*3.1. Heparin and Heparan Sulfate for Anticancer Drug Delivery*

Hep and low-molecular-weight heparins (LMWHs) are widely used as a clinical anticoagulant due to their ability to bind with and inhibit the serine-threonine antithrombin protease [114]. Hep is also studied and used for applications in other therapeutic areas due to its biocompatibility, for example, wound healing, burn injury treatment, inhibition of inflammation, and metastatic spread of tumor cells [115]. Hep's chemical and physical properties connected with the large surface area of its chain and the presence of reactive functional groups allow efficient binding of different anti-tumor agents. Nanoparticles based on Hep can be applied as efficient anticancer agent carriers with versatile surface chemistry for functionalization and the introduction of biomolecules [116]. Some of the Hep derivatives are used to deliver imaging agents, such as iron oxide nanoparticles, to detect tumor cells in humans [117]. Sodium deoxycholate (DOC)-conjugated Hep derivatives (DOC-heparin) were used to prepare nanoparticles for in vivo tumor targeting and inhibition of angiogenesis based on chemical conjugation and the EPR effect [118]. More substantial anti-tumor effects of the DOC-heparin were achieved in

animal studies compared to Hep alone. Obtained results confirmed that the conjugated Hep retained its ability to inhibit binding with the angiogenic factors, inducing a significant decrease in endothelial tubular formation. In a separate study, dendronized Hep–doxorubicin (Dox) conjugates were prepared, exhibiting a combination of Dox and Hep features and characterized as pH-sensitive drug delivery vehicles [119]. The prepared nanoparticles showed potent anti-tumor activity, induced apoptosis, and significant antiangiogenesis effects in the 4T1 breast tumor model. Additionally, dendronized Hep and the derived nanoparticles with the loaded drug demonstrated no significant toxicity to the healthy organs of both tumor-bearing and healthy mice, which was confirmed by histological analysis.

Park et al. first attached low molecular weight Hep to stearylamine to obtain amphiphilic polymer that was used to prepare self-assembled micelle-like nanoparticles, loaded with docetaxel in their hydrophobic core. The obtained preparation was tested in MCF-7 and MDAMD 231 human breast cancer cells. This approach demonstrated that Hep retained about 30% of its anticoagulant activity, increased the half-life time of docetaxel in the novel preparation used, and significantly inhibited tested cells' viability [120]. Park et al. synthesized an amphiphilic biopolymer made of Hep and deoxycholic acid and prepared nanoparticles loaded with Dox. These nanoparticles were tested for cytotoxicity and anti-tumor effects. The investigated system showed high loading efficiency and a substantial anti-tumor effect [121].

Other studies describe the characteristic properties of Hep-based nanoparticles as potential drug delivery systems, not focusing on specific types of cancer [122].

In summary, Hep is capable of forming nanoparticles upon the introduction of amphiphilic or hydrophobic molecules [116,123,124]. It can also interact with proteins, which leads to the formation of complexes with various biological effects [125,126]. Nevertheless, absorption of blood proteins upon the introduction of Hep nanoparticles into the human body needs to be controlled.
