The crash occurred at the intersection of Via Ca' Raffaelli and Via Tavoleto in Misano Adriatico, near the Misano World Circuit Marco Simoncelli where Hayden had raced many times. Hayden was traveling west on Via Ca' Raffaelli at approximately 14:00 local time when he was struck by a Peugeot 206 CC as he entered the intersection to cross Via Tavoleto. A home surveillance camera installed near the intersection recorded the entire accident.
According to investigation findings released in September 2017, the car was traveling at approximately 73 kilometres per hour in a 50 kilometres per hour zone, and there was no indication that the driver braked before the collision. Hayden had been traveling at 20 kilometres per hour and entered the intersection without stopping at a stop sign. The investigation apportioned 70 percent of the blame to the driver and 30 percent to Hayden.
The driver stated he was traveling to work when Hayden suddenly appeared in front of him. The impact was severe enough to completely shatter the windshield and dent the roof of the car. Hayden's bicycle was found in a nearby ditch with its frame snapped in half.
Earlier that morning, Hayden had been running with fellow former MotoGP champion Kevin Schwantz, who had been competing in the same World Superbike round at Misano that Hayden was participating in. Schwantz declined Hayden's invitation to join him for the afternoon bicycle ride, as he did not have a bicycle with him at the time.
Hayden was taken first to Rimini hospital and subsequently transferred to the major trauma unit at the Maurizio Bufalini Hospital in Cesena. The extent of his injuries was described as polytrauma: a traumatic brain injury resulting in severe cerebral damage, a broken femur, broken pelvis, and multiple fractured vertebrae. He was placed on life support in an intensive care unit. No surgery was performed due to the severity of the cerebral damage. Hayden died on 22 May 2017, five days after the accident, without any signs of recovery.
At the time of the accident, Hayden was competing in his second season in the Superbike World Championship with the Red Bull Honda World Superbike Team, having moved to Superbike racing in 2016 after thirteen seasons in MotoGP. He had been in 13th place in the championship standings. His final race had been the second race of the Italian Round at Misano on 14 May 2017, three days before the accident, where he finished 12th.
Hayden had ended his MotoGP career in 2015 after a stint with the Aspar Team, following five largely difficult seasons with Ducati from 2009 to 2013 and one season with the Aspar Honda outfit in 2014. He had won three MotoGP races in his career and the 2006 World Championship, achieved by defeating Valentino Rossi's five-year consecutive title streak by just five points in a dramatically contested final round at Valencia.
The motorcycle racing world responded to Hayden's death with an outpouring of tributes. His family had deep roots in American motorcycle racing: his brothers Tommy and Roger Lee Hayden were also professional racers, and his father Earl had used the same number 69 before Nicky. The number 69 was chosen, Earl Hayden joked, because it was still readable when he frequently ended up upside down in the dirt.
In 2017, the AMA Horizon Award was renamed the Nicky Hayden AMA Horizon Award in his honor. On 8 June 2018, the City of Owensboro, Kentucky unveiled a bronze memorial sculpture commissioned by the city and the Hayden family, created by sculptor George Lundeen. The mayor declared 9 June as Nicky Hayden Day in the city. During the 2018 Red Bull Grand Prix of the Americas at Circuit of the Americas, turn 18 was officially renamed Hayden Hill. Hayden was inducted into the AMA Motorcycle Hall of Fame in 2018, with the traditional five-year retirement rule waived in recognition of his contribution to the sport, and into the Motorsports Hall of Fame of America in 2021.