Scission-Enhanced Molecular Imaging (SEMI).

Positron emission tomography (PET) imaging methods have advanced our understanding of human biology, while targeted radiotherapeutic drug treatments are now routinely used clinically. The field is expected to grow considerably based on an expanding repertoire of available affinity ligands, radionuclides, conjugation chemistries, and their FDA approvals. With this increasing use, strategies for dose reduction have become of high interest to protect patients from unnecessary and off-target toxicity. Here, we describe a simple and powerful method, scission-enhanced molecular imaging (SEMI). The technique allows for rapid corporeal elimination of radionuclides once imaging or theranostic treatment is completed and relies on "click-to-release" bioorthogonal linkers.

Extended Pharmacokinetics Improve Site-Specific Prodrug Activation Using Radiation.

Radiotherapy is commonly used to treat cancer, and localized energy deposited by radiotherapy has the potential to chemically uncage prodrugs; however, it has been challenging to demonstrate prodrug activation that is both sustained in vivo and truly localized to tumors without affecting off-target tissues. To address this, we developed a series of novel phenyl-azide-caged, radiation-activated chemotherapy drug-conjugates alongside a computational framework for understanding corresponding pharmacokinetic and pharmacodynamic (PK/PD) behaviors. We especially focused on an albumin-bound prodrug of monomethyl auristatin E (MMAE) and found it blocked tumor growth in mice, delivered a 130-fold greater amount of activated drug to irradiated tumor versus unirradiated tissue, was 7.5-fold more efficient than a non albumin-bound prodrug, and showed no appreciable toxicity compared to free or cathepsin-activatable drugs. These data guided computational modeling of drug action, which indicated that extended pharmacokinetics can improve localized and cumulative drug activation, especially for payloads with low vascular permeability and diffusivity and particularly in patients receiving daily treatments of conventional radiotherapy for weeks. This work thus offers a quantitative PK/PD framework and proof-of-principle experimental demonstration of how extending prodrug circulation can improve its localized activity in vivo.

Sustained and Localized Drug Depot Release Using Radiation-Activated Scintillating Nanoparticles.

Clinical treatment of cancer commonly incorporates X-ray radiation therapy (XRT), and developing spatially precise radiation-activatable drug delivery strategies may improve XRT efficacy while limiting off-target toxicities associated with systemically administered drugs. Nevertheless, achieving this has been challenging thus far because strategies typically rely on radical species with short lifespans, and the inherent nature of hypoxic and acidic tumor microenvironments may encourage spatially heterogeneous effects. It is hypothesized that the challenge could be bypassed by using scintillating nanoparticles that emit light upon X-ray absorption, locally forming therapeutic drug depots in tumor tissues. Thus a nanoparticle platform (Scintillating nanoparticle Drug Depot; SciDD) that enables the local release of cytotoxic payloads only after activation by XRT is developed, thereby limiting off-target toxicity. As a proof-of-principle, SciDD is used to deliver a microtubule-destabilizing payload MMAE (monomethyl auristatin E). With as little as a 2 Gy local irradiation to tumors, MMAE payloads are released effectively to kill tumor cells. XRT-mediated drug release is demonstrated in multiple mouse cancer models and showed efficacy over XRT alone (p < 0.0001). This work shows that SciDD can act as a local drug depot with spatiotemporally controlled release of cancer therapeutics.

Thyroid Cancers Exhibit Oncogene-Enhanced Macropinocytosis that Is Restrained by IGF1R and Promote Albumin-Drug Conjugate Response.

PURPOSE: Oncogene-driven macropinocytosis fuels nutrient scavenging in some cancer types, yet whether this occurs in thyroid cancers with prominent MAPK-ERK and PI3K pathway mutations remains unclear. We hypothesized that understanding links between thyroid cancer signaling and macropinocytosis might uncover new therapeutic strategies. EXPERIMENTAL DESIGN: Macropinocytosis was assessed across cells derived from papillary thyroid cancer (PTC), follicular thyroid cancer (FTC), non-malignant follicular thyroid, and aggressive anaplastic thyroid cancer (ATC), by imaging fluorescent dextran and serum albumin. The impacts of ectopic BRAFV600E and mutant RAS, genetic PTEN silencing, and inhibitors targeting RET, BRAF, and MEK kinases were quantified. BrafV600E p53-/- ATC tumors in immunocompetent mice were used to measure efficacy of an albumin-drug conjugate comprising microtubule-destabilizing monomethyl auristatin E (MMAE) linked to serum albumin via a cathepsin-cleavable peptide (Alb-vc-MMAE). RESULTS: FTC and ATC cells showed greater macropinocytosis than non-malignant and PTC cells. ATC tumors accumulated albumin at 8.8% injected dose per gram tissue. Alb-vc-MMAE, but not MMAE alone, reduced tumor size by >90% (P < 0.01). ATC macropinocytosis depended on MAPK/ERK activity and nutrient signaling, and increased by up to 230% with metformin, phenformin, or inhibition of IGF1Ri in monoculture but not in vivo. Macrophages also accumulated albumin and express the cognate IGF1R ligand, IGF1, which reduced ATC responsiveness to IGF1Ri. CONCLUSIONS: These findings identify regulated oncogene-driven macropinocytosis in thyroid cancers and demonstrate the potential of designing albumin-bound drugs to efficiently treat them.

