Recent developments in treatments for kidney cancer

There are 338,000 new cases of kidney cancer per year worldwide, and the incidence has increased in recent years (Ferley et al., 2012). Although the majority of Renal Cell Carcinoma (RCC) cases are sporadic and affect those over 50 years old, 2-3% of cases are caused by inherited conditions such as BHD, VHL, HLRCC and TSC and are associated with an earlier onset (Randall et al., 2014). These inherited forms of RCC have provided great insights into the genetics of sporadic cancer – for example 75% of RCC cases are associated with mutations in the VHL gene.

The majority of small local RCC tumours can be surgically removal. However these treatments are not without risk and complete nephrectomies leave patients with severely reduced kidney function. The development of selective drug treatments that target only cancerous cells could therefore increase patient quality of life.

Current treatments for advanced or metastasised RCC are focused on counteracting the metabolic and angiogenic changes associated with the VHL/HIF pathway: Tyrosine Kinase Inhibitors (TKI) sunitinib, sorafenib and axitinib inhibit VEGF and PDGF signalling, increased as a result of aberrant HIF signalling, to limit angiogenesis; and the mTOR inhibitor everolimus, and derivatives, reduce mTOR and downstream HIF signalling to minimise tumour growth and the metabolic shift from oxidative phosphorylation to glycolysis. Despite these advances only half of kidney cancer patients are currently expected to survive past 10 years (Chow et al., 2010), so more advanced and effective treatments are sorely required.

This week’s blog is a brief review of several recent reports on the development of new treatments for RCC and updates on current treatments.

  • Englerin A, a purified molecule from Phyllanthus engleri bark, selectively kills cancer cells (Ratnayake et al., 2009) by increasing intracellular calcium levels (Sulzmaier et al., (2012). Akbulut et al., (2015) now report that activation of the Transient Receptor Potential cation channels TRPC4 and TRPC5 induces calcium influx resulting in rapid death of tumourigenic cells, thereby identifying novel drug targets for the selectively treatment of cancerous kidney cells.
  • Hall et al., (2014) reported that TRPM3 cation channel activation promotes clear cell RCC tumour growth by activating autophagy. Increased intracellular calcium levels remove miR-214-inhibition of autophagy, however this pathway can be blocked by mefenamic acid (MFA). The identified drug targets could form the basis of an effective combination treatment targeting the upregulation of autophagy and tumourigenic angiogenesis.
  • Additional targets identified through genetic and biochemical studies of tumours and tumour cell lines include HIF2α, chromatin regulators SETD2, BAP1 and PBRM, MET in papillary type I tumours, and inhibitors that can limit glucose uptake in tumours that have switched to aerobic glycolysis as the main source of energy (reviewed in Srinivasan et al., 2015).

In addition to this new research, preliminary data from several clinical trials has recently been released.

  • A trial assessing a combination of dalantercept and axitinib for treatment of advanced RCC reported encouraging results. In part 1 of the phase II trial dalantercept, and ALK-1 inhibitor, in combination with the multiple TKI axitinib, showed a partial response and prolonged disease control in patients. A larger phase II part 2 trial will assess if the combination therapy enhances progression free survival more than treatment with axitinib alone.
  • The ASSURE clinical trial reported that adjuvant use of sorafenib and sunitinib did not significant improve disease free survival or overall survival, nor reduction time to disease recurrence compared to a placebo in patients following surgical tumour removal. Due to significant side effects both drugs were individually titrated to reduce the high discontinuation rate.
  • Phase II trial results results show lenvatinib, a multi-TKI, alone or in combination with everolimus, prolongs progression free survival significantly more than everolimus treatment alone in patients with advanced or metastatic RCC. The combination therapy inhibits multiple tumourigenic signalling pathways simultaneously reducing the risk of developed drug resistance.
  • A combinatorial trial of Bevacizumab and Erlotinib in HLRCC and sporadic papillary RCC patients reported a 65% and 29% response rate in HLRCC patients and sporadic patients respectively, with the majority of tumours shrinking or remaining stable and extended survival. The enhanced need of these papillary tumours for high levels of glucose makes them particularly susceptible to treatments that impair glucose delivery.

As RCC is really a collection of distinct diseases that occur in the same organ, it is unlikely that any one single therapy will be suitable for all patients. Instead through increased understanding of the tumourigenic pathways and processes in different patient cohorts more personalised targeted therapies (as discussion in this blog post) can be developed.


