During Sunday's loss to Washington, Dinwiddie crumpled to the court after taking an awkward step on a fast break and his left knee buckled.
He was on the court for a few minutes while trainers and coach Tad Boyle came to his side. Dinwiddie was helped to his feet but had to be assisted off the court with his arms around teammates.
"It's a big blow for him, " Colorado coach Tad Boyle said in a statement. "He's worked so hard to put himself in the position he has and help lead this team to where we are today. To have that all taken away from you in one basketball play is ... it's tough."
The 6-foot-6, 200-pound junior leads the 21st-ranked Buffaloes in scoring (14.7) and assists (3.8) per game.
The Buffaloes (14-3, 3-1 Pac-12) were considered a potential Sweet 16 team and might have even been the biggest threat to top-ranked Arizona in the Pac-12 with the versatile Dinwiddie in the fold. Without him, the Buffs could wind up being a team on the fringe of the NCAA tournament.
"We don't have the star system here," Boyle said. "Spencer was our leading scorer and leading assist guy, he led us in steals. There's no question he was important to our team. I'm not trying to minimize this loss, but I just want our players to realize they're here for a reason: they're capable as well. When one guy goes down, the door opens for one, or in this case, maybe two or three more."
Dinwiddie was considered a potential first-round pick in June's NBA draft before the injury, if the California native elected to leave school early.
ESPN's Jeff Goodman contributed to this report.