According to the country's constitution, the more senior of two vice presidents will stand in as president until a new leader is elected by parliament within seven working days, Reuters reported.
This means that Myint Swe, who was the military's appointment for vice-president, will become acting president.
The president is the head of state and government in Myanmar, and under the constitution has far-reaching powers. However, Htin Kyaw's role was more ceremonial because Aung San Suu Kyi has been Myanmar's de facto leader since April 2016.
A constitution drafted by the former junta bars Nobel Peace Prize laureate Suu Kyi from the top office and so she hand-picked Htin Kyaw, a close ally of hers, to become president.
Aung Shin, a spokesman for Suu Kyi's National League for Democracy (NLD) party, said that Htin Kyaw, 71, had stepped down due to ill-health.
The president's office did not give a reason for Htin Kyaw's resignation. In its posting, the president's office said that, according to the constitution, steps would be taken to replace him within seven working days.
"The next president will be an NLD member or one who suits with NLD policy," Aung Shin said. "The current vice-president cannot be the next president, according to the constitution."