  • Akbulut Y, Gaunt HJ, Muraki K, Ludlow MJ, Amer MS, Bruns A, Vasudev NS, Radtke L, Willot M, Hahn S, Seitz T, Ziegler S, Christmann M, Beech DJ, Waldmann H. (-)-Englerin A is a Potent and Selective Activator of TRPC4 and TRPC5 Calcium Channels. Angew Chem Int Ed Engl. 2015 Mar 16;54(12):3787-91. PubMed PMID: 25707820.
  • Chow WH, Dong LM, Devesa SS. Epidemiology and risk factors for kidney cancer. Nat Rev Urol. 2010 May;7(5):245-57. Review. PubMed PMID: 20448658.
  • Ferlay J, Soerjomataram I, Ervik M, et al. GLOBOCAN 2012 v1.0, Cancer Incidence and Mortality Worldwide: IARC CancerBase No. 11 [Internet]. Lyon, France: International Agency for Research on Cancer; 2013. Available from:, accessed March 2015.
  • Hall DP, Cost NG, Hegde S, Kellner E, Mikhaylova O, Stratton Y, Ehmer B, Abplanalp WA, Pandey R, Biesiada J, Harteneck C, Plas DR, Meller J, Czyzyk-Krzeska MF. TRPM3 and miR-204 establish a regulatory circuit that controls oncogenic autophagy in clear cell renal cell carcinoma. Cancer Cell. 2014 Nov 10;26(5):738-53. PubMed PMID: 25517751.
  • Randall JM, Millard F, & Kurzrock R (2014). Molecular aberrations, targeted therapy, and renal cell carcinoma: current state-of-the-art. Cancer metastasis reviews, 33 (4), 1109-24 PMID: 25365943
  • Ratnayake R, Covell D, Ransom TT, Gustafson KR, Beutler JA. Englerin A, a selective inhibitor of renal cancer cell growth, from Phyllanthus engleri. Org Lett. 2009 Jan 1;11(1):57-60. PubMed PMID: 19061394.
  • Srinivasan R, Ricketts CJ, Sourbier C, Linehan WM. New strategies in renal cell carcinoma: targeting the genetic and metabolic basis of disease. Clin Cancer Res. 2015 Jan 1;21(1):10-7. PubMed PMID:25564569.
  • Sulzmaier FJ, Li Z, Nakashige ML, Fash DM, Chain WJ, Ramos JW. Englerin a selectively induces necrosis in human renal cancer cells. PLoS One. 2012;7(10):e48032. PubMed PMID: 23144724.
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Differential effects of HIF-α isoforms on apoptosis in renal carcinoma cell lines

Under hypoxic conditions the activation of HIF transcription factors enables cells to alter their metabolism and avoid stress-induced apoptosis. Aberrant HIF activity in the inherited renal cancers BHD, VHL, HLRCC and TSC, is linked to the expression of growth and pro-angiogenic factors that are important in tumour growth. A new report from Doonachar et al., (2015) focuses on the differential effects of the HIF-1α and HIF-2α isoforms on stress-induced apoptosis in two VHL-deficient renal cell carcinoma (RCC) cell lines.

Under normoxic conditions VHL protein forms part of the E3 ligase complex that targets HIF for degradation thereby limiting the production of HIF-target genes; HIF-1α primarily upregulates glycolytic genes and HIF-2α upregulates angiogenic and growth factors. Homozygous loss-of-function mutations in VHL, both inherited and sporadic, in renal cells results in an increase in either HIF-1α and HIF-2α activity or only HIF-2α activity, and subsequent tumour growth. HIF-2α is therefore seen as a major driver of renal tumourigenesis following the loss of a tumour suppressor such as VHL (Kondo et al., 2003).

VHL has also been shown to help protect renal cells from apoptosis following a range of stresses (Schoenfeld et al., 2000). Doonachar et al., aimed to determine any differential effects of HIFα isoform on apoptosis in VHL-defective cells following cellular stress.

In the VHL-deficient 786-O cell line the transgenic expression of mutant VHL, compared to transgenic wild type VHL, induced expression of HIF-2α (the only isoform this line expresses) and apoptosis following UV exposure. Inhibiting HIF-2α translation with shRNAs, confirmed through reduced target gene GLUT1 expression, marginally reduced apoptosis suggesting a partial protection from UV-induced apoptosis. The loss of HIF-2α in this cell line did alter the rate of cell death induced by glucose starvation or serum withdrawal. Under all stress conditions the expression of functional VHL drastically reduced 786-O cell death confirming it as anti-apoptosis factor.

