The cell adhesion molecule Nectin-4 shows elevated expression in multiple tumor types correlated with poor prognosis. Nectin-4-directed ADCs show efficacy in multiple xenograft tumor models. The ADC enfortumab vedotin (EV) delivers the antimitotic toxin MMAE to Nectin-4-expressing cells via internalization and cleavage of a valine-citrulline dipeptide linker component. Clinical validation of Nectin-4 as a target in urothelial cancer has been demonstrated with EV. In 2021, the FDA-approved EV for patients with locally advanced or metastatic urothelial cancer who had previously received a PD-1 or PD-L1 inhibitor and platinum-containing chemotherapy or were ineligible for cisplatin containing chemotherapy and had received one or more prior lines of therapy. Bicycle toxin conjugates (BTC) are structurally constrained bicyclic peptides conjugated through cleavable linkers to a toxin. Various, well-described linker toxin combinations can be incorporated into the BTC molecule (e.g., BT1718 and BT5528; refs.). They are of low molecular weight (approximately 4-4.5 kDa) and being chemically synthesized can be optimized for appropriate affinity, stability, and solubility relatively simply. Through intravenous administration, high systemic Cmax values can be attained, which, along with BTCs relatively small size, helps drive rapid diffusion into extra-vascular compartment, as reflected in a volume of distribution similar to extracellular fluid. We believe that delivery of a high number of BTCs, each carrying a reduced toxin load (peptide toxin ratio of 1:1) should improve tumor penetration and reduce the impact of the binding site barrier. BTCs show moderate clearance from the systemic vasculature, predominantly by the renal route. This overall profile marks them out from most ADCs and provides the possibility of enhanced clinical efficacy with a wider therapeutic index. BTCs are new therapeutic modality that shows a very different pharmacokinetic and structural profile to classic ADCs, whereas possessing robust tumoricidal properties. BTCs targeting MT1-MMP and EphA2 are currently in clinical trials. BT8009 is the most recent BTC to enter clinical trial. BT8009 is highly selective for Nectin-4 over other nectin family members and an extensive range of cell membrane expressed proteins. It shares the same cleavable linker and toxin combination as EV and can be cleaved by proteases in the TME, releasing cell penetrant MMAE that diffuses into tumor cells, or bystander stromal-supportive cells, evoking cell death, and tumor regression. It shows a robust and dose-dependent antitumor activity in multiple CDX and PDX models, representing lung, breast, bladder, head and neck cancers. Optimal tumor regression is associated with membrane expression of Nectin-4, in conjunction with MMAE sensitivity. Tumor regression rates are comparable for small and large tumors, indicative of deep and rapid penetration throughout the tumor. BT8009 provides tumoricidal activity with several regimens, potentially allowing for clinical titration of dose and dose interval if required.

The cell adhesion molecule Nectin-4 shows elevated expression in multiple tumor types correlated with poor prognosis. Nectin-4-directed ADCs show efficacy in multiple xenograft tumor models. The ADC enfortumab vedotin (EV) delivers the antimitotic toxin MMAE to Nectin-4-expressing cells via internalization and cleavage of a valine-citrulline dipeptide linker component. Clinical validation of Nectin-4 as a target in urothelial cancer has been demonstrated with EV. In 2021, the FDA-approved EV for patients with locally advanced or metastatic urothelial cancer who had previously received a PD-1 or PD-L1 inhibitor and platinum-containing chemotherapy or were ineligible for cisplatin containing chemotherapy and had received one or more prior lines of therapy. Bicycle toxin conjugates (BTC) are structurally constrained bicyclic peptides conjugated through cleavable linkers to a toxin. Various, well-described linker toxin combinations can be incorporated into the BTC molecule (e.g., BT1718 and BT5528; refs.). They are of low molecular weight (approximately 4-4.5 kDa) and being chemically synthesized can be optimized for appropriate affinity, stability, and solubility relatively simply. Through intravenous administration, high systemic Cmax values can be attained, which, along with BTCs relatively small size, helps drive rapid diffusion into extra-vascular compartment, as reflected in a volume of distribution similar to extracellular fluid. We believe that delivery of a high number of BTCs, each carrying a reduced toxin load (peptide toxin ratio of 1:1) should improve tumor penetration and reduce the impact of the binding site barrier. BTCs show moderate clearance from the systemic vasculature, predominantly by the renal route. This overall profile marks them out from most ADCs and provides the possibility of enhanced clinical efficacy with a wider therapeutic index. BTCs are new therapeutic modality that shows a very different pharmacokinetic and structural profile to classic ADCs, whereas possessing robust tumoricidal properties. BTCs targeting MT1-MMP and EphA2 are currently in clinical trials. BT8009 is the most recent BTC to enter clinical trial. BT8009 is highly selective for Nectin-4 over other nectin family members and an extensive range of cell membrane expressed proteins. It shares the same cleavable linker and toxin combination as EV and can be cleaved by proteases in the TME, releasing cell penetrant MMAE that diffuses into tumor cells, or bystander stromal-supportive cells, evoking cell death, and tumor regression. It shows a robust and dose-dependent antitumor activity in multiple CDX and PDX models, representing lung, breast, bladder, head and neck cancers. Optimal tumor regression is associated with membrane expression of Nectin-4, in conjunction with MMAE sensitivity. Tumor regression rates are comparable for small and large tumors, indicative of deep and rapid penetration throughout the tumor. BT8009 provides tumoricidal activity with several regimens, potentially allowing for clinical titration of dose and dose interval if required.

