Crystal forms of a cyclictetrapeptide and processes for preparing

Abstract

The present disclosure provides a novel solid crystalline form of isopropyl ((11S,12S,13S,9S,12S)-9-amino-12-((1-(6-aminohexyl)-5-fluoro-1H-indol-3-yl)methyl)-4,10,13-trioxo-2-oxa-5,11-diaza-1(3,1)-pyrrolidina-7(1,3)-benzenacyclotridecaphane-12-carbonyl)-L-threoninate (Compound 1), or pharmaceutically acceptable salt thereof, compositions comprising Compound 1 (Form 1A and / or Form 2) synthesis thereof, and an improved crystallization process using Form 2 to prepare Form 1. Novel compositions also include Compound 1 solid Form 1, Form 2 and / or other crystalline and amorphous solid forms of Compound 1.

Description

CRYSTAL FORMS OF A CYCLICTETRAPEPTIDE AND PROCESSES FOR PREPARING CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Patent Application Serial No. 63 / 671,544 filed July 15, 2024, the entire contents of which are incorporated by reference herein. REFERENCE TO SEQUENCE LISTING SUBMITTED ELECTRONICALLY

[0002] The contents of the electronic sequence listing (25970-WO-PCT_SL.xml; Size: 4,655 bytes; and Date of Creation: September 25, 2024) are herein incorporated by reference in their entirety. FIELD

[0003] This disclosure relates generally to a novel solid crystalline form of isopropyl ((11S,12S,13S,9S,12S)-9-amino-12-((1-(6-aminohexyl)-5-fluoro-1H-indol-3-yl)methyl)-4,10,13- trioxo-2-oxa-5,11-diaza-1(3,1)-pyrrolidina-7(1,3)-benzenacyclotridecaphane-12-carbonyl)-L- threoninate (Compound 1), or pharmaceutically acceptable salt thereof, compositions comprising Compound 1 (Form 1A and / or Form 2) synthesis thereof, and an improved crystallization process using Form 2 to prepare Form 1. BACKGROUND

[0004] Isopropyl ((11S,12S,13S,9S,12S)-9-amino-12-((1-(6-aminohexyl)-5-fluoro-1H-indol-3- yl)methyl)-4,10,13-trioxo-2-oxa-5,11-diaza-1(3,1)-pyrrolidina-7(1,3)-benzenacyclotridecaphane- 12-carbonyl)-L-threoninate (Compound 1), or pharmaceutically acceptable salt thereof, is a cyclictetrapeptide useful in synthetic processes for making complex polypeptides. The structure of Compound 1 is shown below.

[0005] Compound 1 and the process for synthesizing are described in. See also USSN 63 / 639279, filed 4 / 26 / 24; USSN 63 / 611847, filed 12 / 19 / 23; USSN 63 / 668869, filed 7 / 9 / 2024 Attorney Docket No.25869 (Engineered ATP-Dependent Ligases for the Synthesis of Oligopeptides) and Attorney Docket No.25920 (Engineered Carboxylesterase Enzymes And Methods For Their Use In Macrocyclization Of Non-Canonical Tetrapeptides), both Attorney Docket Nos.25869 and 25920 filed contemporaneously with the instant disclosure, and all incorporated by reference in their entirety.

[0006] The synthesis of Compound 1 is difficult to control through traditional chemical methods of amide formation, resulting in lack of specificity and formation of impurities and undesired byproducts. There remains a need for a simplified and improved manufacturing process for the production of Compound 1.

[0007] As described herein, new crystalline Compound 1 (Form 1) provides the foundation for a simplified crystallization-based purification process. The new crystallization process provides a higher yield, significant reduction in solvent usage, and shorter cycle times. The new process also utilizes standard manufacturing equipment, thereby affording a more productive, greener, and portable manufacturing process. Additionally, the streamlined purification process reduces overall manufacturing time, culminating in cost savings and a reduction in energy consumption. SUMMARY

[0008] The present disclosure provides a novel solid form of Compound 1 (Form 1) bisoxalate salt and a novel crystalline solvate of Compound 1 (Form 2) compositions comprising Compound 1 and / or 2, synthesis of Form 1 and Form 2, and an improved crystallization process using Form 2 to prepare Form 1. Novel compositions also include Compound 1 solid Form 1, Form 2 and / or other crystalline and amorphous solid forms of Compound 1.

[0009] Crystalline Compound 1, (Form 1), may be present in a wet phase (referred to herein as “Form 2”). BRIEF DESCRIPTION OF THE DRAWINGS

[0010] FIG.1 is a graph of a Powder X-Ray Diffraction (“PXRD”) pattern of Compound 1 Form 2 crystalline n-BuOH solvate form, generated using the equipment and methods described herein. The graph plots the intensity of the peaks as defined by counts per second versus the diffraction angle 2 theta (2θ) in degrees.

[0011] FIG.2 is a graph of a Powder X-Ray Diffraction (“PXRD”) pattern of Compound 1 crystalline Form 1 bisoxalate salt, generated using the equipment and methods described herein.The graph plots the intensity of the peaks as defined by counts per second versus the diffraction angle 2 theta (2θ) in degrees. DETAILED DESCRIPTION

[0012] The present disclosure relates to novel solid forms of Compound 1 (Form 1), processes for preparing such, as well as manufacturing processes that utilize Form 2 to prepare Form 1 of Compound 1. The novel solid forms of the instant invention may be a crystalline form, amorphous form or mixture thereof, unless otherwise specified.

[0013] X-ray Powder Diffraction (XRPD) Powder X-ray Diffraction data were acquired on a Panalytical X-pert Pro PW3040 System configured in the Bragg-Brentano configuration and equipped with a Cu radiation source with monochromatization to Kα achieved using a Nickel filter. A fixed slit optical configuration was employed for data acquisition. Data were acquired between 2 and 40° 2θ. Samples were prepared by gently pressing either the wet solid sample or the dry powdered sample of Form 1 onto a shallow cavity zero background silicon holder. Results are seen in Figures 1 and 2.

[0014] The intensity of the peaks (y-axis is in counts per second) were plotted versus the 2 theta angle (x-axis is in degrees 2 theta). In addition, the data were plotted with detector counts normalized for the collection time per step versus the 2-theta angle. Peak locations (on the 2-theta x-axis) consistent with these profiles are displayed in Table 1 (+ / - 0.2° 2 theta).

[0015] In another aspect, crystalline Form 2 is characterized by a powder x-ray diffraction pattern having one or more characteristic diffractions at angles (2 theta ±0.2) listed in Table 1. Table 1: Diffraction peaks and corresponding d-spacings for crystalline Form 2 (wet phase) of Compound 1 Peak Number Position [° Two d-spacing Diagnostic Peak Theta] [Å] Rel. Int. [%] Set25970 Peak Number Position [° Two d-sp Diagnostic Peak Theta] acing [Å] Rel. Int. [%] Set 12 167 53 2472

[0016] In a further aspect, the PXRD peak locations displayed in Table 1 and / or FIG.1 most characteristic of crystalline Form 2 can be selected and grouped as “diagnostic peak sets” to conveniently distinguish this crystalline form from others. Selections of such characteristic peaks are set out in Table 1 in the column labeled Diagnostic Peak Set.

[0017] Thus, in another aspect, there is provided a crystalline Form 2 characterized by a powder x-ray diffraction pattern comprising each of the 2-theta values listed in Diagnostic Peak Set 1 in Table 1, + / - 0.2° 2-theta.

[0018] In another aspect, there is provided a crystalline Form 2 characterized by a powder x-ray diffraction pattern comprising each of the 2-theta values listed in Diagnostic Peak Set 2 in Table 1, + / - 0.2° 2-theta.

[0019] In another aspect, there is provided a crystalline Form 2 characterized by a powder x-ray diffraction pattern comprising each of the 2-theta values listed in Diagnostic Peak Set 3 in Table 1, + / - 0.2° 2-theta.