In a second kidney tumour cell line, RCC10, that expresses both HIFα isoforms, the presence of only mutated VHL resulted in increased expression of HIF-1α and HIF-2α and a greater rate of apoptosis following UV exposure. Inhibiting transcription of either HIF-1α or HIF-2α via shRNA reduced respective protein levels but did not impact on GLUT1 expression suggesting transcriptional redundancy. shRNA-inhibition of HIF-1α increased cell death following UV exposure and glucose starvation suggesting that HIF-1α has an anti-apoptotic role. In contrast shRNA-inhibition of HIF-2α reduced cell death following UV exposure and serum withdrawal, indicative of a pro-apoptotic role. However, as there was also a, potentially compensatory, increase in HIF-1α expression following HIF-2α shRNA-inhibition, it could also be attributed to an increase in HIF-1α anti-apoptotic activity. There was no comparative increase in HIF-2α expression detected following shRNA-inhibition of HIF-1α.

Doonachar et al., conclude that HIFα isoforms seem to have different roles following cellular stress; HIF-1α is generally anti-apoptotic and HIF-2α pro-apoptotic. Further work is needed to determine if differences in isoform function are also relevant in tumour formation and if they could be exploited in treatments.

In contrast to VHL studies, Flcn-null cell lines show no increase in HIF-1α or HIF-2α expression despite a marked increase in activity determined by increased expression of target genes (Preston et al., 2010, Yan et al., 2014). The increase in HIF activity reported in BHD RCCs is thought to be mediated through AMPK and PGC1α activity. Yan et al., propose that FLCN loss increases AMPK and PCG1α activity leading to increased mitochondrial biogenesis, increased ROS production and subsequent activation of HIF signalling. Mutations in FLCN have also been linked to aberrant mTOR activity (Baba et al., 2006), an upstream activator of HIF signalling in tumour cells (Hudson et al., 2002). It is possible that these mechanisms, alongside increased HIF protein stability resulting from reduced VHL-dependent ubiquitination and degradation, drive enhanced HIF signalling in VHL-RCCs.

Increased HIF activity has also been reported in BHD pulmonary cyst epithelial cells. Nishii et al., (2013) detected a mild increase in HIF-1α and VEGF levels, and increased angiogenesis in BHD-lung cysts but not control tissues. As there is no evidence of second hit mutations in lung tissue any changes are more likely due to haploinsufficiency that a complete loss of function. The resulting milder changes in HIF activity could explain the lack of tumourigenic cell proliferation in pulmonary cysts.

Aberrant HIF activity in renal tumours suggests an important role in tumourigenesis. Increased understanding of the expression and targets of HIF-α isoforms under normal and renal tumourigenic conditions will enable identification of new drug targets. In addition, as HIF signalling appears to play a role in other aspects of BHD pathology, such treatments could be applicable to these systems.


  • Baba M, Hong SB, Sharma N, Warren MB, Nickerson ML, Iwamatsu A, Esposito D, Gillette WK, Hopkins RF 3rd, Hartley JL, Furihata M, Oishi S, Zhen W, Burke TR Jr, Linehan WM, Schmidt LS, Zbar B. Folliculin encoded by the BHD gene interacts with a binding protein, FNIP1, and AMPK, and is involved in AMPK and mTOR signaling. Proc Natl Acad Sci U S A. 2006 Oct 17;103(42):15552-7 PubMed PMID: 17028174.
  • Doonachar A, Gallo MD, Doukas D, Pasricha R, Lantsberg I, Schoenfeld AR. Differential effects of HIF-α isoforms on apoptosis in renal carcinoma cell lines. Cancer Cell Int. 2015 Feb 22;15:23. doi: 10.1186/s12935-015-0175-3. eCollection 2015. PubMed PMID: 25729330.
  • Hudson CC, Liu M, Chiang GG, Otterness DM, Loomis DC, Kaper F, Giaccia AJ, Abraham RT. Regulation of hypoxia-inducible factor 1alpha expression and function by the mammalian target of rapamycin. Mol Cell Biol. 2002 Oct;22(20):7004-14. PubMed PMID: 12242281.
  • Kondo K, Kim WY, Lechpammer M, Kaelin WG Jr. Inhibition of HIF2alpha is sufficient to suppress pVHL-defective tumor growth. PLoS Biol. 2003 Dec;1(3):E83. PubMed PMID: 14691554.
  • Nishii T, Tanabe M, Tanaka R, Matsuzawa T, Okudela K, Nozawa A, Nakatani Y, Furuya M. Unique mutation, accelerated mTOR signaling and angiogenesis in the pulmonary cysts of Birt-Hogg-Dubé syndrome. Pathol Int. 2013 Jan;63(1):45-55. PubMed PMID: 23356225.
  • 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. Absence of the Birt-Hogg-Dubé gene product is associated with increased hypoxia-inducible factor transcriptional activity and a loss of metabolic flexibility. Oncogene. 2011 Mar 10;30(10):1159-73 PubMed PMID: 21057536.
  • Schoenfeld AR, Parris T, Eisenberger A, Davidowitz EJ, De Leon M, Talasazan F, Devarajan P, Burk RD. The von Hippel-Lindau tumor suppressor gene protects cells from UV-mediated apoptosis. Oncogene. 2000 Nov 30;19(51):5851-7. PubMed PMID: 11127815.
  • Yan M, Gingras MC, Dunlop EA, Nouët Y, Dupuy F, Jalali Z, Possik E, Coull BJ, Kharitidi D, Dydensborg AB, Faubert B, Kamps M, Sabourin S, Preston RS, Davies DM, Roughead T, Chotard L, van Steensel MA, Jones R, Tee AR, Pause A. The tumor suppressor folliculin regulates AMPK-dependent metabolic transformation. J Clin Invest. 2014 Jun;124(6):2640-50. PubMed PMID: 24762438.
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Drug Repositioning for Rare Diseases

