Three-dimensional printed calcium phosphate bone cement composite scaffolds for bone regeneration, precursor compositions, and methods of printing

Abstract

Disclosed are 3D-printed scaffolds having high bone cement content, and in particular, high hydroxyapatite (HA) content. The disclosed methods and compositions provide the ability to print biocompatible scaffolds having patient-specific geometries with controlled porosity, microstructure, osteoconductivity, and mechanical strength. The scaffolds may be used for in vitro and in vivo craniofacial and dental applications.

What is claimed is: 
     
         1 . A three-dimensional, biocompatible precursor composition for room-temperature printing a three-dimensional (3D) bio-compatible polymer/bone cement composite scaffold, the precursor composition comprising:
 a room-temperature slurry comprising a mixture of:
 a solid phase comprising at least one calcium-containing compound selected from a first group consisting of tetracalcium phosphate (TTCP), dicalcium phosphate anhydrous (DCPA), α-tricalcium phosphate (α-TCP), β-tricalcium phosphate (β-TCP), amorphous calcium phosphate (ACP), octacalcium phosphate (OCP), dicalcium phosphate dihydrate (DCPD), monocalcium phosphate monohydrate (MCPM), monocalcium phosphate anhydrous (MCPA), calcium sulfate (CaSO 4 ), calcium sulfate hemihydrate (α- or β-CaSO 4 .0.5H 2 O), calcium carbonate (CaCO 3 ), calcium sulfate dihydrate (CaSO 4 .2H 2 O), calcium oxide (CaO), and calcium hydroxide (Ca(OH) 2 ), and 
 a liquid phase comprising a polymer in at least one non-aqueous solvent, wherein the polymer is insoluble or has low solubility in water, 
   
       wherein a weight ratio of the solid phase to the liquid phase is between 0.1 to 1 and 3 to 1. 
     
     
         2 . The precursor composition of  claim 1 , wherein the weight ratio of the solid phase to the liquid phase is between 0.75 to 1 and 3 to 1. 
     
     
         3 . The precursor composition of  claim 1 , wherein the at least one calcium-containing compound in the solid phase comprises at least one of tetracalcium phosphate (TTCP; Ca 4 (PO 4 ) 2 O) and dicalcuim phosphate anhydrous (DCPA; CaHPO 4 ). 
     
     
         4 . The precursor composition of  claim 3 , wherein the TTCP is provided with a particle size in a range from 1 to 17 μm and wherein the DCPA is provided with a particle size in a range from 1 to 5 μm. 
     
     
         5 . The precursor composition of  claim 3 , wherein the TTCP and DCPA are provided with a weight ratio TTCP:DCPA in a range of about 97%:3% to 8%:92%. 
     
     
         6 . The precursor composition of  claim 3 , wherein the solid phase compounds have a Ca/P molar ratio between 1 and 2. 
     
     
         7 . The precursor composition of  claim 1 , wherein the polymer is selected from a polymer group consisting of polyvinyl butyral (PVB), polycaprolactone (PCL), poly lactic-co-glycolic acid (PLGA), Poly(ethylene glycol), Polyvinyl pyrrolidone (PVP), Poly(methyl methacrylate) (PMMA), Polyoxazoline, polyphosphoesters (PPE), Poly-L-latic acid (PLLA), Polyacrylic acid (PAA), and Dextran. 
     
     
         8 . The precursor composition of  claim 1 , wherein the at least one non-aqueous solvent is selected from a solvent group consisting of ethanol (EtOH), tetrahydrofuran (THF), acetic acid, acetone, methanol, 2-propanol, butanol, 2-butoxyethanol, cyclohexanone, benzyl alcohol, 1-methoxy-propanol-2, butyl glycol, n-butyl, acetate, ethyl acetate, N,N-dimethylacetamide, N,N-dimethylformamide, N,N-dimethylsulfoxide, NMP, chloroform, dichloromethane, carbon tetrachloride, benzene, toluene, cyclohexanone and 2-nitropropane, acetone, 2-butanone, ethyl acetate, dimethylformamide, acetonitrile, dichloromethane, chloroform, and ethyl acetate. 
     
     
         9 . The precursor composition of  claim 1 , further comprising a hardening accelerator, wherein the hardening accelerator is selected from a second group consisting of sodium phosphate dibasic, monosodium phosphate, trisodium phosphate, ammonium phosphate, ammonium dihydrogen phosphate, monopotassium phosphate, dipotassium phosphate, tripotassium phosphate, sodium fluoride, potassium fluoride, sodium acetate, potassium oxalate, sodium sulfate, sodium cacodylate, and organic acid, and wherein the organic acid is selected from a third group consisting of glycolic acid, citric acid, tartaric acid, malonic acid, malic acid, and maleic acid. 
     
