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Nagayama. J Cancer Metastasis Treat 2021;7:6 I http://dx.doi.org/10.20517/2394-4722.2020.114 Page 5 of 16

In thyroid
The physiological role for autophagy in thyroid was studied using genetically engineered mice. We
generated the thyroid specific Atg5, a component of the autophagy machinery [Figure 1], KO mice Atg5 flox/flox ;
) . These KO mice were born with an expected Mendelian trait and in euthyroid state
TPO-Cre (Atg5 thyr-KO [21]
during the 1-year-observational period. However, accumulation of ubiquitinated proteins was detected at
4 months, and apoptotic cell death, decreased number of thyroid follicular epithelial cells (thyrocytes), and
increased DNA damages detected by 8-hydroxy-2’-deoxyguanosine (8-OHdG) and p53-binding protein
1 (53BP1) foci were observed at 8 and 12 months, indicating the critical role of basal level of autophagic
activity in thyrocyte survival and homeostasis.

To study the significance of induced autophagy in a stressed condition, we first clarified regulation
of autophagic activity by hormones, finding the positive control by TSH and the negative control by
thyroid hormone T 4 [11] . When KO mice were treated with methimazole and perchlorate, which induce
increased TSH and decreased T levels (as a net effect, induce autophagy), smaller follicle sizes and lower
4
thyroglobulin (TG) contents in thyrocytes were observed as compared to control mice, implying impaired
TG production presumably by diminished nutrient supply due to a lack of autophagy. Altogether, basal
level of autophagy is required for maintenance of cell homeostasis and survival, and induced autophagy,
elicited by elevated TSH and decreased thyroid hormone, supports the provision of building blocks for
sufficient synthesis of proteins including TG.

KO mice with disrupted Vps34 gene, a component of class III PI3K (a more upstream molecule than ATG5
cKO
in autophagic flow; see Figure 1) were also generated . Vps34 flox/flox[11] ; Pax8-Cre (Vps34 ) mice were also
[22]
born normally but died at approximately 1 month of age; they had severe hypothyroidism, impaired TG
iodination, abnormal TG trafficking, defective lysosomal proteolysis, and the presence of macrophages
cKO
eating iodinated TG in the lumen were observed. More pronounced phenotype alterations in Vps34
mice are evident when comparing to Atg5 thyr-KO mice, which may be explained by autophagy-independent
[23]
functions of autophagy-related proteins . Indeed, the findings in the latter seem to be at least in part
attributed to the role for VPS34 in vesicular trafficking rather than autophagic activity.

AUTOPHAGY IN CANCER
Cancer initiation
In general
It is well accepted that autophagy has the dual effects on cell survival and death, that is, it on one hand
supports cell survival by providing nutrients required for maintaining cellular homeostasis, but on the other
hand promotes cell death, i.e., autophagy-regulated cell death [19,20] . Particularly in cancer, it is proposed that
autophagy suppresses tumor development in the early stage of tumorigenesis, while autophagy promotes
tumor survival and proliferation once a tumor develops . Therefore, numerous preclinical and clinical
[24]
studies on autophagy-targeted treatment mainly focused on autophagy inhibitors [24,25] . However, the data so
far accumulated are not so simple, and even in established cancers, studies show therapeutic effect of not
only autophagy inhibitors but also autophagy inducers on tumor growth [24,26] .

Much evidence indicating the role for autophagic activity in the initial step of cancer development comes
from the studies with genetically engineered mice. Beclin-1 heterozygous KO mice developed spontaneous
lung cancers, liver cancers, and lymphomas, and accelerated the development of HBV-induced liver
premalignant lesions [27,28] . Systemic mosaic Atg5 KO and liver-specific Atg7 KO mice developed benign
liver adenomas [29,30] . Although accumulated p62, commonly observed in all KO mice, is thought to play a
role for tumorigenesis [30,31] , development of cancers in multiple organs in Beclin-1 heterozygous KO mice
as compared to that of benign tumors only in liver in systemic mosaic Atg5 KO and liver-specific Atg7 KO
mice may indicate autophagy-independent anti-tumorigenic function of BECLIN-1; for example, BECLIN-