A recent Findacure meeting focused on the importance and progression of drug repositioning in rare diseases. Dr Bruce Bloom, Dr Mike Briggs and Dr Farid Khan, discussed ongoing repositioning work, methods to identify new drugs and targets and the need for collaborations to drive new research and trials. Cures Within Reach are launching a new interactive platform CureAccelerator to help form such collaborations. The session’s presentations are available here.

There are over 6000 rare diseases1 recognised worldwide, and as the development of a new drug is thought to take between 10-15 years2 and cost approximately $1.2 billion USD3, the limited market appeal of rare disease or orphan drugs reduces investment incentive.  The Orphan Drug Act of 1983 was passed in the US to increase incentive and, to date, 486 orphan drugs have been approved4.  The remaining 90-95% of rare diseases do not have any specific treatments.

An alternative approach is to reposition (or repurpose) some of the 4000 existing drugs approved for human use worldwide (Huang et al., 2011). Existing early phase safety and efficacy data for these drugs means repositioning studies can be cheaper, safer and faster than conventional drug development.

BHD is a rare disease for which no preventative treatments are available and lists only three trials for BHD. Two long running studies at the National Cancer Institute are looking at the clinical, genetic and molecular basis of BHD and all heritable urologic disorders, and a third, funded by the Myrovlytis Trust, assessed the use of topical rapamycin to treat fibrofolliculomas (discussed here).

Rapamycin, an mTOR inhibitor, is a repositioned drug: first licenced as an immunosuppressant it is now used as a treatment for a range of diseases including cancers including advanced kidney cancer in BHD patients. The role of mTOR activity in fibrofolliculoma pathology is less understood. This could explain why the clinical trial did not find evidence of topical rapamycin as an effective treatment for BHD fibrofolliculomas (Gijezan et al., 2014).

FLCN loss also disrupts other pathways including HIF signalling; increased HIF signalling is seen in FLCN-null cell lines (Preston et al., 2011) as a result of altered AMPK activity, and could be a driving force in the development of renal tumours. HIF-1 hyperactivity is also seen in sporadic renal tumours which lead to the development of HIF-1 inhibitors (Onnis et al., 2009, Welsh et al., 2013). If BHD pathologies are linked to aberrant HIF-1 activity these drugs may be able to provide the basis for new treatments.

A drug screen, funded by the Myrovlytis Trust, using a BHD-kidney cell line (Yang et al., 2008) identified Mithramycin, an antineoplastic antibiotic, as selectively toxic to FLCN null cells (Lu et al., 2011). Used to treat testicular cancer, leukaemia and Paget’s disease, Mithramycin could potentially, after further research, be a viable drug for repositioning trials in BHD.

The pathology of BHD fibrofolliculomas and pulmonary cysts is less well understood than FLCN-associated renal tumours. This potentially limits the development of new treatments based on aberrant biological pathways or activity. Further research into the mechanisms through which a reduction in FLCN leads to these phenotypes will hopefully enable new drug targets to be identified.

Drug screens, incidental findings, bioscience research and computational biology can all provide insights into potential new treatments for specific diseases or pathways. Greater understanding of the pathways affected in BHD and their roles in pathology will enable researchers to identify more likely drug targets. Repositioning of known drugs could then prove useful in discovering safe and effective treatments for BHD and other rare disease patients in a shorter time span than conventional methods.

