One Step Closer to Clinical Translation: Enhanced Tumor Targeting of [99mTc]Tc-DB4 and [111In]In-SG4 in Mice Treated with Entresto

Background: Peptide radioligands may serve as radionuclide carriers to tumor sites overexpressing their cognate receptor for diagnostic or therapeutic purposes. Treatment of mice with the neprilysin (NEP)-inhibitor phosphoramidon was previously shown to improve the metabolic stability and tumor uptake of biodegradable radiopeptides. Aiming to clinical translation of this methodology, we herein investigated the impact of the approved pill Entresto, releasing the potent NEP-inhibitor LBQ657 in vivo, on the stability and tumor uptake of two radiopeptides. Methods: The metabolic stability of [99mTc]Tc-DB4 (DB4, N4-Pro-Gln-Arg-Tyr-Gly-Asn-Gln-Trp-Ala-Val-Gly-His-Leu-Nle-NH2) and [111In]In-SG4 (SG4, DOTA-DGlu-Ala-Tyr-Gly-Trp-Nle-Asp-Phe-NH2) was tested in LBQ657/Entresto-treated mice vs. untreated controls. The uptake in gastrin-releasing peptide receptor (GRPR)-, or cholecystokinin subtype 2 receptor (CCK2R)-positive tumors respectively, was compared between LBQ657/Entresto-treated mice and untreated controls. Results: LBQ657/Entresto treatment induced marked stabilization of [99mTc] Tc-DB4 and [111In]In-SG4 in peripheral mice blood, resulting in equally enhanced tumor uptake at 4 h post-injection. Accordingly, the [99mTc]Tc-DB4 uptake of 7.13 ± 1.76%IA/g in PC-3 tumors increased to 16.17 ± 0.71/17.50 ± 3.70%IA/g (LBQ657/Entresto) and the [111In]In-SG4 uptake of 3.07 ± 0.87%IA/g in A431-CCK2R(+) tumors to 8.11 ± 1.45/9.61 ± 1.70%IA/g. Findings were visualized by SPECT/CT. Conclusions: This study has shown the efficacy of Entresto to notably improve the profile of [99mTc]Tc-DB4 and [111In]In-SG4 in mice, paving the way for clinical translation of this approach.

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Key-Protease Inhibition Regimens Promote Tumor Targeting of Neurotensin Radioligands

Pharmaceutics ◽

10.3390/pharmaceutics12060528 ◽

2020 ◽

Vol 12 (6) ◽

pp. 528

Author(s):

Panagiotis Kanellopoulos ◽

Aikaterini Kaloudi ◽

Marion de Jong ◽

Eric P. Krenning ◽

Berthold A. Nock ◽

...

Keyword(s):

Tumor Targeting ◽

Ace Inhibitor ◽

Tumor Imaging ◽

Colon Adenocarcinoma ◽

Ace Inhibition ◽

Tumor Uptake ◽

Protease Inhibition ◽

Structural Modifications

Neurotensin subtype 1 receptors (NTS1R) represent attractive molecular targets for directing radiolabeled neurotensin (NT) analogs to tumor lesions for diagnostic and therapeutic purposes. This approach has been largely undermined by the rapid in vivo degradation of linear NT-based radioligands. Herein, we aim to increase the tumor targeting of three 99mTc-labeled NT analogs by the in-situ inhibition of two key proteases involved in their catabolism. DT1 ([N4-Gly7]NT(7-13)), DT5 ([N4-βAla7,Dab9]NT(7-13)), and DT6 ([N4-βAla7,Dab9,Tle12]]NT(7-13)) were labeled with 99mTc. Their profiles were investigated in NTS1R-positive colon adenocarcinoma WiDr cells and mice treated or not with the neprilysin (NEP)-inhibitor phosphoramidon (PA) and/or the angiotensin converting enzyme (ACE)-inhibitor lisinopril (Lis). Structural modifications led to the partial stabilization of 99mTc-DT6 in peripheral mice blood (55.1 ± 3.9% intact), whereas 99mTc-DT1 and 99mTc-DT5 were totally degraded within 5 min. Coinjection of PA and/or Lis significantly stabilized all three analogs, leading to a remarkable enhancement of tumor uptake for 99mTc-DT1 and 99mTc-DT5, but was less effective in the case of poorly internalizing 99mTc-DT6. In conclusion, NEP and/or ACE inhibition represents a powerful tool to improve tumor targeting and the overall pharmacokinetics of NT-based radioligands, and warrants further validation in the field of NTS1R-targeted tumor imaging and therapy.