The cell adhesion molecule Nectin-4 shows elevated expression in multiple tumor types correlated with poor prognosis. Nectin-4-directed ADCs show efficacy in multiple xenograft tumor models. The ADC enfortumab vedotin (EV) delivers the antimitotic toxin MMAE to Nectin-4-expressing cells via internalization and cleavage of a valine-citrulline dipeptide linker component. Clinical validation of Nectin-4 as a target in urothelial cancer has been demonstrated with EV. In 2021, the FDA-approved EV for patients with locally advanced or metastatic urothelial cancer who had previously received a PD-1 or PD-L1 inhibitor and platinum-containing chemotherapy or were ineligible for cisplatin containing chemotherapy and had received one or more prior lines of therapy. Bicycle toxin conjugates (BTC) are structurally constrained bicyclic peptides conjugated through cleavable linkers to a toxin. Various, well-described linker toxin combinations can be incorporated into the BTC molecule (e.g., BT1718 and BT5528; refs.). They are of low molecular weight (approximately 4-4.5 kDa) and being chemically synthesized can be optimized for appropriate affinity, stability, and solubility relatively simply. Through intravenous administration, high systemic Cmax values can be attained, which, along with BTCs relatively small size, helps drive rapid diffusion into extra-vascular compartment, as reflected in a volume of distribution similar to extracellular fluid. We believe that delivery of a high number of BTCs, each carrying a reduced toxin load (peptide toxin ratio of 1:1) should improve tumor penetration and reduce the impact of the binding site barrier. BTCs show moderate clearance from the systemic vasculature, predominantly by the renal route. This overall profile marks them out from most ADCs and provides the possibility of enhanced clinical efficacy with a wider therapeutic index. BTCs are new therapeutic modality that shows a very different pharmacokinetic and structural profile to classic ADCs, whereas possessing robust tumoricidal properties. BTCs targeting MT1-MMP and EphA2 are currently in clinical trials. BT8009 is the most recent BTC to enter clinical trial. BT8009 is highly selective for Nectin-4 over other nectin family members and an extensive range of cell membrane expressed proteins. It shares the same cleavable linker and toxin combination as EV and can be cleaved by proteases in the TME, releasing cell penetrant MMAE that diffuses into tumor cells, or bystander stromal-supportive cells, evoking cell death, and tumor regression. It shows a robust and dose-dependent antitumor activity in multiple CDX and PDX models, representing lung, breast, bladder, head and neck cancers. Optimal tumor regression is associated with membrane expression of Nectin-4, in conjunction with MMAE sensitivity. Tumor regression rates are comparable for small and large tumors, indicative of deep and rapid penetration throughout the tumor. BT8009 provides tumoricidal activity with several regimens, potentially allowing for clinical titration of dose and dose interval if required.

