Circulation Half-life Research Articles

DNA Base Pairing-Inspired Supramolecular Nanodrug Camouflaged by Cancer-Cell Membrane for Osteosarcoma Treatment.

Osteosarcoma (OS) is one of the most common bone malignant tumors which mainly develops in adolescents. Although neoadjuvant chemotherapy has improved the prognosis of patients, numerous chemotherapeutic challenges still limit their use. Here, inspired by the Watson-Crick base pairing in nucleic acids, hydrophobic (methotrexate) and hydrophilic (floxuridine) chemo-drugs are mixed and self-assembled into M:F nanoparticles (M:F NPs) through molecular recognition. Then, the obtained NPs are co-extruded with membranes derived from OS cells to form cancer-cell membrane-coated NPs (CCNPs). With protected membranes at the outer layer, CCNPs are highly stable in both physiological and weak acid tumor conditions and possess homologous tumor targeted capability. Furthermore, the proteomic analysis first identifies over 400 proteins reserved in CCNPs, most of them participatingin tumor cell targeting and adhesion processes. In vitro studies reveal that CCNPs significantly inhibit the PI3K/AKT/mTOR pathway, which promotes cell apoptosis and cell cycle arrest. More importantly, cell membrane camouflage significantly prolongs the circulation half-life of CCNPs, elevates the drug accumulation at tumor sites, and promotes anti-tumor efficacy in vivo. As a convenient and effective strategy to construct a biomimetic NP with high drug loading ratio, the CCNPs provide new potentials for precise and synergistic antitumor treatment.

Abstract 4189: Evaluation of implantable cytokine factories in combination with checkpoint inhibitors for eradication of malignant pleural mesothelioma (MPM) tumors in mice with safe and predictable dosing in non-human primates

Abstract Pro-inflammatory cytokines have been approved as a cancer immunotherapy for treatment of metastatic melanoma and renal carcinoma for over 30 years. However, widespread use of cytokine therapy in the clinic is limited by its short half-life in circulation and the associated toxicities that emerge as a result of high systemic exposure. To overcome these limitations, we developed a clinically translatable cytokine delivery platform, called cytokine factories, composed of genetically engineered epithelial cells encapsulated in biocompatible polymers. These cells are able to produce a wide range of natural cytokines (IL2, IL7, IL10, or IL12) and allow for controlled and predictable dosing in vivo. In vivo administration of cytokine factories created a high local cytokine concentration (IP space) without substantial leakage into the systemic circulation. Local, or tumor adjacent, administration of pro-inflammatory (IL-2-based) cytokine factories caused reduction of tumor burden by 70% after only 1 week of treatment when delivered as a monotherapy to mice with malignant pleural mesothelioma (MPM). Importantly, when administered in combination with local anti-PD1 injections, these cytokine factories lead to eradication of these highly aggressive tumors in 7/7 treated mice. To validate the translatability of this platform, we evaluated the safety profile in non-human primates. Significantly, this platform produced local and systemic T cell biomarker profiles that predict efficacy without renal, liver, or general toxicity in non-human primates. Our findings demonstrate the safety and efficacy of cytokine factories in preclinical animal models and provide rationale for future clinical testing for the treatment of metastatic peritoneal cancers in humans. Citation Format: Amanda Nash, Samira Aghlara-Fotovat, Andrea Hernandez, Bertha Castillo, Alexander Lu, Aarthi Pugazenthi, Peter Rios, Sofia Ghani, Ira Joshi, Douglas Isa, Chunyu Xu, Rahul Sheth, Weiyi Peng, Jose Oberholzer, Amir Jazaeri, Hee-Jin Jang, Bryan Burt, Hyun-Sung Lee, Ravi Ghanta, Omid Veiseh. Evaluation of implantable cytokine factories in combination with checkpoint inhibitors for eradication of malignant pleural mesothelioma (MPM) tumors in mice with safe and predictable dosing in non-human primates [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4189.

