quantitative ELISA colorimetric kit (Anaspec, Fremont, CA) was used to estimate the amount of Aβ42 in plasma and CSF in rats. Instructions described in the kit were followed. The human Aβ42 standard was reconstituted with 1 mL peptide standard reconstitution buffer and was aliquoted 100 μL per vial. Serial dilution of the human Aβ42 standard was done to get 0-500 pg/mL by diluting the standard solution (originally 1 μg/mL) in sample dilution buffer. Detection antibody was diluted 200-fold in the detection antibody dilution buffer. Fifty μl of the above antibody solution for each well to be run in the assay was prepared. The mouse monoclonal anti-Aβ42 coated plate which came with the kit was labelled for duplicate samples and standards. One hundred μl per well of the diluted Aβ42 standard in duplicate including blank, was loaded onto the wells. Rat plasma and CSF samples were diluted 1:20 and 1:4 respectively with 1X sample dilution buffer to avoid sample matrix effects. Protease inhibitor cocktail with phenylmethylsulfonyl fluoride was added to all samples to avoid protein degradation (instructions in the kit were followed). Diluted standards and samples were incubated in plate overnight at 4°C. After incubation and washing, 100 μL of the TMB substrate solution was added into each well. The plate was tapped gently and incubated at room temperature for 10-15 min. Fifty μl of the stop solution was added into each well and the plate tapped gently. The absorbance (OD) at 450 nm was measured. A calibration curve using linear regression curve-fitting was plotted for the standards and utilized to determine the concentration of human Aβ42 in the tested samples.

Y-Maze Test

The apparatus is a Y-shaped maze made of black Plexiglas with three arms terminating (A, B, and C arm) in small chambers, with equal angles between all arms.^19-21 The animal was initially placed in one arm of the chamber and allowed to move freely around the apparatus, while the software recorded the sequence and number of arm entries for each rat once per minute for 6 min. This test is designed to estimate the short-term memory of the last arms entered. The percentage of triads in which all three arms are represented was recorded as an alternation, which means testing successive rat entries into three arms per triad. For instance, the following arms entries in triads are recorded as alternation: ABC, CAB, BCA, etc. The entries which are not an alternation include ABA, ACA, BAB, BCB, CAC, etc. The total number of possible alternations is the number of arm entries minus two. The percent alternation score was calculated as a ratio of the actual number of alternations to the total number of possible alternations and multiplied by 100.^19-21

Fear Conditioning

Pavlovian fear conditioning is a highly efficient way to study learning and memory in rats.^22,23 In this study, we used cue-plus-contextual fear conditioning design. Rats were placed in a fear conditioning apparatus for about 2 min, then a 30 s acoustic tone was delivered. During the last 2 s of the tone, a mild foot shock was delivered through the floor grid of the apparatus. This pairing protocol was repeated two more times with each tone-shock pairing separated by a period of 2 min. Video was recorded and the software scored freezing behavior. When trained in this fashion, the rat learns the training chamber (the context) is a place to be feared. This component of the learning is referred to as contextual fear conditioning. Twenty-four hours after training, rats were placed in the same chambers. Freezing behavior during the first 5 min in the chamber was monitored as a measure of contextual recall of the aversive stimulus (shock). Subsequently, the same 30 s tone was delivered but no foot shock was performed. Freezing behavior in the 30 s after delivery of the conditioned stimulus (tone) was recorded as a measure of associative recall. All measures were compared between treated, untreated and wild type groups.

Semi Quantitative Assessment of Aβ by Immunohistochemistry on Stained Brain Sections from AD Rats

Digital slides consisting of brain sections stained with Aβ immunohistochemical staining and 3,3′-diaminobenzidine (DAB) were analyzed. The sections consisted of a sagittal cut of the brain and a semi-quantitative evaluation of the distribution of Aβ was performed. The presence and distribution of Aβ plaques was assessed in samples from 17 transgenic rats and 2 wild-type rats. Entire sagittal brain sections were stained with Aβ immunohistochemical staining. The evaluation focused on the quantification of the amyloid plaques (extracellular). All areas of the brain were evaluated, and the neuroanatomical distribution was noted based on the Allen brain atlas. For eight samples, the distribution of the intraneuronal accumulation of Aβ was performed.