The cell adhesion molecule Nectin-4 shows elevated expression in multiple tumor types correlated with poor prognosis. Nectin-4-directed ADCs show efficacy in multiple xenograft tumor models. The ADC enfortumab vedotin (EV) delivers the antimitotic toxin MMAE to Nectin-4-expressing cells via internalization and cleavage of a valine-citrulline dipeptide linker component. Clinical validation of Nectin-4 as a target in urothelial cancer has been demonstrated with EV. In 2021, the FDA-approved EV for patients with locally advanced or metastatic urothelial cancer who had previously received a PD-1 or PD-L1 inhibitor and platinum-containing chemotherapy or were ineligible for cisplatin containing chemotherapy and had received one or more prior lines of therapy. Bicycle toxin conjugates (BTC) are structurally constrained bicyclic peptides conjugated through cleavable linkers to a toxin. Various, well-described linker toxin combinations can be incorporated into the BTC molecule (e.g., BT1718 and BT5528; refs.). They are of low molecular weight (approximately 4-4.5 kDa) and being chemically synthesized can be optimized for appropriate affinity, stability, and solubility relatively simply. Through intravenous administration, high systemic Cmax values can be attained, which, along with BTCs relatively small size, helps drive rapid diffusion into extra-vascular compartment, as reflected in a volume of distribution similar to extracellular fluid. We believe that delivery of a high number of BTCs, each carrying a reduced toxin load (peptide toxin ratio of 1:1) should improve tumor penetration and reduce the impact of the binding site barrier. BTCs show moderate clearance from the systemic vasculature, predominantly by the renal route. This overall profile marks them out from most ADCs and provides the possibility of enhanced clinical efficacy with a wider therapeutic index. BTCs are new therapeutic modality that shows a very different pharmacokinetic and structural profile to classic ADCs, whereas possessing robust tumoricidal properties. BTCs targeting MT1-MMP and EphA2 are currently in clinical trials. BT8009 is the most recent BTC to enter clinical trial. BT8009 is highly selective for Nectin-4 over other nectin family members and an extensive range of cell membrane expressed proteins. It shares the same cleavable linker and toxin combination as EV and can be cleaved by proteases in the TME, releasing cell penetrant MMAE that diffuses into tumor cells, or bystander stromal-supportive cells, evoking cell death, and tumor regression. It shows a robust and dose-dependent antitumor activity in multiple CDX and PDX models, representing lung, breast, bladder, head and neck cancers. Optimal tumor regression is associated with membrane expression of Nectin-4, in conjunction with MMAE sensitivity. Tumor regression rates are comparable for small and large tumors, indicative of deep and rapid penetration throughout the tumor. BT8009 provides tumoricidal activity with several regimens, potentially allowing for clinical titration of dose and dose interval if required.

The cell adhesion molecule Nectin-4 shows elevated expression in multiple tumor types correlated with poor prognosis. Nectin-4-directed ADCs show efficacy in multiple xenograft tumor models. The ADC enfortumab vedotin (EV) delivers the antimitotic toxin MMAE to Nectin-4-expressing cells via internalization and cleavage of a valine-citrulline dipeptide linker component. Clinical validation of Nectin-4 as a target in urothelial cancer has been demonstrated with EV. In 2021, the FDA-approved EV for patients with locally advanced or metastatic urothelial cancer who had previously received a PD-1 or PD-L1 inhibitor and platinum-containing chemotherapy or were ineligible for cisplatin containing chemotherapy and had received one or more prior lines of therapy. Bicycle toxin conjugates (BTC) are structurally constrained bicyclic peptides conjugated through cleavable linkers to a toxin. Various, well-described linker toxin combinations can be incorporated into the BTC molecule (e.g., BT1718 and BT5528; refs.). They are of low molecular weight (approximately 4-4.5 kDa) and being chemically synthesized can be optimized for appropriate affinity, stability, and solubility relatively simply. Through intravenous administration, high systemic Cmax values can be attained, which, along with BTCs relatively small size, helps drive rapid diffusion into extra-vascular compartment, as reflected in a volume of distribution similar to extracellular fluid. We believe that delivery of a high number of BTCs, each carrying a reduced toxin load (peptide toxin ratio of 1:1) should improve tumor penetration and reduce the impact of the binding site barrier. BTCs show moderate clearance from the systemic vasculature, predominantly by the renal route. This overall profile marks them out from most ADCs and provides the possibility of enhanced clinical efficacy with a wider therapeutic index. BTCs are new therapeutic modality that shows a very different pharmacokinetic and structural profile to classic ADCs, whereas possessing robust tumoricidal properties. BTCs targeting MT1-MMP and EphA2 are currently in clinical trials. BT8009 is the most recent BTC to enter clinical trial. BT8009 is highly selective for Nectin-4 over other nectin family members and an extensive range of cell membrane expressed proteins. It shares the same cleavable linker and toxin combination as EV and can be cleaved by proteases in the TME, releasing cell penetrant MMAE that diffuses into tumor cells, or bystander stromal-supportive cells, evoking cell death, and tumor regression. It shows a robust and dose-dependent antitumor activity in multiple CDX and PDX models, representing lung, breast, bladder, head and neck cancers. Optimal tumor regression is associated with membrane expression of Nectin-4, in conjunction with MMAE sensitivity. Tumor regression rates are comparable for small and large tumors, indicative of deep and rapid penetration throughout the tumor. BT8009 provides tumoricidal activity with several regimens, potentially allowing for clinical titration of dose and dose interval if required.

