Cardinal Oswald Gracias is speaking after a judge in the state of Raipur found there wasn't enough evidence to convict Dinesh Dhurv, 19, and Jitendra Pathak, 25, of the attack in June 2015.
The nun was found tied and gagged at the missionary-run medical center shes works at in the city by colleagues.
At the time police said they did not think robbery was a motive for the attack, given that a significant amount of money was found after the attack.
This has led to reports the attackers targeted the nun because of her Christian faith. Advocacy charities have said attacks against believers by Hindu nationalist extremists have risen in India in recent years.
The archbishop also says Indian police failed to collect evidence properly, allowing the nun's attackers to escape justice.
According to Vatican Radio, Cardinal Oswald said: "This acquittal once again brings to our attention the problem of violence against women.
"It is a huge setback for all of us working for the rights and dignity of women, in particular victims of violence.
He also accused Indian police of having a "halfhearted attitude" to collecting evidence in the nun's rape case.
In March 2015, another nun in West Bengal was also raped by robbers.