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Oncology
Major limitations in the effective treatment of neurological cancer include systemic cytotoxicity of chemotherapy, inaccessibility, and inoperability. The capability to successfully target a drug to the tumor site(s) without incurring serious side effects—especially in the case of aggressive tumors, such as glioblastoma and neuroblastoma—would represent a significant breakthrough in therapy. Orthotopic systems, capable of storing and releasing proteins over a prolonged period at the site of a tumor, that utilize nanoparticles, liposomes, and hydrogels have been proposed. One candidate for drug delivery is Micro-Fragmented Adipose Tissue (MFAT). Easily obtained from the patient by abdominal subcutaneous liposuction (autologous), and with a high content of Mesenchymal Stem Cells (MSCs), mechanically derived nanofat is a natural tissue graft with a structural scaffold organization. It has a well-preserved stromal vascular fraction and a prolonged capacity to secrete anti-tumorigenic concentrations of pre-absorbed chemotherapeutics within extracellular vesicles.
- MSC
- MFAT
- biomaterials
1. Introduction
Human Micro-Fragmented Adipose Tissue (MFAT) has been intensively studied in the last few years in a wide variety of research fields, showing great potential regarding its anti-inflammatory, anti-proliferative, anti-apoptotic, pro-regenerative, and, most recently, its drug delivery capabilities. Most of these properties of MFAT are associated with the presence of Mesenchymal Stem Cells (MSCs) found within the fat. MFAT is produced mechanically as liposuction-derived fat, using an enclosed sterile micronizer, such as the Lipogems or Lipocell systems. It consists of a heterogeneous mixture of adipocytes, MSCs (pericytes), endothelial cells (EC), cells belonging to the immune system, and connective tissue cells such as fibroblasts, all of which are supported by an extracellular matrix (ECM) composed of collagen fibers, polysaccharides (glycosaminoglycans), and other proteins [1,2,3].
The clinical applications in which adipose-tissue-derived MSCs, and/or MFAT, have demonstrated effectiveness include various surgical specialties [4,5]. For example, menopausal women suffering from lichen sclerosis became asymptomatic concomitant with complete epithelial regeneration of the vulva, over a period of more than a year after multiple subdermal targeted injections of autologous MFAT [6]. Whilst, in a case study, a 35-year-old woman was successfully treated for vesicouterine fistula using MFAT injection together with endoscopic repair, resulting in complete resolution of the pathology [7].
MFAT has proven effective in the treatment of perianal fistulas with Crohn’s disease. Laureti et al. treated 15 patients with persistent complex fistulizing perianal Crohn’s disease. Twenty-four weeks after a single targeted injection of MFAT, 10 of the patients had complete clinical and radiographic remission with no adverse events reported, demonstrating that this could be a novel and potentially effective minimally invasive treatment for inflammatory bowel disease [8].
MFAT has been used most frequently in the field of orthopedic surgery, with promising published clinical studies in the treatment of joint injuries and osteoarthritis (OA) in animals (e.g., racehorses and dogs) and humans. For example, Giorgini et al. treated 49 patients with moderate to severe knee OA using a combination of arthroscopy and a single injection of MFAT. In this 2-year retrospective single-center study, long-term significant improvement as measured with the knee injury and OA outcome score (KOOS) was shown [9].
These and other studies demonstrate that the use of autologous MFAT injection is an innovative and safe method for treating inflammatory-based disease [5,10,11,12]. This procedure is characterized by its simplicity, affordability, speed, minimal invasiveness, one-step application, low risk of complications, and compliance with regulatory guidelines [4].
2. Anti-Inflammatory and Other Anti-Cancer Beneficial Properties of MFAT
2.1. MFAT Extraction and Characterization
MFAT is prepared from abdominal adipose tissue obtained through liposuction using an enclosed sterile process, involving micronization to nanofragments of around 0.3–1 mm in diameter and cleaning to remove excess oil, haematological cells, and fibrous tissue. The most commonly used kit is called LipoGems S.p.A. Firstly, a mini liposuction is carried out in the abdominal region following an injection of Klein solution in a locally anesthetized donor, and approximately 100–200 mL of fat is removed. Next, the fat is ‘sieved’ with a filter set (0.7 mm diameter holes) and homogenized in an enclosed sterile tube, held under low pressure in the presence of ball bearings, and finally washed in a large volume of saline, resulting in the production of approximately 10 mL of MFAT per 100 mL of original extract. The processed MFAT consists primarily of clusters of perivascular cells held together in the form of micrografts with adipocytes, which can be collected in a Luer Lock syringe and injected locally where needed [3,19].