  • Gijezen LM, Vernooij M, Martens H, Oduber CE, Henquet CJ, Starink TM, Prins MH, Menko FH, Nelemans PJ, van Steensel MA. Topical rapamycin as a treatment for fibrofolliculomas in Birt-Hogg-Dubé syndrome: a double-blind placebo-controlled randomized split-face trial. PLoS One. 2014 Jun 9;9(6):e99071. PubMed PMID: 24910976.
  • Huang, R., Southall, N., Wang, Y., Yasgar, A., Shinn, P., Jadhav, A., … Austin, C. P. (2011). The NCGC Pharmaceutical Collection: A comprehensive resource of clinically approved drugs enabling repurposing and chemical genomics.Science Translational Medicine3(80). PMID: 21525397
  • Lu X, Wei W, Fenton J, Nahorski MS, Rabai E, Reiman A, Seabra L, Nagy Z, Latif F, Maher ER. Therapeutic targeting the loss of the birt-hogg-dube suppressor gene. Mol Cancer Ther. 2011 Jan;10(1):80-9. PubMed PMID: 21220493.
  • Onnis B, Rapisarda A, Melillo G. Development of HIF-1 inhibitors for cancer therapy. J Cell Mol Med. 2009 Sep;13(9A):2780-6. PubMed PMID: 19674190.
  • 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. Absence of the Birt-Hogg-Dubé gene product is associated with increased hypoxia-inducible factor transcriptional activity and a loss of metabolic flexibility. Oncogene. 2011 Mar 10;30(10):1159-73. PubMed PMID:21057536.
  • Welsh SJ, Dale AG, Lombardo CM, Valentine H, de la Fuente M, Schatzlein A, Neidle S. Inhibition of the hypoxia-inducible factor pathway by a G-quadruplex binding small molecule. Sci Rep. 2013 Sep 30;3:2799. PubMed PMID: 24165797.
  • Yang Y, Padilla-Nash HM, Vira MA, Abu-Asab MS, Val D, Worrell R, Tsokos M, Merino MJ, Pavlovich CP, Ried T, Linehan WM, Vocke CD. The UOK 257 cell line: a novel model for studies of the human Birt-Hogg-Dubé gene pathway. Cancer Genet Cytogenet. 2008 Jan 15;180(2):100-9. PubMed PMID: 18206534.


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Distinctive expression patterns of FLCN and GPNMB in BHD renal tumours

As discussed on this blog previously, developing histological screening techniques for renal cell carcinomas (RCCs) associated with BHD is important for early diagnosis. Individuals with folliculin (FLCN) mutations are more likely to develop multiple bilateral renal tumours (Zbar et al., 2002, Pavlovich et al., 2002). A misdiagnosis of sporadic RCC may compromise future treatment and wellbeing of the patient and other affected family members. Currently there are no known histological markers to distinguish between all subtypes of sporadic and FLCN-associated tumours.

A new report by Furuya et al., (2015) addressed this by analysed expression of FLCN and one of its downstream targets GlycoProtein Non-Metatastic B (GPNMB) in normal and neoplastic tissue to determine if they could be used to aid differential diagnosis in RCC samples. FLCN is expressed in normal kidney tissue, including in those who carry heterozygous FLCN mutations, but is not detectable in BHD tumours (Warren et al., 2004). In comparison GPNMB is not typically expressed in kidney tissue but is expressed in a range of neoplastic tissues (Hong et al. 2010).

Furuya et al. analysed 27 tumours from 18 unrelated, except for a parent and child pair, Japanese BHD patients: 12 chromophobe RCCs; six hybrid oncocytoma/chromophobe tumours (HOCTs); three papillary RCCs; and two clear cell RCCs (ccRCC). For seven of these tumours sections of non-neoplastic kidney were also frozen for comparison. Expression of FLCN and GPNMB in the BHD-associated tumours was compared to 62 sporadic renal tumours from an unreported number of patients. All patients were medically examined for the classical BHD symptoms – pulmonary cysts, pneumothorax, fibrofolliculomas and multifocal/hybrid RCC – and a family history obtained. One of the sporadic group also had pulmonary cysts so a diagnosis of BHD was ruled out by genetic testing. The rest of this group did not show any BHD manifestations or have any family history so were assumed to not carry FLCN mutations.

FLCN expression in the tumour samples was assessed by western blot and immunohistochemistry using a monoclonal (D14G9) and previously unpublished polyclonal (ab93196) antibody respectively. The western blot results showed a clear loss of FLCN in BHD-associated but not sporadic tumours or non-neoplastic BHD tissue. In addition the typical strong nuclear staining for FLCN was seen in the majority of sporadic tumours whereas the majority of BHD-associated tumours showed only cytoplasmic or no staining. It is suggested that the presence of cytoplasmic staining should be considered a potential BHD indicator.