[0020] In another aspect, there is provided a crystalline Form 2 characterized by a powder x-ray diffraction pattern comprising each of the 2-theta values listed in Diagnostic Peak Set 4 in Table 1, + / - 0.2° 2-theta.

[0021] In another aspect, there is provided crystalline Compound 1 ((n-BuOH) solvate (wet phase) Form 2)) characterized by a powder x-ray diffraction pattern comprising each of the 2- theta values in Table 1, + / - 0.2° 2-theta of 4.4, 7.6, 8.8, 9.6, 11.6, and 13.8.

[0022] In another aspect, there is provided crystalline Compound 1 ((n-BuOH) solvate (wet phase) Form 2)) characterized by a powder x-ray diffraction pattern comprising each of the 2- theta values listed in Table 1, + / - 0.2° 2-theta of 4.4, 5.8, 7.6, 8.8, 9.6, 10.1, 11.6, 13.8, 15.3, 15.9, 16.3, 16.7, 17.0, 17.6, 18.1, 19.6, 20.3, 20.7, 21.4, 22.4, 23.0, 23.6, 24.0, 24.9, 25.2, and 25.7.

[0023] In another aspect, there is provided crystalline Compound 1 ((n-BuOH) solvate (wet phase) Form 2)) characterized by a powder x-ray diffraction pattern comprising each of the 2- theta values listed in Table 1, + / - 0.2° 2-theta of 4.4, 5.8, 7.6, 8.8, 9.6, 10.1, 11.6, 13.8, 15.3, 15.9, 16.3, 16.7, 17.0, 17.6, 18.1, 19.2, 19.6, 20.3, 20.7, 21.4, 22.1, 22.4, 23.0, 23.6, 24.0, 24.9, 25.2, 25.7, 26.6, 28.0, 28.3, 29.4, and 29.9.

[0024] In another aspect, there is provided crystalline Compound 1 ((n-BuOH) solvate (wet phase) Form 2)) characterized by a powder x-ray diffraction pattern comprising each of the 2- theta values listed in Table 1, + / - 0.2° 2-theta of 4.4, 5.8, 7.6, 8.8, 9.6, 10.1, 11.6, 13.8, 15.3, 15.9, 16.3, 16.7, 17.0, 17.6, 18.1, 19.2, 19.6, 20.3, 20.7, 21.4, 22.1, 22.4, 23.0, 23.6, 24.0, 24.9, 25.2, 25.7, 26.6, 28.0, 28.3, 29.4, 29.9, 31.2, 32.0, 33.2, 34.5, 37.3, 38.3.

[0025] In another aspect, crystalline Compound 1 (Form 1) is characterized by a powder x-ray diffraction pattern having one or more characteristic diffractions at angles (2 theta ±0.2) listed in Table 2. Table 2: Diffraction peaks and corresponding d-spacings for crystalline Form 1(dry phase) of Compound 1 Peak Number Position [° Two d-spacing [Å] Rel. Int. [%] Diagnostic PeakPeak Number Position [° Two d- Diagnostic Peak Theta] spacing [Å] Rel. Int. [%] Set 473 121 231Peak Number Position [° Two d-spacing [Å] Rel. Int Diagnostic Peak Theta] . [%] Set 49 303 30 424characteristic of crystalline Form 1 can be selected and grouped as “diagnostic peak sets” to conveniently distinguish this crystalline form from others. Selections of such characteristic peaks are set out in Table 2 in the column labeled Diagnostic Peak Set.

[0027] Thus, in another aspect, there is provided a crystalline Form 1 characterized by a powder x-ray diffraction pattern comprising each of the 2-theta values listed in Diagnostic Peak Set 1 in Table 2, + / - 0.2° 2-theta.

[0028] In another aspect, there is provided a crystalline Form 1 characterized by a powder x-ray diffraction pattern comprising each of the 2-theta values listed in Diagnostic Peak Set 2 in Table 2, + / - 0.2° 2-theta.

[0029] In another aspect, there is provided a crystalline Form 1 characterized by a powder x-ray diffraction pattern comprising each of the 2-theta values listed in Diagnostic Peak Set 3 in Table 2, + / - 0.2° 2-theta.

[0030] In another aspect, there is provided a crystalline Form 1 characterized by a powder x-ray diffraction pattern comprising each of the 2-theta values listed in Diagnostic Peak Set 4 in Table 2, + / - 0.2° 2-theta.

[0031] In another aspect, there is provided crystalline Compound 1 ((bisoxalate (dry phase) Form 1)) characterized by a powder x-ray diffraction pattern comprising each of the 2-theta values in Table 2, + / - 0.2° 2-theta of 4.4, 4.8, 7.3, 7.7, 8.7, and 9.5.

[0032] In another aspect, there is provided crystalline Compound 1 ((bisoxalate (dry phase) Form 1)) characterized by a powder x-ray diffraction pattern comprising each of the 2-theta values in Table 2, + / - 0.2° 2-theta of 4.4, 4.8, 6.0, 7.3, 7.7, 8.7, 9.5, 10.2, 11.2, 12.0, 12.5, 13.1, 14.0, 14.3, 14.8, 15.4, 15.9, 16.1, 16.2, 16.6, 17.1, 17.5, 18.3, 18.5, 19.0, 19.2, and 19.7.

[0033] In another aspect, there is provided crystalline Compound 1 ((bisoxalate (dry phase) Form 1)) characterized by a powder x-ray diffraction pattern comprising each of the 2-theta values in Table 2, + / - 0.2° 2-theta of 4.4, 4.8, 6.0, 7.3, 7.7, 8.7, 9.5, 10.2, 11.2, 12.0, 12.5, 13.1,14.0, 14.3, 14.8, 15.4, 15.9, 16.1, 16.2, 16.6, 17.1, 17.5, 18.3, 18.5, 19.0, 19.2, 19.7, 20.0, 20.5, 20.7, 21.5, 21.8, 22.1, 22.4, 22.8, 23.2, 24.2, 24.6, 25.2, 26.3, 26.4, 27.1, 27.4, 27.6, 28.1, 28.7, 29.1, and 29.6.

[0034] In another aspect, there is provided crystalline Compound 1 ((bisoxalate (dry phase) Form 1)) characterized by a powder x-ray diffraction pattern comprising each of the 2-theta values in Table 2, + / - 0.2° 2-theta of 4.4, 4.8, 6.0, 7.3, 7.7, 8.7, 9.5, 10.2, 11.2, 12.0, 12.5, 13.1, 14.0, 14.3, 14.8, 15.4, 15.9, 16.1, 16.2, 16.6, 17.1, 17.5, 18.3, 18.5, 19.0, 19.2, 19.7, 20.0, 20.5, 20.7, 21.5, 21.8, 22.1, 22.4, 22.8, 23.2, 24.2, 24.6, 25.2, 26.3, 26.4, 27.1, 27.4, 27.6, 28.1, 28.7, 29.1, 29.6, 30.3, 30.9, 31.5, 32.3, 34.0, 34.7, and 38.8.

[0035] In another aspect, crystalline Compound 1, Form 2, is obtained by a process comprising the steps of: a) combining a solution containing Compound 1 as a free base having a temperature of about 15°C to about 35 °C, with oxalic acid solution comprising water, oxalic acid, and a miscible solvent to form a slurry; b) filtering the slurry to produce a wet cake while maintaining a temperature of about 10°C to about 20°C; c) washing the wet cake with (n-BuOH); d) filtering to obtain wet solids of Compound 1 (Form 2).

[0036] In an embodiment of this disclosure for making Compound 1, Form 2, the amount of oxalic acid solution combined with the solution of Compound 1 free base in step a) results in from about 1.00 to about 2.5 equivalents (eq.) of oxalic acid relative to Compound 1 free base added to the mixture, preferably from about 1.30 to about 2.3, more preferably 1.35 to about 2.25.

