Japan's Nikkei 225 fell 0.8%. Hong Kong's Hang Seng Index dropped 0.3%, while China's Shanghai Composite Index declined 0.1%.
Markets in South Korea were closed for a public holiday.
Asian stocks rose earlier in the week ahead of scheduled trade talks between top US and Chinese negotiators in Washington on Thursday and Friday. But renewed strains between the two sides have overshadowed those discussions.
On Tuesday, US Secretary of State Mike Pompeo announced visa restrictions on Chinese officials suspected of being involved in the detention and human rights abuses of millions of Uyghur Muslims and other minority groups in China's Xinjiang region.
That came a day after the US State Department blacklisted 28 Chinese entities over similar allegations.
Those moves have angered Beijing.
"We urge the US to correct its mistakes at once and stop its interference in China's internal affairs," the Chinese embassy in the United States said on Twitter.
Some of the Chinese tech companies on the US blacklist rebounded Wednesday. Meiya Pico, a digital forensics firm, rallied 4.6% in Shenzhen after a 1.6% loss Tuesday.
IFlytek, a voice recognition software developer, was up about 1.5% in Shenzhen after falling 2.7% Tuesday.
Both companies have said that the blacklist will not have a major effect on their operations.
Surveillance camera makers Hikvision and Dahua Technology, meanwhile, have suspended trading of their shares since Tuesday.
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