Total Synthesis of [Ψ[C(NH)NH]Tpg⁴]Vancomycin and its (4-Chlorobiphenyl)methyl Derivative: Impact of Peripheral Modifications on Vancomycin Analogues Redesigned for Dual d‑Ala‑d‑Ala and d‑Ala‑d‑Lac Binding

The total synthesis of two key analogues of vancomycin containing single-atom exchanges in the binding pocket (residue 4 amidine and thioamide) are disclosed as well as their peripherally modified (4-chlorobiphenyl)­methyl (CBP) derivatives. Their assessment indicates that combined pocket amidine and CBP peripherally modified analogues exhibit a remarkable spectrum of antimicrobial activity (VSSA, MRSA, VanA and VanB VRE) and impressive potencies (MIC = 0.06–0.005 μg/mL) against both vancomycin-sensitive and -resistant bacteria and likely benefit from two independent and synergistic mechanisms of action. Like vancomycin, such analogues are likely to display especially durable antibiotic activity not prone to rapidly acquired clinical resistance

Silver(I)-Promoted Conversion of Thioamides to Amidines: Divergent Synthesis of a Key Series of Vancomycin Aglycon Residue 4 Amidines That Clarify Binding Behavior to Model Ligands

Development of a general Ag­(I)-promoted reaction for the conversion of thioamides to amidines is disclosed. This reaction was employed to prepare a key series of vancomycin aglycon residue 4 substituted amidines that were used to clarify their interaction with model ligands of peptidoglycan precursors and explore their resulting impact on antimicrobial properties

Total Synthesis of [Ψ[C(S)NH]Tpg⁴]Vancomycin Aglycon, [Ψ[C(NH)NH]Tpg⁴]Vancomycin Aglycon, and Related Key Compounds: Reengineering Vancomycin for Dual d-Ala-d-Ala and d-Ala-d-Lac Binding

The total synthesis of [Ψ­[C­(S)­NH]­Tpg⁴]­vancomycin aglycon (<b>8</b>) and its unique AgOAc-promoted single-step conversion to [Ψ­[C­(NH)­NH]­Tpg⁴]­vancomycin aglycon (<b>7</b>), conducted on a fully deprotected substrate, are disclosed. The synthetic approach not only permits access to <b>7</b>, but it also allows late-stage access to related residue 4 derivatives, alternative access to [Ψ­[CH₂NH]­Tpg⁴]­vancomycin aglycon (<b>6</b>) from a common late-stage intermediate, and provides authentic residue 4 thioamide and amidine derivatives of the vancomycin aglycon that will facilitate ongoing efforts on their semisynthetic preparation. In addition to early stage residue 4 thioamide introduction, allowing differentiation of one of seven amide bonds central to the vancomycin core structure, the approach relied on two aromatic nucleophilic substitution reactions for formation of the 16-membered diaryl ethers in the CD/DE ring systems, an effective macrolactamization for closure of the 12-membered biaryl AB ring system, and the defined order of CD, AB, and DE ring closures. This order of ring closures follows their increasing ease of thermal atropisomer equilibration, permitting the recycling of any newly generated unnatural atropisomer under progressively milder thermal conditions where the atropoisomer stereochemistry already set is not impacted. Full details of the evaluation of <b>7</b> and <b>8</b> along with several related key synthetic compounds containing the core residue 4 amidine and thioamide modifications are reported. The binding affinity of compounds containing the residue 4 amidine with the model d-Ala-d-Ala ligand <b>2</b> was found to be only 2–3 times less than the vancomycin aglycon (<b>5</b>), and this binding affinity is maintained with the model d-Ala-d-Lac ligand <b>4</b>, representing a nearly 600-fold increase in affinity relative to the vancomycin aglycon. Importantly, the amidines display effective dual, balanced binding affinity for both ligands (<i>K</i>_a <b>2</b>/<b>4</b> = 0.9–1.05), and they exhibit potent antimicrobial activity against VanA resistant bacteria (E. faecalis, VanA VRE) at a level accurately reflecting these binding characteristics (MIC = 0.3–0.6 μg/mL), charting a rational approach forward in the development of antibiotics for the treatment of vancomycin-resistant bacterial infections. In sharp contrast, <b>8</b> and related residue 4 thioamides failed to bind either <b>2</b> or <b>4</b> to any appreciable extent, do not exhibit antimicrobial activity, and serve to further underscore the remarkable behavior of the residue 4 amidines

Total Synthesis of [Ψ[C(S)NH]Tpg⁴]Vancomycin Aglycon, [Ψ[C(NH)NH]Tpg⁴]Vancomycin Aglycon, and Related Key Compounds: Reengineering Vancomycin for Dual d-Ala-d-Ala and d-Ala-d-Lac Binding

