In 1943, the Axis powers decide on a show of strength to bully neutral Turkey into joining the war on their side. Their target is 2000 British soldiers who are marooned on the island of Keros in the Aegean. Rescue by the Royal Navy is impossible because of massive radar-directed guns on the nearby island of Navarone. Time is short, because the Germans are expected to launch an assault on the British forces.
Efforts to destroy the guns by bombing have proved fruitless. So that six destroyers can pick up the stranded men, Commodore Jensen of Allied intelligence activates a team of saboteurs to sail to Navarone and destroy the guns. Led by Major Roy Franklin (Anthony Quayle), they are Captain Keith Mallory (Gregory Peck); Andrea Stavrou (Anthony Quinn), a former Colonel in the defeated Greek army; Corporal Miller (David Niven), an explosives expert; Greek-American Spyros Pappadimos (James Darren); and "Butcher" Brown (Stanley Baker), an engineer and expert knife fighter.
Disguised as Greek fishermen on a decrepit boat, they sail across the Ageaen Sea. They are intercepted by a German boat and boarded. They attack and kill all the Germans and sink the patrol boat. During the remainder of the voyage, Mallory confides to Franklin that Andrea has sworn to kill him after the war, because he was inadvertently responsible for the deaths of Andrea's wife and children.
In a violent storm, the ship is wrecked and they lose part of their equipment, but manage to land on the island. Led by Mallory, who was recruited for his climbing skills, they scale the 'unclimbable' cliff. But Franklin is badly injured; the injury later becoming infected with gangrene. They find that the cliff is in fact guarded after all. Miller, a friend of Franklin, suggests that they leave Franklin to be "well cared for" by the enemy. Mallory, who now assumes command of the mission, feels that Franklin would be forced to reveal their plans, so he orders two men to carry the injured man on a stretcher.
Franklin tries to commit suicide, but Mallory lies to him, saying that their mission has been "scrubbed" and that a major naval attack will be mounted on Navarone. They rendezvous with local resistance workers, Spyros's sister Maria (Irene Papas) and her friend Anna (Gia Scala), who is supposedly mute after German torture.
The mission is continually dogged by Germans, and they are captured by Lieutenant Muesel (Walter Gotell) in the town of Mandrakos, when they try to find a doctor for Franklin. Muesel and Hauptmann Sessler (George Mikell) of the SS fail to persuade the commandos to tell them where Miller's explosives are. Andrea pretends to betray the others and surprises the Germans, allowing the group to overpower their captors. They escape in German uniform, but leave Franklin behind so that he can receive medical attention.
In due course, Franklin is injected with scolpolamine and gives up the false "information", as Mallory had hoped. German units are deployed away from the guns and in the direction of the supposed invasion point.
Miller discovers that most of his explosives have been sabotaged and deduces that Anna is the saboteur. It transpires that she is not mute after all and was not tortured by the Nazis; but she agreed to become an informer in exchange for her release. She pleads that she was coerced by the Germans into treachery, but Mallory decides she must be silenced. It is Maria who shoots her dead.
The team splits up: Mallory and Miller go for the guns, while Andrea and Pappadimos create a distraction in the city; Maria and Brown are assigned to steal a boat for their escape.
Mallory and Miller make their way into the heavily fortified gun emplacements. Locking the main entrance behind them — which sets off an alarm — they set obvious explosives on the guns and hide more below an elevator leading to the guns. The Germans cut through the thick emplacement doors, as Mallory and Miller make their escape by diving into the sea, reaching the stolen boat. But Pappadimos and Brown have been killed and Andrea is wounded. Mallory saves him, pulling him into the boat; thus voiding the 'blood feud' between them.
The Allied destroyers appear on schedule. The Germans remove the explosives planted on the guns and begin to fire on the passing Allied flotilla. The first salvo falls short. The second brackets the lead ship. However, just as the guns are prepared to fire again, the elevator descends low enough to trigger the hidden explosives. The guns and fortifications are destroyed in a spectacular explosion. Franklin is able to hear it from his hospital bed.
Andrea, who has fallen in love with Maria, decides to return to Navarone with her. Mallory and Miller observe the aftermath of the destruction from a destroyer.
The film was a major Box Office success and the top grossing film of 1961 earning a net profit of $18,500,000. As a result, MacLean reunited Mallory, Miller, and Andrea in the best-seller Force 10 from Navarone, the only sequel of his long writing career, in 1968. That was in turn filmed as the significantly different Force 10 from Navarone in 1978 by British director Guy Hamilton, a veteran of several James Bond films. Despite a cast that included Robert Shaw, Edward Fox, and Harrison Ford, it was a critical and commercial failure.