Skin Cancer Research Goes Digital: Looking for Biomarkers within the Droplets

Abstract

Skin cancer, which includes the most frequent malignant non-melanoma carcinomas (basal cell carcinoma, BCC, and squamous cell carcinoma, SCC), along with the difficult to treat cutaneous melanoma (CM), pose important worldwide issues for the health care system. Despite the improved anti-cancer armamentarium and the latest scientific achievements, many skin cancer patients fail to respond to therapies, due to the remarkable heterogeneity of cutaneous tumors, calling for even more sophisticated biomarker discovery and patient monitoring approaches. Droplet digital polymerase chain reaction (ddPCR), a robust method for detecting and quantifying low-abundance nucleic acids, has recently emerged as a powerful technology for skin cancer analysis in tissue and liquid biopsies (LBs). The ddPCR method, being capable of analyzing various biological samples, has proved to be efficient in studying variations in gene sequences, including copy number variations (CNVs) and point mutations, DNA methylation, circulatory miRNome, and transcriptome dynamics. Moreover, ddPCR can be designed as a dynamic platform for individualized cancer detection and monitoring therapy efficacy. Here, we present the latest scientific studies applying ddPCR in dermato-oncology, highlighting the potential of this technology for skin cancer biomarker discovery and validation in the context of personalized medicine. The benefits and challenges associated with ddPCR implementation in the clinical setting, mainly when analyzing LBs, are also discussed.

Skin Cancer Pathobiology at a Glance: A Focus on Imaging Techniques and Their Potential for Improved Diagnosis and Surveillance in Clinical Cohorts

“…Similarly with CLSM, TPEF allows the visualization of both endogenous (nicotinamide adenine dinucleotide (NADH), flavin adenine dinucleotide (FAD), melanin, keratin, and so on) and exogenous fluorophores, being exploited in fluorescence lifetime imaging for a plethora of experimental approaches [ 3 ]. Extrinsic contrast agents, such as organic fluorophores and genetically expressed fluorescent proteins, are suitable for exploring the skin’s structure and biochemistry with multiphoton microscopy, whereas endogenous contrast agents help investigate the main components of the skin [ 4 ]. Flavin proteins and NAD(P)H are mainly located in mitochondria, but NAD(P)H may also be present in the cytosol.…”

“…Flavin proteins and NAD(P)H are mainly located in mitochondria, but NAD(P)H may also be present in the cytosol. Nonetheless, the fluorescence resulting from reduced pyridine nucleotides and oxidized flavin proteins is very useful in portraying the keratinocytes [ 4 ]. Besides, the endogenous fluorescence of keratin can be exploited to emphasize the presence of this structural protein in the stratum corneum, where it is abundant [ 5 ].…”

“…One of the most important things to consider is the skin’s stratified structure, with different structural layers having very different refractive indices. Briefly, in multiphoton imaging, this multi-layered structure with varying indices of refraction causes spherical aberration and distorts the excitation focus, resulting in signal loss and a reduction in image resolution [ 4 ]. It influences the imaging depth as well.…”

“…The difference in penetration depth between the brain and the skin may be explained by the fact that deep brain imaging targets relatively larger vascular structures. In contrast, skin imaging focuses more on detailed features such as the morphology of individual cells or elastin fibers [ 4 ]. Additionally, skin imaging is based on endogenous fluorophores such as NAD(P)H, which emit at approximately 450 nm, a wavelength that has a short mean free path in tissues, while imaging deep in the brain is often based on exogenous fluorophores emitting more at red wavelengths [ 4 ].…”

“…By far, cumulative exposure to solar UV radiation (UVA and UVB) remains the most prominent risk factor for skin cancer since it leads to genomic instability, inflammation, and the emergence of cutaneous tumors in humans, even decades after initial exposure [ 2 , 3 ]. Based on the cell of origin and clinical behavior, cutaneous tumors are subdivided into two main categories: cutaneous melanoma (CM) and non-melanoma skin cancers (NMSC) [ 4 ]. CM, which evolves from the malignant transformation of melanocytes, the pigment-producing cells in the dermis, accounts for only 4% of skin cancer cases; hence, if left untreated or caught in a late stage, CMs are challenging to treat and potentially life-threatening, being responsible for more than 75% of skin cancer-related deaths [ 5 ].…”