[0037] In an embodiment of this disclosure, the resultant slurry from step a) undergoes an annealing step wherein the slurry is heated to about 30°C-50°C, preferably about 35°C-40,°C, aged, additional oxalic acid solution added, and mixture cooled to about 5°C-25 °C. A subembodiment of this aspect of the disclosure is realized when the amount of additional oxalic acid solution results in from about 0.005 to about 0.75 equivalents (eq.) of oxalic acid relative to Compound 1 free base, preferably 0.10 to about 0.60 eq., added to the mixture. Another subembodiment of this aspect of the disclosure is realized when the additional oxalic acid solution is added over a period of about 1 to about 10 hours, preferably about 2 to about 8 hours.

[0038] Another subembodiment of this aspect of the disclosure is realized when the annealing step is optionally repeated at least once. A non-limiting example of the annealing step conducted two times is illustrated as follows:First Annealing Step – Slurry from step a) heated to about 35°C to about 45 °C and held at temperature for no less than 30 minutes. Mixture is cooled to about 30°C to about 40°C, then aged for 1 hour. Oxalic acid solution (0.15 eq) is added to over a period of 3 hours. Then, batch is cooled to about 15°C to about 25 °C, over 3 hours, and aged for 30 minutes. Second Annealing Step – Mixture from first Heating and Cooling Step is heated to about 35°C to about 45 °C, and held at temperature for no less than 30 minutes. Mixture is cooled to about 30°C to about 40°C, then aged for 1 hour. Oxalic acid (0.55 eq) is added over a period of 8 hours and aged for 1 hour, then cooled to about 7°C to about 13 °C over 6 hours.

[0039] In an embodiment of this aspect of the disclosure, the slurry from step a) undergoes at least one annealing step. Still in another subembodiment of this aspect of the disclosure, the slurry undergoes two or more annealing steps.

[0040] In a further embodiment of this disclosure, the first miscible solvent in oxalic acid solution is one that dissolves the oxalic acid with some water present. Examples of first miscible solvents include acetonitrile, isopropyl alcohol, tert-amyl alcohol, 1-propanol, ethanol, tert-butyl alcohol, dioxane, methyl-tert-butyl ether (MTBE), tetrahydrofuran (THF), or mixtures thereof. In a further embodiment, the miscible solvent in oxalic acid solution of step a) is MTBE. Another aspect of this embodiment is realized when crystal solid Compound 1 (Form 1) is obtained by drying the wet solids of Compound 1 (Form 2) of step f), to produce dry solids of Compound 1 (Form 1). An aspect of this embodiment is realized when the drying is done with nitrogen and vacuum for at least 10 hours at a temperature of about 20°C to about 30 °C.

[0041] In another aspect, crystalline Compound 1, Form 1, is obtained by a process comprising the steps of: i) combining a solution containing Compound 1 as a free base having a temperature of about 15°C to about 35 °C, with oxalic acid solution comprising water, oxalic acid, and a first miscible solvent to form a slurry; ii) adding Compound 1 ( Form 1) seeds, adding additional oxalic acid solution; iii) filtering resultant slurry to produce a wet cake; iv) washing the wet cake with (n-BuOH) forming a slurry; v) filtering and washing the slurry with a second miscible solvent to produce wet solids of Compound 1 (Form 2) vi) drying the wet crystals of Compound 1 (Form 2) to produce a solid form of Compound 1 (Form 1).

[0042] In another embodiment of this disclosure, the amount of oxalic acid solution combined with the solution of Compound 1 free base in step i) results in from about 1.00 to about 2.5equivalents (eq.) of oxalic acid relative to Compound 1 free base added to the mixture, preferably from about 1.30 to about 1.6, more preferably about 1.35 to about 1.55.

[0043] In another embodiment of this disclosure, the amount of seeds in step ii) added to the mixture is from about 0 to about 5 wt%, preferably from about 0.3 to about 1 wt%. A subembodiment of this aspect of the disclosure is realized when after addition of the seeds, the temperature is heated to about 25°C to about 45 °C. Another subembodiment of this aspect of the disclosure is realized when the amount of oxalic acid solution added in step ii) results in from about 0.005 to about 1.5 equivalents (eq.) of oxalic acid relative to Compound 1 free base, preferably 0.08 to about 0.75 eq., added to the mixture. Another subembodiment of this aspect of the disclosure is realized when the amount of oxalic acid solution added over a period of about 0.5 to about 12 hours. Another subembodiment of this aspect of the disclosure is realized when the mixture is cooled to about 10°C to about 30 °C, preferably, to about 13°C to about 27 °C after addition of the oxalic acid solution. In a further embodiment, wet solids of Compound 1 solvate (Form 2), dry solids of Compound 1 (Form 1) or a mixture thereof can be used to seed in step ii) above.

[0044] In another embodiment of this disclosure, the mixture from step ii) undergoes an annealing step wherein the slurry is heated to about 30°C-50°C, preferably about 32°C-48,°C, then cooled to 27°C-43°C, aged, additional oxalic acid solution added, and mixture cooled to about 5°C-30 °C, preferably about 13°C-27 °C. A subembodiment of this aspect of the disclosure is realized when the amount of additional oxalic acid solution results in from about 0.10 to about 0.20 equivalents (eq.) of oxalic acid relative to Compound 1 free base, preferably 0.12 to about 0.17 eq., added to the mixture. Another subembodiment of this aspect of the disclosure is realized when the additional oxalic acid solution is added over a period of about 0.5 to about 10 hours, preferably about 0.5 to about 8 hours.

[0045] Another subembodiment of this aspect of the disclosure is realized when the annealing step is optionally repeated at least once. Another subembodiment of this aspect of the disclosure is realized when the annealing step is optionally repeated two or more times.

[0046] Another embodiment of this disclosure is realized in step iii) when the temperature is maintained a temperature of about 10°C to about 20°C.

[0047] An aspect of this embodiment is realized when crystal solid Compound 1 (Form 1) is obtained by drying the washed Compound 1 solvate (Form 2) of step iv), to produce dry solids of Compound 1 (Form 1). An aspect of this embodiment is realized when the drying is done with nitrogen and vacuum for at least 10 hours at a temperature of about 20°C to about 30 °C.25970

[0048] These manufacturing methods are useful for the manufacture of compositions comprising Compound 1 in solid form.

[0049] In an embodiment, Examples of the second miscible solvents include acetonitrile, isopropyl alcohol, tert-amyl alcohol, 1-propanol, ethanol, tert-butyl alcohol, dioxane, methyl- tert-butyl ether (MTBE), tetrahydrofuran (THF), or mixtures thereof. In a further embodiment, the second miscible solvent in oxalic acid solution of step a) is MTBE.

[0050] Compositions of the instant invention can include a Compound 1 (Form 1A) and Compound 1 (Form 2), obtained by methods described herein.

[0051] In some embodiments, the order of addition of reactants is not critical. The reactants may be added together at the same time to a solvent (e.g., monophasic solvent, biphasic aqueous co-solvent system, and the like), or alternatively, some of the reactants may be added separately, and some together at different time points.

[0052] The instant invention provides novel solid forms that are used in a simplified manufacturing process to produce macrocyclic peptides and other complex molecules. The manufacturing process, shown herein, results in a higher yield, significant reduction in solvent usage, shorter cycle times, and utilizes standard manufacturing equipment. Thereby affording a more productive, greener, and portable manufacturing process. Additionally, a streamlined purification process would reduce overall manufacturing time, culminating in cost savings and a reduction in energy consumption.

[0053] In addition, the manufacturing method involves an improved crystallization process for making Compound 1 Form 1 and Form 2. The improved crystallization process results in significantly larger particle growth, which in in turn leads to increased filtration rates. This culminates in reduced manufacturing time, cost savings, and a reduction in energy consumption. Definitions

[0054] The terms “effective amount” or “therapeutically effective amount” means the amount of the subject compound that will elicit the biological or medical response of a tissue, system, animal or human that is being sought by the researcher, veterinarian, medical doctor or other clinician.