Abstract 1077: Stability and safety evaluation of ELU001, a targeted C’Dot drug conjugate for the potential treatment of folate receptor alpha-overexpressing cancers

Abstract While several antibody-drug conjugates (ADCs) have shown efficacy in the treatment of solid tumors ADCs remain limited in their therapeutic efficacy due to long circulatory half-life and limited tumor penetration. Ultra-small (sub-10nm) C’Dot-drug-conjugates (CDCs) directly address this, exhibiting deep penetration and retention in solid tumor models, with limited systemic exposure following rapid renal clearance. CDCs can also achieve greater potency by delivering over 10x more payload and by binding to tumor cells with a higher avidity allowing effective killing of tumors with broader range of moderate to high expression of target antigens. ELU001 is a CDC that targets folate receptor alpha (FRα) overexpressing tumors via folic acid moieties on its surface and delivers an average of 21 exatecan topoisomerase-1 inhibitor molecules as its payload. FRα is overexpressed on a variety of tumors including ovarian, endometrial, triple negative breast and non-small cell lung, but is minimally expressed on normal tissues making it an attractive tumor-associated antigen for targeted drug delivery. ELU001 is highly stable in plasma and elicits antitumor efficacy in a variety of cell line and PDX-derived tumor models both in vitro and in vivo. In 15-day repeat dose toxicology and toxicokinetic studies performed in Wistar Han rats and Beagle dogs, ELU001 was well tolerated at up to 0.87 mg/kg/day in rats and 0.174 mg/kg/day in dogs based upon conjugated exatecan concentration when administered on a QWx3 schedule via a 1-hour infusion. Observed dose-related toxicities for both species were limited to the bone marrow and GI tract - the same organs as those observed when free payload (exatecan) was administered suggesting that the delivery of exatecan conjugated to the CDC did not broaden the tissue toxicity profile. Observed toxicities were recovered or substantially reduced by the end of a two-week recovery period. No drug-related hepatic, renal, pulmonary or ocular toxicities were observed and there were no drug-related deaths in the repeat dose toxicity study. TK parameters, estimated in the 15-day GLP studies revealed similar plasma exposure values in males and females for ELU001, Total Exatecan (conjugated and released) and Released Exatecan for rats and dogs, respectively. ELU001 exhibited an average circulatory half-life ranging from approximately 15 to 20 hrs in rats and 24 to 29 hrs in dogs with no accumulation of ELU001, Total Exatecan or Free Exatecan observed from day 1 to day 15. Based upon AUC0-last (hr*ng/ml) released payload levels in the circulation were less than approximately 0.3% and 0.1% of the total payload levels in the rat and the dog respectively. No ELU001 anti-drug antibodies were induced in either species. In summary, ELU001 has a favorable nonclinical safety/TK profile and is currently under evaluation in a clinical safety study - NCT05001282. Citation Format: Gregory P. Adams, Kai Ma, Feng Chen, Marion Scocca, Aranapakam Venkatesan, Cathy Reddick, Mary Hilgart, Thomas Gardinier, Fei Wu, Melik Turker, Peiming Chen, Tin Khor, Vaibhav Patel, Eliel Bayever. Stability and safety evaluation of ELU001, a targeted C’Dot drug conjugate for the potential treatment of folate receptor alpha-overexpressing cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1077.

TP-CSO: A Triptolide Prodrug for Pancreatic Cancer Treatment.

Triptolide (TP) is a potential drug candidate for the treatment of cancer, but its use was hampered by its systemic toxicity and poor water solubility. Hence, a TP-CSO prodrug was synthesized by conjugating TP to chitosan oligosaccharide (CSO), and characterized by 1H NMR, FTIR, DSC and XRD analyses. The TP-CSO containing about 4 wt% of TP exhibited excellent water solubility (15 mg/mL) compared to TP (0.017 mg/mL). Compared with TP, the pharmacokinetics of the conjugate after oral administration showed a three-fold increase in the half-life in the blood circulation and a 3.2-fold increase in AUC (0–∞). The orally administered TP-CSO could more effectively inhibit tumor progression but with much lower systemic toxicity compared with TP, indicating significant potential for further clinical trials. In conclusion, CSO-based conjugate systems may be useful as a platform for the oral delivery of other sparingly soluble drugs.

ELU-FRα-1: A study to evaluate ELU001 in patients with solid tumors that overexpress folate receptor alpha (FRα).

