Over the past decade, evidence from the scientific and medical communities has demonstrated that nanobiotechnology and nanomedicine have tremendous potential to affect numerous aspects of cancer and other disorders in term of early diagnosis and targeted therapy. The utilization of nanotechnology for the development of new Nano-carrier systems has the potential to offer improved targeted delivery through increased solubility and sustained retention and more importantly active targeting. One of the major advantages of this innovative technology is its unique multifunctional characteristics. Targeted delivery of drug incorporated nanoparticles, through conjugation of site-specific cell surface markers, such as tumor-specific antibodies or ligands, which can enhance the efficacy of the anticancer drug and reduce the side effects. Additionally, multifunctional characteristics of the Nano-carrier system would allow for simultaneous imaging of tumor mass, targeted drug delivery and monitoring (Theranostics).

A summary of recent progress in nanotechnology as it relates to nanoparticles and drug delivery will be reviewed. Nano Nutraceuticals using combination of various natural products provide a great potential in diseases prevention. Additionally, various Nanomedicine approaches for the detection and treatment of various types of organ specific delivery, vascular targeting, and vaccine will be briefly discussed.

Reference: Mousa SA, Bawa R, and Audette GF (Editors): The Road from Nanomedicine to Precision Medicine, Jenny Stanford Publishing (2020) – 1200+ pages, 36 chapters [ISBN 978-981-4800-59-4 (Hardcover), 978-0-429-29501-0 (eBook)].

Systemic Lupus Erythematosus (SLE) is an autoimmune disease which presents clinical manifestations in different organs and presenting autoantibodies targeting its own body. The pathogenesis of SLE is not yet fully understood. However, there is no proper biomarker to diagnose SLE and to measure disease activity. Proteomics based precision medicine approaches aims to improve diagnostics and prognostics of SLE. Components of the complement system have been associated with improved diagnosis of SLE patients.

We present a proteomics driven study to address and identify potential proteomic markers for patient subtyping. Plasma samples from the four different SLE patient groups (low vs. high SLEDAI; low vs. high C3gd) were selected based on clinical scores from the SLE patients (n=40). Plasma samples were analyzed by quantitative proteome analysis tandem mass spectrometry and low-abundance regulatory proteins using multiplex analysis. Autoantigen profiles were determined by 1536plex Immunome array. Our findings identify patients with the high SLEDAI scores were diagnosed with SLE at an earlier age of, while the patients with the low SLEDAI score were diagnosed at a later age. Proteomics analysis enabled quantitative assessment of patient group specific changes enabling subtyping of the four patient groups by principle component analysis. When compared between C3dghigh – C3dglow groups, 7 proteins were found to be upregulated and 43 proteins were downregulated. There were 16 upregulated proteins and 10 downregulated proteins in SLEDAIhigh – SLEDAIlow protein comparison.

From our experiments, we found the differences in key parameters of the blood test such as age of diagnosis, anti-dsDNA concentration, lymphocyte count of C1q concentration. The list of upregulated and downregulated proteins was obtained in the proteome analysis of these patient groups. These proteins can be valuable to understand subtypes of SLE patients as well as serve as potential diagnostic biomarkers for SLE.

Kynurenic acid (KYNA) is a metabolite of the L-tryptophan (TRP)-kynurenine (KYN) pathway which has been shown to possess neuroprotective and antidepressant-like properties. The intracerebroventricular (i.c.v.) administration of kynurenic acid (KYNA) triggered antidepressant-like effects at least in part through the serotonin 5-hydroxytryptamine (5-HT) type 2 receptors, D₂, D₃, D₄ dopamine receptors, and gamma-aminobutyric acid subunit A (GABA_A) receptors in modified forced swimming test (FST) of mouse. However, KYNA is impermeable to the blood-brain-barrier (BBB) and thus it is probably a cause that the peripheral administration of KYNA did not exhibit an antidepressant-like effect.

New KYNA analogues were designed to overcome the BBB in attempt to deliver neuroprotective KYNA molecules to the central nervous system (CNS). The antidepressant-like effects of KYNA analogues SZR-72, SZR-81, and SZR-104 were studied i.c.v. and intraperitoneally (i.p.) with a modified FST.

The FST revealed that i.c.v. administration of SZR-81 significantly decreased immobility and significantly increased swimming time, which suggested that its antidepressant-like effects were triggered through the serotonin 5-HT nervous system. SZR-72 and SZR-104 did not significantly change either immobility, climbing, or swimming time, suggesting SZR-72and SZR-104 did not have antidepressant-like effects. Furthermore, i.p. administration of SZR-81 did not change either immobility, climbing, or swimming time like i.c.v. administration.