[0055] "Pharmaceutically acceptable carriers" for therapeutic use are well known in the pharmaceutical art, and are described, for example, in Remingtons Pharmaceutical Sciences, Mack Publishing Co. (A.R. Gennaro edit.1985). For example, sterile saline and phosphate- buffered saline at physiological pH may be used. Preservatives, stabilizers, dyes and even flavoring agents may be provided in the pharmaceutical composition. For example, sodium25970 benzoate, sorbic acid and esters of p-hydroxybenzoic acid may be added as preservatives. rd. at 1449. In addition, antioxidants and suspending agents may be used.

[0056] "Pharmaceutically acceptable salt" refers to salts of the compounds of the present invention derived from the combination of such compounds and an organic or inorganic acid (acid addition salts) or an organic or inorganic base (base addition salts). Examples of pharmaceutically acceptable salt include but not limited to those described in for example: "Handbook of Pharmaceutical Salts, Properties, Selection, and Use", P. Heinrich Stahl and Camille G. Wermuth (Eds.), Published by VHCA (Switzerland) and Wiley- VCH (FRG), 2002. The compounds of the present invention may be used in either the free base or salt forms, with both forms being considered as being within the scope of the present invention.

[0057] “Annealing” refers to heating up and cooling down slurry crystals and is generally conducted to improve morphology. Seeded Crystallization Process

[0058] The process for making Compound 1 crystals (Form 1) using seeds can be made as generally described in the following paragraphs and illustrated in Examples 1 and 3.

[0059] The final organic layer solution containing Compound 1 free base, as described in Example 1 below, is transferred through an inline 10 micron filter to a vessel for crystallization (reactor 1) and the temperature adjusted to from about 15°C to about 35 °C. Prepared in a separate vessel (reactor 2) is a solution of oxalic acid that is made, for example, by dissolving 1.75-2.25 eq of oxalic acid in 6-10 L / kg of an MTBE solution with 0.38-0.62 %wt water, agitating and aging to ensure complete dissolution.

[0060] Oxalic acid solution from reactor 2 is charged into reactor 1, so that a total of about 1.39-1.51 equivalents of oxalic acid are transferred into reactor 1. Compound 1 (Form 1 as a bisoxalate salt) about 0.0044-0.0056 kg / kg is added as seeds into the first reactor. Reactor 1 is heated to from about 27°C to about 43 °C and an additional 0.08-1.2 equivalents of oxalic acid from the solution from reactor 2 is charged into reactor 1 over a period of 0.5-8 hours. Reactor 1 is cooled to from about 13°C to about 27 °C.

[0061] Reactor 1 is then heated to from about 32°C to about 48 °C and aged for no less than 30 minutes. The mixture is cooled to from about 27°C to about 43°C and 0.12-0.17 eq of oxalic acid from the solution from reactor 2 is charged into reactor 1 over a period of 0.5-8 hours. The mixture is then cooled to from about 13°C to about 27 °C and aged for 30 minutes.

[0062] Reactor 1 is then heated to from about 32°C to about 48 °C and aged for no less than 30 minutes. The mixture is cooled to from about 27°C to about43°C and 0.5-0.6 eq of oxalic acid25970 from the solution from reactor 2 is charged into reactor 1 over a period of 4-12 hours. The mixture is aged for no less than 30 min, then cooled to from about 2°C -18 °C and the final slurry aged for no less than 30 min.

[0063] The batch is then isolated in a jacketed filter funnel at from about 5°C to about 25 °C. A displacement wash of the cake with 2.65-3.35 L / kg of n-butanol is performed. A second displacement wash of the cake with 2.65-3.35 L / kg of MTBE is formed and the isolator jacket temperature is adjusted to from about 15°C to about 35 °C. A slurry wash with 5.5-6.5 L / kg of MTBE with 0.6-1.4 % water v / v solution for no less than 15 minutes is performed. A second slurry wash 5.5-6.5 L / kg of MTBE with 0.6-1.4 % water v / v solution for no less than 15 minutes is performed. A third displacement wash with 2.65-3.35 L / kg of MTBE with 0.6-1.4 % water v / v solution is performed.

[0064] Drying of the cake is initiated by blowing nitrogen for no less than 1 hour. Static drying with nitrogen and vacuum is began for no less than 5 hours with a jacket temperature of from about 20°C to about 30 °C. A third slurry wash with 2.65-3.35 L / kg of MTBE with 0.6-1.4 % water v / v solution for no less than 15 minutes is performed. A fourth displacement wash with 2.65-3.35 L / kg MTBE with 0.6-1.4 % water v / v solution is performed. Nitrogen is blown through the cake to deliquor it. After deliquoring the cake, further drying is completed with nitrogen and vacuum at from about 15°C to about 35 °C with intermittent agitation for no less than 24 hrs. The final dry cake is then discharged and packaged. Unseeded Crystallization Process

[0065] The process for making Compound 1 crystals (Form 1) can be made as generally described in the following paragraphs.

[0066] The final organic layer solution containing Compound 1 free base, as described in Examples 1 and 3, is transferred through an inline 10 micron filter to a vessel for crystallization (reactor 1) and the temperature adjusted to from about 20°C to about 30 °C. Prepared in a separate vessel (reactor 2) is a solution of oxalic acid that is made, for example by dissolving 2.00 eq of oxalic acid in 8.3 L / kg of an MTBE solution with 0.5 %wt water, agitating and aging the solution to ensure complete dissolution].

[0067] Oxalic acid solution from reactor 2 is charged into reactor 1, so that a total of about 1.45 equivalents of oxalic acid are transferred into reactor 1 and the temperature is adjusted to from about 30°C to about 40 °C and held there for no less than 30 minutes. Additional 0.1 equivalents of oxalic acid from the solution from reactor 2 is charged into reactor 1 over a period of 2 hours. The reactor is then cooled to from about 15°C to about 25 °C over 4 hours, then aged for an additional hour.25970

[0068] Reactor 1 is then heated to from about 35°C to about 45 °C and held at temperature for no less than 30 minutes. The mixture is cooled to from about 30°C to about 40°C, aged for 1 hour and 0.15 eq of oxalic acid from the solution from reactor 2 is charged into reactor 1 over a period of 3 hours. The batch is cooled to from about 15°C to about 25 °C over 3 hours, and aged for 30 minutes.

[0069] Reactor 1 is heated to from about 35°C to about 45 °C, held at temperature for no less than 30 minutes, then cooled to from about 30°C to about 40°C, and aged for 1 hour. Oxalic acid (0.55 eq) from the solution from reactor 2 is charged into reactor 1 over a period of 8 hours, aged for 1 hour, then cooled to from about 7°C to about 13 °C over 6 hours. The final slurry is aged for no less than 1 hour.

[0070] The batch is isolated in a jacketed filter funnel at from about 10°C to about 20 °C. A displacement wash of the cake with 3 L / kg of n-butanol is performed. A second displacement wash of the cake with 3 L / kg of MTBE is performed and the isolator jacket temperature is adjusted to from about 20°C to about 30 °C. A slurry wash with 6 L / kg of MTBE with 1 % water v / v solution for 2 hours is performed. A second slurry wash 6 L / kg of MTBE with 1 % water v / v solution for 2 hours is performed. A third displacement wash with 3 L / kg of MTBE with 1 % water v / v solution is performed.

[0071] Drying of the cake is initiated by blowing nitrogen for at least 2 hours. Static drying with nitrogen and vacuum for at least 10 hours with a jacket temperature of from about 20 to about 30 °C is then began. A third slurry wash is performed with 3 L / kg of MTBE with 1 % water v / v solution for 2 hours. A fourth displacement wash is performed with 3 L / kg MTBE with 1 % water v / v solution. Nitrogen is blown through the cake to deliquor it. After deliquoring the cake, further drying is completed with nitrogen and vacuum at from about 20 to about 30 °C with intermittent agitation for 75 hours. The final dry cake is then discharged and packaged.