The cell adhesion molecule Nectin-4 shows elevated expression in multiple tumor types correlated with poor prognosis. Nectin-4-directed ADCs show efficacy in multiple xenograft tumor models. The ADC enfortumab vedotin (EV) delivers the antimitotic toxin MMAE to Nectin-4-expressing cells via internalization and cleavage of a valine-citrulline dipeptide linker component. Clinical validation of Nectin-4 as a target in urothelial cancer has been demonstrated with EV. In 2021, the FDA-approved EV for patients with locally advanced or metastatic urothelial cancer who had previously received a PD-1 or PD-L1 inhibitor and platinum-containing chemotherapy or were ineligible for cisplatin containing chemotherapy and had received one or more prior lines of therapy. Bicycle toxin conjugates (BTC) are structurally constrained bicyclic peptides conjugated through cleavable linkers to a toxin. Various, well-described linker toxin combinations can be incorporated into the BTC molecule (e.g., BT1718 and BT5528; refs.). They are of low molecular weight (approximately 4-4.5 kDa) and being chemically synthesized can be optimized for appropriate affinity, stability, and solubility relatively simply. Through intravenous administration, high systemic Cmax values can be attained, which, along with BTCs relatively small size, helps drive rapid diffusion into extra-vascular compartment, as reflected in a volume of distribution similar to extracellular fluid. We believe that delivery of a high number of BTCs, each carrying a reduced toxin load (peptide toxin ratio of 1:1) should improve tumor penetration and reduce the impact of the binding site barrier. BTCs show moderate clearance from the systemic vasculature, predominantly by the renal route. This overall profile marks them out from most ADCs and provides the possibility of enhanced clinical efficacy with a wider therapeutic index. BTCs are new therapeutic modality that shows a very different pharmacokinetic and structural profile to classic ADCs, whereas possessing robust tumoricidal properties. BTCs targeting MT1-MMP and EphA2 are currently in clinical trials. BT8009 is the most recent BTC to enter clinical trial. BT8009 is highly selective for Nectin-4 over other nectin family members and an extensive range of cell membrane expressed proteins. It shares the same cleavable linker and toxin combination as EV and can be cleaved by proteases in the TME, releasing cell penetrant MMAE that diffuses into tumor cells, or bystander stromal-supportive cells, evoking cell death, and tumor regression. It shows a robust and dose-dependent antitumor activity in multiple CDX and PDX models, representing lung, breast, bladder, head and neck cancers. Optimal tumor regression is associated with membrane expression of Nectin-4, in conjunction with MMAE sensitivity. Tumor regression rates are comparable for small and large tumors, indicative of deep and rapid penetration throughout the tumor. BT8009 provides tumoricidal activity with several regimens, potentially allowing for clinical titration of dose and dose interval if required.

Nectin-4 is a cell adhesion molecule from the Nectin and Nectin-like family. It is a clinically validated tumor target and has been reported to be highly expressed in a wide range of solid tumors, including bladder, esophageal, pancreatic, and lung, but with limited distribution in healthy tissues.An antibody drug conjugate (PADCEV, enfortumab vedotin) that targets Nectin-4 was recently approved for the treatment of bladder cancer following the generation of positive data in a number of clinical studies. Herein we describe the discovery via phage display and subsequent chemical optimization of a Nectin-4 binding Bicycle and its incorporation into BT8009, a BTC that is currently under clinical evaluation. A detailed report of the pharmacologic properties of BT8009 has recently been described.In this, BT8009 shows potent efficacy in multiple tumor models, including patient-derived xenografts, across a variety of tumor indications and is well-tolerated in preclinical safety studies. In several models it demonstrated superior or equivalent efficacy to an analogue of the ADC PADCEV.

Nectin-4 is a cell adhesion molecule from the Nectin and Nectin-like family. It is a clinically validated tumor target and has been reported to be highly expressed in a wide range of solid tumors, including bladder, esophageal, pancreatic, and lung, but with limited distribution in healthy tissues.An antibody drug conjugate (PADCEV, enfortumab vedotin) that targets Nectin-4 was recently approved for the treatment of bladder cancer following the generation of positive data in a number of clinical studies. Herein we describe the discovery via phage display and subsequent chemical optimization of a Nectin-4 binding Bicycle and its incorporation into BT8009, a BTC that is currently under clinical evaluation. A detailed report of the pharmacologic properties of BT8009 has recently been described.In this, BT8009 shows potent efficacy in multiple tumor models, including patient-derived xenografts, across a variety of tumor indications and is well-tolerated in preclinical safety studies. In several models it demonstrated superior or equivalent efficacy to an analogue of the ADC PADCEV.