The presence of GPNMB protein was confirmed by western blot and immunohistochemistry, but mRNA levels were also quantified by quantitative RT-PCR. The markedly higher expression (up to 23-fold) detected in BHD tumours by qRT-PCR correlated with the intensity of the western band seen. GPNMB was barely detectable in the sporadic tumour samples and non-neoplastic BHD renal tissue. Immunostaining showed positive staining in BHD-associated tumours excluding the ccRCC samples and one papillary sample. The majority of sporadic samples were in contrast negative with the exception of 50% of sporadic chromophobe RCC samples which showed weak GPNMB staining.

It was also possible to detect low expression of GPNMB in small nodules of “non-neoplastic” BHD kidney tissue which could indicate the sites of future tumour development. As the formation of multiple small tumours is characteristic of BHD this could also help in differential diagnosis.

The germline FLCN mutations for each patient was determined from a peripheral blood sample; the most common (50%) mutation was duplication in the hypermutable cytosine tract in exon 11 and four patients had the same GATG deletion in exon 13. In addition Furuya et al., attempted to detect secondary somatic mutations in a number of the tumour samples. A second mutation was found in six individual tumours and a loss of heterozygosity in a further two. Only one of these secondary mutations has so far been reported as a germline mutation in a BHD patient. The other mutations may therefore not be pathogenic but as all would result in a frameshift it is more likely they are new unique mutations.

The results from this work suggest that staining for both a lack of FLCN and a gain of GPNMB could help to distinguish sporadic oncocytomas, chromophobe and papillary RCC from BHD-associated HOCTS, chromophobe and papillary RCCs. It would not be sufficient however to definitively classify a ccRCC as FLCN-related, but a lack of nuclear FLCN staining could identify cases appropriate for genetic testing.

  • Furuya M, Hong SB, Tanaka R, Kuroda N, Nagashima Y, Nagahama K, Suyama T, Yao M, & Nakatani Y (2015). Distinctive expression patterns of glycoprotein non-metastatic B and folliculin in renal tumors in patients with Birt-Hogg-Dubé syndrome. Cancer science PMID: 25594584
  • Hong SB, Oh H, Valera VA, Baba M, Schmidt LS, Linehan WM. Inactivation of the FLCN tumor suppressor gene induces TFE3 transcriptional activity by increasing its nuclear localization. PLoS One. 2010 Dec 29;5(12):e15793. doi:10.1371/journal.pone.0015793. PMID: 21209915.
  • Pavlovich CP, Walther MM, Eyler RA, Hewitt SM, Zbar B, Linehan WM, Merino MJ. Renal tumors in the Birt-Hogg-Dubé syndrome. Am J Surg Pathol. 2002 Dec;26(12):1542-52. PubMed PMID: 12459621.
  • Warren MB, Torres-Cabala CA, Turner ML, Merino MJ, Matrosova VY, Nickerson ML, Ma W, Linehan WM, Zbar B, Schmidt LS. Expression of Birt-Hogg-Dubé gene mRNA in normal and neoplastic human tissues. Mod Pathol. 2004 Aug;17(8):998-1011. PMID: 15143337.
  • Zbar B, Alvord WG, Glenn G, Turner M, Pavlovich CP, Schmidt L, Walther M, Choyke P, Weirich G, Hewitt SM, Duray P, Gabril F, Greenberg C, Merino MJ, Toro J, Linehan WM. Risk of renal and colonic neoplasms and spontaneous pneumothorax in the Birt-Hogg-Dubé syndrome. Cancer Epidemiol Biomarkers Prev. 2002 Apr;11(4):393-400. PubMed PMID: 11927500.


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Rare Disease Day 2015: Living with a Rare Disease

There are over 6000 rare diseases known worldwide. Whilst individually these diseases only affect a small number of people it is estimated that, assessed cumulatively, 1 in 17 people will be affected by a rare disease in their lifetime. To raise awareness of these diseases, every year since 2008, at the end of February there has been a Rare Disease Day.

The theme for Rare Disease Day 2015 is focused on the daily lives of patients, families and caregivers who are Living with a Rare Disease. Limited access to healthcare professionals knowledgeable about these rare and typically complex diseases often results in family members and friends becoming the main source of support and care. Rare Disease Day 2015 is a tribute to all the family and friends whose lives are affected by rare disease and who stand day-by-day and hand-in-hand with rare disease patients.

The lack of specific knowledge from local care authorities can lead to the patients and families becoming relative experts in their disease, often providing the information to new medical professionals when required. One invaluable source of information and support for rare disease patients is a disease-specific patient organisation. Although these can vary in the level of expertise and resources, many families find comfort in knowing that they are not alone in their diagnosis.