TPS3158 Background: ELU001 is a novel, first-in-class, new molecular entity described as a C’Dot Drug Conjugate (CDC). ELU001 consists of ̃12 folic acid targeting moieties and ̃22 exatecan topoisomerase-1 inhibitor payloads on Cathepsin-B cleavable linkers covalently bound to the surface of each silicon core/polyethylene glycol C’Dot nanoparticle. CDCs are small in size (̃6 nm), have a greater ability to penetrate into and through tumors as compared to ADCs, and are rapidly eliminated by the kidneys. The rapid systemic elimination is expected to lead to less toxicity than is observed with targeting platforms like ADCs that have a longer half-life in circulation. ELU001’s high avidity is believed to promote internalization into FRα overexpressing cancer cells, selectively delivering it’s ̃22 molecules of payload. The first-in-human clinical trial, ELU-FRα-1, is currently recruiting patients that have advanced, recurrent or refractory FRα overexpressing tumors considered to be topoisomerase 1 inhibitor-sensitive based on the literature, and, in the opinion of the Investigator, have no satisfactory therapeutic options available. Methods: This is a Phase 1 / 2 multicenter, open label clinical trial with two parts: Part 1 Dose Escalation and Part 2 Tumor Group Expansion Cohort(s). In Part 1, patients with cancer types with a high likelihood of having FRα overexpressing tumors based on historical data, (specifically, ovarian, endometrial, colorectal, gastric, gastroesophageal junction, triple negative breast, or non-small cell lung cancers, or cholangiocarcinoma), will be enrolled to the study. Patients will receive ELU001 on a weekly dose regimen (QW) (once a week for 3 weeks, 1 week rest) or every other week dose regimen (Q2W, with no rest between cycles). Retrospective analysis of FRα expression will be determined. Part 2 uses a Simon’s Two-Stage design to evaluate 4-6 tumor group expansion cohorts, each consisting of patients with specific tumor types (to be identified based on emerging data) that overexpress FRα (prospectively determined prior to enrollment). The primary objective for Part 1 is to identify the MTD/RP2D and for Part 2 is ORR. Dose Escalation will recruit about 25 patients per dose regimen (QW or Q2W). Dose Expansion will recruit about 15 patients per tumor group expansion cohort. Secondary objectives are anti-tumor activity (DOR, PFS, TFST, PFS2, OS), frequency, severity and tolerability of adverse events, PK, ADA, and FRα expression assessments. The study is actively enrolling in the US – Cohorts 1-2 have been completed without DLT. Enrollment to Cohort 3 began in December 2021. Clinical trial information: NCT05001282 .

Overcoming the Shortcomings of Peptide-Based Therapeutics

Peptides have traditionally been perceived as poor drug candidates due to unfavorable characteristics mainly regarding their pharmacokinetic behavior, including plasma stability, membrane permeability and circulation half-life. Nonetheless, in recent years, general strategies to tackle those shortcomings have been established, and peptides are subsequently gaining increasing interest as drugs due to their unique ability to combine the advantages of antibodies and small molecules. Macrocyclic peptides are a special focus of drug development efforts due to their ability to address so called ‘undruggable’ targets characterized by large and flat protein surfaces lacking binding pockets. Here, the main strategies developed to date for adapting peptides for clinical use are summarized, which may soon help usher in an age highly shaped by peptide-based therapeutics. Nonetheless, limited membrane permeability is still to overcome before peptide therapeutics will be broadly accepted.

TransCon IL-2 β/γ, a novel long-acting prodrug with sustained release of an IL-2Rβ/γ-selective IL-2 analog, demonstrates improved pharmacokinetics and profound expansion of cytotoxic immune cells in non-human primates.

