Prosecutors say Sayfullo Saipov was behind the wheel of a rental truck when he plowed into people along the West Side Highway.
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Last week a grand jury returned a 22-count indictment against Saipov.
Saipov, 29, an Uzbek who had been living in New Jersey, appeared in grey and shackled as he entered his plea. He indicated to the judge he understood English, but relied on an interpreter during his arraignment.
Federal prosecutors said Saipov drove a truck he rented two hours earlier into pedestrians and cyclists, killing mainly foreign tourists. Court records said he wanted to fly the ISIS flag from the truck but thought better of it so he wouldn't attract attention. Federal prosecutors called the attack a "calculated act of terrorism" inspired by the numerous ISIS videos Saipov watched on his mobile phone.
Saipov is charged with murder in the aid of racketeering, attempted murder in aid of racketeering, providing and attempting to provide material support to a designated foreign terrorist organization and violence and destruction of motor vehicles.
His public defender had no comment at the conclusion of Tuesday's brief appearance.
Saipov's next court appearance is January 23.
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He could face the death penalty if convicted.
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