A cleavable cytolysin–neuropeptide Y bioconjugate enables specific drug delivery and demonstrates intracellular mode of action

Ahrens, Verena M.; Kostelnik, Katja B.; Rennert, Robert C.; Böhme, David; Kalkhof, Stefan; Kosel, David; Weber, Lutz; Bergen, Martin von; Beck‐Sickinger, Annette G.

doi:10.1016/j.jconrel.2015.04.037

Cited by 11 publications

(16 citation statements)

References 48 publications

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“…In a first step, we aimed to verify selective Y 2 R signaling by treating the cells either with Y 2 R specific agonist Ahx 5-24 -NPY or Y 1 R specific agonist F 7 , P 34 -NPY. Stimulation of the cells with the selective Y 2 R agonist Ahx [5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22][23][24] -NPY resulted in a robust receptor activation (Fig. 4d, dark grey curve, EC 50 : 41.9 nM), whereas no Ca 2+signal was detectable when the cells were treated with selective Y 1 R agonist F 7 , P 34 -NPY (Fig.…”

Section: High G Protein Turnover Of Y 2 R Is Terminated By Depleted Imentioning

confidence: 99%

“…These mechanisms are of great interest since many pharmacological agents targeting GPCR display diminished effectiveness over time [4][5][6][7]. Moreover, such knowledge is required for applications that use GPCR as a shuttle system for intracellular drug delivery [8,9]. To date, approximately 35% of approved drugs address GPCR and the number of promising targets increases steadily [10].…”

Section: Introductionmentioning

confidence: 99%

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Unusually persistent Gαi-signaling of the neuropeptide Y2 receptor depletes cellular Gi/o pools and leads to a Gi-refractory state

et al. 2020

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Background: A sensitive balance between receptor activation and desensitization is crucial for cellular homeostasis. Like many other GPCR, the human neuropeptide Y 2 receptor (hY 2 R) undergoes ligand dependent activation and internalization into intracellular compartments, followed by recycling to the plasma membrane. This receptor is involved in the pathophysiology of distinct diseases e.g. epilepsy and cancer progression and conveys anorexigenic signals which makes it an interesting and promising anti-obesity target. However, Y 2 R desensitization was observed after daily treatment with a selective PYY 13-36 analog in vivo by a yet unknown mechanism. Materials: We studied the desensitization and activatability of recycled Y 2 R in transiently transfected HEK293 cells as well as in endogenously Y 2 R expressing SH-SY5Y and SMS-KAN cells. Results were evaluated by one-way ANOVA and Tukey post test. Results: We observed strong desensitization of the Y 2 R in a second round of stimulation despite its reappearance at the membrane. Already the first activation of the Y 2 R leads to depletion of the functional cellular Gα i/o protein pool and consequently desensitizes the linked signal transduction pathways, independent of receptor internalization. This desensitization also extends to other Gα i/o-coupled GPCR and can be detected in transfected HEK293 as well as in SH-SY5Y and SMS-KAN cell lines, both expressing the Y 2 R endogenously. By overexpression of chimeric Gα qi proteins in a model system, activation has been rescued, which identifies a critical role of the G protein status for cellular signaling. Furthermore, Y 2 R displays strong allosteric coupling to inhibitory G proteins in radioligand binding assays, and loses 10-fold affinity in the G protein-depleted state observed after activation, which can be largely abrogated by overexpression of the Gα i-subunit. Conclusion: The unusually persistent Gα i-signaling of the Y 2 R leads to a state of cellular desensitization of the inhibitory Gα i-pathway. The strong allosteric effects of the Y 2 R-Gα i-interaction might be a mechanism that contributes to the burst of Gα i-signaling, but also serves as a mechanism to limit the Y 2-mediated signaling after recycling. Thus, the cell is left in a refractory state, preventing further Gα i-signaling of the Y 2 R itself but also other Gα i/o-coupled receptors by simply controlling the repertoire of downstream effectors.

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Section: High G Protein Turnover Of Y 2 R Is Terminated By Depleted Imentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Unusually persistent Gαi-signaling of the neuropeptide Y2 receptor depletes cellular Gi/o pools and leads to a Gi-refractory state

et al. 2020

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“…For hY 1 R, a number of selective full‐length ligands with 36 amino acids have been reported, most prominently [Phe 7 ,Pro 34 ]NPY . Since 2001, hY 1 R has been known to be overexpressed in breast tumour tissue, making selective agonists even more relevant for use as shuttles for the site‐specific delivery of cytotoxic drugs . The preference of an agonist over an antagonist for that purpose lies in the ability of the agonist to induce internalisation of the receptor–peptide complex, thereby delivering the conjugated drug into the cell .…”