2563 Background: Recombinant interleukin-2 (IL-2, aldesleukin) is an approved cancer immunotherapy but causes severe side effects including cytokine release syndrome (CRS) and vascular leak syndrome (VLS). This is believed to be due to activation of IL-2Rα+ endothelial cells and inflammatory eosinophils as well as high Cmax due to the short half-life requiring frequent high dose administrations. Also, potent activation of immunosuppressive IL-2Rα+ regulatory T cells (Tregs) may limit IL-2’s efficacy. TransCon IL-2 β/γ is a novel long-acting prodrug with sustained release of an IL-2Rβ/γ-selective IL-2 analog designed to optimally address these shortcomings. Methods: Naïve male and female cynomolgus monkeys received 2-3 doses of TransCon IL-2 β/γ intravenously Q2W. Blood samples were taken to assess plasma concentrations of IL-2 β/γ (active drug), serum cytokines, hematology and immunophenotyping. Histopathology was also performed. Results: Administration of TransCon IL-2 β/γ induced robust increases in absolute lymphocyte counts (ALC), with no/minor increases in eosinophil counts. The increase in ALC was primarily driven by potent activation and expansion of CD8+ T cells, NK cells and γδ T cells, relative to CD4+ T cells and Tregs. At doses with maximum pharmacodynamic (PD) responses, TransCon IL-2 β/γ induced on average > 20-fold ALC increases, > 19-fold CD8+ T cell increases, > 50-fold Effector Memory CD8+ T cell increases, > 18-fold NK cell increases and > 400-fold γδ T cell increases compared to pre-dose levels. These changes corresponded with a low Cmax and a long effective half-life ( > 30 hours) for IL-2 β/γ. A clear induction of soluble CD25 and chemokine MCP-1 was seen after dosing, while levels of IL-5, IL-6, IFN-γ and TNF-α remained low. Further, histopathology showed no signs of vascular damage, tissue necrosis, pulmonary edema or eosinophilia. Conclusions: In cynomolgus monkeys, TransCon IL-2 β/γ demonstrated remarkable PD responses in CD8+ T cells, NK cells and γδ T cells over Tregs and eosinophils. No signs of VLS or CRS were observed. These results are likely due to the selective IL-2Rβ/γ bias along with the slow and sustained release of IL-2 β/γ resulting in low Cmax and long circulatory half-life enabled by the TransCon prodrug technology. The massive expansion of γδ T cells, which are known to possess considerable anti-tumor activity, is to our knowledge a unique finding for TransCon IL-2 β/γ among IL-2 variants in clinical development. Altogether, the responses seen in monkeys are suggestive of a potentially substantial improvement in the therapeutic index beyond what has been achieved by aldesleukin or new IL-2 variants to date. TransCon IL-2 β/γ is currently being evaluated in a clinical Phase 1/2 trial as monotherapy and in combination with pembrolizumab (NCT05081609).

Simultaneous Delivery of Doxorubicin and Protease Inhibitor Derivative to Solid Tumors via Star-Shaped Polymer Nanomedicines Overcomes P-gp- and STAT3-Mediated Chemoresistance.

The derivative of protease inhibitor ritonavir (5-methyl-4-oxohexanoic acid ritonavir ester; RD) was recently recognized as a potent P-gp inhibitor and cancerostatic drug inhibiting the proteasome and STAT3 signaling. Therefore, we designed high-molecular-weight HPMA copolymer conjugates with a PAMAM dendrimer core bearing both doxorubicin (Dox) and RD (Star-RD + Dox) to increase the circulation half-life to maximize simultaneous delivery of Dox and RD into the tumor. Star-RD inhibited P-gp activity, potently sensitizing both low- and high-P-gp-expressing cancer cells to the cytostatic and proapoptotic activity of Dox in vitro. Star-RD + Dox possessed higher cytostatic and proapoptotic activities compared to Star-Dox and the equivalent mixture of Star-Dox and Star-RD in vitro. Star-RD + Dox efficiently inhibited STAT3 signaling and induced caspase-3 activation and DNA fragmentation in cancer cells in vivo. Importantly, Star-RD + Dox was found to have superior antitumor activity in terms of tumor growth inhibition and increased survival of mice bearing P-gp-expressing tumors.

Gold Nanorod-Assisted Photothermal Therapy and Improvement Strategies.

Noble metal nanoparticles have been sought after in cancer nanomedicine during the past two decades, owing to the unique localized surface plasmon resonance that induces strong absorption and scattering properties of the nanoparticles. A popular application of noble metal nanoparticles is photothermal therapy, which destroys cancer cells by heat generated by laser irradiation of the nanoparticles. Gold nanorods have stood out as one of the major types of noble metal nanoparticles for photothermal therapy due to the facile tuning of their optical properties in the tissue penetrative near infrared region, strong photothermal conversion efficiency, and long blood circulation half-life after surface modification with stealthy polymers. In this review, we will summarize the optical properties of gold nanorods and their applications in photothermal therapy. We will also discuss the recent strategies to improve gold nanorod-assisted photothermal therapy through combination with chemotherapy and photodynamic therapy.