Results

Procedural Feasibility and Capturing of Aβ₄₂ From Whole Blood of AD Rat by Extracorporeal Amytrapper Catheter

Figure 1 depicts the in vivo experimental setup for testing the Amytrapper catheter in McGill-R-Thy1-APP Alzheimer’s model rats. These rats were implanted with VABs to facilitate blood circulation through the Amytrapper catheter and maintain long-term patency. The VAB’s closed-system design is the preferred method for repeated blood sampling procedures in rats, as it minimizes stress responses associated with traditional methods.²⁴

Analysis of catheters after treatment revealed bound Aβ42 levels from rat whole blood which was circulated during treatment. Compared to control uncoated (mock) catheters, Amytrapper coated catheters showed a 42% increase (P-value <0.001, Figure 2A) in Aβ42 content. Plasma Aβ42 levels from rats were measured before treatment, after the third treatment, and at the end of the study after sixth treatment (Figure 2B). A reduction in plasma Aβ42 levels and the increased bound Aβ42 observed on catheters strongly suggest a shift of Aβ42 from blood to catheters. Even after third treatment, plasma levels of Aβ42 decreased by around 50 pg/mL, reaching around 100 pg/mL. Importantly, this reduction aligns with the clinical significance of Amytrapper catheters, as circulating Aβ42 levels in human AD patients typically range from 350-400 pg/mL.²⁵ These findings suggest that the Amytrapper catheter effectively shifts Aβ42 from blood plasma to the device, potentially offering a robust amyloid removal system for AD treatment.

Figure 2.

Amytrapper Catheter Capture Aβ42 From Whole Blood in vivo in AD Rats. (A) Aβ42 From AD Rat Whole Blood Bound to Amytrapper Coated and Control Uncoated (Mock) Catheters at the End of the Treatment. After Treatment, Amytrapper Catheters and Mock Catheters Were Used for the Measurement of Catheter-Bound Aβ Through Designed Immunosorbent Assay. The Experiment Aimed to Determine the Effectiveness of Amytrapper-Coated Catheters in Capturing Aβ42 From the Bloodstream. Statistically Significant Difference Between the Mock Catheter (n = 20 Assay) and Amytrapper Catheter Treatment Groups (n = 41 Assay) is Expressed as ***P < 0.001, Cohen’s d = 3.496416, Utilizing Unpaired T-test. (B) Plasma Aβ42 From Rats before and after Catheter Treatments as Measured by Sensolyte® Quantitative ELISA kit. Plasma Aβ42 Levels before Mock Catheter (n = 7) or Amytrapper Catheter Treatment (n = 16) are Displayed as Blue Columns. Plasma Aβ42 Levels after third Mock Catheter (n = 6) or Amytrapper Catheter Treatment (n = 14) are Displayed as Yellow Columns. Plasma Aβ42 Levels after sixth Mock Catheter (n = 5) or Amytrapper Catheter Treatment (n = 6) are Displayed as Orange Columns. Statistically Significant Difference is Arrived by Comparing the Same Treatment (third Treatment and sixth Treatment) Between Mock Catheter Groups and Amytrapper Catheter Groups Using the Unpaired T-test. Statistically Significant Difference Between the Mock and Amytrapper Catheter Treatment Groups is Expressed (*P < 0.05, **P < 0.01 Utilizing Unpaired T-test)

Neurobehavioral Improvements in AD Rat Treated With Extracorporeal Amytrapper Catheter

The retention of amyloid by Amytrapper catheter was sufficient to induce behavioral changes in rats (n = 5/group) as measured by the Y-maze test. This test is a well-established method for assessing spatial working memory in rodents. In the Y-maze, rats are placed in a three-armed maze and allowed to freely explore. Their exploratory behavior is tracked, with a higher percentage of alterations in arm choices reflecting better spatial working memory. The results, expressed as percent alterations (Figure 3A), suggest that Amytrapper catheter treatment improved the spatial working memory of AD model rats compared to the mock catheter treatment group.

