Many drug-resistant tumors and cancer stem cells (CSC) express elevated levels of CD44 receptor, a cellular glycoprotein binding hyaluronic acid (HA). Here, we report the synthesis of nanogel-drug conjugates based on membranotropic cholesteryl-HA (CHA) for efficient targeting and suppression of drug-resistant tumors. These conjugates significantly increased the bioavailability of poorly soluble drugs with previously reported activity against CSC, such as etoposide, salinomycin, and curcumin. The small nanogel particles (diam. 20–40 nm) with a hydrophobic core and high drug loads (up to 20%) formed after ultrasonication and demonstrated a sustained drug release following the hydrolysis of biodegradable ester linkage. Importantly, CHA-drug nanogels demonstrated 2–7 times higher cytotoxicity in CD44-expressing drug-resistant human breast and pancreatic adenocarcinoma cells compared to free drugs and non-modified HA-drug conjugates. These nanogels were efficiently internalized via CD44 receptor-mediated endocytosis and simultaneous interaction with the cancer cell membrane. Anchoring by cholesterol moieties in the cellular membrane after nanogel unfolding evidently caused more efficient drug accumulation in cancer cells compared to non-modified HA-drug conjugates. CHA-drug nanogels were able to penetrate multicellular cancer spheroids and displayed higher cytotoxic effect in the system modeling tumor environment than both free drugs and HA-drug conjugates. In conclusion, the proposed design of nanogel-drug conjugates allowed us to significantly enhance drug bioavailability, cancer cell targeting, and the treatment efficacy against drug-resistant cancer cells and multicellular spheroids.

Tumor-associated macrophages (TAMs) form approximately 50% of tumor mass. TAMs were shown to promote tumor growth by suppressing immunocompetent cells, inducing neovascularization and supporting cancer stem cells. TAMs retain mobility in tumor mass, which can potentially be employed for better intratumoral biodistribution of nanocarriers and effective tumor growth inhibition. Due to the importance of TAMs, they are increasingly becoming principal targets of novel therapeutic approaches. In this review, we compare features of macrophages and TAMs that are essential for TAM-directed therapies, and illustrate the advantages of nanomedicine that are related to the preferential capture of nanocarriers by Mφ in the process of drug delivery. We discuss recent efforts in reprogramming or inhibiting tumor-protecting properties of TAMs, and potential strategies to increase efficacy of conventional chemotherapy by combining with macrophage-associated delivery of nanodrugs.

Inherent or therapy-induced drug resistance is a major clinical setback in cancer treatment. The extensive usage of cytotoxic nucleobases and nucleoside analogs in chemotherapy also results in the development of specific mechanisms of drug resistance; such as nucleoside transport or activation deficiencies. These drugs are prodrugs; and being converted into the active mono-, di- and triphosphates inside cancer cells following administration, they affect nucleic acid synthesis, nucleotide metabolism, or sensitivity to apoptosis. Previously, we have actively promoted the idea that the nanodelivery of active nucleotide species, e.g. 5′-triphosphates of nucleoside analogs, can enhance drug efficacy and reduce nonspecific toxicity. In this study we report the development of a novel type of drug nanoformulations, polymeric conjugates of nucleoside analogs, which are capable of the efficient transport and sustained release of phosphorylated drugs. These drug conjugates have been synthesized, starting from cholesterol-modified mucoadhesive polyvinyl alcohol or biodegradable dextrin, by covalent attachment of nucleoside analogs through a tetraphosphate linker. Association of cholesterol moieties in aqueous media resulted in intramolecular polymer folding and the formation of small nanogel particles containing 0.5 mmol/g of a 5′-phosphorylated nucleoside analog, e.g. 5-fluoro-2′-deoxyuridine (floxuridine, FdU), an active metabolite of anticancer drug 5-fluorouracyl (5-FU). The polymeric conjugates demonstrated rapid enzymatic release of floxuridine 5′-phosphate and much slower drug release under hydrolytic conditions (pH 1.0–7.4). Among the panel of cancer cell lines, all studied polymeric FdU-conjugates demonstrated an up to 50 times increased cytotoxicity in human prostate cancer PC-3, breast cancer MCF-7 and MDA-MB-231 cells, and more than 100 times higher efficacy against cytarabine-resistant human T-lymphoma (CEM/araC/8) and gemcitabine-resistant follicular lymphoma (RL7/G) cells as compared to free drugs. In the initial in vivo screening, both PC-3 and RL7/G subcutaneous tumor xenograft models showed enhanced sensitivity to sustained drug release from polymeric FdU-conjugate after peritumoral injections and significant tumor growth inhibition. All these data demonstrate a remarkable clinical potential of novel polymeric conjugates of phosphorylated nucleoside analogs, especially as new therapeutic agents against drug-resistant tumors.

A majority of nanoencapsulated drugs that have shown promise in cancer chemotherapy are administered intravenously. Development of effective oral nanoformulations presents a very challenging medical goal. Here, we describe successful applications of innovative polymeric nanogels in the form of conjugates with activated nucleoside analogs for oral administration in cancer chemotherapy. Previously, we reported the synthesis of amphiphilic polyvinyl alcohol and dextrin-based nanogel conjugates with the phosphorylated 5-FU nucleoside Floxuridine and demonstrated their enhanced activity against regular and drug-resistant cancers[1]. In this study, we synthesized and evaluated oral applications of nanogel conjugates of a protected Gemcitabine, the drug never used in oral therapies. These conjugates were able to quickly release an active form of the drug (Gemcitabine 5′-mono-, di- and triphosphates) by specific enzymatic activities, or slowly during hydrolysis. Gemcitabine conjugates demonstrated up to 127 times higher in vitro efficacy than the free drug against various cancer cells, including the lines resistant to nucleoside analogs. Surprisingly, these nanogel-drug conjugates were relatively stable in gastric conditions and able to actively penetrate through the gastrointestinal barrier based on permeability studies in Caco-2 cell model. In tumor xenograft models of several drug-resistant human cancers, we observed an efficient inhibition of tumor growth and extended the life-span of the animals by 4 times that of the control with orally treated Gemcitabine- or Floxuridine-nanogel conjugates. Thus, we have demonstrated a potential of therapeutic nanogel conjugates with the activated and stabilized Gemcitabine as a successful oral drug form against Gemcitabine-resistant and other drug-resistant tumors.