     
         10 . The precursor composition of  claim 9 , wherein the hardening accelerator makes up to 10% of the solid phase by weight. 
     
     
         11 . The precursor composition of  claim 9 , wherein the scaffold is printed submerged in an aqueous bath. 
     
     
         12 . The precursor composition of  claim 1 , wherein the scaffold is printed submerged in an aqueous bath and wherein a hardening accelerator is added to the aqueous bath. 
     
     
         13 . The precursor composition of  claim 1 , wherein the solid phase further comprises a compound selected from a fourth group consisting of a carbonate, chloride (Cl), an alkali metal fluoride, an alkaline earth metal fluoride, a silver-based fluoride, stannous fluoride, ammonium fluoride, a quaternary ammonium fluoride, a fluorosilicate, and a monofluorophosphate. 
     
     
         14 . The precursor composition of  claim 1 , wherein the slurry further comprises a bioactive material selected from a fifth group consisting of osteoblasts, osteoclasts, osteids, endothelial cells, endothelial progenitor cells, neutrophils, macrophages, and combinations thereof. 
     
     
         15 . A precursor composition that, when printed at room temperature, forms a 3D bio-compatible polymer/calcium phosphate cement composite scaffold, the precursor composition comprising:
 a room temperature slurry comprising:
 a solid phase comprising a mixture of one or more calcium phosphate compounds selected from a first group consisting of TTCP, α-TCP and β-TCP, ACP, OCP, DCPA, DCPD, MCPM, and MCPA, and 
 a liquid phase comprising a polymer in a solvent, the solvent being selected from a first solvent group consisting of ethanol (EtOH) and tetrahydrofuran (THF); and 
   a hardening accelerator, which is added during printing of the polymer/calcium phosphate cement composite scaffold.   
     
     
         16 . The precursor composition of  claim 15 , wherein the hardening accelerator is selected from a second group consisting of sodium phosphate dibasic (Na 2 HPO 4 ), ammonium phosphate ((NH 4 ) 3 PO 4 ), potassium phosphate dibasic (K 2 HPO 4 ), sodium fluoride (NaF), potassium fluoride (KF), sodium acetate, potassium oxalate (C 2 K 2 O 4 ), sodium sulfate (Na 2 SO 4 ), and sodium cacodylate. 
     
     
         17 . The precursor composition of  claim 15 , wherein the precursor composition is printed in an aqueous bath. 
     
     
         18 . The precursor composition of  claim 17 , wherein the hardening accelerator is added to the aqueous bath during scaffold printing. 
     
     
         19 . The precursor composition of  claim 15 ,
 wherein the polymer is selected from a first polymer group consisting of polyvinyl butyral (PVB) and polycaprolactone (PCL);   wherein the PVB is dissolved in a solvent selected from a second solvent group consisting of acetic acid, acetone, methanol, 2-propanol, butanol, 2-butoxyethanol, cyclohexanone, benzyl alcohol, 1-methoxy-propanol-2, butyl glycol, n-butyl, acetate, ethyl acetate, N,N-dimethylacetamide, N,N-dimethylformamide, N,N-dimethylsulfoxide, and NMP; and   wherein the PCL is dissolved in a solvent selected from a third solvent group consisting of chloroform, dichloromethane, carbon tetrachloride, benzene, toluene, cyclohexanone, 2-nitropropane, acetone, 2-butanone, ethyl acetate, dimethylformamide, and acetonitrile.   
     
     
         20 . The precursor composition of  claim 15 , wherein the solid phase compounds comprise TTCP and DCPA and have a Ca/P molar ratio in a range from 1.33 to 1.90. 
     
     
         21 . The precursor composition of  claim 15 , further comprising a compound selected from a fourth group consisting of a carbonate, chloride (Cl), an alkali metal fluoride, an alkaline earth metal fluoride, a silver-based fluoride, stannous fluoride, ammonium fluoride, a quaternary ammonium fluoride, a fluorosilicate, and a monofluorophosphate. 
     
     
         22 . A computer-controlled method for room temperature 3D printing a biocompatible, composition-controlled scaffold, the method comprising:
 preparing a solid phase composition comprising a calcium-containing cement powder;   preparing a liquid phase composition comprising a polymer material dissolved in a solvent;   homogeneously mixing the solid phase composition and the liquid phase composition to create a homogeneous, bio-compatible slurry;   disposing the slurry in a reservoir system coupled to a printing nozzle system, the printing nozzle system comprising at least one printing nozzle;   submerging a printing substrate in a liquid bath disposed below the printing nozzle;   under control of a computer, operating a motor to extrude the slurry, at room temperature, from the reservoir system through the printing nozzle system and to cause relative x, y, and z displacement between the printing nozzle system and the printing substrate;   employing a hardening accelerator to assist formation of the biocompatible, composition-controlled scaffold; and   maintaining the 3D printing scaffold fully submerged in the liquid bath during an entire 3D printing process.   
     