The BHD foundation website was established to help patients, families and doctors understand BHD from a more scientific stance, including the dissemination of new research. It also provides a primary contact for new BHD patients needing information on testing or local doctors. Complementary to this are the active facebook BHD groups which continue to be a wonderful source of mutual support and information from families affected by BHD regarding testing, symptoms, treatments and day-to-day issues associated with having a rare disease.

BHD has only been recognised as a disease since 1977. However thanks to the continuing work of a small, but increasing, number of clinicians and researchers worldwide, it has been possible to identify the gene associated with BHD, Follciulin, and to begin to understand its functions in the cell. Other research has allowed for the identification of more BHD patients and saved lives by allowing earlier treatment. The more we can understand about BHD and the interactions of folliculin in both normal and pathological situations the closer we will come to alleviating BHD symptoms and develop targeted treatments. They may not know every individual BHD patient in the world but our BHD researchers and clinicians are standing with them to try and make a difference.

There are Rare Disease Day events happening around the world, but if you are planning your own event and want help raising awareness get in touch with the Rare Disease Day team. You can also help spread awareness across social media by joining in with the Rare Disease Day Thunderclap.

Clicking on the images below will take you to Rare Disease Day Events homepages

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Characterisation of renal tumours in patients with Birt-Hogg-Dubé Syndrome

Due to mutations in their folliculin (FLCN) gene Birt-Hogg-Dubé (BHD) syndrome patients have a greater risk of developing renal cell carcinomas (RCC) than others (Zbar et al., 2002, Houweling et al., 2011). Unlike sporadic cases of RCC, where the majority are classified as clear cell RCC (ccRCC), studies of FLCN-related tumours have found that the majority are either chromophobe RCCs (34%) or hybrid oncocytoma/chromophobe tumours (HOCTs, 50%) with fewer cases of ccRCC (9%) and only rare occurrences of renal oncocytomas or papillary RCCs (Pavlovich et al., 2002). It is also common for multiple tumours of different subtypes to develop on the same kidney. Using current histology methods diagnosing a tumour as being related to BHD rather than sporadic can be difficult, especially in undiagnosed BHD cases where the genetic background will be unknown.

A new paper by Iribe et al., (2015) has investigated if the immunohistochemical profile of BHD-related tumours is sufficiently distinct from sporadic RCC tumours to be of help in classification. They suggest that a panel of markers including Carbonic Anhydrase IX (CA-IX), Kidney specific cadherin (Ksp-cadherin), Cytokeratin 7 (CK7) and CD82 could help in the screening for FLCN-related RCCs and enable the correct classification of some different subtypes.

In total 32 tumours, from 17 BHD patients, were analysed by in situ hybridisation. The tumours had already been classified as chromophobe RCCs (n=14), HOCTs (n=15) or ccRCC (n=3). Expression of the markers in these samples was compared to sporadic chromophobe RCC, ccRCCs and oncocytomas – the presence of FLCN mutations in these controls was not excluded but, based on a lack of other BHD pathologies, was assumed.

Most of the FLCN-related chromophobe RCC and HOCTs analysed were Ksp-cadherin+, CD82+ and CA-IX-, a profile similar to sporadic chromophobe RCCs. Unfortunately the results reported in this paper indicate that it would be difficult to distinguish between sporadic and FLCN-associated chromophobe RCC using this marker panel. However sporadic chromophobe RCC samples showed significantly higher expression of CK7 than FLCN-associated HOCTs. Additionally FLCN-associated HOCTs showed significantly greater expression of Ksp-cadherin and CD82 compared to sporadic oncocytomas. These differences indicate that the suggested panel of markers would be useful for distinguishing FLCN-associated HOCTs from sporadic chromophobe RCCs and oncocytomas.

All of the studied ccRCC samples, both BHD-associated and sporadic, stained positive for CA-IX but negative for the other markers. Therefore the panel suggested would be unsuitable for distinguishing FLCN-associated ccRCC from sporadic ccRCC. It would however be useful in determining if the clear cell-looking foci seen in some FLCN-associated RCCs are true ccRCC based on CA-IX+ expression or in fact Ksp-cadherin+, CK7+ and CD82+ HOCT or chromophobe cells.

Although the presence of HOCTs is more classically related to BHD, ccRCC and chromophobe RCC tumours also develop in BHD patients and therefore it is important to be able to distinguish between sporadic cases and those associated with FLCN mutations. A FLCN mutation will increase the likelihood of more tumours developing and the need for continual monitoring. The markers suggested by Iribe et al., are suitable for distinguishing FLCN-associated HOCTs from sporadic tumours but are insufficient to make this distinction for other RCC subtypes. Additional work from the group (Fuyura et al., 2015) will be able to add to the screening protocol and will be the topic of this blog in a few weeks.