Section: Discussionmentioning

confidence: 99%

“…The preference of an agonist over an antagonist for that purpose lies in the ability of the agonist to induce internalisation of the receptor–peptide complex, thereby delivering the conjugated drug into the cell . For [Phe 7 ,Pro 34 ]NPY, this was already successfully applied by conjugation of cytotoxic tubulysin‐derived peptides or methotrexate, both of which were selectively delivered into hY 1 R‐overexpressing (tumour) cells and induced apoptosis. Another approach was the coupling of carbaborane clusters to agonist peptides to enhance the boron concentration in tumour cells for boron neutron capture therapy .…”

Section: Discussionmentioning

confidence: 99%

Carbaboranylation of Truncated C‐Terminal Neuropeptide Y Analogue Leads to Full hY₁ Receptor Agonism

et al. 2018

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The human Y receptor is overexpressed in breast tumour cells and is, therefore, a valuable target for site-selective drug delivery. The well-established hY R-selective ligand [Phe7,Pro34]NPY has been used to couple to drugs but its length of 36 amino acids also implies complex synthesis and high production costs. Therefore, shorter ligands are desirable. However, truncated versions of neuropeptide Y (NPY) usually result in reduced binding, antagonists, or only partial G-protein-biased agonists. Herein, we report on a nonamer peptide derived from the C terminus of NPY that is modified by a carbaborane, which is able to activate hY R fully in terms of G-protein activation but also arrestin recruitment and internalisation. We provide evidence that this unique behaviour is due to the bulky nature of the carbaborane cluster, which might address a specific second binding pocket in hY R that is crucial for arrestin recruitment. Thus, this study helps in deciphering ligand-induced onset of different pathways in hY R mediated signalling.

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“…[ 185 ] To selectively target breast cancer cells, the hY 1 R‐preferring [F 7 ,P 34 ]‐NPY was conjugated at its Lys 4 side‐chain to the anti‐microtubule agent cytolysine by a disulfide linker. [ 186 ] The cytolysine‐PDC exerted selective and strong cytotoxicity on hY 1 R‐expressing cell lines, in contrast to the unspecific activity of free cytolysine. In addition, proteomic analysis revealed an identical mode of action of cytolysine delivered by the PDC compared to free cytolysine in the hY 1 R‐expressing breast cancer cell lines MDA‐MB‐468 and MCF‐7.…”

Section: Variation Of the Drug Cargo In Receptor‐targeting Peptide‐drmentioning

confidence: 99%

Targeting of peptide‐binding receptors on cancer cells with peptide‐drug conjugates

2020

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Specifically addressing cell surface molecules on cancer cells facilitates targeted cancer therapies that offer the potential to selectively destroy malignant cells, while sparing healthy tissue. Thus, undesired side-effects in tumor patients are highly reduced. Peptide-binding receptors are frequently overexpressed on cancer cells and therefore promising targets for selective tumor therapy. In this review, peptidebinding receptors for anti-cancer drug delivery are summarized with a focus on peptide ligands as delivery agents. In the first part, some of the most studied peptidebinding receptors are presented, and the ghrelin receptor and the Y 1 receptor are introduced as more recent targets for cancer therapy. Furthermore, nonpeptidic small molecules for receptor targeting on cancer cells are outlined. In the second part, peptide conjugates for the delivery of therapeutic cargos in cancer therapy are described.The essential properties of receptor-targeting peptides are specified, and recent developments in the fields of classical peptide-drug conjugates with toxic agents, radiolabeled peptides for radionuclide therapy, and boronated peptides for boron neutron capture therapy are presented. K E Y W O R D SBNCT, G protein-coupled receptor, internalization, peptide drug conjugate, tumor targeting

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A cleavable cytolysin–neuropeptide Y bioconjugate enables specific drug delivery and demonstrates intracellular mode of action

Cited by 11 publications

References 48 publications

Unusually persistent Gαi-signaling of the neuropeptide Y2 receptor depletes cellular Gi/o pools and leads to a Gi-refractory state

Unusually persistent Gαi-signaling of the neuropeptide Y2 receptor depletes cellular Gi/o pools and leads to a Gi-refractory state

Carbaboranylation of Truncated C‐Terminal Neuropeptide Y Analogue Leads to Full hY₁ Receptor Agonism

Targeting of peptide‐binding receptors on cancer cells with peptide‐drug conjugates

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A cleavable cytolysin–neuropeptide Y bioconjugate enables specific drug delivery and demonstrates intracellular mode of action

Cited by 11 publications

References 48 publications

Unusually persistent Gαi-signaling of the neuropeptide Y2 receptor depletes cellular Gi/o pools and leads to a Gi-refractory state

Unusually persistent Gαi-signaling of the neuropeptide Y2 receptor depletes cellular Gi/o pools and leads to a Gi-refractory state

Carbaboranylation of Truncated C‐Terminal Neuropeptide Y Analogue Leads to Full hY1 Receptor Agonism

Targeting of peptide‐binding receptors on cancer cells with peptide‐drug conjugates

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Carbaboranylation of Truncated C‐Terminal Neuropeptide Y Analogue Leads to Full hY₁ Receptor Agonism