Figure 3.

Behavioral Improvements in AD Rats Treated With Amytrapper Catheter. (A) Y Maze Analyses of Rat Behavior (Expressed in Percent Alterations) Showing Wild Type (n = 5) as Baseline Control and AD Model Rats after Mock Catheter (n = 3) or Amytrapper Catheter (n = 3) Procedure. This Y-Maze Test is Designed to Estimate the Short-Term Memory Related to the Last Arms Entered, which Emphasizes that the Test Subject Does not Repeat Entering the Arm which was Already Entered Most Recently. The Percentage of Triads in which all Three Arms are Entered by the Testing Animal was Recognized as an Alternation. The Percent Alternation Score was Calculated as a Ratio of the Actual Number of Alternations to the Total Number of Possible Alternations and Multiplied by 100. A Higher Alternation Score Suggests Better Short-Term Memory. The Unpaired Student’s T-test was Used to Determine Statistical Significance (*P < 0.05). (B) Fear Conditioning (Contextual) Displays the Percentage of Freezing Behavior in Different Groups of Rats. This Test is Used to Study the Associated Learning and Memory of the Tested Rats. After Training, the Rat Learns the Training Chamber (the Context) is a Place to be Feared. The Fear-Related Freezing Behaviors in Response to the Conditioned Stimulus Were Recorded in This Test. This Graph Shows the Percentage of Freezing Behavior in Wild Type Rats (WT), Alzheimer’s Disease (AD) Rats Treated With a Mock Catheter ( n = 10), and AD Rats Treated With an Amytrapper Catheter ( n = 6). A One-way ANOVA Followed by Tukey’s Post-hoc Test was Used to Determine Statistical Significance (*P < 0.05)

During contextual fear conditioning, we observed a difference between AD-mock catheter treated rats and WT rats (non-diseased rats), but no significant difference was observed between AD-Amytrapper coated catheter and WT controls (Figure 3B). As AD rats treated with Amytrapper coated catheter performed similar to WT non-diseased animals, these data suggest a positive impact of the catheter-delivered Amytrapper treatment on contextual memory and recall, even if the performance of the two AD groups is not significantly different.

Decrease in Aβ Density/Accumulation Brain Sections from AD Rats Treated With Amytrapper Coated Catheter

Immunohistochemical staining for Aβ and DAB were analyzed on sagittal sections of the rat brain and a semi-quantitative evaluation of the distribution of Aβ was performed. In the AD rats from the mock catheter treated group and the Amytrapper catheter treated group, there was no difference in the distribution or expression of the Aβ signal among animals or groups. Most of the Aβ immune-positive staining seen was made of intraneuronal positive signal, characterizing the intracellular accumulation of Aβ. The intraneuronal signal was present in the cerebral cortex, cerebellar nuclei, thalamus, anterior hypothalamic nuclei, pons and also slightly distributed across all parts of the brain. In the AD rats from the control (mock) catheter treated group, there were positive amyloid plaques in the cerebral cortex compared to no positive amyloid plaques in the rats treated with Amytrapper coated catheters (representative images are shown in Figure 4B and 4B’). The wild-type rats had no positive Aβ signal throughout the sections, the neurons and neuroprenchyma were devoid of any immunopositivity in these animals (Figure 4A).

Figure 4.

Digital Slide Sections Consisting of Brain Sections Stained With Aβ Immunohistochemical Staining and DAB. The Sections Consist of a Sagittal Cut of the Brain and a Semi-quantitative Evaluation of the Distribution of Aβ was Performed. Comparison of Aβ Staining Between Wild Type (WT) and AD Rat Hippocampus, no Staining was Observed for WT Rats (A). Representative Section of Positive Amyloid Plaque in the Cerebral Cortex of AD Rats (B) and (B′) was Displayed. Scale Bars in 4B and 4B′ Represent 50 μm. Scale Bars in 4A Represent 1 mm

Cerebrospinal Fluid Aβ₄₂ Levels from AD Rat by Extracorporeal Amytrapper Catheter