Nectin-4 is a cell adhesion molecule from the Nectin and Nectin-like family. It is a clinically validated tumor target and has been reported to be highly expressed in a wide range of solid tumors, including bladder, esophageal, pancreatic, and lung, but with limited distribution in healthy tissues.An antibody drug conjugate (PADCEV, enfortumab vedotin) that targets Nectin-4 was recently approved for the treatment of bladder cancer following the generation of positive data in a number of clinical studies. Herein we describe the discovery via phage display and subsequent chemical optimization of a Nectin-4 binding Bicycle and its incorporation into BT8009, a BTC that is currently under clinical evaluation. A detailed report of the pharmacologic properties of BT8009 has recently been described.In this, BT8009 shows potent efficacy in multiple tumor models, including patient-derived xenografts, across a variety of tumor indications and is well-tolerated in preclinical safety studies. In several models it demonstrated superior or equivalent efficacy to an analogue of the ADC PADCEV.

Folate receptor α (FRα) and vanilloid subfamily member 6 of transient receptor potential channels (TRPV6) are potential promising therapeutic targets due to their high expression level in many solid tumors including ovarian cancer. CBP-1008 is a first-in-class (FIC) bi-specific ligand drug conjugate targeting FRα and TRPV6 carrying monomethyl auristatin E (MMAE) as payload.

Folate receptor α (FRα) and vanilloid subfamily member 6 of transient receptor potential channels (TRPV6) are potential promising therapeutic targets due to their high expression level in many solid tumors including ovarian cancer. CBP-1008 is a first-in-class (FIC) bi-specific ligand drug conjugate targeting FRα and TRPV6 carrying monomethyl auristatin E (MMAE) as payload.

Folate receptor α (FRα) and vanilloid subfamily member 6 of transient receptor potential channels (TRPV6) are potential promising therapeutic targets due to their high expression level in many solid tumors including ovarian cancer. CBP-1008 is a first-in-class (FIC) bi-specific ligand drug conjugate targeting FRα and TRPV6 carrying monomethyl auristatin E (MMAE) as payload.

Folate receptor α (FRα) and vanilloid subfamily member 6 of transient receptor potential channels (TRPV6) are potential promising therapeutic targets due to their high expression level in many solid tumors including ovarian cancer. CBP-1008 is a first-in-class (FIC) bi-specific ligand drug conjugate targeting FRα and TRPV6 carrying monomethyl auristatin E (MMAE) as payload.

Folate receptor α (FRα) and vanilloid subfamily member 6 of transient receptor potential channels (TRPV6) are potential promising therapeutic targets due to their high expression level in many solid tumors including ovarian cancer. CBP-1008 is a first-in-class (FIC) bi-specific ligand drug conjugate targeting FRα and TRPV6 carrying monomethyl auristatin E (MMAE) as payload.

Folate receptor α (FRα) and vanilloid subfamily member 6 of transient receptor potential channels (TRPV6) are potential promising therapeutic targets due to their high expression level in many solid tumors including ovarian cancer. CBP-1008 is a first-in-class (FIC) bi-specific ligand drug conjugate targeting FRα and TRPV6 carrying monomethyl auristatin E (MMAE) as payload.

Folate receptor α (FRα) and vanilloid subfamily member 6 of transient receptor potential channels (TRPV6) are potential promising therapeutic targets due to their high expression level in many solid tumors including ovarian cancer. CBP-1008 is a first-in-class (FIC) bi-specific ligand drug conjugate targeting FRα and TRPV6 carrying monomethyl auristatin E (MMAE) as payload.

Folate receptor α (FRα) and vanilloid subfamily member 6 of transient receptor potential channels (TRPV6) are potential promising therapeutic targets due to their high expression level in many solid tumors including ovarian cancer. CBP-1008 is a first-in-class (FIC) bi-specific ligand drug conjugate targeting FRα and TRPV6 carrying monomethyl auristatin E (MMAE) as payload.

Folate receptor α (FRα) and vanilloid subfamily member 6 of transient receptor potential channels (TRPV6) are potential promising therapeutic targets due to their high expression level in many solid tumors including ovarian cancer. CBP-1008 is a first-in-class (FIC) bi-specific ligand drug conjugate targeting FRα and TRPV6 carrying monomethyl auristatin E (MMAE) as payload.

Folate receptor α (FRα) and vanilloid subfamily member 6 of transient receptor potential channels (TRPV6) are potential promising therapeutic targets due to their high expression level in many solid tumors including ovarian cancer. CBP-1008 is a first-in-class (FIC) bi-specific ligand drug conjugate targeting FRα and TRPV6 carrying monomethyl auristatin E (MMAE) as payload.

