The known dihydroazafluoranthene 6 is elaborated to the σ-homo-o-benzoquinone monoacetal 9 which reacts with trifluoroacetic acid to give the title alkaloid 1.

Regiocontrolled total synthesis of imerubrine—the first total synthesis of a tropoloisoquinoline alkaloid

Remote functionalization of SCH 39166: discovery of potent and selective benzazepine dopamine D1 receptor antagonists.

Thermolysis of the title compound 8 produced not only the expected azafluoranthene 7 but also the regioisomeric compound 23 and its dihydro counterpart 22. A mechanism for the formation of compounds 7, 22 and 23 from precursor 8 has been advanced. Reaction of salt 8 with copper bronze in aqueous sulfuric acid produced good yields of dihydroazafluoranthene 29 together with small amounts of the reduction product 30. The tubulin binding properties of the azafluoranthenes 7, 22, 23, 24 and 29 have been determined.

Studies directed towards total syntheses of the tropoloisoquinoline alkaloids grandirubrine and imerubrine. Part 2. Thermolysis of 8,9-dihydro-2-hydroxy-3,10,11,12-tetra methoxyisoquino[2,1-c][1,2,3]benzotriazin-7-ium chloride: competitive modes of cyclisation leading to the indeno[1,2,3-ij]isoquinoline (azafluoranthene) skeleton

The (almost) unique combination of nucleofugacity and polarizability of the iodide atom has made alkyl (sp 3 ) iodides, particularly, vulnerable to displacement by (polarizable) carbon nucleophiles. Carbon–iodide bond characteristics have been parleyed into a range of transition metal-mediated carbon–carbon bond forming processes utilizing alkenyl- or aryl (sp 2 ) iodides. This chapter discusses the prospects for alkynyl (sp) iodides in carbon–carbon bond forming reactions. The introduction of the more electrophilic alkynyliodonium salts triggered an upsurge in interest in carbon–carbon bond-forming transformations via the (sp)-carbon–I bond. These organoiodine species participate in nucleophilic addition reactions. The chapter also discusses the amine substituents that brought the nitrogen's nucleophilicity within the acceptable range for alkynyliodonium salt addition to be in a position to develop new strategy-level transformations that could be applied to alkaloid synthesis. One implicit benefit of such a strategy is convergency because only the alkynyliodonium salt approach to reactive alkylidenecarbene intermediates permits the introduction of the carbene's substituents from different sources.

Chapter 6 - Alkynyliodonium Salts in Organic Synthesis

Studies Directed Towards Total Syntheses of the Tropoloisoquinoline Alkaloids Grandirubrine (XIVa) and Imerubrine (XIVb). Part 1. Preparation of Two 4,5,6‐Trimethoxycyclopent(ij)isoquinolin‐7‐ones and Their Response to Robinson Annulation Conditions.

In connection with efforts to develop total syntheses of the tropoloisoquinoline alkaloids grandirubrine (1) and imerubrine (2), preparations of tricyclic ketone (8) and the related ethoxycarbonyl system (47) are described. Of the various approaches to (8) which were investigated, that proceeding from the known 8-bromo-5,6,7-trimethoxyisoquinoline (27) proved most efficient. Thus, conversion of (27), over three steps, into the Reissert anion (37), followed by alkylation of the latter with methyl iodide or ethyl bromoacetate, afforded (38) and (46), respectively. Treatment of compound (38) with sodium ethoxide resulted in regeneration of the isoquinoline nucleus and provided the C1-substituted compound (40). Upon reaction with lithium diisopropylamide, isoquinoline (40) underwent Dieckmann-type cyclization to give compound (8). A similar reaction sequence involving diester (46) afforded enamine (47), the X-ray crystal structure of which is reported. Efforts to effect Robinson annulation of (8) or (47) with methyl vinyl ketone (or a synthetic equivalent), and thereby produce the tetracyclic compound (10), have been unsuccessful.

Studies Directed Towards Total Syntheses of the Tropoloisoquinoline Alkaloids Grandirubrine and Imerubrine. I. Preparation of Two 4,5,6-Trimethoxycyclopent[ij]isoquinolin-7-ones and Their Response to Robinson Annulation Conditions

Designed loop extension followed by combinatorial screening confers high specificity to a broad matrix metalloproteinaseinhibitor.