[0072] The invention is illustrated in the following generic schemes and the examples in the experimental Section that follows. This section is set forth to aid in an understanding of the invention but is not intended to and should not be construed to limit in any way the invention as set forth in the claims which follow thereafter.

[0073] The compounds of the present invention were prepared by the general methods outlined in the synthetic schemes.25970 EXAMPLES

[0074] The disclosed process for Compound 1, crystallization of Compound 1, and crystal forms of Compound 1 are not to be limited in scope by the specific embodiments and examples described herein. Indeed, various modifications of the disclosure in addition to those described will become apparent to those skilled in the art from the foregoing description and accompanying figures. Such modifications are intended to fall within the scope of the appended claims.

[0075] All references (e.g., publications or patents or patent applications) cited herein are incorporated herein by reference in their entirety and for all purposes to the same extent as if each individual reference (e.g., publication or patent or patent application) was specifically and individually indicated to be incorporated by reference in its entirety for all purposes. Other embodiments are within the following claims.

[0076] In reference to the present disclosure, the technical and scientific terms used in the descriptions herein will have the meanings commonly understood by one of ordinary skill in the art, unless specifically defined otherwise. Abbreviations NaPi Sodium phosphate HTS High throughput screening25970 UHPLC Ultra high-performance liquid chromatography wt weight HTP Hi h th h tIntermediate Example 1(71.5 mg), dipeptide 2c (oxalate salt (111.3 mg)), amino acid monomer 1f hydrochloride salt [Amino- Phe.HCl] (64.3 mg), magnesium chloride hexahydrate (61 mg), and adenosine 5'-monophosphate monohydrate [AMP] (5.9 mg) was dissolved in 2.8 mL water. Sodium hydroxide (5 N solution in water) was then added to adjust pH to 7.5. Next, sodium hexametaphosphate (122.4 mg), amino acid monomer 1d hydrobromide salt [Trp-Linker.HBr] (99.5 mg), tergitol 15-s-9TM(90 mg) was added. Sodium hydroxide (5 N solution in water) was then added to adjust pH to 8.0. To initiate the reaction, 30 mL PPK22 (20 g / L) and 30 mL amino acid ligase Trp-ligase SEQ ID NO: 1 (50 g / L) was added. The reaction mixture was agitated at 20°C -25 °C for ca.24 hrs to give Tripeptide 3d as a slurry. MS [M+H]+C35H55FN5O8, 692.4029; Found: 692.4030.25970 Intermediate Example 2

[0078] Tripeptide 3d (346 mg) was dissolved in 0.1 M HEPES (pH 7.5) to make a 150 mM stock solution at 20 °C. PPK22 (20 mg) lyophilized powder was dissolved in 1 mL 0.1 M HEPES (pH 7.5) to make a 20 mg / mL stock solution. Phe-ligase SEQ ID NO.2 (50mg) lyophilized powder was dissolved in 1 mL 0.1 M HEPES (pH 7.5) to make a 50 mg / mL stock solution.1 M HEPES (pH 8.0) was purchased from Thermo Fisher.

[0079] To a 2-mL glass vial, 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (71.5 mg), monomer 1f hydrochloride salt (64.3 mg), magnesium chloride hexahydrate (61 mg), and adenosine 5'-monophosphate monohydrate (5.9 mg) was dissolved in 2.8 mL water. Sodium hydroxide (5 N solution in water) was then added to adjust pH to 7.7. Next, 150 mL tripeptide 3d (150 mM) was added, followed by sodium hexametaphosphate (9.2 mg) and tergitol 15-s-9TM(90 mg). To initiate the reaction, 30 mL PPK22 (20 g / L) and 60 mL Phe-ligase SEQ ID NO.2 (50 g / L) was added. The reaction mixture was agitated at 20°C -25 °C for ca.6 hrs to give tetrapeptide 3k as a slurry, MS [M+H]+C45H67FN7O9, 868.4979; Found: 868. Intermediate Example 3probe was added 15 mL of NMP and 5.20 g (54.0 mmol) of solid sodium tert-butoxide. To this solution was added 3.00 g (13.5 mmol) of solid 5-fluorotryptophan (Compound 3) and the sides of the flask rinsed with 6 mL of NMP. To the mixture was added 0.073 mL of water and the mixture was25970 warmed to 45 ºC for 1.5 h. In a separate flask was added 5.2 g (14.2 mmol) of 6-Bromohexan-1- aminium 4-methylbenzenesulfonate (Compound 2A) and 10 mL of NMP and the mixture was warmed to 45 ºC to give a homogeneous solution which was added to the above mixture. The mixture is aged for 1 h, and then at this point and the pH of the mixture was adjusted to between 4-9 by the addition of 48% aqueous HBr solution. The mixture is then cooled to below 5 °C to allow for crystal growth of Compound 1d. The temperature is returned to 20 °C and then is diluted with 30 mL of ½ saturated aqueous NaBr which was prepared by dissolving 49.0 g of NaBr in 100 mL of water. The temperature of the reaction mixture was raised between 50-55 ºC, and the resulting slurry was aged at 50 °C overnight, and then cooled to 5 °C over a period of 5 h. The slurry was filtered and washed with 8:11-PrOH / water and dried under vacuum at 55 °C overnight to give Compound 1d: mp 246 ºC (DSC);1H NMR (DMSO-d6, 500 MHz) δ 7.87 (br s, 5H), 7.41 (dd, 1H, J = 8.9 and 4.4 Hz), 7.37 (dd, 1H, J = 10.1 and 2.5 Hz), 7.28 (s, 1H), 6.94 (td, 1H, J = 9.2 and 2.5 Hz), 4.12 (t, 2H, J = 6.4 Hz), 3.55 (t, 2H, J = 5.5 Hz), 3.30 (br s, 2H), 3.10 (qd, 2H, J = 15.0 and 5.5 Hz), 2.71 (m, 2H), 1.73 (m, 2H), 1.49 (m, 2H), 1.37-1.21 (m, 2H), 1.13 (m, 2H);13C NMR (DMSO-d6, 125 MHz) δ 171.5, 156.9 (d, J = 231.0 Hz), 132.7, 129.7, 128.1 (d, J = 9.8 Hz), 110.6 (d, J = 10.0 Hz), 109.0 (d, J = 26.4 Hz), 108.5 (d, J = 4.8 Hz), 103.7 (d, J = 22.0 Hz), 54.5, 45.4, 38.5, 29.4, 26.8, 26.6, 25.6, 25.3;19F NMR (DMSO-d6, 471 MHz) δ -125.6. HRMS Cacld. For C17H15FNO2: 322.1931 [M + H]. Found: 322.1925 [M + H]. See USSN 63 / 611847, filed 12 / 19 / 23; Attorney Docket 25886, incorporated herein in its entirety. Intermediate Example 4 8 Batch procedure:

[0081] A 1-L pyrex bottle was charged with L-3-cyanophenylalanine (Compound 4g: See (Knittel et al., 1990 Pept. Res.3:176-181; US Patent Publication US2006 / 0142305; Chien et al., 2018 J. Med. Chem.61:7358-7373) (34.00 g, 1.0 equiv.), isopropanol (245 mL), water (163 mL) and 12 M aqueous HCl solution (29.6 mL), and stirred until all solids had dissolved. The obtained solution was transferred into a 1-L autoclave followed by adding 10% Pd / C catalyst (1.7 g). The autoclave was sealed and purged with nitrogen gas three times. Next, it was purged25970 with hydrogen gas three times, the reaction pressure set to 400 psi and the temperature to 25oC, before aging the reaction overnight. When the reaction was complete, the autoclave was vented and the atmosphere inerted by purging three times with nitrogen gas. Agitation was stopped and the solution discharged into a 1-L Pyrex bottle. The reaction mixture was then filtered with a funnel under vacuum to obtain a solution containing Compound 1f. See also USSN 63 / 639279, filed 4 / 26 / 24; Attorney Docket 25931, incorporated herein in its entirety. Intermediate Example 5 (2S,3S)-3-(2-(tert-butoxy)-2-oxoethoxy)-2-(((2S,3R)-3-hydroxy-1-isopropoxy-1-oxobutan-2- yl)carbamoyl)pyrrolidine (batch hydrogenation)

