Hypoxia regulation ensures cell survival and growth in low oxygen environments. HIF signalling is a well-established element of this regulation but is also associated with tumourigenesis in BHD, VHL, HLRCC, TSC, and sporadic cancers. New research from Lee et al., (2015) has identified a second, HIF-independent, hypoxia response which can modify cell survival and growth signalling pathways – the lactate-induced activation of NDRG3-mediated signalling.
Oxygen-dependent hydroxylation of HIF-1α and HIF-2α by prolyl hydroxylase domain (PHD) enzymes and ubiquitination by pVHL results in proteosomal degradation inhibiting HIF signalling. Immunoprecipitation identified NDRG3, a protein previously implicated in cell proliferation and migration signalling (Melotte et al., 2010), as also interacting with PHD2. Knockdown of either PHD2 or pVHL increased NDRG3 protein levels, indicating normoxic post-translational degradation by PHD2/VHL (Lee et al., 2015).
NDRG3 protein levels increased in multiple cell types in response to hypoxia and were correlated with increased angiogenesis, anti-apoptotic, motility and proliferative (but not metabolic) gene expression. A critical role in these processes was confirmed when reduction in NDRG3 abolished the hypoxia-induced expression of pro-angiogenic factors, increased cellular apoptosis under prolonged hypoxia and decreased tumour cell growth.
NDRG3-accumulation in hypoxia occurs after HIF activation and is correlated with HIF-1α-mediated cellular lactate production; NDRG3 activity is not directly HIF-dependent but is coupled to the HIF-mediated hypoxia response. Lee et al. determined that lactate physically binds to NDRG3 blocking VHL-mediated ubiquitination and proteosomal degradation. Further work is required to determine if lactate is also inhibiting the PHD2-mediated hydroxylation required for VHL binding.
NDRG3 increased phosphorylation of c-Raf and B-RAF suggesting a role in activation of the Raf-ERK signalling pathway – associated with promoting cell growth and increased expression of angiogenic markers in hypoxia and tumours (Roberts & Der, 2007). Lee et al., propose two phases in prolonged hypoxia that enable cells to cope: initially HIF-mediated changes to metabolism increase the production of biosynthetic building blocks and results in lactate build-up, then NDRG3-mediated signalling pathways provide cues for cellular growth and angiogenesis.
However, this self-sufficient mechanism that enables recovery from hypoxia, can also be used by tumour cells enabling continued growth. Lee et al., reported increased levels of NDRG3 and ERK1/2 signalling molecules in engrafted tumours and patient hepatocellular carcinoma samples (n=25/103) supporting a role in tumour development. Given the importance of pVHL in NDRG3 regulation, the NDRG3-Raf-ERK signalling pathway could also be playing a role in VHL-associated tumourigenesis.
Cancer cells commonly show metabolic changes and increased dependency on glycolysis leading to increased lactate production. Lactate can be an alternative energy source and an inducer of tumour angiogenesis (Doherty & Cleveland, 2013), but lactate dehydrogenase inhibition can suppress tumour growth (Le et al., 2010). Determining NDRG3 as a mediator of lactate-induced responses has identified a new therapeutic target; targeting NDRG3 in combination with HIF signalling could increase treatment efficiency by reducing apoptotic-escape via a HIF-independent pathway (Lee et al., 2015).
NDRG3 targeting treatments or combination treatments could prove effective for BHD as a loss of FLCN has been linked to increased lactate production (Preston et al., 2011) and ERK1/2 signalling (Baba et al., 2008, Hudon et al., 2010). In addition, impairing lactate production in UOK257 (FLCN-null) cells reduces cell growth (Preston et al., 2011) which could indicate an important role for NDRG3-Raf-ERK signalling in BHD tumourigenesis. Treatments that either directly target NDRG3 or that block lactate accumulation could therefore be beneficial in the treatment of a wide range of cancers.
- Baba M, Furihata M, Hong SB, Tessarollo L, Haines DC, Southon E, Patel V, Igarashi P, Alvord WG, Leighty R, Yao M, Bernardo M, Ileva L, Choyke P, Warren MB, Zbar B, Linehan WM, Schmidt LS (2008). Kidney-targeted Birt-Hogg-Dube gene inactivation in a mouse model: Erk1/2 and Akt-mTOR activation, cell hyperproliferation, and polycystic kidneys. J Natl Cancer Inst. Jan 16;100(2):140-54. PubMed PMID: 18182616.
- Doherty JR, Cleveland JL (2013). Targeting lactate metabolism for cancer therapeutics. J Clin Invest. Sep;123(9):3685-92. Review. PubMed PMID: 23999443.
- Hudon V, Sabourin S, Dydensborg AB, Kottis V, Ghazi A, Paquet M, Crosby K, Pomerleau V, Uetani N, Pause A (2010). Renal tumour suppressor function of the Birt-Hogg-Dubé syndrome gene product folliculin. J Med Genet. Mar;47(3):182-9. PubMed PMID: 19843504.
- Le A, Cooper CR, Gouw AM, Dinavahi R, Maitra A, Deck LM, Royer RE, Vander Jagt DL, Semenza GL, Dang CV (2010). Inhibition of lactate dehydrogenase A induces oxidative stress and inhibits tumor progression. Proc Natl Acad Sci U S A. Feb 2;107(5):2037-42. PubMed PMID: 20133848.
- Lee DC, Sohn HA, Park ZY, Oh S, Kang YK, Lee KM, Kang M, Jang YJ, Yang SJ, Hong YK, Noh H, Kim JA, Kim DJ, Bae KH, Kim DM, Chung SJ, Yoo HS, Yu DY, Park KC, & Yeom YI (2015). A lactate-induced response to hypoxia. Cell, 161 (3), 595-609 PMID: 25892225.
- Melotte V, Qu X, Ongenaert M, van Criekinge W, de Bruïne AP, Baldwin HS, van Engeland M (2010). The N-myc downstream regulated gene (NDRG) family: diverse functions, multiple applications. FASEB J. Nov;24(11):4153-66. Review. PubMed PMID: 20667976.
- Preston RS, Philp A, Claessens T, Gijezen L, Dydensborg AB, Dunlop EA, Harper KT, Brinkhuizen T, Menko FH, Davies DM, Land SC, Pause A, Baar K, van Steensel MA, Tee AR (2011). Absence of the Birt-Hogg-Dubé gene product is associated with increased hypoxia-inducible factor transcriptional activity and a loss of metabolic flexibility. Oncogene. Mar 10;30(10):1159-73. PubMed PMID: 21057536.
- Roberts PJ, Der CJ (2007). Targeting the Raf-MEK-ERK mitogen-activated protein kinase cascade for the treatment of cancer. Oncogene. May 14;26(22):3291-310. Review. PubMed PMID: 17496923.