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Proliferation of primary human hepatocytes and prevention of hepatitis B virus reinfection efficiently deplete nuclear cccDNA in vivo

Gut ◽

10.1136/gutjnl-2016-312162 ◽

2017 ◽

Vol 67 (3) ◽

pp. 542-552 ◽

Cited By ~ 58

Author(s):

Lena Allweiss ◽

Tassilo Volz ◽

Katja Giersch ◽

Janine Kah ◽

Giuseppina Raffa ◽

...

Keyword(s):

Acute Infection ◽

In Situ Hybridisation ◽

Human Hepatocytes ◽

Human Hepatocyte ◽

Therapeutic Interventions ◽

Viral Spread ◽

Primary Human Hepatocytes ◽

The Stability ◽

The Impact

ObjectiveThe stability of the covalently closed circular DNA (cccDNA) in nuclei of non-dividing hepatocytes represents a key determinant of HBV persistence. Contrarily, studies with animal hepadnaviruses indicated that hepatocyte turnover can reduce cccDNA loads but knowledge on the proliferative capacity of HBV-infected primary human hepatocytes (PHHs) in vivo and the fate of cccDNA in dividing PHHs is still lacking. This study aimed to determine the impact of human hepatocyte division on cccDNA stability in vivo.MethodsPHH proliferation was triggered by serially transplanting hepatocytes from HBV-infected humanised mice into naïve recipients. Cell proliferation and virological changes were assessed by quantitative PCR, immunofluorescence and RNA in situ hybridisation. Viral integrations were analysed by gel separation and deep sequencing.ResultsPHH proliferation strongly reduced all infection markers, including cccDNA (median 2.4 log/PHH). Remarkably, cell division appeared to cause cccDNA dilution among daughter cells and intrahepatic cccDNA loss. Nevertheless, HBV survived in sporadic non-proliferating human hepatocytes, so that virological markers rebounded as hepatocyte expansion relented. This was due to reinfection of quiescent PHHs since treatment with the entry inhibitor myrcludex-B or nucleoside analogues blocked viral spread and intrahepatic cccDNA accumulation. Viral integrations were detected both in donors and recipient mice but did not appear to contribute to antigen production.ConclusionsWe demonstrate that human hepatocyte division even without involvement of cytolytic mechanisms triggers substantial cccDNA loss. This process may be fundamental to resolve self-limiting acute infection and should be considered in future therapeutic interventions along with entry inhibition strategies.

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Tackling the opioid crisis - development of kappa-opioid receptor peptide agonists for safer analgesic therapy

10.22541/au.163940789.98987711/v1 ◽

2021 ◽

Author(s):

Rink-Jan Lohman ◽

Karnaker Reddy Tupally ◽

Ajit Kandale ◽

Peter Cabot ◽

Harendra Parekh

Keyword(s):