     
         23 . The computer-controlled method of  claim 22 , wherein the liquid bath is an aqueous solution containing the hardening accelerator, wherein the hardening accelerator is selected from a first group consisting of sodium phosphate dibasic (Na 2 HPO 4 ), ammonium phosphate ((NH 4 ) 3 PO 4 ), potassium phosphate dibasic (K 2 HPO 4 ), sodium fluoride (NaF), potassium fluoride (KF), sodium acetate, potassium oxalate (C 2 K 2 O 4 ), sodium sulfate (Na 2 SO 4 ), and sodium cacodylate. 
     
     
         24 . The computer-controlled method of  claim 22 , further comprising adding to the solid phase composition, a compound selected from a second group consisting of carbonate (NaHCO 3 ), chloride (Cl), and sodium fluoride (NaF). 
     
     
         25 . The computer-controlled method of  claim 22 ,
 wherein the polymer in the slurry is selected from a polymer group consisting of: PVB, Polycaprolactone (PCL), Poly Lactic-co-Glycolic acid (PLGA), Poly-L-lactic acid (PLLA), Poly(ethylene glycol), Polyvinyl pyrrolidone (PVP), Poly(methyl methacrylate) (PMMA), Polyoxazoline, polyphosphoesters (PPE), and Dextran;   wherein the PVB is dissolved in a solvent selected from a first solvent group consisting of acetic acid, acetone, methanol, ethanol, 2-propanol, butanol, 2-butoxyethanol, cyclohexanone, benzyl alcohol, 1-methoxy-propanol-2, butyl glycol, n-butyl, acetate, ethyl acetate, N,N-dimethylacetamide, N,N-dimethylformamide, N,N-dimethylsulfoxide, NMP, and THF;   wherein the PCL is dissolved in a in a solvent selected from a second solvent group consisting of THF, chloroform, dichloromethane, carbon tetrachloride, benzene, toluene, cyclohexanone and 2-nitropropane, acetone, 2-butanone, ethyl acetate, dimethylformamide, and acetonitrile;   wherein the PLGA is dissolved in a in a solvent selected from a third solvent group consisting of THF, acetone, ethyl acetate, and chlorinated solvents;   wherein the PLLA is dissolved in a solvent selected from a fourth solvent group consisting of chloroform and dichloromethane (DCM);   wherein the PLLA is dissolved in a solvent selected from a fifth solvent group consisting of acetone, dichloromethane, ethanol (95%), methanol, benzene, glycerin, and glycols;   wherein the PVP is dissolved in a solvent selected from a sixth solvent group consisting of methanol and ethanol;   wherein the PMMA is dissolved in a solvent selected from a seventh solvent group consisting of THF, methanol, and ethanol;   wherein the PMMA is dissolved in a solvent selected from an eighth solvent group consisting of toluene, dichloromethane, chloroform, and acetone;   wherein the PPE is dissolved in a solvent selected from a ninth solvent group consisting of THF, acetonitrile, chloroform, ethyl acetate, and Poly((lactide-co-ethylene glycol)-co-ethyloxyphosphate)); and   wherein the Dextran is dissolved in a solvent selected from a tenth solvent group consisting of methyl sulfide, formamide, ethylene glycol, and glycerol.   
     
     
         26 . The computer-controlled method of  claim 22 , further comprising disposing the hardening accelerator in the reservoir system, the liquid bath, and both the reservoir system and the liquid bath. 
     
     
         27 . The computer-controlled method of  claim 22 , further comprising disposing in the reservoir system, the liquid bath, and both the reservoir system and the liquid bath, one or more compounds chosen from a third group consisting of carbonate (NaHCO 3 ), sodium chloride (NaCl), an alkali metal fluoride, an alkaline earth metal fluoride, a silver-based fluoride, stannous fluoride, ammonium fluoride, a quaternary ammonium fluoride, a fluorosilicate, and a monofluorophosphate. 
     
     
         28 . The computer-controlled method of  claim 22 , further comprising incorporating cells during printing, the cells being selected from a group consisting of osteoblasts, osteoclasts, osteoids, endothelial cells, endothelial progenitor cells, neutrophils, macrophages, and combinations thereof. 
     
     
         29 . The computer-controlled method of  claim 22 , further comprising incorporating cells into the printed scaffold, the cells being selected from a group consisting of osteoblasts, osteoclasts, osteoids, endothelial cells, endothelial progenitor cells, neutrophils, macrophages, and combinations thereof.