Matrix metalloproteinases (MMPs) are key drivers of various diseases, including cancer. While several antibodies against MMPs are in development, our goal is to construct therapeutic anti-MMP inhibitors based on a natural broad MMP inhibitor, tissue inhibitor of metalloproteinases-2 (N-TIMP2). To confer high binding specificity toward one MMP type, we extend one of the N-TIMP2 loops, allowing it to interact with the non-conserved MMP surface. Multiple computational designs of the loop were used to design a focused library for yeast surface display, which was sorted for high binding to the target MMP-14 and low binding to off-target MMP-3. Deep sequencing of the two selected populations followed by comparative data analysis was used to identify the most promising variants, which were expressed, purified, and tested for inhibition of MMP-14 and off-target MMPs. Our best N-TIMP2 variant exhibited 29 pM binding affinity to MMP-14 and 2.4 µM affinity to MMP-3, 7500-fold more specific than WT N-TIMP2. Furthermore, the variant inhibited cell invasion with increased potency relative to WT N-TIMP2 in two breast cancer cell lines. We obtained the engineered variant high-accuracy model by including NGS data as input to AlphaFold multiple sequence alignment (MSA). Modeling results together with experimental mutagenesis demonstrate that the loop packs tightly against non-conserved residues on MMP-14 and clashes with MMP-3. This study demonstrates that introduction of loop extensions into inhibitors to stretch to the non-conserved surface of the target proteins is an attractive strategy for conferring high binding specificity in design of MMP inhibitors and other therapeutic proteins.

A broad matrix metalloproteinase inhibitor with designed loop extension exhibits ultrahigh specificity for MMP-14

Matrix metalloproteinase-9 (MMP-9) is an endopeptidase that remodels the extracellular matrix and has been implicated as a major driver in cancer metastasis. Hence, there is a high demand for MMP-9 inhibitors for therapeutic purposes. For such drug design efforts, large amounts of MMP-9 are required. Yet, the catalytic domain of MMP-9 (MMP-9Cat) is an intrinsically unstable enzyme that tends to auto-cleave within minutes, making it difficult to use in drug design experiments and other biophysical studies. We set our goal to design MMP-9Cat variant that is active but stable to autocleavage. For this purpose, we first identified potential autocleavage sites on MMP-9Cat using mass spectroscopy and then eliminated the autocleavage site by predicting mutations that minimize autocleavage potential without reducing enzyme stability. Four computationally designed MMP-9Cat variants were experimentally constructed and evaluated for auto-cleavage and enzyme activity. Our best variant, Des2, with 2 mutations, was as active as the wild-type enzyme but did not exhibit auto-cleavage after seven days of incubation at 37°C. This MMP-9Cat variant, with an identical to MMP- 9Cat WT active site, is an ideal candidate for drug design experiments targeting MMP-9 and enzyme crystallization experiments. The developed strategy for MMP-9CAT stabilization could be applied to redesign of other proteases to improve their stability for various biotechnological applications.

/pdf/computational-design-of-matrix-metalloprotenaise-9-mmp-9-176i8ue7.pdf

A. Bonadio

Papers

Studies Directed Towards Total Syntheses of the Tropoloisoquinoline Alkaloids Grandirubrine and Imerubrine. I. Preparation of Two 4,5,6-Trimethoxycyclopent[ij]isoquinolin-7-ones and Their Response to Robinson Annulation Conditions

Designed loop extension followed by combinatorial screening confers high specificity to a broad matrix metalloproteinaseinhibitor.

A broad matrix metalloproteinase inhibitor with designed loop extension exhibits ultrahigh specificity for MMP-14

Computational design of Matrix Metalloprotenaise-9 (MMP-9) resistant to autocleavage

Studies Directed Towards Total Syntheses of the Tropoloisoquinoline Alkaloids Grandirubrine (XIVa) and Imerubrine (XIVb). Part 1. Preparation of Two 4,5,6‐Trimethoxycyclopent(ij)isoquinolin‐7‐ones and Their Response to Robinson Annulation Conditions.