[0082] To a 3- -3-(2-(tert-butoxy)-2-oxoethoxy)-2-(((2S,3R)-3-hydroxy-1-isopropoxy-1-oxobutan-2- yl)carbamoyl)pyrrolidine-1-carboxylate (~40.0 g, 1:1:0.04 MTBE / iPrOH / water, 390 mL) and 5% Pd / C (2 g). A balloon of H2was attached, and the headspace of the flask was flushed thoroughly with H2. The reaction flask was placed under H2 atmosphere, and the reaction mixture was then heated to 55°C. The reaction mixture was aged for 4 h. After aging, the reaction mixture was poured onto a pad of diatomaceous earth (CELITE® 545, available from Millipore Sigma, 20 g) that had been wetted with iPrOH:MTBE 1:1. The filtrate was collected, and the filter pad was washed with 1:1 iPrOH / MTBE (40 mL). The wash was combined with the filtrate, and the combined liquids were concentrated to provide (2S,3S)-3-(2-(tert-butoxy)-2-oxoethoxy)-2- (((2S,3R)-3-hydroxy-1-isopropoxy-1-oxobutan-2-yl)carbamoyl)pyrrolidine. See USSN 63 / 668,869, filed 07 / 09 / 2024; incorporated herein in its entirety.25970 Example 1 – Compound 11f bishydrochloride salt (19.86 kg) were dissolved in 930 L water at 25 °C. Sodium Hydroxide (10 N solution in water) was then added to adjust pH to 7.5. Dipeptide 2c oxalate salt (38.20 kg) was then added and dissolved. Sodium hydroxide (10 N solution in water) was again added to adjust pH back to 7.5. Sodium hexametaphosphate (42.22 kg), adenosine 5'-monophosphate monohydrate (2.52 kg), tergitol 15-s-9TM(27.6 kg), compound 1d hydrobromide salt (29.84 kg) and magnesium chloride hexahydrate (18.70 kg) were sequentially added. Sodium hydroxide (10 N solution in water) was then added to adjust pH to 8.0. PPK22 (0.20 kg), Trp-ligase SEQ ID NO.1 (0.30 kg) and Phe-ligase SEQ ID NO.2 (1.8 kg) were then sequentially added. During the course of the reaction, the pH was adjusted at ca.10 h (target pH 7.5) and ca.20 h (target pH 7.9) with sodium hydroxide (10 N solution in water). The resulting reaction mixture was agitated at 25 °C for 48 h to give tetrapeptide 3k as a slurry. Analysis confirmed residual tripeptide 3d ≤ 4.0 area% and residual compound 1d ≤ 0.1 area% by UPLC, the pH was adjusted to 8.0 with sodium hydroxide (10 N solution in water). Carboxyesterase SEQ ID NO.3 (0.20 kg) was added, and the resulting reaction mixture was agitated at 25 °C for 12 h, yielding Compound 1 as a slurry: MS [M+H]+C41H57FN7O8794.4; Found: 794.4.25970 Example 2 – Compound 1 Bisoxalate crystallization procedure40.3 kg) was charged to the vessel to adjust pH to 3.74. The reaction was then aged for 69 h at 20 °C before the pH was adjusted to 7.44 with NaOH (10N, 88.6 kg). Acid washed Celite 545 (29.00 kg) and 1-butanol (411.1 kg) were then charged to the vessel and stirred for 2 h. The resulting mixture was then filtered through a pressure filter and the filtrate was collected. The vessel was rinsed with additional 1-butanol (102.7 kg) which was then filtered through the waste cake and collected. The combined filtrates were then charged back to the vessel, stirred for 30 min and allowed to settle for 2 h. The aqueous layer was then cut to waste and potassium bicarbonate (15 % w / v in water, 723.2 kg) was added to the vessel. After stirring for 30 mins, the mixture was allowed to settle for 1 hour and the aqueous layer was cut to waste. Water (603.4 kg) was added to the vessel and the resulting mixture was stirred for 30 min and allowed to settle for 1 h. The aqueous layer was cut to waste and oxalic acid (2.40 kg) was added to the vessel and aged for 2 h to dissolve. The organic layer was drummed and sampled for assay yield (43.2 kg macrocyclic peptide Compound 1 as a freebase, 78.9% assay yield).

[0085] The resulting solution was then transferred through an inline 10 micron filter to a second vessel for crystallization. At 25 °C, a solution of oxalic acid was charged (126.9 kg of 3.69 wt% oxalic acid in 99.5:0.5 MTBE:water, v:v) to bring total oxalic acid equivalents in batch to 1.45 with respect to the compound 1 freebase. The resulting solution was seeded with macrocyclic peptide 1 as bis-oxalate salt (0.22 kg, 0.5 wt%) and aged for 1 h then warmed to 35 °C and aged 30 min further. Oxalic acid solution (13.4 kg of 3.69 wt% oxalic acid in 99.5:0.5 MTBE:water, v:v) was charged over 2 h at 35 °C, then cooled slowly to 20 °C over 4 h. After ageing at 20 °C for 1 hour, the slurry was heated to 40 °C, aged 30 mins, then cooled to 35 °C and aged for 1 h. Oxalic acid solution (20.1 kg of 3.69 wt% oxalic acid in 99.5:0.5 MTBE:water, v:v) was charged over 34 h at 35 °C, then cooled to 20 °C over 3 h. After ageing at 20 °C for 30 min, the slurry was heated to 40 °C and aged 30 mins. The slurry was then returned to 35 °C and aged for25970 1 h. Oxalic acid solution (73.6 kg of 3.69 wt% oxalic acid in 99.5:0.5 MTBE:water, v:v) was charged over 8 h at 35 °C, then cooled to 10 °C over 6 h. After ageing for 3 days at 10 °C, the slurry was then warmed to 15 °C and transferred to an agitated filter dryer and filtered. A displacement wash of 1.75:1 v:v 1-butanol:MTBE (66.0 kg) was charged and the cake was deliquored. A second displacement wash of MTBE (93.3 kg) was charged and the cake was again deliquored. A slurry wash of wet MTBE (186.6 kg of 99:1 MTBE:water v:v) was charged and the batch was agitated for 2 hours followed by deliquoring.

[0086] The cake was then dried under a flow of nitrogen. A slurry wash of wet MTBE (186.6 kg of 99:1 MTBE:water v:v) was charged and the batch was agitated for 2 h followed by deliquoring. A final displacement wash of the cake was then executed with wet MTBE (93.3 kg of 99:1 MTBE:water v:v). After deliquoring, the cake was dried under a flow of nitrogen with intermittent agitation at 25 °C. The desired macrocyclic peptide Compound 1 bisoxalate salt was isolated as a solid (52.2 kg).

