**4. New targets and technologies**

#### **4.1. Targeting the immune system in AD**

The vast majority of passive immunotherapeutic approaches in AD have targeted Aβ and tau; this is a natural outcome of the primacy of these proteins as the principal pathological hallmarks of the disease. The association of mutations of APP (and proteins that modulate its generation, such as presenilin-1) to familial AD, and the high degree of correlation between tau pathological development and cognition, strengthen the validity of these two proteins as important causative disease agents. However, new approaches, primarily targeting immunomodulatory proteins, are also currently under development.

The presence of neuroinflammatory processes and signatures in AD has been well established, but the exact role they play in disease etiology, or whether neuroinflammation has a primarily protective or harmful role, has not been clear (reviewed in [94]). Studies examining the complement cascade have helped to understand this duality. The synaptic pruning activity carried out by microglia is regulated by complement [95]. The initiating protein of the classical complement cascade, C1q, is enriched in the developing mouse CNS and localizes to synapses; genetic ablation of this protein results in misregulated innervation due to increased presence of synapses [96]. While C1q is normally downregulated after development, it is elevated in normal aging [97] and disease, including AD [98]. In a transgenic APP mouse, C1q localizes to synapses, and is required for pathological synapse loss. Treatment of C1q knockout mice with oligomeric Aβ displayed no synaptic loss, indicating that C1q is a required mediator of Aβ-induced toxicity. Interestingly, an anti-C1q antibody rescued Aβ-induced synaptotoxicity *in vivo*, and LTP impairment *in situ*, when compared to isotype control [99]. These data hinted at the promise of C1q immunotherapy to provide protective benefits by neutralizing a key mediator of Aβ-induced microglial overactivation, which results in synaptic loss. The anti-C1q antibody used in this study has been developed into a human therapeutic, and is beginning clinical trials (clinicaltrials.gov; Identifier: NCT03010046) [100].

mechanisms may be different than those proposed for Aβ immunotherapy. There remain conflicting details from the studies presented here, such as the relative contribution of microglialmediated phagocytosis, the relative importance of eTau-mediated Aβ production, the extent of trans-synaptic transmission in transgenic mice with widespread expression in the brain, and the optimal epitope to target. Gaining a clearer understanding of these factors continues

Clinical trials with Aβ immunotherapies have demonstrated the importance of proper clinical diagnosis, patient selection, sensitive cognition tests, and effective biomarkers to monitor efficacy and disease progression. Though some general commonalities may exist in the clinical design of Aβ and tau passive immunotherapy trials, there are substantial differences in the targets and any potential clinical development approaches. In contrast to Aβ, there are a number of non-AD tauopathies such as progressive supranuclear palsy (PSP) [89] and frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17) [90] that may provide alternative clinical development pathways to test novel tau-directed therapeutic approaches. In contrast to AD, these diseases present pathological signatures composed almost uniformly of tau and neurofibrillary tangles; in addition, FTDP-17 is an autosomal dominant disorder, genetically validating the causative role of tau. Diagnosis of these and other tauopathies have historically been made solely based on clinicopathology; due to the difficulty of diagnosis from to the overlap of symptomologies with other neurodegenerative disorders, as well as the lack of clear biomarkers, diagnosis is only confirmed at autopsy [91]. Modern tau PET imaging agents are currently under clinical investigation [92]; while early generations of tau PET tracers displayed nonspecificity and suboptimal binding and PK characteristics, the newest class of tracers display improved specificity, PK properties, and may allow for improved diagnosis in tauopathies as well as an ability to monitor tau pathology in

The vast majority of passive immunotherapeutic approaches in AD have targeted Aβ and tau; this is a natural outcome of the primacy of these proteins as the principal pathological hallmarks of the disease. The association of mutations of APP (and proteins that modulate its generation, such as presenilin-1) to familial AD, and the high degree of correlation between tau pathological development and cognition, strengthen the validity of these two proteins as important causative disease agents. However, new approaches, primarily targeting immuno-

The presence of neuroinflammatory processes and signatures in AD has been well established, but the exact role they play in disease etiology, or whether neuroinflammation has a primarily protective or harmful role, has not been clear (reviewed in [94]). Studies examining the complement cascade have helped to understand this duality. The synaptic pruning activity carried out by microglia is regulated by complement [95]. The initiating protein of the

are a current research focus.

138 Alzheimer's Disease - The 21st Century Challenge

AD clinical trials [93].

**4. New targets and technologies**

**4.1. Targeting the immune system in AD**

modulatory proteins, are also currently under development.

The mounting evidence of involvement of the adaptive immune system in restraining the advance of AD pathology has opened the possibility of directing passive immunotherapies to the periphery, which considerably eases the challenge of achieving sufficient drug exposure in the CNS to affect pathology. Microglia resident in the brain are known to be recruited to sites of injury such as senile plaques, but the finding that peripherally-derived bone marrow stem cells are able to enter the CNS, and differentiate into microglia [101, 102], was the first direct evidence that repopulation and recruitment of microglia from the periphery was an active process. This finding was extended to AD mouse models with the finding that peripherally-recruited microglia are mobilized by Aβ, recruited to the site of senile plaques, and are able to clear plaques via phagocytosis [103]. The protective role of these immune cells in the presence of AD-like pathology was confirmed with the observation that (a) knocking out the chemokine receptor CCR2 in an APP-transgenic mouse resulted in decreased recruitment of monocytes to Aβ plaques [104], and (b) the specific ablation of bone-marrow derived cells via diphtheria-toxin receptor expression resulted in increased Aβ plaques [105]. Furthermore, increasing trafficking of macrophages by inhibiting the normally immunosuppressive regulatory T-cells through pharmacologic or genetic methods results in reduced Aβ pathology [106].