Combination of αPD-1 and extended half-life IL-2 is effective against the GL261 glioma and uniquely reverses GBM-associated immunosuppression

Abstract Glioblastoma multiforme (GBM) is a deadly CNS malignancy with an average survival of ~12–18 months post diagnosis. The unique immune environment of the brain and the immunosuppressive features of GBM complicate the use of immunotherapies against these tumors. We aimed to promote the efficacy of αPD-1 against the GL261 murine model of GBM by supplementing this therapy with an engineered IL-2 cytokine. IL-2 therapy has been used clinically for metastatic melanoma and renal cancer, but its short half-life in circulation necessitated high doses to keep the drug concentration sufficiently high. Fusing IL-2 to the mouse serum albumin molecule (MSA-IL-2), extends the cytokine’s half-life. Neither αPD-1 nor MSA-IL-2 are effective monotherapies for GL261 tumors, but the combination provides durable tumor clearance and prevents subsequent rechallenge. Most strikingly, this combination therapy cleared established GL261 tumors even in mice incapable of MHC class I restricted antigen presentation. Instead, therapeutic efficacy was abrogated by the depletion of CD4 T cells. Our combination therapy also reversed multiple key features of GBM-associated peripheral immune suppression, including depressed CD4 T cell counts and dampened MHC class II expression. Our combination therapy’s ability to overcome these immune derangements associated with glioma growth and to function independent of MHC class I restricted antigen presentation enhance the appeal of this therapy for future translational adaptation. This work was supported by the National Institutes of Health (R01 NS103212) and the Mayo Clinic-Koch Institute Cancer Solutions Team Grant

Tumor Adjacent IL2 Cytokine Factories Eradicates Ovarian Cancer in Mice Through Cytotoxic T-cell Activation with Safe and Predictable Dosing in Non-human Primates

Abstract Recombinant IL-2 has been approved for the treatment of melanoma and renal cancer. However, IL-2 therapy is limited by its short half-life in circulation and the associated toxicities that emerge as a result of high systemic exposure. To overcome these limitations, we developed a localized cytokine LOCOcyteTM platform composed of allogeneic and engineered cells which produce potent immune effector molecules for local delivery with temporal regulation of activity. Administration of AVB-001 in mice and non-human primates demonstrated that IL2 local concentration (IP space) was 100x higher than the systemic concentration. Additional studies in mice demonstrated dose-dependent levels of IL2 in the intraperitoneal cavity in mice. Treatment of peritoneal tumors in mice using a single administration of AVB-001 provided sustained eradication of peritoneal tumors in an ovarian cancer mouse model. Our data in mice confirmed local increases in activation (CD25+CD8+) (8.6 fold) and proliferation (Ki67+CD8+)(3.4 fold) of cytotoxic T cells within the IP space in comparison to sham mice. Finally, a single administration of AVB-001 led to therapeutic levels of IL2 in the IP space in NHP and produced local and systemic T cell biomarker profiles that predict efficacy, AVB-001 was well tolerated by all animals with no abnormal clinical observations during the study. Our findings demonstrate that the LOCOcyteTM platform has a dose-dependent PK/PD effect, is safe and efficacious in animal models and enables itself to deliver a diverse set of cytokines alone or in combination which is presently explored. Combined, our pre-clinical results provide a rationale for testing AVB-001 in ovarian cancer patients.

Tumor micro-environment targeted collagenase-modified albumin nanoparticles for improved drug delivery

Gemcitabine is a deoxycytidine analogue used as an adjunct therapy in the treatment of various solid tumors. The therapeutic efficacy of this drug is compromised because of its short circulation half-life (∼17 min), fast metabolic degradation and dose dependent adverse effects. Also, there are various barriers in the tumor microenvironment which limit the drug delivery to the solid tumor. Keeping these aspects into consideration, in the present investigation, protease (collagenase) coated gemcitabine-loaded human serum albumin (HSA) nanoparticles (CG-HSANPs) were prepared to improve the drug delivery to the solid tumor compared with uncoated gemcitabine-loaded HSA nanoparticles (G-HSANPs). The desolvation technique was used to develop nanocarriers, which were evaluated for physicochemical properties such as average particle diameter, surface charge, morphology, drug loading and in vitro drug release. The average particle diameter of G-HSANPs and CG-HSANPs was found to be 160.5 ± 4.1 and 181.7 ± 2.54 nm, respectively. The morphology of nanocarrier was investigated by transmission electron microscopy (TEM) and the nanoparticles were also evaluated using differential scanning calorimetry (DSC). The in vitro antiproliferative activity showed comparable cytotoxicity of CG-HSANPs, G-HSANPs and native drug. The fluorescence microscopy showed uptake of collagenase coated HSA nanoparticles (C-HSANPs) and uncoated HSA nanoparticles (HSANPs) in MiaPaCa-2 cells and there was higher nuclear fragmentation, generation of reactive oxygen species and perturbation in mitochondrial membrane potential of CG-HSANPs compared to the native drug. Further, C-HSANPs exhibited improved tumor spheroid penetration as compare to HSANPs. The present study demonstrated the potential of CG-HSANPs as a carrier for improved drug delivery.

