**1. Introduction**

Cutaneous squamous cell carcinoma (cSCC) together with basal cell carcinoma (BCC) represent the most frequent non-melanoma skin cancers [1]. Unlike BCC, cSCC may exhibit an aggressive behavior with a great ability to metastasize to any organ in the body. The most important risk factors associated with cSCC are sun exposure, fair skin phototype, age (mainly diagnosed in middle-aged and older adults), certain beta human papillomavirus

**Citation:** Tampa, M.; Nicolae, I.; Mitran, C.I.; Mitran, M.I.; Ene, C.; Matei, C.; Georgescu, S.R.; Ene, C.D. Serum Sialylation Changes in Actinic Keratosis and Cutaneous Squamous Cell Carcinoma Patients. *J. Pers. Med.* **2021**, *11*, 1027. https://doi.org/ 10.3390/jpm11101027

Academic Editor: Jun Fang

Received: 13 September 2021 Accepted: 12 October 2021 Published: 15 October 2021

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(HPV) types [2], and immunosuppression [2,3]. It is well known that cSCC arises on damaged skin, on sites characterized by chronic inflammation such as scars or burns [4]. Sun-exposed keratinocytes produce a wide range of molecules (e.g., inflammatory cytokines, chemokines, growth factors, etc.) that induce increased vascular permeability and the recruitment of immune cells such as neutrophils, macrophages, etc. However, UV radiation leads to the depletion of Langerhans cells in the epidermis [5]. UV radiation promotes the formation of an inflammatory milieu that contributes to skin carcinogenesis [6]. Chronic inflammation is a cofactor for tumor development and induces local immunosuppression facilitating tumor invasiveness and metastasis. Immunocompromised individuals have a 65- to 250-fold increased risk of developing a cSCC [4,7]. Regarding HPV infection, several studies suggest the role of beta HPV types in the pathogenesis of cSCC [4,8–10]. Beta HPV DNA has been identified in cSCC samples and antibodies against HPV have been detected in the serum of cSCC patients [4]. The main factors predicting a poor outcome are depth of invasion higher than 2 mm, a low degree of differentiation, location in high-risk areas (face, ear, hands, feet, genitalia), perineural involvement, and the presence of multiple tumors [11–13]. cSCC can occur de novo or from the histological transformation of preexisting actinic keratoses (AK), which represent the intradermal proliferation of dysplastic keratinocytes and display potential for malignant transformation into non-melanoma skin cancer, giving rise especially to SCC [14,15].

Carcinogenesis is frequently associated with abnormal sialylation of glycoproteins and glycolipids as a consequence of changes in the activity of sialyltransferases and sialidases [16–19]. Sialic acids are negatively charged sugars that commonly are coupled to the terminal carbohydrate chains of glycoproteins and glycolipids [20]. Aberrant expression of sialic acid plays a crucial role in tumor aggressiveness by promoting cell proliferation, cell-cell interaction, cell migration, angiogenesis, and tumor metastasis [17,19,21–23]. Total serum sialic acid (TSA) and lipid-bound sialic acid (LSA) are significantly elevated in skin cancers [18,24]. Hypersilalylation influences immune cell responses. Sialic acid-binding receptors such as Siglecs modulate the activity of immune cells in the tumor microenvironment, leading to an abnormal inflammatory response [25]. Siglecs are involved in tumor progression and immune evasion, for example, engagement of Siglec-9 or Siglec-E on neutrophils prevents neutrophil-mediated killing of malignant cells [26].

Sialylation is a process mainly governed by sialyltransferases and sialidases [27]. The transfer of sialic acids is modulated by sialyltransferases, a group of glycosyltransferases divided into four families: β-galactoside α2,3-sialyltransferases (ST3Gal-I-VI), β-galactoside α2,6-sialyltransferases (ST6Gal-I and -II), GalNAc α2,6-sialyltransferases (ST6GalNAc-I-VI), and α2,8-sialyltransferases (ST8Sia-I-VI) [28]. During cell differentiation and neoplastic transformation, the expression of sialyltransferases undergoes substantial alterations resulting in phenotypic changes [27–30]. Sialyltransferases are well known as crucial modulators of several important processes such as cell-cell communication, cell-matrix interaction, cell adhesion, cell signaling, and trafficking [29]. Sialidases, or neuraminidases, are glycohydrolytic enzymes that catalyze the hydrolysis of α−glycosidically linked sialic acid residues from carbohydrate groups of glycoproteins and glycolipids [31]. To date, four types of human sialidases have been identified: NEU1, NEU2, NEU3, and NEU4 [32]. In cancer, the alteration of sialidase activity was associated with cell proliferation, invasion, and metastasis [33].

We have previously investigated sialoglycoconjugate abnormalities and anti-ganglioside immune response as possible mechanisms involved in oncogenesis (cutaneous melanoma [23], clear cell renal cell carcinoma [34]), autoimmune diseases (systemic lupus erythematosus, lupus nephritis [35]), and diabetes [36]. Increased sialylation in melanoma cells could represent an event associated with the progression of cutaneous melanoma [17]. Sialoglycoconjugates may promote processes that are involved in the modulation of host immune and inflammatory responses [37]. Sialylation, manifested as the overexpression of TSA, LSA, and orosomucoid, has been shown to be an early, well-expressed event in the initial stages of clear cell renal cell carcinoma [34]. In the medical literature, there are few studies

that have analyzed sialylation in cSCC and AK [29,38–40]. Therefore, the data are scarce and inconclusive. The aim of our study is to investigate the sialyation in patients with cSCC and patients with AK and find reliable serum parameters useful in the diagnosis of cSCC. To achieve these goals, we have investigated both the levels of sialic acid (TSA and LSA) and the levels of the enzymes involved in its metabolism (ST6GalI and NEU3). In addition, we have analyzed the relationship between the studied parameters and the histological characteristics of the tumor (diameter, depth of invasion, Ki67, and ulceration).