Previous studies have established a well-documented phenomenon in progressive AD: a decrease in cerebrospinal fluid (CSF) Aβ42 levels.^26,27 This reduction is attributed to amyloid deposition and plaque formation within the brain. After two months of treatment, AD rats treated with the Amytrapper catheter displayed similar, and potentially slightly higher, levels of Aβ42 in the CSF compared to the control group (Figure 5). These observations can be interpreted within the context of Aβ42 dynamics in AD. Immunohistochemistry of brain slices from untreated animals (Figure 4) suggests a trend of Aβ42 accumulation, potentially reflecting its movement towards brain tissue and subsequent plaque formation. In contrast, the maintained CSF Aβ42 levels in the Amytrapper catheter treated group suggest a potential shift in the equilibrium between CSF and blood Aβ42. The continuous removal of Aβ42 by the catheter may alter this equilibrium, preventing its movement towards the brain and plaque formation. Collectively, these findings suggest that the Amytrapper catheter may influence Aβ42 distribution, potentially leading to reduced brain plaque formation at late initial stages of AD progression.

Figure 5.

Cerebrospinal Fluid Aβ42 Levels in Mock and Amytrapper Catheter-Treated AD Rats. CSF Aβ42 From Rats before and after Catheter Treatments was Measured Using the Sensolyte® Quantitative ELISA kit. Following Two Months of Treatment, Rats Treated With the Amytrapper Catheter (n = 6) Displayed Slightly Higher Levels of Aβ42 in the CSF Compared to the Mock Catheter (n = 5) as Control Group (P > 0.05; n.s, Utilizing Unpaired T-test)

Discussion

The peripheral Aβ clearance hypothesis offers an alternative approach to targeting AD. It centers on the concept that peripheral Aβ levels play a crucial role in regulating Aβ clearance from the central nervous system, where Aβ accumulation contributes to neuronal death through the formation of oligomers, protofibrils, and fibrils. This hypothesis is supported by the dynamic equilibrium of Aβ between the central nervous system and the periphery.^12,28 Studies, including passive immunization with an antibody recognizing Aβ1−12 in mouse models like Tg2576, have demonstrated cognitive improvement by tipping the balance of Aβ towards the periphery.¹⁰ Additionally, results from a phase II study using therapeutic apheresis to replace albumin with Albutein® 5% showed safety, mobilization of plasma Aβ, and stabilization of cognitive abilities.¹¹ Moreover, several other studies have indicated that removing peripheral amyloid leads to the depletion of brain amyloid and plaques, further supporting the amyloid theory of Alzheimer’s disease progression.^12-14 The benefits observed in whole blood exchange studies, which led to the depletion of cerebral amyloid plaques alongside behavioral improvements, highlight the potential of investigating novel amyloid-binding compounds and procedures to delay or prevent the progression of Alzheimer’s disease.²⁹ Importantly, the recent FDA approvals of aducanumab, lecanemab, and donanemab represent a significant advancement in AD treatment. These monoclonal antibodies all target the reduction of Aβ, a pathological hallmark of AD. Notably, these therapies demonstrate promise in improving cognitive function in patients, suggesting a potential link between Aβ clearance and positive clinical outcomes. These findings provide strong support for the Aβ hypothesis of AD and encourage further development of disease-modifying therapies targeting Aβ for the treatment of AD.¹⁴ However, these immune-therapies come with significant risky side effects such as ARIA-H, etc. Recent studies, including a January 2024 meta-analysis, suggest that monoclonal antibody treatments for AD may carry significant risks that could outweigh their potential benefits.⁷ Experts are weighing these risks carefully when considering the use of these treatments for AD patients.

In order to safely target peripheral Aβ in developing disease-modifying AD treatments, in a preliminary proof-of-concept study we have previously shown that the Amytrapper catheter captured amyloid from plasma of a small number of AD patients in vitro¹³ Here we expand on that study and test the Amytrapper catheter as an extracorporeal device on a rat model of AD in vivo. We show for these AD rats a significant reduction in blood Aβ levels and behavioral improvements compared to controls. We further show that the current study, supported by observations from our previous studies, opens doorways for novel therapeutic interventions focused on safe and efficient removal of Aβ from the periphery.