[0087] 1H NMR (600 MHz, 6:4 acetonitrile-d3:D2O) δ 7.87 (1H, d, J = 7.9 Hz), 7.55 (1H, dd, J = 8.3, 4.3 Hz), 7.39-7.34 (3H, m), 7.25-7.23 (2H, m), 7.16 (1H, s), 6.98 (1H, td, J = 9.3, 2.5 Hz), 6.88 (1H, s), 5.00 (1H, hept, J = 6.2 Hz), 4.91 (1H, dd, J = 7.6, 5.6 Hz), 4.72 (1H, dd, J = 14.7, 8.3 Hz), 4.58 (1H, s), 4.28-4.27 (2H, m), 4.23 (1H, dd, J = 9.5, 3.7 Hz), 4.18 (1H, d, J = 15.8 Hz), 4.17 (1H, dd, J = 14.7, 4.0 Hz), 4.17 (1H, s), 4.10 (1H, d, J = 16.1 Hz), 4.07 (2H, t, J = 7.4 Hz), 3.97 (1H, dd, J = 14.7, 4.0 Hz), 3.71 (1H, q, J = 9.4 Hz), 3.27 (1H, dd, J = 13.8, 3.6 Hz), 3.10-3.03 (4H, m), 2.88-2.85 (2H, m), 1.96 (2H, m), 1.74 (2H, pent, J = 7.4 Hz), 1.55 (2H, pent, J = 7.6 Hz), 1.33-1.31 (2H, m), 1.26-1.24 (2H, m), 1.25 (3H, d, J = 6.2 Hz), 1.23 (3H, d, J = 6.2 Hz), 1.16 (3H, d, J = 6.2 Hz).13C NMR (150 MHz, 6:4 acetonitrile-d3:D2O) δ 171.80, 171.79, 171.60, 170.18, 168.03, 166.41 (4 oxalate carbons), 158.40 (d, JC-F = 232.2 Hz), 139.68, 134.72, 133.73, 130.70, 130.42, 129.66, 129.38, 128.97 (d, JC-F = 10.0 Hz), 128.79, 111.78 (d, JC-F = 10.0 Hz), 110.40 (d, JC-F= 26.5 Hz), 108.72 (d, JC-F= 4.4 Hz), 104.21 (d, JC-F= 23.2 Hz), 81.62, 70.94, 68.16, 67.82, 65.19, 59.51, 54.42, 52.01, 46.79, 45.79, 42.74, 40.23, 37.36, 31.33, 30.59, 29.13, 27.60, 26.79, 26.43, 21.93, 21.86, 20.28. MS [M+H]+C41H57FN7O8794.4; Found: 794.4. See Attorney Docket 25971, filed contemporaneously with the instant application and incorporated herein in its entirety.25970 Example 31f bishydrochloride salt (19.86 kg) was dissolved in 920 L water at 25 °C. Sodium hydroxide (10 N solution in water) was then added to adjust pH to 7.6. Dipeptide 2c oxalate salt (38.16 kg) was then added and dissolved. Sodium hydroxide (10 N solution in water) was again added to adjust pH back to 7.4. Sodium hexametaphosphate (42.24 kg), Adenosine 5'-monophosphate monohydrate (2.52 kg), Tergitol 15-s-9TM(27.6 kg), Compound 1d hydrobromide salt (29.9 kg) and magnesium chloride hexahydrate (18.72 kg) were sequentially added. Sodium hydroxide (10 N solution in water) was then added to adjust pH to 8.0. Polyphosphate kinase PPK22 (0.2 kg), ligase Trp-Ligase SEQ ID NO.1 (0.3 kg) and Phe-ligase SEQ ID NO.2 (1.84 kg) were then sequentially added. During the course of the reaction, the pH was adjusted at10 h (target pH 7.5) and 20 h (target pH 8.0) with sodium hydroxide (10 N solution in water). The resulting reaction mixture was agitated at 25 °C for 48 h to give tetrapeptide 3k as a slurry. The pH was adjusted to 7.8 with sodium hydroxide (10 N solution in water). Carboxylesterase SEQ ID NO.3 (0.2 kg) was added, and the resulting reaction mixture was agitated at 25 °C for 8 h, yielding the macrocylic peptide compound 1 as a slurry. MS [M+H]+C41H57FN7O8794.4; Found: 794.4.25970 Example 4was Phosphoric acid (85%, 40.3 kg) was charged to the vessel to adjust pH to 3.6. The reaction was then aged for 4 h at 20 °C before the pH was adjusted to 7.44 with NaOH (10N, 88.6 kg). Acid washed Celite 545 (29.0 kg) and 1-butanol (11 kg) were then charged to the vessel and stirred for 2 h. The resulting mixture was then filtered through a pressure filter and the filtrate was collected. The vessel was rinsed with additional 1-butanol (102.7 kg) which was then filtered through the waste cake and collected. The combined filtrates were then charged back to the vessel, stirred for 0.5 h and allowed to settle for 2 h. The aqueous layer was then cut to waste and potassium bicarbonate (15 % w / v in water, 723.3 kg) was added to the vessel. After stirring for 30 mins, the mixture was allowed to settle for 1 h and the aqueous layer was cut to waste. Water (603.4 kg) was added to the vessel and the resulting mixture was stirred for 30 mins and allowed to settle for 2 h. The aqueous layer was cut to waste and oxalic acid (2.42 kg) was added to the vessel and aged for 2 h to dissolve. The organic layer was drummed off.

[0090] The resulting solution was then transferred through an inline 10 micron filter to a second vessel for crystallization. At 25 °C, a solution of oxalic acid was charged (124.4 kg of 3.69 wt% oxalic acid in 99.5:0.5 MTBE:water, v:v). The resulting solution was seeded with macrocyclic peptide 1 as bis-oxalate salt (0.22 kg, 0.5 wt%) and aged for 30 min then warmed to 35 °C and aged 30 min further. Oxalic acid solution (19.8 kg of 3.69 wt% oxalic acid in 99.5:0.5 MTBE:water, v:v) was charged over 3 h at 35 °C, then cooled slowly to 20 °C over 4 h. After ageing at 20 °C for 1 h, the slurry was heated to 40 °C, aged 30 min, then cooled to 35 °C and aged for 1 h. Oxalic acid solution (20.1 kg of 3.69 wt% oxalic acid in 99.5:0.5 MTBE:water, v:v) was charged over 3 h at 35 °C, then cooled to 20 °C over 3 h. After ageing at 20 °C for 30 min, the slurry was heated to 40 °C and aged 30 min. The slurry was then returned to 35 °C and aged for 1 h. Oxalic acid solution (72.6 kg of 3.69 wt% oxalic acid in 99.5:0.5 MTBE:water, v:v) was charged over 8 h at 35 °C, aged at 35 °C for 1 h, then cooled to 10 °C over 6 h. Afterageing for 9 h at 10 °C, the slurry was then warmed to 15 °C and transferred to an agitated filter dryer and filtered. A displacement wash of 1-butanol (100.8 kg) was charged and the cake was deliquored. A second displacement wash of MTBE (92.1 kg) was charged and the cake was again deliquored. A slurry wash of wet MTBE (184.2 kg of 99:1 MTBE:water v:v) was charged and the batch was agitated for 2 h followed by deliquoring. A slurry wash of wet MTBE (184.2 kg of 99:1 MTBE:water v:v) was charged and the batch was agitated for 2 h followed by deliquoring. A displacement wash of wet MTBE (92.1 kg of 99:1 MTBE:water v:v) was charged and the batch was deliquored.

[0091] The cake was then dried under a flow of nitrogen. A slurry wash of wet MTBE (184.2 kg of 99:1 MTBE:water v:v) was charged and the batch was agitated for 2 h followed by deliquoring. A final displacement wash of the cake was then executed with wet MTBE (92.1 kg of 99:1 MTBE:water v:v). After deliquoring, the cake was dried under a flow of nitrogen with intermittent agitation at 25 °C to provide the desired macrocyclic peptide product 1 bisoxalate salt (48.8 kg).

[0092] The disclosed subject matter is not to be limited in scope by the specific embodiments and examples described herein. Indeed, various modifications of the disclosure in addition to those described will become apparent to those skilled in the art from the foregoing description and accompanying figures. Such modifications are intended to fall within the scope of the appended claims.

[0093] All references (e.g., publications or patents or patent applications) cited herein are incorporated herein by reference in their entirety and for all purposes to the same extent as if each individual reference (e.g., publication or patent or patent application) was specifically and individually indicated to be incorporated by reference in its entirety for all purposes. Other embodiments are within the following claims. Table 3 Sequences SEQ ID Sequence NO: T I G M E KI G H GSEQ ID Sequence NO: 3 MHHHHHHLSTARKPSSYELPVADPATVGMSRDKLQLVGDKVQSLIRENRIAGASVMVTRKGKIVYSE F T H T N

Claims

25970 WHAT IS CLAIMED IS:

1. Crystalline Compound 1: of Form 2 that is pattern havingdiffractions at angles (2-theta and 13.

8.

