Nazmuddin M, Stammes MA, Klink PC, Vernes MK, Bakker J, Langermans JAM, van Laar T, Philippens IHCHM (2025) Stereotactic lesioning of cholinergic cells by injection of ME20.4 Saporin in the nucleus basalis of Meynert in a rhesus monkey (Macaca mulatta). J Neuropathol Exp Neurol nlaf081. doi: 10.1093/jnen/nlaf081 PMID: 40673943
Objective: To describe a procedure to inject ME20.4-SAP, an immunotoxin that specifically binds to and depletes cholinergic neurons stereotactically into the nucleus basalis of Meynert (NBM) of a rhesus monkey (Macaca mulatta).
Summary: A digital non-human primate brain atlas was co-registered to the brain of the monkey. A custom-designed cranial chamber was also implanted to the skull to guide the injection. The effects of the ME20.4-SAP injections were evaluated in vivo with PET-CT using [18F]-FEOBV as a radiotracer. This approach yielded reliable spatial accuracy and successful delivery of ME20.4-SAP into the NBM. This saporin-mediated selective destruction of cholinergic neurons in the NBM, using MRI-guidance and a cranial chamber, offers a promising method to study the pathophysiology of NBM degeneration and possible therapeutic interventions.
Usage: The first dose was chosen based on previous NBM lesioning works in common marmosets where infusing 1.4 μg ME20.4-SAP (Cat. #IT-15, in a concentration of 0.20 μg/μl) into each side of the NBM produced partial NBM depletion. At the second injection session, 5 μg ME20.4-SAP (in 0.5 μg/μl solution) was administered into each NBM side.
Limonta P, Chiaramonte R, Casati L (2024) Unveiling the dynamic interplay between cancer stem cells and the tumor microenvironment in melanoma: Implications for novel therapeutic strategies. Cancers (Basel) 16(16):2861. doi: 10.3390/cancers16162861 PMID: 39199632
Objective: To review the bidirectional communication between melanoma cancer stem cells (CSCs) and the tumor microenvironment, highlighting its role in drug resistance and tumor relapse.
Summary: Melanoma CSCs evade immune surveillance and recruit immune cells with immunosuppressive and tumor-promoting properties, establishing a supportive microenvironment. They also transfer stemness and aggressive traits to neighboring non-CSCs, driving tumor progression and metastasis. Targeting these interactions may offer novel therapeutic strategies for combating melanoma.
Usage: This review publication highlights the usage of Anti-CD271-SAP and CD133-SAP in previous publications.
Nodal FR, Leach ND, Keating P, Dahmen JC, Zhao D, King AJ, Bajo VM (2024) Neural processing in the primary auditory cortex following cholinergic lesions of the basal forebrain in ferrets. Hear Res 447:109025. doi: 10.1016/j.heares.2024.109025 PMID: 38733712
Objective: To explore whether behavioral deficits are associated with changes in the response properties of cortical neurons, neural activity was recorded in the primary auditory cortex (A1) of anesthetized ferrets in which cholinergic inputs had been reduced
Summary: Results show that while ACh is required for behavioral adaptation to altered spatial cues, it is not required for maintenance of the spectral and spatial response properties of A1
Usage: Bilateral injections of the immunotoxin ME20.4-SAP in the nucleus basalis (NB)
Ngo M, Han A, Lakatos A, Sahoo D, Hachey S, Weiskopf K, Beck A, Weissman I, Boiko A (2016) Antibody therapy targeting CD47 and CD271 effectively suppresses melanoma metastasis in patient-derived xenografts. Cell Rep 16:1701-1716. doi: 10.1016/j.celrep.2016.07.004
Summary: The high rate of metastasis and recurrence among melanoma patients indicates the presence of cells within melanoma that have the ability to both initiate metastatic programs and bypass immune recognition. The authors identified CD47 as a regulator of melanoma tumor metastasis and immune evasion. The study involved antibody-mediated blockade of CD47 coupled with targeting of CD271+ melanoma cells by way of ME20.4-SAP (Cat. #IT-15). Mice bearing human melanoma tumor (M213 or M727) were randomized into four treatment groups with one of those groups receiving treatment with ME20.4-SAP. 1 ug in 50 ul volumes were injected directly into the center mass of the tumor once every 2 days. A therapeutic effect was observed where tumor metastasis in patient-derived xenografts was strongly inhibited when treated with the combination of antibody-mediated blockade of CD47 and targeted with ME20.4-SAP.
Bajo V, Leach N, Cordery P, Nodal F, King A (2014) The cholinergic basal forebrain in the ferret and its inputs to the auditory cortex. Eur J Neurosci 40:2922-2940. doi: 10.1111/ejn.12653 PMID: 24945075
Summary: The ferret has become a more common animal model in auditory neuroscience. Unlike rodent models, however, anatomical data describing the organization of the basal forebrain cholinergic system and its projections to the auditory cortex have not been well characterized. Using a variety of methods the authors mapped the architecture of the ferret basal forebrain. IHC was done with several antibodies including anti-ChAT (Cat. #AB-N34AP; 1:1000) and anti-NGFr (Cat. #AB-N07; 1:500). Animals also received 17 μg of ME20.4-SAP (Cat. #IT-15) in a total of 17 injections into the ectosylvian gyrus. The results indicate that acetylcholine is most likely involved in modulation of auditory processing.