Antibody-Incorporated Nanomedicines for Cancer Therapy.

Antibody-based cancer therapy, one of the most significant therapeutic strategies, has achieved considerable success and progress over the past decades. Nevertheless, obstacles including limited tumor penetration, short circulation half-lives, undesired immunogenicity, and off-target side effects remain to be overcome for the antibody-based cancer treatment. Owing to the rapid development of nanotechnology, antibody-containing nanomedicines that have been extensively explored to overcome these obstacles have already demonstrated enhanced anticancer efficacy and clinical translation potential. This review intends to offer an overview of the advancements of antibody-incorporated nanoparticulate systems in cancer treatment, together with the nontrivial challenges faced by these next-generation nanomedicines. Diverse strategies of antibody immobilization, formats of antibodies, types of cancer-associated antigens, and anticancer mechanisms of antibody-containing nanomedicines are provided and discussed in this review, with an emphasis on the latest applications. The current limitations and future research directions on antibody-containing nanomedicines are also discussed from different perspectives to provide new insights into the construction of anticancer nanomedicines.

Enhancing Circulation and Tumor Accumulation of Carboplatin via an Erythrocyte-Anchored Prodrug Strategy.

The short circulatory half-lives and low tumor accumulation of carboplatin greatly limit the drug's efficacy in vivo. Herein, we address these challenges by using a prodrug strategy and present the rational design of a novel platinum(IV) anticancer prodrug that can hitchhike on erythrocytes. This prodrug, designated as ERY1-PtIV , can bind to erythrocytes efficiently and stably, possessing a circulatory half-life 18.5 times longer than that of carboplatin in mice. This elongated circulatory half-life enables platinum to accumulate at levels 7.7 times higher than with carboplatin, with steady levels in the tumors. As a consequence, the ERY1-PtIV prodrug is proved to exhibit significantly enhanced antitumor activity and reduced side effects compared with carboplatin. Collectively, our novel approach highlights an efficient strategy to utilize intrinsic erythrocytes as auto-binding carriers to enhance the tumor accumulation and subsequent antitumor efficacy of platinum drugs.

Bioorthogonal PEGylation Prolongs the Elimination Half-Life of N-TIMP2 While Retaining MMP Inhibition.

Tissue inhibitors of metalloproteinases (TIMPs) are natural inhibitors of the matrix metalloproteinase (MMP) family of proteins, whose members are key regulators of the proteolysis of extracellular matrix components and hence of multiple biological processes. In particular, imbalanced activity of matrix metalloproteinase-14 (MMP-14) may lead to the development of cancer and cardiovascular and other diseases. This study aimed to engineer TIMP2, one of the four homologous TIMPs, as a potential therapeutic by virtue of its ability to bind to the active-site Zn2+ of MMP-14. However, the susceptibility to degradation of TIMP2 and its small size, which results in a short circulation half-life, limit its use as a therapeutic. PEGylation was thus used to improve the pharmacokinetic profile of TIMP2. PEGylation of the MMP-targeting N-terminal domain of TIMP2 (N-TIMP2), via either cysteine or lysine residues, resulted in a significant decrease in N-TIMP2 affinity toward MMP-14 or multisite conjugation and conjugate heterogeneity, respectively. Our strategy designed to address this problem was based on incorporating a noncanonical amino acid (NCAA) into N-TIMP2 to enable site-specific mono-PEGylation. The first step was to incorporate the NCAA propargyl lysine (PrK) at position S31 in N-TIMP2, which does not interfere with the N-TIMP2-MMP-14 binding interface. Thereafter, site-specific PEGylation was achieved via a click chemistry reaction between N-TIMP2-S31PrK and PEG-azide-20K. Inhibition studies showed that PEGylated N-TIMP2-S31PrK did indeed retain its inhibitory activity toward MMP-14. The modified protein also showed improved serum stability vs non-PEGylated N-TIMP2. In vivo pharmacokinetic studies in mice revealed a significant 8-fold increase in the elimination half-life of PEGylated N-TIMP2 vs the non-PEGylated protein. This study shows that site-specific bioorthogonal mono-PEGylation extends the half-life of N-TIMP2 without impairing its biological activity, thereby highlighting the advantage of this strategy for generating potent PEGylated proteins.