The total synthesis of [Ψ­[C­(S)­NH]­Tpg⁴]­vancomycin aglycon (<b>8</b>) and its unique AgOAc-promoted single-step conversion to [Ψ­[C­(NH)­NH]­Tpg⁴]­vancomycin aglycon (<b>7</b>), conducted on a fully deprotected substrate, are disclosed. The synthetic approach not only permits access to <b>7</b>, but it also allows late-stage access to related residue 4 derivatives, alternative access to [Ψ­[CH₂NH]­Tpg⁴]­vancomycin aglycon (<b>6</b>) from a common late-stage intermediate, and provides authentic residue 4 thioamide and amidine derivatives of the vancomycin aglycon that will facilitate ongoing efforts on their semisynthetic preparation. In addition to early stage residue 4 thioamide introduction, allowing differentiation of one of seven amide bonds central to the vancomycin core structure, the approach relied on two aromatic nucleophilic substitution reactions for formation of the 16-membered diaryl ethers in the CD/DE ring systems, an effective macrolactamization for closure of the 12-membered biaryl AB ring system, and the defined order of CD, AB, and DE ring closures. This order of ring closures follows their increasing ease of thermal atropisomer equilibration, permitting the recycling of any newly generated unnatural atropisomer under progressively milder thermal conditions where the atropoisomer stereochemistry already set is not impacted. Full details of the evaluation of <b>7</b> and <b>8</b> along with several related key synthetic compounds containing the core residue 4 amidine and thioamide modifications are reported. The binding affinity of compounds containing the residue 4 amidine with the model d-Ala-d-Ala ligand <b>2</b> was found to be only 2–3 times less than the vancomycin aglycon (<b>5</b>), and this binding affinity is maintained with the model d-Ala-d-Lac ligand <b>4</b>, representing a nearly 600-fold increase in affinity relative to the vancomycin aglycon. Importantly, the amidines display effective dual, balanced binding affinity for both ligands (<i>K</i>_a <b>2</b>/<b>4</b> = 0.9–1.05), and they exhibit potent antimicrobial activity against VanA resistant bacteria (E. faecalis, VanA VRE) at a level accurately reflecting these binding characteristics (MIC = 0.3–0.6 μg/mL), charting a rational approach forward in the development of antibiotics for the treatment of vancomycin-resistant bacterial infections. In sharp contrast, <b>8</b> and related residue 4 thioamides failed to bind either <b>2</b> or <b>4</b> to any appreciable extent, do not exhibit antimicrobial activity, and serve to further underscore the remarkable behavior of the residue 4 amidines

Potent GCN2 Inhibitor Capable of Reversing MDSC-Driven T Cell Suppression Demonstrates In Vivo Efficacy as a Single Agent and in Combination with Anti-Angiogenesis Therapy

General control nonderepressible 2 (GCN2) protein kinase is a cellular stress sensor within the tumor microenvironment (TME), whose signaling cascade has been proposed to contribute to immune escape in tumors. Herein, we report the discovery of cell-potent GCN2 inhibitors with excellent selectivity against its closely related Integrated Stress Response (ISR) family members heme-regulated inhibitor kinase (HRI), protein kinase R (PKR), and (PKR)-like endoplasmic reticulum kinase (PERK), as well as good kinome-wide selectivity and favorable PK. In mice, compound 39 engages GCN2 at levels ≥80% with an oral dose of 15 mg/kg BID. We also demonstrate the ability of compound 39 to alleviate MDSC-related T cell suppression and restore T cell proliferation, similar to the effect seen in MDSCs from GCN2 knockout mice. In the LL2 syngeneic mouse model, compound 39 demonstrates significant tumor growth inhibition (TGI) as a single agent. Furthermore, TGI mediated by anti-VEGFR was enhanced by treatment with compound 39 demonstrating the complementarity of these two mechanisms

Potent GCN2 Inhibitor Capable of Reversing MDSC-Driven T Cell Suppression Demonstrates In Vivo Efficacy as a Single Agent and in Combination with Anti-Angiogenesis Therapy

General control nonderepressible 2 (GCN2) protein kinase is a cellular stress sensor within the tumor microenvironment (TME), whose signaling cascade has been proposed to contribute to immune escape in tumors. Herein, we report the discovery of cell-potent GCN2 inhibitors with excellent selectivity against its closely related Integrated Stress Response (ISR) family members heme-regulated inhibitor kinase (HRI), protein kinase R (PKR), and (PKR)-like endoplasmic reticulum kinase (PERK), as well as good kinome-wide selectivity and favorable PK. In mice, compound 39 engages GCN2 at levels ≥80% with an oral dose of 15 mg/kg BID. We also demonstrate the ability of compound 39 to alleviate MDSC-related T cell suppression and restore T cell proliferation, similar to the effect seen in MDSCs from GCN2 knockout mice. In the LL2 syngeneic mouse model, compound 39 demonstrates significant tumor growth inhibition (TGI) as a single agent. Furthermore, TGI mediated by anti-VEGFR was enhanced by treatment with compound 39 demonstrating the complementarity of these two mechanisms