Radiation Cleaved Drug-Conjugate Linkers Enable Local Payload Release.

Conjugation of therapeutic payloads to biologics including antibodies and albumin can enhance the selectively of drug delivery to solid tumors. However, achieving activity in tumors while avoiding healthy tissues remains a challenge, and payload activity in off-target tissues can cause toxicity for many such drug-conjugates. Here, we address this issue by presenting a drug-conjugate linker strategy that releases an active therapeutic payload upon exposure to ionizing radiation. Localized X-ray irradiation at clinically relevant doses (8 Gy) yields 50% drug (doxorubicin or monomethyl auristatin E, MMAE) release under hypoxic conditions that are traditionally associated with radiotherapy resistance. As proof-of-principle, we apply the approach to antibody- and albumin-drug conjugates and achieve >2000-fold enhanced MMAE cytotoxicity upon irradiation. Overall, this work establishes ionizing radiation as a strategy for spatially localized cancer drug delivery.

Deciphering albumin-directed drug delivery by imaging.

Albumin is the most abundant plasma protein, exhibits extended circulating half-life, and its properties have long been exploited for diagnostics and therapies. Many drugs intrinsically bind albumin or have been designed to do so, yet questions remain about true rate limiting factors that govern albumin-based transport and their pharmacological impacts, particularly in advanced solid cancers. Imaging techniques have been central to quantifying - at a molecular and single-cell level - the impact of mechanisms such as phagocytic immune cell signaling, FcRn-mediated recycling, oncogene-driven macropinocytosis, and albumin-drug interactions on spatial albumin deposition and related pharmacology. Macroscopic imaging of albumin-binding probes quantifies vessel structure, permeability, and supports efficiently targeted molecular imaging. Albumin-based imaging in patients and animal disease models thus offers a strategy to understand mechanisms, guide drug development and personalize treatments.

Zwitterionic Polymer Electroplating Facilitates the Preparation of Electrode Surfaces for Biosensing.

Surface chemistry critically affects the diagnostic performance of biosensors. An ideal sensor surface should be resistant to nonspecific protein adsorption, yet be conducive to analytical responses. Here a new polymeric material, zwitterionic polypyrrole (ZiPPy), is reported to produce optimal surface condition for biosensing electrodes. ZiPPy combines two unique advantages: the zwitterionic function that efficiently hydrates electrode surface, hindering nonspecific binding of hydrophobic proteins; and the pyrrole backbone, which enables rapid (<7 min), controlled deposition of ZiPPy through electropolymerization. ZiPPy-coated electrodes show lower electrochemical impedance and less nonspecific protein adsorption (low fouling), outperforming bare and polypyrrole-coated electrodes. Moreover, affinity ligands for target biomarkers can be immobilized together with ZiPPy in a single-step electropolymerization. ZiPPy-coated electrodes are developed with specificity for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The prepared sensor detects SARS-CoV-2 antibodies in human saliva down to 50 ng mL-1 , without the need for sample purification or secondary labeling.

Macrophage-Targeted Therapy Unlocks Antitumoral Cross-talk between IFNγ-Secreting Lymphocytes and IL12-Producing Dendritic Cells.

Macrophages often abound within tumors, express colony-stimulating factor 1 receptor (CSF1R), and are linked to adverse patient survival. Drugs blocking CSF1R signaling have been used to suppress tumor-promoting macrophage responses; however, their mechanisms of action remain incompletely understood. Here, we assessed the lung tumor immune microenvironment in mice treated with BLZ945, a prototypical small-molecule CSF1R inhibitor, using single-cell RNA sequencing and mechanistic validation approaches. We showed that tumor control was not caused by CSF1R+ cell depletion; instead, CSF1R targeting reshaped the CSF1R+ cell landscape, which unlocked cross-talk between antitumoral CSF1R- cells. These cells included IFNγ-producing natural killer and T cells, and an IL12-producing dendritic cell subset, denoted as DC3, which were all necessary for CSF1R inhibitor-mediated lung tumor control. These data indicate that CSF1R targeting can activate a cardinal cross-talk between cells that are not macrophages and that are essential to mediate the effects of T cell-targeted immunotherapies and promote antitumor immunity.See related Spotlight by Burrello and de Visser, p. 4.