The presence of germline FLCN mutations was confirmed in the Iribe et al., cohort however there was no identified pattern between mutation and tumour subtypes. Based on the Knudson two-hit hypothesis, tumourgenesis only occurs if both alleles of a tumour suppressor gene are mutated. Previous work by Vocke et al., (2005) identified such second-hit mutation in 70% of BHD-RCC tumours analysed. Such analysis was not completed in this study but it could be that larger analysis of such secondary mutations could provide clues to RCC subtype development in different patients. This could help with our understanding of tumourgenesis in BHD but also impact on the development of more targeted treatments.

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Pulmonologists should be more aware of Birt-Hogg-Dubé Syndrome

When BHD was first described in 1977 it was based on the presence of characteristic skin lumps (Birt et al., 1977). In addition BHD is now known to be associated with increased risk of renal cancer, the development of lung cysts and an associated increased risk in pneumothorax. In 2002, the association of BHD with mutations in the folliculin (FLCN) gene provided the definitive mechanism for BHD diagnosis (Nickerson et al., 2002). As such all patients suspected of having BHD should undergo genetic testing to confirm this diagnosis.

Pneumothorax in BHD patients is associated with the development of pulmonary cysts. Numerous research and clinical groups have confirmed high prevalence (up to 90%) of lung cysts in BHD patients (Toro et al., 2008, Agarwal et al., 2011). Whilst the development of lung cysts or blebs is not limited to BHD patients – pulmonary blebs are also associated with other pulmonary disease and smoking – the distribution and formation of the cysts seen in BHD is quite distinct. In BHD patients it is typical to find multiple, thin-walled, elliptical or lentiform, well-defined but with no internal structure, air-filled cysts. These cysts are also predominantly found in the medial or basal sections of the lung and are often subpleural (Tobino et al., 2009, Johannesma et al., 2014d). Toro et al., (2007) reported that BHD patients with a family history of spontaneous pneumothorax and more pulmonary cysts were significantly more likely to develop pneumothoraces.

BHD is underdiagnosed due to a lack of awareness in the medical profession of such a rare disease in combination with its variable presentation and onset. Only a small percentage of BHD diagnoses are suspected based on the presentation of pulmonary pathologies alone. However over the last few years there have been increasing numbers of such case studies where individuals have presented with spontaneous pneumothorax and genetic testing was used to confirm a BHD diagnosis (Predina et al., 2011, Auerbach et al., 2014, Kilinceret al., 2014, Johannesma et al., 2013, Johannesma et al., 2014c, Johannesma et al., 2014fArdilouze et al., 2015).

It has been estimated that BHD patients have a 50-times higher risk of developing pneumothorax than non-BHD individuals (Zbar et al., 2002). Two large cohort studies also looked at the incidence of BHD in cases of primary spontaneous pneumothorax (PSP) and found that approximately 5-10% of PSP patients carried FLCN mutations (Ren et al.,2008, Johannesma et al., 2014f). PSP incidence is 1.2-6/100,000 for women and 7.4-18/100,000 for men, annually (Luh, 2010)) and the global population is 7×109.  This suggests that the total number of PSP annually is 300,000-800,000, with 15,000-80,000 of those patients carrying FLCN mutations.  These suggested figures are vastly more than the current number of diagnosed BHD patients, which might  be  a result of unintentional cohort sampling bias or due to the difficultly of establishing accurate statistics when dealing with such a small sample population as BHD patients. The true number of un- or mis-diagnosed BHD patients is difficult to estimate however it is clear that many BHD patients are being misdiagnosed as PSP patients. This will continue to be the case unless there is more awareness of BHD as a potential cause of spontaneous pneumothorax. The variation in BHD presentation makes it is highly important, especially with younger patients, to obtain a detailed family history with regards to BHD pathologies as these can contain additional differential diagnostic information.

New diagnosis guidelines suggested by Gupta et al., (2013) would allow diagnosis of BHD to be based on the presence of “characteristic BHD-cysts” on a high resolution CT scan in combination with fibrofolliculomas, BHD-related renal cancer, a first or second degree relative with confirmed BHD, or a positive genetic test. Whilst the numerous individual case and cohort studies suggest that the presence of characteristic lung cysts is indicative of BHD, it is only via genetic testing that the diagnosis can be confirmed. Identifying these patients, and any other affected family members, as early as possible is important as it will impact on future monitoring and treatment. Therefore it is of great importance that awareness of BHD and its pulmonary presentation is increased in pulmonologists and associated professions.

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