Aims: Nodule-in-nodule (N/N) hepatocellular carcinoma (HCC) is a convincing proof of multistep hepatocarcinogenesis. In this lesion, an inner HCC develops within an outer, more differentiated, tumour, which can be rapidly taken over by the former so that N/N HCC is rarely detected., Methods and Results: Ten resected N/N HCCs arising in cirrhotic background and characterized: (i) as outer lesions by early (n = 3) and G1 (n = 7) HCC; (ii) as inner lesions by G1 (n = 3) and G2 (n = 7) HCC. The largest/smallest diameters of outer and inner nodules were, respectively, 20/6 mm and 16/4 mm. We investigated vascular (CD34 and endocan), hepatocellular VEGF, GS, GPC3, HSP70 and CHC) and molecular (TERT promoter and β-catenin) changes taking place from the outer neoplastic compartment to the inner neoplastic compartment (INC). A diffuse pattern of CD34+ capillarized vessels and focal endocan immunoreactivity were major distinctive features acquired in the INC; VEGF immunoreactivity was inversely related to CD34 staining. A gain in the number of cells immunoreactive for GPC3, HSP70, and CHC, but not of GS-immunoreactive cells, also occurred in the INC. TERT promoter mutations were seen in half of the cases in both compartments, whereas β-catenin mutations were more rarely detectable., Conclusions: Major phenotypic changes take place in the INC of N/N HCC. TERT promoter mutations take place frequently and very early, and, in contrast to β-catenin mutations, do not appear to be acquired during N/N growth. These findings suggest that inner nodules represent a step further along the pathway of tumour progression, in contrast to earlier, simply initiated, lesions, and that complete neovascularisation predicts a change in HCC biology., (© 2018 John Wiley & Sons Ltd.)