Current progress, strategy, and prospects of PD-1/PDL-1 immune checkpoint biosensing platforms for cancer diagnostics, therapy monitoring, and drug screening.

Recent technological advancements in testing and monitoring instrumentation have greatly contributed to the progress in cancer treatment by surgical, chemotherapeutic and radiotherapeutic interventions. However, the mortality rate still remains high, calling for the development of new treatment strategies with higher efficacy. Extensive efforts driven in this direction have included broadening of early cancer screening and applying innovative theranostic nanotechnologies. They have been supported by platforms introduced to enable the detection and monitoring of cancer biomarkers, inhibitors, and other agents, able to slow down cancer progression and prevent metastasis. Despite of the well-recognized principles of the immune checkpoint blockade, the efficacy of immunotherapy achieved so far does not meet the well-founded expectations. For a successful cancer treatment, highly sensitive, robust, and inexpensive multiplex biosensors have to be designed to aid in the biomarkers monitoring and in the development of new inhibitors. In this review, we provide an overview of the efforts undertaken to aid in the development and monitoring of anticancer immunotherapy, based on the programmed cell-death immune checkpoint (PD-1/PDL-1) blockade, by designing biosensors for the detection of relevant cancer biomarkers and their inhibitors screening. This review also emphasizes alternative targets made by exosomes carrying PD-L1 overexpressed in cancer cells and passed into the excreted exosomes. Evaluated are also novel targeted drug delivery nanocarriers, providing simultaneous biosensing, thereby contributing to the emerging immune checkpoint cancer therapy. On the basis of the current trends and the emerging technologies, future perspectives of cancer diagnostics and treatment monitoring using biosensing platforms are projected.

Mitochondrial Dynamics in Neurodegenerative Diseases: Unraveling the Role of Fusion and Fission Processes.

Neurodegenerative diseases (NDs) are a diverse group of disorders characterized by the progressive degeneration and death of neurons, leading to a range of neurological symptoms. Despite the heterogeneity of these conditions, a common denominator is the implication of mitochondrial dysfunction in their pathogenesis. Mitochondria play a crucial role in creating biomolecules, providing energy through adenosine triphosphate (ATP) generated by oxidative phosphorylation (OXPHOS), and producing reactive oxygen species (ROS). When they're not functioning correctly, becoming fragmented and losing their membrane potential, they contribute to these diseases. In this review, we explore how mitochondria fuse and undergo fission, especially in the context of NDs. We discuss the genetic and protein mutations linked to these diseases and how they impact mitochondrial dynamics. We also look at the key regulatory proteins in fusion (MFN1, MFN2, and OPA1) and fission (DRP1 and FIS1), including their post-translational modifications. Furthermore, we highlight potential drugs that can influence mitochondrial dynamics. By unpacking these complex processes, we aim to direct research towards treatments that can improve life quality for people with these challenging conditions.

Gated Resonance Energy Transfer (gRET) Controlled by Programmed Death Protein Ligand 1.

The resonance energy transfer (RET) between an excited fluorescent probe molecule and a plasmonic nanoparticle (AuNP) has been investigated to evaluate the effect of protein molecules on the RET efficiency. We have found that the energy transfer to a functionalized AuNP can be modulated by a sub-monolayer film of programmed death-ligand 1 (PD-L1) protein. The interactions of PD-L1 with AuNP@Cit involve incorporation of the protein in AuNP shell and formation of a submonolayer adsorption film with voids enabling gated surface plasmon resonance energy transfer (SPRET). A model of the gated-RET system based on the protein size, estimated using Fisher-Polikarpov-Craievich density approximation, has been developed and can be utilized for other proteins, with minimum data requirement, as well. The value of the equilibrium constant KL determined for the Langmuir isotherm is high: KL = 1.27 × 108 M-1, enabling highly sensitive control of the gated-RET by PD-L1. Thus, with the gated-RET technique, one can determine PD-L1 within the dynamic range, extending from 1.2 to 50 nM. Moreover, we have found that the Gibbs free energy for PD-L1 binding to AuNP@Cit is -46.26 kJ/mol (-11.05 kcal/mol), indicating a strong adsorption with supramolecular interactions. The proposed gated-RET system, with the fluorescence intensity of the fluorophore probe molecule modulated by plasmonic quenching with AuNP and shielding of energy transfer by the adsorbed PD-L1 can be further developed for determination of PD-L1 in pharmaceutical formulations for immune checkpoint control in cancer therapy.

Monitoring of dynamic ATP level changes by oligomycin-modulated ATP synthase inhibition in SW480 cancer cells using fluorescent "On-Off" switching DNA aptamer.

Adenosine triphosphate (ATP) is the main energy source in cells and an important biomolecule participating in cellular reactions in living organisms. Since the ATP level changes dynamically reflecting the development of a debilitating disease or carcinogenesis, we have focused in this work on monitoring of the oligomycin (OMC)-modulated ATP synthase inhibition using a fluorescent-switching DNA aptamer designed for the detection of ATP (Apt(ATP)), as the model for studies of dynamic ATP level variation. The behavior of the ATP aptamer has been characterized using fluorescence spectroscopy. The Intramolecular fluorescence resonance energy transfer (iFRET) operates in the proposed aptamer from the FAM dye moiety to guanines of the aptamer G-quadruplex when the target ATP is present and binds to the aptamer changing its conformation. The iFRET process enables the detection of ATP down to the limit of detection, LOD = 17 µM, without resorting to any extra chemi-amplification schemes. The selectivity coefficients for relevant interferent triphosphates (UTP, GTP, and CTP) are low for the same concentration as that of ATP. We have demonstrated an efficient transfection of intact cells and OMC-treated SW480 colon cancer cells with Apt(ATP), using microscopic imaging, iFRET measurements, and cell viability testing with MTT method. The applicability of the switching DNA aptamer for the analysis of real samples, obtained by lysis of SW480 cells, was also tested. The proposed Apt(ATP) may be considered as a viable candidate for utilization in measurements of dynamic ATP level modulation in cells in different stages of cancer development and testing of new drugs in pharmacological studies. Graphical abstract.