Nodal FR, Leach ND, Keating P, Dahmen JC, King AJ, Bajo VM (2013) Spatial processing in the primary auditory cortex following cholinergic lesions of the basal forebrain in ferrets. Neuroscience 2013 Abstracts 353.09. Society for Neuroscience, San Diego, CA.
Summary: Cortical acetylcholine release has been implicated in different cognitive functions, including perceptual learning. We have recently shown that cortical cholinergic innervation is necessary for normal sound localization accuracy in ferrets, as well as for their ability to adapt with training to altered spatial cues (Leach et al., 2013, J Neurosci 33:6659-71). To explore whether these behavioral deficits are associated with changes in the spatial sensitivity of cortical neurons, we recorded neural activity in the primary auditory cortex (A1) from three animals in which cholinergic inputs had previously been reduced by making bilateral injections of the immunotoxin ME20.4-SAP in the nucleus basalis (NB). Neural activity was recorded from 146 penetrations in the left and right A1 under anesthesia (medetomidine/ketamine) using Neuronexus multi-site silicon probes. Histological analysis after the recording sessions revealed a mean loss of cholinergic neurons in the NB of 89.3±7.1% when compared to control animals, as well as a significant reduction in cholinergic fiber density across the auditory cortex, including the middle ectosylvian gyrus where A1 is located. On the basis of the location of the penetrations and electrophysiological characterization of the neural responses, which typically exhibited a mean latency of ≤20 ms, frequency tuning and onset responses with occasional weaker offset responses, we were able to assign the recordings to A1. The distribution of unit best frequencies was used to ensure that the tonotopic axis of A1 was evenly sampled. Spatial tuning was determined using virtual acoustic space stimuli comprising 200 ms broadband noise presented at three different levels (56, 70 and 84 dB SPL) from 12 locations separated by 30° in azimuth. Most of the units were broadly tuned, responding to all the virtual sound locations tested. Their spatial preferences were quantified by calculating the centroid direction vector from the variation in spike count with stimulus location within the onset response. This revealed a contralateral preference for most units, with the majority of the centroid azimuths located within the frontal hemifield. These data are consistent with the distribution of azimuth tuning previously described in the ferret, and initial comparisons with control animals have not shown any differences in spatial sensitivity in the animals with cholinergic lesions. Reduced cholinergic release therefore does not appear to influence the spatial response properties of A1 neurons in anesthetized animals, suggesting that any effects on sensory coding may only become apparent during behavior.
Leach ND, Nodal FR, Cordery PM, King AJ, Bajo VM (2013) Cortical cholinergic input is required for normal auditory perception and experience-dependent plasticity in adult ferrets. J Neurosci 33(15):6659-6671. doi: 10.1523/JNEUROSCI.5039-12.2013
Summary: In order to study how cholinergic input from the nucleus basalis affects auditory perception and learning, the authors injected a total of 35.2 ng of ME20.4-SAP (Cat. #IT-15) into the nucleus basalis in each hemisphere of ferrets. Based on several learning tasks, the data suggest that these cholinergic inputs aid in the perception of sound source location and aid in the adaptation of the auditory system to changes in spatial cues.
Summary: Episodic memory is controlled by several interconnected brain structures. The order in which these structures sustain damage can affect the processes lost. In this work the authors performed numerous bilateral injections of ME20.4-SAP (Cat. #IT-15) into the infero-temporal cortex, the medial surface of the temporal lobe, the perirhinal and entorhinal cortex, and the temporal pole of monkeys. These injections totaled 2.2-2.5 μg of conjugate. The results indicate that loss of cortical acetylcholine function will interfere with adaptation to memory impairments caused by structural damage in episodic memory centers.
Croxson PL, Kyriazis DA, Baxter MG (2011) Cholinergic modulation of a specific memory function of prefrontal cortex. Nat Neurosci 14(12):1510-1512. doi: 10.1038/nn.2971
Summary: The authors investigated loss of acetylcholine in the large and highly differentiated PFC’s of rhesus monkeys. The monkeys received 80-92 20-ng injections of ME20.4-SAP (Cat. #IT-15) per hemisphere. Lesioned animals were severely impaired on tasks involving spatial working memory.
Bajo Lorenzana VM Leach ND, Cordery PM, Nodal FR, King AJ (2011) The cholinergic basal forebrain in the ferret and its inputs to the auditory cortex. IBRO 2011 Abstracts International Brain Research Organization, Florence, Italy.
Summary: Projections from the NB to the auditory cortex were investigated by injecting tracers into the NB itself (n=5), or by applying tracer deposits to the surface of the auditory cortex (n=4). Tracers included Rhodamine, Fluorescein, Cascade Blue, as well as the cholinergic immunotoxin ME20.4-SAP. Both ME20.4-SAP injections in the auditory cortex and epipial tracer deposits revealed that NB provides the main cholinergic input to the cortex, and that this projection is predominantly ipsilateral.