Micelle/Hydrogel Composite as a "Natural Self-Emulsifying Reversible Hybrid Hydrogel (N'SERH)" Enhances the Oral Bioavailability of Free (Unconjugated) Resveratrol.

The poor oral bioavailability, rapid biotransformation to less active metabolites, and fast elimination from systemic circulation have been identified as the major limitations responsible for the clinical insignificance of many drug candidates and phytonutrients. Despite the technological advancements in the nanoformulations of synthetic drugs, there exist many challenges for nutritional therapy, due to the regulatory issues, use of high levels of synthetic emulsifiers and polymers, low stability, low loading levels, mainly liquid state, etc. Herein, we report the characterization and human pharmacokinetics of a natural self-emulsifying hybrid-hydrogel formulation of trans-resveratrol prepared by uniformly impregnating resveratrol micelles into the fenugreek galactomannan hydrogel scaffold to form a water-soluble micelle/hydrogel composite in powder form (RF-20). Fourier transform infrared spectroscopy (FTIR), powder X-ray diffraction (PXRD), scanning electron microscopy (SEM), particle size analysis by dynamic light scattering (DLS), and transmission electron microscopy (TEM) demonstrated the uniform impregnation of resveratrol micelles within the galactomannan hydrogel matrix to form a soluble (average particle size of 172.0 ± 10.4 nm and −21.0 ± 2.5 mV zeta potential) and amorphous powder form with smooth and translucent surface morphology for RF-20, with no chemical alterations. Upon pharmacokinetic studies on healthy human subjects (n = 16) following a randomized, double-blinded, placebo-controlled, 2-arm, 4-sequence crossover design and tandem mass spectrometry (UPLC-ESI-MS/MS), 80 mg of trans-resveratrol from RF-20 provided enhanced free resveratrol bioavailability and pharmacokinetic properties compared to the unformulated resveratrol with 98% purity. The enhancement in bioavailability was more when supplemented in sachet (12.98-fold) form than the capsule (10.48-fold) with improved absorption (Cmax = 50.97 ± 15.82 ng/mL), circulation half-life (t1/2 = 7.01 ± 1.44 h), and sustained delivery (Tmax = 4.71 ± 0.73 h), as compared to the unformulated form (Cmax = 15.07 ± 5.10 ng/mL; t1/2 = 1.58 ± 0.65 h; Tmax = 1.21 ± 0.42 h).

Albumin-binding as a universal strategy for half-life extension

With the development of recombinant DNA technology, a growing number of peptides and proteins are being applied to clinical treatment of diseases. However, a huge challenge needed to be addressed for many therapeutic peptides or proteins is that their half-lives in circulation is short due to rapid renal clearance and enzymatic degradation. To overcome the challenge, half-life extension strategies have been extensively developed. Albumin-binding strategy, whether covalent or non-covalent, represent a widely applied and highly successful half-life extension strategy, and many products have been marketed or tested in clinical trials. This review focuses on the utilization of albumin for half-life prolongation and emphasizes on the albumin-binding drugs already on the market or in the clinical stage. We summarize the related techniques including genetic fusion, chemical conjugation, non-covalent binding fatty acid and protein moieties.

Emerging Albumin-Binding Anticancer Drugs for Tumor-Targeted Drug Delivery: Current Understandings and Clinical Translation.

Albumin has shown remarkable promise as a natural drug carrier by improving pharmacokinetic (PK) profiles of anticancer drugs for tumor-targeted delivery. The exogenous or endogenous albumin enhances the circulatory half-lives of anticancer drugs and passively target the tumors by the enhanced permeability and retention (EPR) effect. Thus, the albumin-based drug delivery leads to a potent antitumor efficacy in various preclinical models, and several candidates have been evaluated clinically. The most successful example is Abraxane, an exogenous human serum albumin (HSA)-bound paclitaxel formulation approved by the FDA and used to treat locally advanced or metastatic tumors. However, additional clinical translation of exogenous albumin formulations has not been approved to date because of their unexpectedly low delivery efficiency, which can increase the risk of systemic toxicity. To overcome these limitations, several prodrugs binding endogenous albumin covalently have been investigated owing to distinct advantages for a safe and more effective drug delivery. In this review, we give account of the different albumin-based drug delivery systems, from laboratory investigations to clinical applications, and their potential challenges, and the outlook for clinical translation is discussed. In addition, recent advances and progress of albumin-binding drugs to move more closely to the clinical settings are outlined.