Here we describe the development of BT5528, a bicyclic peptide (Bicycle) conjugated to the auristatin derivative maleimidocaproyl-monomethyl auristatin E to generate the Bicycle toxin conjugate BT5528. There are two potential mechanism of BT5528 bystander activity: extracellular linker cleavage and toxin penetration into neighboring cells, or receptor internalization and intracellular linker cleavage followed by release of cell penetrant toxin from lysed cells. The available data do not allow us to distinguish between these two mechanisms and it seems likely that BT5528 activity is mediated by toxin release following a combination of intracellular and extracellular linker cleavage.

Here we describe the development of BT5528, a bicyclic peptide (Bicycle) conjugated to the auristatin derivative maleimidocaproyl-monomethyl auristatin E to generate the Bicycle toxin conjugate BT5528. There are two potential mechanism of BT5528 bystander activity: extracellular linker cleavage and toxin penetration into neighboring cells, or receptor internalization and intracellular linker cleavage followed by release of cell penetrant toxin from lysed cells. The available data do not allow us to distinguish between these two mechanisms and it seems likely that BT5528 activity is mediated by toxin release following a combination of intracellular and extracellular linker cleavage.

Here, we show that the replacement of the Val-Ala-p-aminobenzyloxycarbamate linker with the Gly-Pro-Leu-Gly-p-aminobenzyloxycarbamate (GPLG-PABC) sequence as enzymatically cleavable linker in the SMDC bearing the cyclo[DKP-isoDGR] αVβ3 integrin ligand as tumor homing moiety and the monomethyl auristatin E (MMAE) as cytotoxic payload led to a 4-fold more potent anti-tumoral effect of the final conjugate on different cancer cell lines. In addition, the synthesized conjugate resulted to be significantly more potent than the free MMAE when tested following the kiss-and-run protocol, and the relative potency were clearly consistent with the expression of the αVβ3 integrin receptor in the considered cancer cell lines. In vitro enzymatic cleavage tests showed that the GPLG-PABC linker is cleaved by lysosomal enzymes, and that the released drug is observable already after 15 min of incubation. Although additional data are needed to fully characterize the releasing capacity of GPLG-PABC linker, our findings are of therapeutic significance since we are introducing an alternative to other well-established enzymatically sensitive peptide sequences that might be used in the future for generating more efficient and less toxic drug delivery systems.

Here, we show that the replacement of the Val-Ala-p-aminobenzyloxycarbamate linker with the Gly-Pro-Leu-Gly-p-aminobenzyloxycarbamate (GPLG-PABC) sequence as enzymatically cleavable linker in the SMDC bearing the cyclo[DKP-isoDGR] αVβ3 integrin ligand as tumor homing moiety and the monomethyl auristatin E (MMAE) as cytotoxic payload led to a 4-fold more potent anti-tumoral effect of the final conjugate on different cancer cell lines. In addition, the synthesized conjugate resulted to be significantly more potent than the free MMAE when tested following the kiss-and-run protocol, and the relative potency were clearly consistent with the expression of the αVβ3 integrin receptor in the considered cancer cell lines. In vitro enzymatic cleavage tests showed that the GPLG-PABC linker is cleaved by lysosomal enzymes, and that the released drug is observable already after 15 min of incubation. Although additional data are needed to fully characterize the releasing capacity of GPLG-PABC linker, our findings are of therapeutic significance since we are introducing an alternative to other well-established enzymatically sensitive peptide sequences that might be used in the future for generating more efficient and less toxic drug delivery systems.

Here, we show that the replacement of the Val-Ala-p-aminobenzyloxycarbamate linker with the Gly-Pro-Leu-Gly-p-aminobenzyloxycarbamate (GPLG-PABC) sequence as enzymatically cleavable linker in the SMDC bearing the cyclo[DKP-isoDGR] αVβ3 integrin ligand as tumor homing moiety and the monomethyl auristatin E (MMAE) as cytotoxic payload led to a 4-fold more potent anti-tumoral effect of the final conjugate on different cancer cell lines. In addition, the synthesized conjugate resulted to be significantly more potent than the free MMAE when tested following the kiss-and-run protocol, and the relative potency were clearly consistent with the expression of the αVβ3 integrin receptor in the considered cancer cell lines. In vitro enzymatic cleavage tests showed that the GPLG-PABC linker is cleaved by lysosomal enzymes, and that the released drug is observable already after 15 min of incubation. Although additional data are needed to fully characterize the releasing capacity of GPLG-PABC linker, our findings are of therapeutic significance since we are introducing an alternative to other well-established enzymatically sensitive peptide sequences that might be used in the future for generating more efficient and less toxic drug delivery systems.