Side Effects ◽

Opioid Receptor ◽

Specific Activity ◽

Opioid Analgesic ◽

Kappa Opioid Receptor ◽

Metabolic Stability ◽

Clinical Translation ◽

Kappa Opioid

The kappa opioid receptor (KOPr) has exceptional potential as an analgesic target, seemingly devoid of the many peripheral side-effects of Mu receptors. Kappa-selective, small molecule pharmaceutical agents have been developed, but centrally mediated side effects have the limited their clinical translation. Here, we modify an active endogenous Dynorphin peptide with the aim of improving drug-likeness and developing safer KOPr agonists for clinical use. Using rational, iterative design and modern peptide chemistry, we developed a series of potent, selective and metabolically stable peptides from Dynorphin 1-7. Peptides were assessed for cAMP-modulation against Kappa, Mu and Delta opioid receptors, metabolic stability, KOPr specificity and binding, and interrogated for in vitro desensitisation and pERK signalling capability. Finally, lead peptides were evaluated for efficacy in Freund’s complete adjuvant rat model of inflammatory nociception. A library of 70 peptides was synthesised and assessed for pharmacological and metabolic stability factors. At least 10 peptide candidates showed low nanomolar activity (˂50 nM) in a cAMP assay, specificity for KORr, and plasma half-life >60 min, with 6 candidates also stable in trypsin. None of the selected peptides showed pERK activity, with a bias towards cAMP signalling. In vivo, KA305 and KA311 showed anti-nociception opioid receptor-specific activity comparable to morphine and U50 844. These highly potent and metabolically stable peptides are promising opioid analgesic leads for clinical translation. Since they are biased peptide KOPr agonists, it is plausible they lack many of the most significant side effects, such as tolerance, addiction, sedation and euphoria/dysphoria, common to opioid analgesics.

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PET FES measures in vivo pharmacodynamics of endocrine therapy

Journal of Clinical Oncology ◽

10.1200/jco.2007.25.18_suppl.14110 ◽

2007 ◽

Vol 25 (18_suppl) ◽

pp. 14110-14110

Author(s):

H. M. Linden ◽

D. A. Mankoff ◽

K. A. Krohn ◽

J. M. Link ◽

S. Stekhova ◽

...

Keyword(s):

Breast Cancer ◽

Hormonal Therapy ◽

Receptor Expression ◽

Tumor Uptake ◽

Hormonal Manipulation ◽

Hormone Sensitive ◽

Initiation Of Therapy ◽

The Impact

14110 Background: Many clinical options are available for management of hormone sensitive breast cancer, including agents which lower estrogen levels such as aromatase inhibitors (AIs) and agents with block ligand binding to receptor such as tamoxifen (TAM) or fulvestrant (FUL). Estrogen receptor (ER) function is essential for sensitivity to hormonal manipulation in breast cancer treatment. We and others have previously shown that functional ER imaging using PET FES predicts response to hormonal therapy using a quantitative threshold of SUV >1.5. Herein we report that FES PET provides a unique insight into in vivo pharmacodynamics of ER therapy. We hypothesized that the impact of therapy on estradiol binding to ER, measured by FES PET, differs between AIs and ER antagonists, and that early changes in receptor expression/occupancy show efficacy of drug at the tumor target. Methods: Patients undergoing treatment with AI, TAM, or FUL underwent baseline PET FDG and FES, and follow-up PET FES imaging at 2–8 weeks post initiation of therapy. Results: We observed the following changes in FES uptake on hormonal therapy: Mean percent change in FES SUV were 54% decline for TAM and FUL vs. 14% decline for AI treated patients (p<.001). Patients on TAM showed complete blockade of tumor FES uptake on therapy (5/5 with residual SUV <1.5), whereas patients on FUL had variable uptake and incomplete blockade at tumor sites in most patients (4/11 with residual SUV <1.5) (p < .05 FUL vs. TAM), despite consistent blockade of uterine FES uptake in patients where the uterus was visualized pre-FUL. Patients on AI therapy (n=14) had variable tumor uptake following treatment initiation. Conclusions: PET FES effectively monitors the in vivo activity of therapy. Estrogen blocking therapies result in a greater change in tumoral estradiol binding than in ligand depletion. TAM effectively blocks uptake of FES as would be predicted by the mechanism of action of this agent. However, FUL (while blocking uterine uptake) incompletely blocks tumor uptake, providing a mechanism to explain reduced activity of this agent in some patients. Ongoing analysis is designed to assess whether early changes in FES predict response or clinical benefit. No significant financial relationships to disclose.