The projected target device will be an extracorporeal processing system designed to detoxify the blood. It will include a medically viable catheter (Amytrapper catheter), which contains a peptide based capturing agent designed to have a high binding affinity for circulating Aβ, as previously described.¹³ This active pharmaceutical ingredient (API) is a retro-inverso tetramer peptide (RI peptide sequence: KWGEWTGR) conjugated to PEG. Unlike other protein-based Aβ-binding ligands, its D-amino acid composition makes it protease-resistant, offering a longer shelf life. Additionally, this peptide can be synthesized in large quantities under stringent quality control processes, minimizing batch-to-batch variations and ensuring cost-effective manufacturing. Furthermore, the extracorporeal processing system will include a pressure-controlled pump and monitor system to regulate blood flow (Figure 6). Blood will be pushed through Amytrapper catheter where amyloid protein will be captured by affinity binding to the API. Purified blood will re-enter the body. In this process, the API always binds to the catheter. In vitro proof-of-concept studies have yielded promising results.¹³ Supported by our observations in this study with the transgenic rat AD model, Amytrapper catheter has the potential to be used to treat patients where removal of Aβ is a therapeutic option to improve their living conditions. Theoretical estimation suggests that the approximate Aβ binding capacity is at least 10-50 times the circulating levels of Aβ.¹⁴ We have consistently shown that removal of circulating amyloid led to the depletion of brain amyloid because of the inherent homeostatic balancing of amyloid levels between circulation and brain.¹⁴ Improvised catheter could turn out to be a robust amyloid removal system to handle the circulating levels of Aβ (usually Aβ range: 350-400 pg/mL) in plasma of AD patients.²⁵ In our study on AD rats, we observed that, compared to control uncoated (mock) catheters, Amytrapper coated catheters were able to capture about 25% of Aβ42. In the extracorporeal processing system, a medically viable Amytrapper catheter with the pegylated RI peptide will replace the cartridges used in clinical hemodialysis. With the Amytrapper catheter in-line, AD patients will undergo hemodialysis treatment in the same way as done for renal disease patients with surgically placed Arterio-Venous Fistula for vascular access (Figure 6). Our API coated catheter will replace the dialyzer and can be custom retrofitted for single/multiple pass-through for incoming arterial blood for amyloid capture. Pre- and post- procedure Aβ measurements in blood and CSF should give a quick readout of the effectiveness.

Figure 6.

A Schematic Showing Patient With Arterio-Venous Fistula for Vascular Access and Dialysis Device With Amytrapper Catheter. This Represents a Putative Design of an Extracorporeal Processing System With Amytrapper Catheter

We believe that this catheter would remove circulating Aβ and thus would aid in Alzheimer’s disease modification. While there are other Aβ removal options available,^30-32 the proposed extracorporeal depletion approach is safer and enables faster results to achieve clinical benefit in AD patients. We have demonstrated that the amytrapper approach does not lead to any leaching of the amytrapper peptide even after 24 h exposure to the circulating buffer.^13,14 The new catheter device, while serving an unmet medical need, will complement existing AD treatments as a disease modifying strategy.

There are several limitations to this study that should be acknowledged. While rat models have been instrumental in advancing our understanding of Alzheimer’s disease (AD), they possess certain inherent limitations when used to model complex human neurodegenerative disorders. One notable concern is the use of transgenic rats that overexpress mutant human genes, such as hAPP751. While this approach can induce AD-like pathological features, it often results in non-physiological levels of protein expression. These artificially elevated levels may not accurately mimic the gradual and multifactorial progression of AD in humans, potentially leading to results that are not fully representative of the human condition. Additionally, rodent models may not capture the full spectrum of cognitive and behavioral symptoms observed in human patients, nor the influence of age-related co-morbidities and genetic diversity.