Elucidation of the biology of inhibitory signaling pathways and proteins such as Programmed cell death protein 1 (PD-1) and cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), known as immune checkpoints, led to the development of antibody therapeutics for use in cancer (reviewed in [107]). These therapies function by neutralizing immune checkpoints and activating T-cells, which prompts antitumor activity. The characterization of checkpoint signaling pathways, along with the findings that peripheral immune cells modulate AD-like pathology in a regulatory T-cell (Treg)-dependent manner, has prompted examination of the PD1/PD-L1 axis in AD. In a recent study, AD transgenic mice were treated with an anti-PD1 antibody to blockade the PD1/PD-L1 axis. Remarkably, checkpoint blockade in this model resulted in substantial rescue of performance in a behavioral assay of memory and cognition after a single dose, and mice exhibited decreases in Aβ pathology with only two dose administrations [108]. The effect on pathology was observed even in mice with profound amyloid burden. While the findings of a profound effect on functional measures after such a short dose regimen are very exciting, they should be taken with a note of caution. A follow-up study, carried out by three pharmaceutical companies using three transgenic models and numerous PD-L1 antibodies was attempted to recapitulate these results. Despite peripheral immune activation, in all instances neither reductions in Aβ pathology nor infiltration of peripheral monocytes were detected [109]. Further studies are needed to elucidate the potential of checkpoint modulation.

**5. Conclusions and future perspectives**

and comments provided by Enchi Liu and Ellen Rose.

PJD and WZ are employees of Prothena Biosciences.

\*Address all correspondence to: pdolan@prothena.com

Prothena Biosciences, South San Francisco, CA, USA

Neurology. 2017;**16**(11):877-897

treating AD.

**Acknowledgements**

**Conflict of interest**

**Author details**

**References**

Philip J. Dolan\* and Wagner Zago

AD provides a monumentally challenging drug development landscape. The uncertainty about disease etiology, variability in patient genetics and disease progression, and difficulties in early diagnosis are all but a noncomprehensive list of hurdles to developing effective drugs. Though development of therapeutics to slow or halt AD disease progression, including passive immunotherapeutics, have not yet yielded clinical benefit, the prospect of applying lessons learned in the clinic towards validated targets such as Aβ and tau provides optimism for future success. In addition, our understanding of the mechanisms of other principal contributing factors to disease progression will provide a variety of new targets to explore. Combined with advances in drug technology to increase the availability of biomolecules in the CNS, these clinical and biological advances offer great promise around future success in

Passive Immunotherapy in Alzheimer's Disease http://dx.doi.org/10.5772/intechopen.76299 141

There is a wealth of excellent studies in the areas of study covered by this chapter—unfortunately, we were unable to list them all. We thank the many researchers not recognized here who have contributed greatly to the field. We would also like to thank the thoughtful review

[1] Group GBDNDC. Global, regional, and national burden of neurological disorders during 1990-2015: A systematic analysis for the Global Burden of Disease Study 2015. Lancet

#### **4.2. Increasing blood-brain barrier (BBB) penetrance for passive immunotherapeutics**

A significant barrier in the development of passive immunotherapeutics for AD is the low percentage of circulating antibody that crosses the BBB. Animal studies have indicated that ~0.1–0.5% of IgG enters the CSF from the periphery [110, 111], which is borne out by preclinical [112] and clinical [113, 114] data obtained with antibodies tested for use in AD. This has led to trials with increasing amounts of antibody administered to patients ([82]; clinicaltrials.gov, Identifier NCT03318523) with the hope of delivering sufficient amounts of antibody to the CNS to achieve a clinical effect. There are, however, indications that concentrations of antibodies are higher in brain parenchyma than what is present in CSF. The chimeric form of aducanumab reported brain:plasma AUC ratios when tested in a transgenic APP model of 1.3% [38]. This is in agreement with the finding that the concentration of protein analyte present in the interstitial fluid is approximately 10-fold higher than in the ISF [62, 115]. This could be due to the rapid turnover of CSF volume [116] compared to ISF, longer elimination times of antibodies in brain parenchyma compared with CSF, or increased residence time due to target-mediated binding. Nevertheless, methods and technologies to increase BBB penetrance of biomolecules urgently need to be applied to antibodies and other proteins.

One of the more promising approaches to increase penetrance of protein therapeutics into the brain utilize endogenous receptors that transcytose between the brain and periphery, such as transferrin receptor (TfR) [117], insulin receptor [118], and LDL receptor-related protein 1 (LRP1) [119]. Protein engineering approaches feature fusion of the therapeutic molecule to proteins, ligands, or peptides that bind these receptors and facilitate transcytosis across the BBB (reviewed in [120]). One of the best understood receptor-mediated delivery systems is the use of TfR, though a similar path has been taken in the development of technologies that utilize insulin receptor. Increased brain uptake of transferrin/antibody fusion proteins were detected in rats [121], though the relatively large size (~80 kDa) of full-length transferrin make this impractical for biotherapeutic use. The detection of increased transcytosis of anti-TfR antibodies and antibody fragments [122, 123], and later advances in antibody generation technologies, enabled bispecific antibodies that bind TfR as well as target [124]. As understanding of the transcytotic properties of TfR binding moieties have increased, so has the understanding of how best to incorporate properties to ensure delivery to the brain. For example, reducing TfR affinity improves delivery, as a low affinity anti-TfR moiety will release from the receptor faster than a high affinity moiety [124]. As receptor-binding fusions enter the clinic, further questions regarding safety and distribution changes brought about by higher CNS concentrations will need to be continually addressed [125, 126]. Work continues to identify receptors that may be useful for increasing BBB concentrations of antibodies to allow engagement with wider range of drug targets [127, 128].
