The 24 year-old Millwall striker capped an energetic performance with a well-placed header early in the second half but Ireland's much-improved defence was undone by Leeds United midfielder Mateusz Klich, who equalised three minutes from time.
Having conceded four against Wales, Ireland set its stall out to defend doggedly against a Poland side keen to make amends for a disappointing World Cup, and O'Neill's three-man back line worked hard to frustrate Napoli forward Arkadiusz Milik.
The signs were not good for Ireland when Milik was allowed a free-header from Marcin Kaminski's flick-on after 10 minutes and guided it just wide to the left of Darren Randolph's goal.
The lively O'Brien lifted a shot high and wide from close range before the half-hour mark and Milik thumped a drive into Randolph's midriff five minutes later but the Ireland defence held firm at the break.
Ireland came out on the front foot in the second half and within eight minutes of the restart O'Brien broke the deadlock, glancing the ball over Wojciech Szczesny with a header from Callum O'Dowda's superb in-swinging cross.
Poland boss Jerzy Brzeczek made five changes to try and turn the game in front of an impatient home crowd, yet what looked to be its best chance was wasted when Damian Szymanski headed wide.
Klich broke Ireland hearts with just five minutes left on the clock, playing a neat one-two with Milik before beating Randolph down to his left and taking the sheen off O'Brien's night.