Here, we show that the replacement of the Val-Ala-p-aminobenzyloxycarbamate linker with the Gly-Pro-Leu-Gly-p-aminobenzyloxycarbamate (GPLG-PABC) sequence as enzymatically cleavable linker in the SMDC bearing the cyclo[DKP-isoDGR] αVβ3 integrin ligand as tumor homing moiety and the monomethyl auristatin E (MMAE) as cytotoxic payload led to a 4-fold more potent anti-tumoral effect of the final conjugate on different cancer cell lines. In addition, the synthesized conjugate resulted to be significantly more potent than the free MMAE when tested following the kiss-and-run protocol, and the relative potency were clearly consistent with the expression of the αVβ3 integrin receptor in the considered cancer cell lines. In vitro enzymatic cleavage tests showed that the GPLG-PABC linker is cleaved by lysosomal enzymes, and that the released drug is observable already after 15 min of incubation. Although additional data are needed to fully characterize the releasing capacity of GPLG-PABC linker, our findings are of therapeutic significance since we are introducing an alternative to other well-established enzymatically sensitive peptide sequences that might be used in the future for generating more efficient and less toxic drug delivery systems.

Ovarian cancer is the most deadly female gynaecological malignancy in developed countries and new treatments are urgently needed. The luteinising hormone releasing hormone (LHRH) peptide drug conjugate Zoptarelin doxorubicin is one such potential new drug modality that entered clinical trials for treating LHRH receptor-positive gynaecological cancers. However, development stopped after disappointing Phase 3 results in 2017. We believe the lack of efficacy was due to linker instability and payload potency. In this work, we replaced its linker-toxin with vedotin (MC-VC-PABC-MMAE), yielding the novel peptide drug conjugate D-Cys6-LHRH vedotin. A GI50 and cell specificity comparison against cancerous and non-cancerous ovarian cell lines showed significantly superior bioactivity and selectivity over Zoptarelin doxorubicin (GI50 4 vs. 453 nM) and other chemotherapeutic drugs used for treating ovarian cancers. Our results suggest D-Cys6-LHRH vedotin can potentially be used as a treatment for ovarian cancer.

Ovarian cancer is the most deadly female gynaecological malignancy in developed countries and new treatments are urgently needed. The luteinising hormone releasing hormone (LHRH) peptide drug conjugate Zoptarelin doxorubicin is one such potential new drug modality that entered clinical trials for treating LHRH receptor-positive gynaecological cancers. However, development stopped after disappointing Phase 3 results in 2017. We believe the lack of efficacy was due to linker instability and payload potency. In this work, we replaced its linker-toxin with vedotin (MC-VC-PABC-MMAE), yielding the novel peptide drug conjugate D-Cys6-LHRH vedotin. A GI50 and cell specificity comparison against cancerous and non-cancerous ovarian cell lines showed significantly superior bioactivity and selectivity over Zoptarelin doxorubicin (GI50 4 vs. 453 nM) and other chemotherapeutic drugs used for treating ovarian cancers. Our results suggest D-Cys6-LHRH vedotin can potentially be used as a treatment for ovarian cancer.

Ovarian cancer is the most deadly female gynaecological malignancy in developed countries and new treatments are urgently needed. The luteinising hormone releasing hormone (LHRH) peptide drug conjugate Zoptarelin doxorubicin is one such potential new drug modality that entered clinical trials for treating LHRH receptor-positive gynaecological cancers. However, development stopped after disappointing Phase 3 results in 2017. We believe the lack of efficacy was due to linker instability and payload potency. In this work, we replaced its linker-toxin with vedotin (MC-VC-PABC-MMAE), yielding the novel peptide drug conjugate D-Cys6-LHRH vedotin. A GI50 and cell specificity comparison against cancerous and non-cancerous ovarian cell lines showed significantly superior bioactivity and selectivity over Zoptarelin doxorubicin (GI50 4 vs. 453 nM) and other chemotherapeutic drugs used for treating ovarian cancers. Our results suggest D-Cys6-LHRH vedotin can potentially be used as a treatment for ovarian cancer.

Ovarian cancer is the most deadly female gynaecological malignancy in developed countries and new treatments are urgently needed. The luteinising hormone releasing hormone (LHRH) peptide drug conjugate Zoptarelin doxorubicin is one such potential new drug modality that entered clinical trials for treating LHRH receptor-positive gynaecological cancers. However, development stopped after disappointing Phase 3 results in 2017. We believe the lack of efficacy was due to linker instability and payload potency. In this work, we replaced its linker-toxin with vedotin (MC-VC-PABC-MMAE), yielding the novel peptide drug conjugate D-Cys6-LHRH vedotin. A GI50 and cell specificity comparison against cancerous and non-cancerous ovarian cell lines showed significantly superior bioactivity and selectivity over Zoptarelin doxorubicin (GI50 4 vs. 453 nM) and other chemotherapeutic drugs used for treating ovarian cancers. Our results suggest D-Cys6-LHRH vedotin can potentially be used as a treatment for ovarian cancer.