Two of the KYNA analogues SZR-72and SZR-104 may not have the antidepressant-like properties and SZR-81 may not cross the BBB or be modified on the way to the CNS. Novel KYNA analogues are expected to be designed in search of potent antidepressants which reserve antidepressant properties, cross the BBB, and target in the CNS.

Antibody-drug conjugates (ADCs) are used in anticancer therapy with some limitations due to their molecular properties. An alternative to monoclonal antibodies is the affibody, composed of 58 aminoacids, with lower binding affinities, small size, and rapid blood clearance and tissue distribution. We investigated the in vitro efficacy of a novel anti-HER2 ZHER2:2891 affibody conjugated to a cytotoxic drug auristatin E (MMAE) in HER2-positive human cancer cells. Adenocarcinoma cell line SK-BR-3, expressing high levels of HER2, and mammary gland adenocarcinoma MDA-MB-231, expressing basal levels of HER2, were treated with ZHER2:2891DCS-MMAE and trastuzumab (as reference compound).

ZHER2:2891DCS-MMAE induced a significant time-dependent toxic effect in SK-BR-3 cells. A 30% reduction in cell viability was found after 10 min exposure at 7 nM with an IC50 of 80.2 nM. On the contrary, MDA-MB-231 cells were not affected by the affibody complex. The HER2-specific cytotoxic effect of the ZHER2:2891DCS-MMAE has been also confirmed by measuring apoptosis by flow cytometry. In SK-BR-3 cells, increasing concentrations of conjugated affibody, induced cell death after 10 min of treatment with the strongest effect observed after 48 hours. Also, treatment with ZHER2:2891DCS-MMAE reduced (up to 50%) HER2 expression at both mRNA and protein levels in SK-BR-3 cells after 24 hours of treatment.

In conclusion, the cytotoxic conjugate based on the anti-HER2 affibody and MMAE efficiently interacts with HER2 over-expressing cancer cells, allowing the selective and specific delivery of the cytotoxic payload. The basal HER2 expressing cells are not affected most probably due to a lower uptake of drug conjugate. This confirms that affibody may be used to target HER2 overexpressing cells while sparing normal cells.

Supramolecular protein assembly, such as cages, rings, and tubes can be used as reaction vessels and molecular templates for applications in catalysis, imaging and drug delivery.¹ Ferritin (Ft) is a member of ferroxidase family of enzymes that forms a spherical nanocage. It sequesters iron by concentrating it in its internal cavity for storage and detoxification.² Ft is very promising as a drug loading and releasing system since it is non-immunogenic, bio-compatible, highly stable, soluble in the bloodstream. It can be internalized via Ft-binding receptors (like transferrin receptor 1³ or Scara5⁴) that are over-expressed in a variety of malignant cells. Well established metallodrugs like cisplatin,⁵ carboplatin,⁶ a Pt-terpyridine compound,⁷ three different gold-based anticancer compounds (Auoxo3, Au2phen, Auoxo4),^8-9 a bimetallic Pt-Au compound¹⁰, a bimetallic Pt-As compound and a di-ruthenium complex¹¹ have been trapped within the Ft nanocages, taking advantage of the alkaline pH disassembly/reassembly protocol. The drug-loaded nanocomposites have been characterized by circular dichroism, to evaluate the protein secondary structure content upon drug encapsulation, and by UV-vis absorption spectroscopy, to assess the drug loading within the protein cage. ICP-MS allowed to quantify the amount of drug trapped inside the nanocage and to define metal/Ft stoichiometry, while X-ray crystallography unveiled the metallodrugs binding sites on the proteins structure and the nature of the interaction of the different compounds with Ft. The compounds often degrade upon encapsulation within the protein cage; metal-containing fragments coordinate Cys or His side chains. However, the coordination does not affect the overall cytotoxicity of the encapsulated anticancer agents, since many molecules of the metallodrugs remain trapped in the bulk within the inner core of the cage. Biological activity studies show that, even though the presence of the cage reduces the overall toxicity of the metallodrugs, the adducts of the selected compounds with Ft are generally more selective towards cancer cells than to non-malignant cells. Altogether these data indicate that encapsulation of metal-based drugs within Ft nanocages is a promising strategy to deliver these molecules to their final targets.¹²

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