Morphic forms of hexadecyloxypropyl-phosphonate esters and methods of synthesis thereof

1. A method of synthesizing morphic Form II of Compound 1:

comprising:

(i) contacting a mixture of (S)—N 1 -[(2-hydroxy-3-triphenylmethoxy)propyl]cytosine (Compound 2) with P-[[[4-methylphenyl)sulfonyl]oxy]methyl]-, mono[3-(hexadecyloxy)propyl]ester, sodium salt (Compound 4) in the presence of a suitable base and in a suitable organic solvent, at a suitable reaction temperature;

(ii) extracting the crude product with a suitable organic solvent and concentrating the mixture to give crude phosphonic acid, [[(S)-2-(4-amino-2-oxo-1(2H)-pyrimidinyl)-1-(hydroxymethyl)-2-(triphenylmethoxy)ethyl]methyl]mono[3-(hexadecyl oxy)propyl] ester (Compound 3);

(iii) diluting crude Compound 3 in methanol and re-concentrating the resulting solution to remove the residual suitable organic solvent of step (ii);

(iv) contacting crude Compound 3 with a suitable deprotecting agent in an organic solvent to give crude Compound 1; and

(v) recrystallizing crude Compound 1 using a suitable solvent system.

2. The method of claim 1 , wherein the suitable base of step (i) is a metal alkoxide selected from the group consisting of magnesium di-tert-butoxide, sodium tert-butoxide, lithium tert-butoxide, sodium tert-amyl alkoxide, potassium tert-butoxide, and sodium methoxide.

3. The method of claim 1 , wherein the suitable base of step (i) is a metal hydride selected from the group consisting of sodium hydride and potassium hydride.

4. The method of claim 1 , wherein the suitable base of step (i) is lithium bis(trimethylsilyl)amide.

5. The method of claim 1 , wherein the suitable organic solvent of step (i) is selected from the group consisting of N,N-dimethylformamide, N, N-dimethylacetamide, dimethylsulfoxide, and 1-methyl-2-pyrrolidinone.

6. The method of claim 1 , wherein the suitable reaction temperature of step (i) is between about 50° C. to about 110° C.

7. The method of claim 1 , wherein step (i) proceeds within about 2-4 hours.

8. The method of claim 1 , wherein the suitable organic solvent of step (ii) is selected from the group consisting of ethyl acetate, isopropyl acetate, and dichloromethane.

9. The method of claim 1 , wherein the suitable deprotecting agent of step (iv) is hydrogen chloride or acetyl chloride.

10. The method of claim 1 , wherein the organic solvent of step (iv) is methanol.

11. The method of claim 1 , wherein the suitable solvent system of step (v) is selected from the group consisting of methanol/acetone/water, ethanol, and methanol.

12. The method of claim 1 , wherein the suitable base of step (i) is magnesium tert-butoxide, the suitable solvent of step (i) is N,N-dimethylformamide, and the suitable reaction temperature of step (i) is about 75-85° C.

13. The method of claim 1 , wherein the suitable organic solvent of step (ii) is isopropyl acetate.

14. The method of claim 1 , wherein the suitable deprotecting agent of step (iv) is gaseous hydrogen chloride.

15. The method of claim 1 , wherein the crude Compound 1 is recrystallized three times from methanol, once from n-heptane and methanol, and again from methanol.

16. A method of synthesizing morphic Form II of Compound 1:

comprising:

a. heating a mixture of (S)-N1-[(2-hydroxy-3-triphenylmethoxy)propyl]cytosine (Compound 2), P-[[[4-methylphenyl)sulfonyl]oxy]methyl]-, mono[3-(hexadecyloxy)propyl]ester, sodium salt (Compound 4), magnesium tert-butoxide, and dimethylformamide;

b. cooling and adding isopropyl acetate;

c. washing sequentially with HCl solution and NaCl solution;

d. diluting the concentrate with methanol and forming a mixture containing phosphonic acid, [[(S)-2-(4-amino-2-oxo-1(2H)-pyrimidinyl)-1-(hydroxymethyl)-2-(triphenylmethoxy)ethyl]methyl]mono[3-(hexadecyloxy)propyl]ester (Compound 3);