Gemfibrozil derivatives as activators of soluble guanylyl cyclase - A structure-activity study.

Previous studies demonstrated that anti-hyperlipidemic drug gemfibrozil acts as NO- and heme-independent activator of NO receptor soluble guanylyl cyclase. A series of new gemfibrozil derivatives were synthesized and evaluated for sGC activation. The structure-activity relationship study identified the positions in gemfibrozil's scaffold that are detrimental for sGC activation and those that are amendable for optimizing modifications. Compared with gemfibrozil, compounds 7c and 15b were more potent activators of cGMP-forming activity of purified sGC and exhibited enhanced relaxation of preconstricted mouse thoracic aorta rings. These studies established the overall framework needed for futher improvement of sGC activators based on gemfibrozil scaffold.

Design and Catalyzed Activation of Mycophenolic Acid Prodrugs.

Mycophenolic acid (MPA) and its morpholino ester prodrug mycophenolate mofetil (MMF) are widely used in solid organ transplantation. These drugs prevent rejection due to their potent inhibition of inosine-5'-monophosphate dehydrogenase (IMPDH), an enzyme vital for lymphocyte proliferation. As a strategy to provide localized immunosuppression in cell transplantation, four mycophenolic acid prodrugs designed to release MPA by two distinct mechanisms were synthesized and characterized. A nitrobenzyl ether prodrug was effectively converted to MPA upon exposure to bacterial nitroreductase, while a propargyl ether was converted to the active drug by immobilized Pd0 nanoparticles. In vitro, both prodrugs were inactive against IMPDH and exhibited reduced toxicity relative to the active drug, suggesting their potential for providing localized immunosuppression.

SMALL MOLECULE IMAGING AGENT FOR MUTANT KRAS G12C.

Multiple potent covalent inhibitors for mutant KRAS G12C have been described and some are in clinical trials. These small molecule inhibitors potentially allow for companion imaging probe development, thereby expanding the chemical biology toolkit to investigate mutant KRAS biology. Herein, we synthesized and tested a series of fluorescent companion imaging drugs (CID) for KRAS G12C, using two scaffolds, ARS-1323 and AMG-510. We created four fluorescent derivatives of each by attaching BODIPY dyes. We found that two fluorescent derivatives (BODIPY FL and BODIPY TMR) of ARS-1323 bind mutant KRAS and can be used for biochemical binding screens. Unfortunately, these drugs could not be used as direct imaging agents in cells, likely because of non-specific membrane labeling. To circumvent this challenge, we then used a two step procedure in cancer cells where an ARS-1323 alkyne is used for target binding followed by fluorescence imaging after in situ click chemsitry with picolyl azide Alexa Fluor 647. We show that this approach can be used to image mutant KRAS G12C directly in cells. Given the current lack of mutant KRAS G12C specific antibodies, these reagents could be useful for specific fluorescence imaging.

Ex-vivo generation of drug-eluting islets improves transplant outcomes by inhibiting TLR4-Mediated NFkB upregulation.

The systemic administration of immunosuppressive and anti-inflammatory drugs is routinely employed in organ transplantation to minimize graft rejection and improve graft survival. Localized drug delivery has the potential to improve transplant outcomes by providing sustained exposure to efficacious drug concentrations while avoiding systemic immunosuppression and off-target effects. Here, we describe the synthesis of a novel prodrug and its direct covalent conjugation to pancreatic islets via a cleavable linker. Post-transplant, linker hydrolysis results in the release of a potent anti-inflammatory antagonist of TLR4, localized to the site of implantation. This covalent islet modification significantly reduces the time and the minimal effective dose of islets necessary to achieve normoglycemia in a murine transplantation model. In streptozotocin-induced diabetic C57BL/6 mice a syngeneic transplant of ∼100 modified islets achieved a 100% cure rate by the end of a 4-week monitoring period, compared to a 0% cure rate for untreated control islets. Overall, this direct prodrug conjugation to islets is well tolerated and preserves their functionality while affording significantly superior transplant outcomes. The development of drug-eluting tissues that deliver sustained and localized doses of small-molecule therapeutics represents a novel pathway for enhancing success in transplantation.

Synthesis and immunostimulatory activity of substituted TLR7 agonists.

Fifteen new substituted adenines were synthesized as potential TLR7 agonists. These compounds, along with 9 previously reported compounds, were analyzed for TLR7 activity and for the selective stimulation of B cell proliferation. Several functionalized derivatives exhibit significant activity, suggesting their potential for use as vaccine adjuvants.