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Octreotide Conjugates for Tumor Targeting and Imaging

Pharmaceutics ◽

10.3390/pharmaceutics11050220 ◽

2019 ◽

Vol 11 (5) ◽

pp. 220 ◽

Cited By ~ 8

Author(s):

Eduard Figueras ◽

Ana Martins ◽

Adina Borbély ◽

Vadim Le Joncour ◽

Paola Cordella ◽

...

Keyword(s):

Tumor Targeting ◽

Somatostatin Receptors ◽

Cytotoxic Agents ◽

Cyanine Dye ◽

Delivery Vehicle ◽

Efficient Delivery ◽

The Stability ◽

G Protein Coupled

Tumor targeting has emerged as an advantageous approach to improving the efficacy and safety of cytotoxic agents or radiolabeled ligands that do not preferentially accumulate in the tumor tissue. The somatostatin receptors (SSTRs) belong to the G-protein-coupled receptor superfamily and they are overexpressed in many neuroendocrine tumors (NETs). SSTRs can be efficiently targeted with octreotide, a cyclic octapeptide that is derived from native somatostatin. The conjugation of cargoes to octreotide represents an attractive approach for effective tumor targeting. In this study, we conjugated octreotide to cryptophycin, which is a highly cytotoxic depsipeptide, through the protease cleavable Val-Cit dipeptide linker using two different self-immolative moieties. The biological activity was investigated in vitro and the self-immolative part largely influenced the stability of the conjugates. Replacement of cryptophycin by the infrared cyanine dye Cy5.5 was exploited to elucidate the tumor targeting properties of the conjugates in vitro and in vivo. The compound efficiently and selectively internalized in cells overexpressing SSTR2 and accumulated in xenografts for a prolonged time. Our results on the in vivo properties indicate that octreotide may serve as an efficient delivery vehicle for tumor targeting.

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Corynebacterium Species Inhibit Streptococcus pneumoniae Colonization and Infection of the Mouse Airway

Frontiers in Microbiology ◽

10.3389/fmicb.2021.804935 ◽

2022 ◽

Vol 12 ◽

Author(s):

Kadi J. Horn ◽

Alexander C. Jaberi Vivar ◽

Vera Arenas ◽

Sameer Andani ◽

Edward N. Janoff ◽

...

Keyword(s):

Streptococcus Pneumoniae ◽

Respiratory Health ◽

Upper Airway ◽

Inhibitory Effect ◽

Airway Microbiome ◽

The Stability ◽

Independent Effects ◽

The Impact

The stability and composition of the airway microbiome is an important determinant of respiratory health. Some airway bacteria are considered to be beneficial due to their potential to impede the acquisition and persistence of opportunistic bacterial pathogens such as Streptococcus pneumoniae. Among such organisms, the presence of Corynebacterium species correlates with reduced S. pneumoniae in both adults and children, in whom Corynebacterium abundance is predictive of S. pneumoniae infection risk. Previously, Corynebacterium accolens was shown to express a lipase which cleaves host lipids, resulting in the production of fatty acids that inhibit growth of S. pneumoniae in vitro. However, it was unclear whether this mechanism contributes to Corynebacterium-S. pneumoniae interactions in vivo. To address this question, we developed a mouse model for Corynebacterium colonization in which colonization with either C. accolens or another species, Corynebacterium amycolatum, significantly reduced S. pneumoniae acquisition in the upper airway and infection in the lung. Moreover, the lungs of co-infected mice had reduced pro-inflammatory cytokines and inflammatory myeloid cells, indicating resolution of infection-associated inflammation. The inhibitory effect of C. accolens on S. pneumoniae in vivo was mediated by lipase-dependent and independent effects, indicating that both this and other bacterial factors contribute to Corynebacterium-mediated protection in the airway. We also identified a previously uncharacterized bacterial lipase in C. amycolatum that is required for inhibition of S. pneumoniae growth in vitro. Together, these findings demonstrate the protective potential of airway Corynebacterium species and establish a new model for investigating the impact of commensal microbiota, such as Corynebacterium, on maintaining respiratory health.