Another significant limitation of this study is the small sample size. A limited number of subjects can reduce the statistical power of the analysis, making it more difficult to detect meaningful differences or subtle effects. This, in turn, may increase the risk of Type II errors (ie, failing to detect a true effect) and limit the generalizability of the findings. A larger sample size would not only enhance the reliability of the results but also allow for subgroup analyses and more robust conclusions. Future studies should consider employing larger, more diverse cohorts and complementary animal or in vitro models to strengthen the translational relevance of the findings.

Conclusion

This study further supports the potential of the extracorporeal system as a disease-modifying treatment for Alzheimer’s disease (AD) through the removal of circulating amyloid-beta (Aβ).¹³ Utilizing an in vivo rat model of AD, the current findings demonstrate that the Amytrapper catheter effectively captures Aβ from the bloodstream.¹³ These promising results provide a strong rationale for continued investigation and support further preclinical and clinical development of the Amytrapper catheter as a novel therapeutic strategy targeting peripheral Aβ clearance in AD.

Footnotes

Acknowledgement

Dushyant Mishra’s help in preparing the draft by compiling the available data is gratefully acknowledged. This material is the result of work supported with resources and the use of facilities at the Oklahoma City US Department of Veterans Affairs Medical Center. We also gratefully acknowledge the assistance provided by Dr Veronica Galvan Hart and her laboratory with the behavior studies.

ORCID iDs

Hao Huang

Pazhani Sundaram

Ethical Approval

Rat handling and care procedures were followed according to recommendations and approval of Oklahoma City VA Health Care System Institutional Animal Care and Use Committee (IACUC).

Consent for Publication

All authors consent for this publication.

Author contributions

Pazhani Sundaram (Conceptualization; Supervision; Funding acquisition; Writing – original draft; Writing – review & editing). Rishi Raj Chhipa (Data curation; Formal analysis; Investigation; Methodology; Writing – original draft; Writing – review & editing); Biji T. Kurien (Data curation; Formal analysis; Methodology; Writing – review & editing). Robert H. Scofield (Data curation; Formal analysis; Methodology; Writing – review & editing). Roman F. Wolf (Data curation; Formal analysis; Methodology; Writing – review & editing). Hao Huang (Data curation; Formal analysis; Investigation; Writing – review & editing).

Funding

The author(s) disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: Dr. Pazhani Sundaram was awarded a small business innovative research phase 2 grant #R44 AG0 57327, from National Institute on Aging, National Institutes of Health, USA. The grant provided salary support for Pazhani Sundaram, Hao Huang, and Rishi Raj Chhipa.

Declaration of Conflicting Interest

The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Data Availability Statement

The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.*

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Amytrapper Catheter: A Prototype Extracorporeal Device That Traps Blood Amyloid-β in a Rat Model of Alzheimer’s Disease

Abstract

Keywords

Introduction

Materials and Methods

Rat AD Model

Testing of the Amytrapper Catheter in Vivo

Estimation of Aβ42 Bound to Catheters

Estimation of Aβ42 in Plasma and CSF by ELISA

Y-Maze Test

Fear Conditioning

Semi Quantitative Assessment of Aβ by Immunohistochemistry on Stained Brain Sections from AD Rats

Results

Procedural Feasibility and Capturing of Aβ42 From Whole Blood of AD Rat by Extracorporeal Amytrapper Catheter

Neurobehavioral Improvements in AD Rat Treated With Extracorporeal Amytrapper Catheter

Decrease in Aβ Density/Accumulation Brain Sections from AD Rats Treated With Amytrapper Coated Catheter

Cerebrospinal Fluid Aβ42 Levels from AD Rat by Extracorporeal Amytrapper Catheter

Discussion

Conclusion

Footnotes

Acknowledgement

ORCID iDs

Ethical Approval

Consent for Publication

Author contributions

Funding

Declaration of Conflicting Interest

Data Availability Statement

References

Procedural Feasibility and Capturing of Aβ₄₂ From Whole Blood of AD Rat by Extracorporeal Amytrapper Catheter

Cerebrospinal Fluid Aβ₄₂ Levels from AD Rat by Extracorporeal Amytrapper Catheter