e. diluting the concentrate containing Compound 3 in methanol;

f. adding HCl gas and maintaining temperature below about 20° C.;

g. filtering to remove impurities and preparing slurry in acetone;

h. filtering the slurry and washing with acetone;

i. recrystallizing from methanol;

j. recrystallizing a second time from methanol;

k. recrystallizing a third time from methanol;

l. recrystallizing from n-heptane and methanol;

m. dissolving crude product in methanol;

n. adding to the solution a seed stock of phosphonic acid, [[(S)-2-(4-amino-2-oxo-1(2H)-pyrimidinyl)-1-(hydroxymethyl) ethoxy]methyl]mono[3-(hexadecyloxy)propyl] ester (Compound 1) and stirring; and

o. cooling, filtering, and washing the solution with methanol and drying;

thereby synthesizing morphic Form II Compound 1.

17. A method of synthesizing morphic Form II of Compound 1:

comprising:

a. heating a mixture of (S)—N 1 -[(2-hydroxy-3-triphenylmethoxy)propyl]cytosine (Compound 2), P-[[[4-methylphenyl)sulfonyl]oxy]methyl]-, mono[3-(hexadecyloxy)propyl]ester, sodium salt (Compound 4), magnesium tert-butoxide, and dimethylformamide at between about 75-85° C. for about 3 hours;

b. cooling the mixture to about 25-35° C. and adding isopropyl acetate;

c further cooling the solution to about 15-25° C. and washing sequentially with HCl solution and NaCl solution;

d. removing isopropyl acetate by vacuum distilling the organic phase thereby forming a concentrate;

e. diluting the concentrate with methanol and further removing isopropyl acetate thereby re-concentrating and forming a mixture containing phosphonic acid, [[(S)-2-(4-amino-2-oxo-1(2H)-pyrimidinyl)-1-(hydroxymethyl)-2-(triphenylmethoxy)ethyl]methyl]mono[3-(hexadecyloxy)propyl]ester (Compound 3);

f. diluting the concentrate containing Compound 3 in methanol;

g. adding HCl gas at a rate that maintains the temperature between about 5 and 15° C.;

h. maintaining the reaction about 10-20° C. for 2 hours before filtering to remove solid impurities;

i. diluting the filtrate with water and adjusting pH to about 2.3-2.7 with NaOH;

j. filtering the solids and washing the solids with water before preparing a slurry in acetone at about 35-45° C. for about 1 hour;

k. filtering the slurry and washing with acetone;

l. drying the acetone washed crude product at a temperature of equal to or less than about 40° C. for about 12 hours;

m. heating the crude product at about 60-70° C. in methanol

n. polish filtering, cooling to about 58-62° C., stirring for one hour, cooling to about 48-52° C. for about six hours, then to about 17-23° C. for two hours, filtering, and then washing with methanol;

o. repeating steps l-m twice or more;

p. heating the product in methanol at about 64° C. and slowly adding n-heptane over 40 min while keeping the temperature above 50° C.;

q. holding at a temperature about 55° C. for 30 min and cooling to 40° C. over a 6 h period;

r. stirring at 40° C. for 2 h, then cooling to 20° C. over six hours;

s. stirring for 2 h at 20° C.

t. filtering and washing with n-heptane and methanol and drying under vacuum;

u. dissolving solids in methanol before cooling to about 59-61° C. and then stirring for about 20 minutes;

v. adding to the solution a seed stock of phosphonic acid, [[(S)-2-(4-amino-2-oxo-1(2H)-pyrimidinyl)-1-(hydroxymethyl) ethoxy]methyl]mono[3-(hexadecyloxy)propyl] ester (Compound 1) and stirring for two hours;

w. cooling the solution to about 47-53° C. by stirring for about eight hours, and then stirring for about two hours;

x. cooling the stirred solution further to about 17-23° C. over about six hours, and further stirring for about two hours;

y. filtering and washing the solution with methanol; and

z. drying at a temperature of equal to or less than about 40° C. for about twenty-four hours, thereby synthesizing morphic Form II Compound 1.