2. The crystalline Form 2 according to Claim 1, further characterized by an X-ray powder diffraction (XRPD) pattern having diffractions at angles (2-theta ±0.2) of at least 4.4, 5.8, 7.6, 8.8, 9.6, 10.1, 11.6, 13.8, 15.3, 15.9, 16.3, 16.7, 17.0, 17.6, 18.1, 19.6, 20.3, 20.7, 21.4, 22.4, 23.0, 23.6, 24.0, 24.9, 25.2, and 25.

7.

3. The crystalline Form 2 according to Claim 1, further characterized by an X-ray powder diffraction (XRPD) pattern having diffractions at angles (2-theta ±0.2) of at least 4.4, 5.8, 7.6, 8.8, 9.6, 10.1, 11.6, 13.8, 15.3, 15.9, 16.3, 16.7, 17.0, 17.6, 18.1, 19.2, 19.6, 20.3, 20.7, 21.4, 22.1, 22.4, 23.0, 23.6, 24.0, 24.9, 25.2, 25.7, 26.6, 28.0, 28.3, 29.4, and 29.

9.

4. The crystalline Form 2 according to Claim 1, further characterized by an X-ray powder diffraction (XRPD) pattern having diffractions at angles (2-theta ±0.2) of at least 4.4, 5.8, 7.6, 8.8, 9.6, 10.1, 11.6, 13.8, 15.3, 15.9, 16.3, 16.7, 17.0, 17.6, 18.1, 19.2, 19.6, 20.3, 20.7, 21.4, 22.1, 22.4, 23.0, 23.6, 24.0, 24.9, 25.2, 25.7, 26.6, 28.0, 28.3, 29.4, 29.9, 31.2, 32.0, 33.2, 34.5, 37.3, and 38.

3.

5. The crystalline form according to any of claims 1 to 4 as an n-BuOH solvate.

6. Crystalline Compound 1:25970 of Form 1 that is pattern havingdiffractions at angles (2-theta and 9.

5.

7. The crystalline Form 1 according to Claim 6, further characterized by an X-ray powder diffraction (XRPD) pattern having diffractions at angles (2-theta ±0.2) of at least 4.4, 4.8, 6.0, 7.3, 7.7, 8.7, 9.5, 10.2, 11.2, 12.0, 12.5, 13.1, 14.0, 14.3, 14.8, 15.4, 15.9, 16.1, 16.2, 16.6, 17.1, 17.5, 18.3, 18.5, 19.0, 19.2, and 19.

7.

8. The crystalline Form 1 according to Claim 6, further characterized by an X-ray powder diffraction (XRPD) pattern having diffractions at angles (2-theta ±0.2) of at least 4.4, 4.8, 6.0, 7.3, 7.7, 8.7, 9.5, 10.2, 11.2, 12.0, 12.5, 13.1, 14.0, 14.3, 14.8, 15.4, 15.9, 16.1, 16.2, 16.6, 17.1, 17.5, 18.3, 18.5, 19.0, 19.2, 19.7, 20.0, 20.5, 20.7, 21.5, 21.8, 22.1, 22.4, 22.8, 23.2, 24.2, 24.6, 25.2, 26.3, 26.4, 27.1, 27.4, 27.6, 28.1, 28.7, 29.1, and 29.

6.

9. The crystalline Form 1 according to Claim 6, further characterized by an X-ray powder diffraction (XRPD) pattern having diffractions at angles (2-theta ±0.2) of at least 4.4, 4.8, 6.0, 7.3, 7.7, 8.7, 9.5, 10.2, 11.2, 12.0, 12.5, 13.1, 14.0, 14.3, 14.8, 15.4, 15.9, 16.1, 16.2, 16.6, 17.1, 17.5, 18.3, 18.5, 19.0, 19.2, 19.7, 20.0, 20.5, 20.7, 21.5, 21.8, 22.1, 22.4, 22.8, 23.2, 24.2, 24.6, 25.2, 26.3, 26.4, 27.1, 27.4, 27.6, 28.1, 28.7, 29.1, 29.6, 30.3, 30.9, 31.5, 32.3, 34.0, 34.7, and 38.

8.

10. The crystalline form according to any of claims 1 to 4 as a bisoxalate salt.

11. A process for making crystalline Compound 1, Form 2, comprising the steps of:25970 a) combining a solution containing Compound 1 as a free base having a temperature of about 15°C to about 35 °C, with oxalic acid solution comprising water, oxalic acid, and a miscible solvent to form a slurry; b) filtering the slurry to produce a wet cake while maintaining a temperature of about 10°C to about 20°C; c) washing the wet cake with (n-BuOH); d) filtering and isolating to obtain wet solids of Compound 1 (Form 2).

12. The process according to claim 11 wherein the amount of oxalic acid solution combined with the solution of Compound 1 free base in step a) results in from about 1.00 to about 2.5 equivalents (eq.) of oxalic acid relative to Compound 1 free base added to the mixture.

13. The process according to any one of claims 11 and 12, wherein the resultant slurry from step a) undergoes at least one annealing step wherein the slurry is heated to about 30°C to about 50°C, additional oxalic acid added and cooled to about 5°C to about 25 °C.

14. The process according to any one of claims 11 to 13 wherein the amount of additional oxalic acid solution results in from about 0.005 to about 0.75 equivalents (eq.) of oxalic acid relative to Compound 1 free base added to the mixture.

15. The process according to any one of claims 11 to 14 is realized wherein the additional oxalic acid solution is added over a period of about 1 to about 10 hours.

16. A process for making Compound 1, Form 1, comprising the steps of: i) combining a solution containing Compound 1 as a free base having a temperature of about 15°C to about 35 °C, with oxalic acid solution comprising water, oxalic acid, and a first miscible solvent to form a slurry; ii) adding Compound 1 ( Form 1) seeds, and additional oxalic acid solution; iii) filtering resultant slurry to produce a wet cake; iv) washing the wet cake with (n-BuOH) forming a slurry; v) filtering and washing the slurry with a second miscible solvent to produce wet solids of Compound 1 (Form 2) vi) drying the wet crystals of Compound 1 (Form 2) and isolating to produce a solid form of Compound 1 (Form 1).25970 17. The process according to claim 16 wherein the amount of oxalic acid solution in step i) results in from about 1.00 to about 2.5 equivalents (eq.) of oxalic acid relative to Compound 1, free base.

18. The process according to claims 16 and 17 wherein the amount of seeds in step ii) added is from about 0.3 to about 1.0 wt%.

19. The process according to claims 16 to 18 when the amount of oxalic acid solution added in step ii) results in from about 0.005 to about 1.5 equivalents (eq.) of oxalic acid relative to Compound 1, free base.

20. The process according to claim 16 to 19 wherein the oxalic acid solution is added over a period of about 0.5 to about 12 hours.

21. The process according to any one of claims 16 to 20 wherein slurry in step i) is cooled to about 10°C to about 30 °C, after addition of the oxalic acid solution.

22. The process according to any one of claims 16 to 21 wherein the slurry from step ii) undergoes at least one annealing step wherein the slurry is heated to about 30°C to about 50°C, additional oxalic acid solution is added, and mixture is cooled to about 5°C to about 30 °C.

23. The process according to claim 22 wherein the amount of additional oxalic acid solution results in from about 0.10 to about 0.20 equivalents (eq.) of oxalic acid relative to Compound 1, free base.

24. The process according to claims 22 and 23 wherein the additional oxalic acid solution is added over a period of about 0.5 to about 10 hours.

25. The process according to any one of claims 16 to 24 wherein the temperature in step iii) is maintained at about 10°C to about 20°C.25970 26. The process according to any one of claims 16 to 25 wherein crystal solid Compound 1 (Form 1) is obtained by drying the washed Compound 1 solvate (Form 2) of step iv) with nitrogen and vacuum for at least 10 hours at about 20 to about 30 °C.

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