Many anticancer drugs exhibit high systemic off-target toxicities causing severe side effects. Peptide-drug conjugates (PDCs) that target tumor-specific receptors such as integrin vβ6 are emerging as powerful tools to overcome these challenges. The development of an integrin vβ6-selective PDC was achieved by combining the therapeutic efficacy of the cytotoxic drug monomethyl auristatin E with the selectivity of the vβ6-binding peptide (vβ6-BP) and with the ability of positron emission tomography (PET) imaging by copper-64. The [64Cu]PDC-1 was produced efficiently and in high purity. The PDC exhibited high human serum stability, integrin vβ6-selective internalization, cell binding, and cytotoxicity. Integrin vβ6-selective tumor accumulation of the [64Cu]PDC-1 was visualized with PET-imaging and corroborated by biodistribution, and [64Cu]PDC-1 showed promising in vivo pharmacokinetics. The [natCu]PDC-1 treatment resulted in prolonged survival of mice bearing vβ6 (+) tumors (median survival: 77 days, vs vβ6 (-) tumor group 49 days, and all other control groups 37 days).

Many anticancer drugs exhibit high systemic off-target toxicities causing severe side effects. Peptide-drug conjugates (PDCs) that target tumor-specific receptors such as integrin vβ6 are emerging as powerful tools to overcome these challenges. The development of an integrin vβ6-selective PDC was achieved by combining the therapeutic efficacy of the cytotoxic drug monomethyl auristatin E with the selectivity of the vβ6-binding peptide (vβ6-BP) and with the ability of positron emission tomography (PET) imaging by copper-64. The [64Cu]PDC-1 was produced efficiently and in high purity. The PDC exhibited high human serum stability, integrin vβ6-selective internalization, cell binding, and cytotoxicity. Integrin vβ6-selective tumor accumulation of the [64Cu]PDC-1 was visualized with PET-imaging and corroborated by biodistribution, and [64Cu]PDC-1 showed promising in vivo pharmacokinetics. The [natCu]PDC-1 treatment resulted in prolonged survival of mice bearing vβ6 (+) tumors (median survival: 77 days, vs vβ6 (-) tumor group 49 days, and all other control groups 37 days).

Many anticancer drugs exhibit high systemic off-target toxicities causing severe side effects. Peptide-drug conjugates (PDCs) that target tumor-specific receptors such as integrin vβ6 are emerging as powerful tools to overcome these challenges. The development of an integrin vβ6-selective PDC was achieved by combining the therapeutic efficacy of the cytotoxic drug monomethyl auristatin E with the selectivity of the vβ6-binding peptide (vβ6-BP) and with the ability of positron emission tomography (PET) imaging by copper-64. The [64Cu]PDC-1 was produced efficiently and in high purity. The PDC exhibited high human serum stability, integrin vβ6-selective internalization, cell binding, and cytotoxicity. Integrin vβ6-selective tumor accumulation of the [64Cu]PDC-1 was visualized with PET-imaging and corroborated by biodistribution, and [64Cu]PDC-1 showed promising in vivo pharmacokinetics. The [natCu]PDC-1 treatment resulted in prolonged survival of mice bearing vβ6 (+) tumors (median survival: 77 days, vs vβ6 (-) tumor group 49 days, and all other control groups 37 days).

Many anticancer drugs exhibit high systemic off-target toxicities causing severe side effects. Peptide-drug conjugates (PDCs) that target tumor-specific receptors such as integrin vβ6 are emerging as powerful tools to overcome these challenges. The development of an integrin vβ6-selective PDC was achieved by combining the therapeutic efficacy of the cytotoxic drug monomethyl auristatin E with the selectivity of the vβ6-binding peptide (vβ6-BP) and with the ability of positron emission tomography (PET) imaging by copper-64. The [64Cu]PDC-1 was produced efficiently and in high purity. The PDC exhibited high human serum stability, integrin vβ6-selective internalization, cell binding, and cytotoxicity. Integrin vβ6-selective tumor accumulation of the [64Cu]PDC-1 was visualized with PET-imaging and corroborated by biodistribution, and [64Cu]PDC-1 showed promising in vivo pharmacokinetics. The [natCu]PDC-1 treatment resulted in prolonged survival of mice bearing vβ6 (+) tumors (median survival: 77 days, vs vβ6 (-) tumor group 49 days, and all other control groups 37 days).