In 1554, Cosimo I de' Medici, with the support of Emperor Charles V, launched a grand campaign to conquer Florence's last remaining rival in Tuscany, the Republic of Siena.

The Florentine-Imperial troops were divided into three corps: Federico Barbolani di Montauto, with 800 men, landed in southern Tuscany to conquer the area of Grosseto; Rodolfo Baglioni, with 3,000 men, invaded the Val di Chiana to conquer Chiusi, Pienza, and Montalcino; and the main corps under Medeghino himself, consisting of 4,500 infantry, 20 cannons, and 1,200 sappers, was deployed at Poggibonsi for the main attack against Siena.

After an initial failed assault, the Marquess of Marignano laid siege to the city, although his men were not numerous enough to totally cut it off from the countryside.

On June 21, Strozzi conquered Montecatini, but did not feel confident enough to join in a pitched battle against Medeghino, waiting instead for further French reinforcements which were to arrive at Viareggio.

On July 17, conscious that only a victory in a pitched battle could save the city, he tried a third sally in the Val di Chiana, in the direction of Arezzo, leaving 1,000 infantry and 200 cavalry as a garrison under Blaise de Montluc.

The battle began with the attack of the Medeghino's cavalry wing, whose impetus easily routed its French-Sienese counterpart, which fled towards Foiano.

A chaotic melee ensued but soon the momentum of the Sienese attack began to wane under the fire of the Imperial artillery which also disorganized the Swiss attempting to relieve the first line.

Il Medeghino easily subdued the neighbouring castles in the days following the battle, and was subsequently able to lay a tighter siege to Siena.

Unable to receive substantial supplies and reinforcements from the French, the city surrendered on April 17, 1555 while the remaining Sienese forces withdrew to Montalcino.

The Duke ordered that a large fortress be constructed as a precaution against the risk of further rebellion by the Sienese, and this has presided over the city, from its north (Florentine) side, since its completion in 1563.