MFAT clusters can be cultured directly in a tissue culture medium in order to assess their size, viability, and activity. In addition, MSCs can be isolated following collagenase-I digestion, with agitation over a period of 1 h. The capacity of MSCs to form colony-forming units (CFU), and the level of secretion of cytokines and growth factors, is indicative of their overall activity and that of the parent MFAT [20].
2.2. MSC Paracrine Effects
MSCs play a crucial role in tissue regeneration and act as medicinal signaling cells. These cells can be obtained from various sources like bone marrow, placenta, umbilical cord blood, and adipose tissue. MSCs and pericytes are effectively the same cells, exhibiting comparable marker expressions and, more importantly, demonstrating similar functional characteristics. Pericytes form the smooth muscle cell coating of microvessels in the fat and, during its activation, the pericytes detach from the surface and become MSCs. It is worth noting that both cell types are considered safe for allogeneic transplantation due to their lack of expression of immune-related, membrane-bound molecules [24].
MSCs secrete a diverse array of bioactive molecules that function in a paracrine manner. These molecules play a crucial role in priming and sustaining angiogenic, anti-fibrotic, anti-apoptotic, and immunomodulatory responses within the target tissue. Maximal therapeutic benefit is conferred, since subcutaneous fat represents the tissue with the highest concentration of MSCs, being far superior to bone-marrow-derived sources [3]. MFAT-derived MSCs produce colony-forming units indicating their stemness.
2.2.1. MSCs and Angiogenesis
Published data have shown that MSCs have the potential to mitigate the extent of cerebral infarction following ischemia and contribute to functional restoration. One proposed mechanism is that MSC transplantation after a stroke enhances angiogenesis, by either producing or amplifying endogenous factors essential for blood vessel formation. These factors include vascular endothelial growth factor (VEGF), angiopoietin-1 (Ang-1), placental growth factor (PlGF), and fibroblast growth factor-2 (FGF-2) [25].
The presence of these growth factors can support the development and the effective maturation of vascular structures associated with revascularization, ultimately—for example—in an ischaemic stroke, MSC treatment can result in a possible reduction of the size of the infarcted area.
MSCs derived from bone marrow were engineered to express kringle-5 (an angiogenesis inhibitor from human plasminogen) under the control of early growth factor-1. Systemic intravenous administration of this modified potential therapeutic resulted in a reduction in tumor growth, and it improved survival in a murine glioblastoma xenograft model, suggesting that MSC-based therapies could act in a dual role, blocking tumor vascularization through direct paracrine secretion and through drug delivery strategies [27].
2.2.2. Neuroprotective Effects of MSCs
A study conducted on experimental animals transplanted human umbilical cord-MSCs (hUC-MSCs) into neonatal rats, resulting in reduced tissue damage and infarct volume due to the migration of cells into the periventricular tissue space. This treatment also led to improved motor function in the neonatal rats. In addition, hUC-MSCs demonstrated significant reductions in apoptosis as well as the expression of beclin-2 and caspase-3, which are critical regulators of the apoptotic cascade [28].
2.2.3. Evidence for the Role of MSCs in Tumor Modulation
Considerations must be given to the potential stimulatory effect of MSC-derived growth factors on angiogenesis-dependent glial tumor growth. Angiogenesis is of critical importance for the optimal proliferation and expansion of glioblastoma [31]. A meta-analysis of clinical trials involving the use of angiogenesis inhibitors showed that only bevacizumab, given as a single treatment, resulted in an improved response to temozolomide, with a significant median progression-free survival at 6 months but no overall improvement in survival time, indicating the relevance but limitation of neo-vascular development to the tumor growth [32].
2.2.4. MSC-Derived Exosomes as Cell Free Targeting Therapeutics
Exosomes are microvesicles released from MSCs that contain all the active proteins which contribute to immunomodulation and cellular regeneration. For this reason, attention has been given to their use in therapy due to their smaller size (allowing penetration through the BBB) and allogeneic capacity. In addition, they retain the encapsulated nature and half-life of the parent cell without the accompanying immuno-compromising characteristics (HLA antigen expression) [37].
Do et al. contemplated the use of exosomes derived from MSCs as a mechanism for targeted delivery in brain tumor treatment. The advantages of exosomes over the parent MSCs include preferential accumulation in the brain parenchyma (homing), an improved safety profile (lower risk of induction of tumor growth), and a longer systemic half-life. Whilst direct parenchymal injection resulted in greater incorporation into the brain tissue than with MSCs. They retain the capacity to uptake and deliver therapeutic drugs and can be targeted, for example, via chemical modification to glioblastoma (neuropilin-1) [40].