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Elucidating the Influence of Tumor Presence on the Polymersome Circulation Time in Mice

Pharmaceutics ◽

10.3390/pharmaceutics11050241 ◽

2019 ◽

Vol 11 (5) ◽

pp. 241 ◽

Cited By ~ 7

Author(s):

Robin M. de Kruijff ◽

René Raavé ◽

Annemarie Kip ◽

Janneke Molkenboer-Kuenen ◽

Stefan J. Roobol ◽

...

Keyword(s):

Polyethylene Oxide ◽

Tumor Targeting ◽

Ex Vivo ◽

Circulation Time ◽

Tumor Bearing ◽

Significant Difference ◽

The Impact ◽

Gamma Emitter ◽

Ex Vivo Biodistribution

The use of nanoparticles as tumor-targeting agents is steadily increasing, and the influence of nanoparticle characteristics such as size and stealthiness have been established for a large number of nanocarrier systems. However, not much is known about the impact of tumor presence on nanocarrier circulation times. This paper reports on the influence of tumor presence on the in vivo circulation time and biodistribution of polybutadiene-polyethylene oxide (PBd-PEO) polymersomes. For this purpose, polymersomes were loaded with the gamma-emitter 111In and administered intravenously, followed by timed ex vivo biodistribution. A large reduction in circulation time was observed for tumor-bearing mice, with a circulation half-life of merely 5 min (R2 = 0.98) vs 117 min (R2 = 0.95) in healthy mice. To determine whether the rapid polymersome clearance observed in tumor-bearing mice was mediated by macrophages, chlodronate liposomes were administered to both healthy and tumor-bearing mice prior to the intravenous injection of radiolabeled polymersomes to deplete their macrophages. Pretreatment with chlodronate liposomes depleted macrophages in the spleen and liver and restored the circulation time of the polymersomes with no significant difference in circulation time between healthy mice and tumor-bearing mice pretreated with clodronate liposomes (15.2 ± 1.2% ID/g and 13.6 ± 2.7% ID/g, respectively, at 4 h p.i. with p = 0.3). This indicates that activation of macrophages due to tumor presence indeed affected polymersome clearance rate. Thus, next to particle design, the presence of a tumor can also greatly impact circulation times and should be taken into account when designing studies to evaluate the distribution of polymersomes.

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New insight into the reaction mechanism of carbon disulfide hydrolysis and the impact of H2S with density functional modeling

New Journal of Chemistry ◽

10.1039/c8nj06078a ◽

2019 ◽

Vol 43 (5) ◽

pp. 2347-2352 ◽

Cited By ~ 3

Author(s):

Xin Song ◽

Chi Wang ◽

Khaled A. M. Gasem ◽

Kai Li ◽

Xin Sun ◽

...

Keyword(s):

Reaction Mechanism ◽

Carbon Disulfide ◽

Density Functional ◽

Functional Modeling ◽

One Step ◽

The Stability ◽

The Impact ◽

Insight Into

One-step hydrolysis can easily take place without a catalyst. H2S is beneficial in maintaining the stability of the intermediates.

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Voltammetric Behaviour of Drug Molecules as a Predictor of Metabolic Liabilities

Scientia Pharmaceutica ◽

10.3390/scipharm88040046 ◽

2020 ◽

Vol 88 (4) ◽

pp. 46 ◽

Cited By ~ 1

Author(s):

Hikari Fuchigami ◽

Mandeep K. Bal ◽

Dale A. C. Brownson ◽

Craig E. Banks ◽

Alan M. Jones

Keyword(s):