Indocyanine-type Infrared-820 Encapsulated Polymeric Nanoparticle-Assisted Photothermal Therapy of Cancer.

Organic small-molecule photosensitizers are well-characterized and known for the light-responsive treatment modality including photodynamic therapy. Compared with ultraviolet–visible (UV–vis) light used in conventional photodynamic therapy with organic photosensitizers, near-infrared (NIR) light from 700 to 900 nm is less absorbed and scattered by biological tissue such as hemoglobin, lipids, and water, and thus, the use of NIR excitation can greatly increase the penetration depth and emission. Additionally, NIR light has lower energy than UV–vis that can be beneficial due to less activation of fluorophores present in tissues upon NIR irradiation. However, the low water stability, nonspecific distribution, and short circulation half-life of the organic photosensitizers limit its broad biological application. NIR responsive small-molecule fluorescent agents are the focus of extensive research for combined molecular imaging and hyperthermia. Recently a new class of NIR dye, IR-820 with excitation and emission wavelengths of 710 and 820 nm, has been developed and explored as an alternative platform to overcome some of the limitations of the most commonly used gold nanoparticles for photothermal therapy of cancer. Herein, we synthesized a core–shell biocompatible nanocarrier envelope made up of a phospholipid conjugated with poly(ethylene glycol) as a shell, while poly(lactic glycolic acid) (PLGA) was used as a core to encapsulate IR-820 dye. The IR-820-loaded nanoparticles were prepared by nanoprecipitation and characterized for their physicochemical properties and photothermal efficiency. These nanoparticles were monodispersed and highly stable in physiological pH with the hydrodynamic size of 103 ± 8 nm and polydispersity index of 0.163 ± 0.031. The IR-820-loaded nanocarrier showed excellent biocompatibility in the dark, whereas remarkable phototoxicity was observed with breast cancer cells (MCF-7) upon NIR laser excitation. Therefore, the IR-820-loaded phospholipid mimicking biodegradable lipid-polymer composite nanoparticles could have great potential for cancer theranostics.

The Role of in silico Research in Developing Nanoparticle-Based Therapeutics.

Nanoparticles (NPs) hold great potential as therapeutics, particularly in the realm of drug delivery. They are effective at functional cargo delivery and offer a great degree of amenability that can be used to offset toxic side effects or to target drugs to specific regions in the body. However, there are many challenges associated with the development of NP-based drug formulations that hamper their successful clinical translation. Arguably, the most significant barrier in the way of efficacious NP-based drug delivery systems is the tedious and time-consuming nature of NP formulation—a process that needs to account for downstream effects, such as the onset of potential toxicity or immunogenicity, in vivo biodistribution and overall pharmacokinetic profiles, all while maintaining desirable therapeutic outcomes. Computational and AI-based approaches have shown promise in alleviating some of these restrictions. Via predictive modeling and deep learning, in silico approaches have shown the ability to accurately model NP-membrane interactions and cellular uptake based on minimal data, such as the physicochemical characteristics of a given NP. More importantly, machine learning allows computational models to predict how specific changes could be made to the physicochemical characteristics of a NP to improve functional aspects, such as drug retention or endocytosis. On a larger scale, they are also able to predict the in vivo pharmacokinetics of NP-encapsulated drugs, predicting aspects such as circulatory half-life, toxicity, and biodistribution. However, the convergence of nanomedicine and computational approaches is still in its infancy and limited in its applicability. The interactions between NPs, the encapsulated drug and the body form an intricate network of interactions that cannot be modeled with absolute certainty. Despite this, rapid advancements in the area promise to deliver increasingly powerful tools capable of accelerating the development of advanced nanoscale therapeutics. Here, we describe computational approaches that have been utilized in the field of nanomedicine, focusing on approaches for NP design and engineering.