2.2.5. Adipocyte Activity from MFAT
Adipocytes themselves possess strong anti-inflammatory properties that contribute significantly to the immunomodulatory function of MFAT. More specifically, adiponectin is a highly secretive immunoprotective molecule that synergizes with MSC-secreted cytokines to protect the balance of the local microenvironment [43].
2.2.6. Uptake of Drugs and Engineered Delivery Vehicles
MSCs can also serve as therapeutic carriers and have recently been shown to absorb chemotherapeutic drugs, releasing them in microvesicles at a regular, useful physiological concentration for up to several months.
The intrinsic ability of MSCs to migrate towards tumor sites makes them promising vehicles for the delivery of pharmacological agents. This approach has shown promise in the targeted delivery of oncolytic viruses as potential anticancer therapies and has potential applications in other cancer-related treatments [45].
3. Proof-of-Concept Evidence for Effective Drug Delivery Using MFAT
Alessandri et al. highlighted that both MFAT and its D-MFAT derivative exhibited comparable and effective abilities to incorporate PTX. Both substances rapidly assimilated PTX, with 5 min being sufficient for them to incorporate 90% to 95% of the chemotherapeutic (2 µg/mL), following the priming of 24 exposures to PTX stock solution (6 mg/mL). The majority of PTX was localized within their adipocytes/lipid content and is dependent upon interaction between the drug and hydrophobic nature of fat. Moreover, they both displayed a slow and efficient release of active PTX in its original form in vitro and could be re-loaded with drugs multiple times without loss of potency and without any other metabolites present. This release persisted for at least 2 months, with D-MFAT releasing double the amount of PTX compared to MFAT [13]. The exact mechanism through which MFAT allows the consistent, slow release of chemotherapeutics, and probably other drugs, is still unknown.
Due to its presentation of spontaneous diseases closely resembling human oncology, the domestic dog is regarded as a valuable animal model for assessing novel drugs and therapeutic approaches. In this regard, a case reported the efficiency of MFAT primed with PTX in a 6-year-old dog diagnosed with mesothelioma. Here, Zeira et al. treated a neutered mixed-breed dog weighing 24 kg, which exhibited progressive weakness, loss of appetite, productive cough, abdominal distension, and breathing difficulties for a period of 2 months. In this case, autologous adipose tissue was obtained through lipo-aspiration from the lumbar flanks of the dog. The adipose tissue was then micro-fragmented using a specialized device that allows for minimal manipulation without the need for enzymatic procedures. The micro-fragmented samples were used as a scaffold for PTX (MFAT–PTX). The treatment protocol involved administering 7 mL of MFAT–PTX into the abdominal and thoracic cavity under ultrasound guidance.
Both MFAT and D-MFAT exhibit properties of natural biological scaffolds, capable of absorbing and releasing a substantial amount of drugs such as PTX. In an orthotopic murine model of human neuroblastoma (HTLA-230), the local administration of D-MFAT–PTX at the tumor site (left adrenal gland), following its surgical resection, resulted in the blocking of cancer relapse, whilst treatment with PTX alone only delayed the recurrence up to 22 days. Pharmacokinetic studies revealed that D-MFAT-delivered PTX (subcutaneously) resulted in a high local concentration, significantly reducing the systemic concentration of PTX compared to systemically administered D-MFAT–PTX (intraperitoneally). These findings strongly support that MFAT can serve as a natural biomaterial capable of absorbing and transporting chemotherapeutic drugs. Unexpectedly, even the devitalized derivative (D-MFAT) remained effective, indicating that the presence of viable MSCs or adipocytes in MFAT was not essential for the uptake, release, and anti-cancer effect of the drug [13].
4. Conclusions
MFAT represents an incredible natural scaffold that has the capacity to deliver targeted combination sets of drugs over a concerted period of time, supporting the more positive treatment outcomes in conditions such as glioblastoma, which occurs in surgically challenging anatomical locations. In addition, the high concentration of MSCs within the tissue graft provides both anti-tumoral activity and positive anti-inflammatory immunomodulation. Limitations may include the requirement in chronic disease for multiple semi-invasive graft injections, although MFAT produced from liposuction can be effectively cryopreserved for future use.
This entry is adapted from the peer-reviewed paper 10.3390/ijms241411530
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