Drug Stability ◽

Pharmaceutical Research ◽

Parent Drug ◽

Vital Role ◽

Oxidation Potential ◽

The Body ◽

Metabolic Stability ◽

Drug Molecules ◽

The Stability

Electron transfer plays a vital role in drug metabolism and underlying toxicity mechanisms. Currently, pharmaceutical research relies on pharmacokinetics (PK) and absorption, distribution, metabolism, elimination and toxicity (ADMET) measurements to understand and predict drug reactions in the body. Metabolic stability (and toxicity) prediction in the early phases of the drug discovery and development process is key in identifying a suitable lead compound for optimisation. Voltammetric methods have the potential to overcome the significant barrier of new drug failure rates, by giving insight into phase I metabolism events which can have a direct bearing on the stability and toxicity of the parent drug being dosed. Herein, we report for the first time a data-mining investigation into the voltammetric behaviour of reported drug molecules and their correlation with metabolic stability (indirectly measured via t½), as a potential predictor of drug stability/toxicity in vivo. We observed an inverse relationship between oxidation potential and drug stability. Furthermore, we selected and prepared short- (<10 min) and longer-circulation (>2 h) drug molecules to prospectively survey the relationship between oxidation potential and stability.

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[99mTc]Tc-DB1 Mimics with Different-Length PEG Spacers: Preclinical Comparison in GRPR-Positive Models

Molecules ◽

10.3390/molecules25153418 ◽

2020 ◽

Vol 25 (15) ◽

pp. 3418

Author(s):

Panagiotis Kanellopoulos ◽

Emmanouil Lymperis ◽

Aikaterini Kaloudi ◽

Marion de Jong ◽

Eric P. Krenning ◽

...

Keyword(s):

Severe Combined Immunodeficiency ◽

Tumor Uptake ◽

Single Digit ◽

Spacer Length ◽

Uptake Rates ◽

Specific Uptake ◽

Immunodeficiency Disease ◽

Grpr Antagonist ◽

The Impact

Background: The frequent overexpression of gastrin-releasing peptide receptors (GRPRs) in human cancers provides the rationale for delivering clinically useful radionuclides to tumor sites using peptide carriers. Radiolabeled GRPR antagonists, besides being safer for human use, have often shown higher tumor uptake and faster background clearance than agonists. We herein compared the biological profiles of the GRPR-antagonist-based radiotracers [99mTc]Tc-[N4-PEGx-DPhe6,Leu-NHEt13]BBN(6-13) (N4: 6-(carboxy)-1,4,8,11-tetraazaundecane; PEG: polyethyleneglycol): (i) [99mTc]Tc-DB7 (x = 2), (ii) [99mTc]Tc-DB13 (x = 3), and (iii) [99mTc]Tc-DB14 (x = 4), in GRPR-positive cells and animal models. The impact of in situ neprilysin (NEP)-inhibition on in vivo stability and tumor uptake was also assessed by treatment of mice with phosphoramidon (PA). Methods: The GRPR affinity of DB7/DB13/DB14 was determined in PC-3 cell membranes, and cell binding of the respective [99mTc]Tc-radioligands was assessed in PC-3 cells. Each of [99mTc]Tc-DB7, [99mTc]Tc-DB13, and [99mTc]Tc-DB14 was injected into mice without or with PA coinjection and 5 min blood samples were analyzed by HPLC. Biodistribution was conducted at 4 h postinjection (pi) in severe combined immunodeficiency disease (SCID) mice bearing PC-3 xenografts without or with PA coinjection. Results: DB7, -13, and -14 displayed single-digit nanomolar affinities for GRPR. The uptake rates of [99mTc]Tc-DB7, [99mTc]Tc-DB13, and [99mTc]Tc-DB14 in PC-3 cells was comparable and consistent with a radioantagonist profile. The radiotracers were found to be ≈70% intact in mouse blood and >94% intact after coinjection of PA. Treatment of mice with PA enhanced tumor uptake. Conclusions: The present study showed that increase of PEG-spacer length in the [99mTc]Tc-DB7–[99mTc]Tc-DB13–[99mTc]Tc-DB14 series had little effect on GRPR affinity, specific uptake in PC-3 cells, in vivo stability, or tumor uptake. A significant change in in vivo stability and tumor uptake was observed only after treatment of mice with PA, without compromising the favorably low background radioactivity levels.

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