Finding useful biomarkers to predict efficacy of Sunitinib

Biological markers – or biomarkers – are an area of extreme interest in medicine, as they can be used to diagnose illness, predict the likely course of a disease, or to predict patients’ response to a particular intervention. Diagnostic, prognostic, and predictive biomarkers can be any biological material, such as DNA, RNA, proteins or metabolites.

In order to be useful, biomarkers need to be easily quantifiable, and their presence or absence needs to be highly associated with a biological state or outcome. For example, a commonly used diagnostic biomarker is prostate specific antigen (PSA), which allows prostate health to be inferred from a simple blood test.

There are currently eight FDA-approved systemic treatments available to treat metastatic renal cancers, and more are being developed. However, patients do not respond equally well to the different treatments. Thus finding biomarkers to accurately predict which patients will benefit from which treatment will improve the prognosis for patients with advanced kidney cancer.

A recent study by Motzer et al. (2014) investigated a number of potential biomarkers to predict the efficiacy of the tyrosine kinase inhibitor Sunitinib, a commonly used first line therapy for metastatic renal cancer. This study was performed as part of the Renal EFFECT Trial testing the efficacy of Sunitinib administered on 4/2 schedule of 50 mg/day for 4 weeks, followed by a two week rest period, versus a continuous dosage schedule of 37.5 mg/day. 292 patients were enrolled in this study and continued treatment for up to two years.

Motzer et al. investigated several previously reported biomarkers linked to tyrosine kinase inhibitor efficacy – single nucleotide polymorphisms (SNPs) in VEGF-A and VEGFR3 (Garcia-Donas et al., 2011, Schneider et al., 2008), HIF1α and CA-IX expression in tumours (Dornbusch et al., 2013, Muriel Lopez et al., 2012), and VHL gene inactivation by mutation, gene deletion or promoter methylation (Moore et al., 2011) – and analysed the levels of serum soluble proteins in patients’ blood before and after treatment.

No statistically significant link was observed between the tumour response to treatment and the VEGF-A or VEGFR3 SNPs analysed, or CA-IX tumour expression. Lower HIF1α tumour expression and VHL inactivation was associated with increased time to tumour progression and progression free survival was observed in the patients receiving Sunitinib on the 4/2 schedule, but not the continuous dosing schedule. Finally, low ANG2 and high MMP2 levels in blood serum before treatment were both associated with increased tumour response to Sunitinib treatment.

Defining a set of biomarkers for all systemic kidney cancer therapies will allow clinicians to choose the best drug for each patient. Furthermore, the fact that HIF1α expression and VHL gene inactivation only correlated with outcomes for patients on the 4/2 dosage schedule suggests that biomarkers may also indicate the optimal dosage schedule, thus allowing a personalised medicine approach for patients with advanced kidney cancer. However, this study did not replicate earlier findings linking CA-IX tumour expression, VEGF-A SNPs and VEGFR3 SNPs to treatment response, suggesting that finding reliable biomarkers will be a difficult undertaking and that multiple biomarkers will be required in order to accurately predict treatment efficacy.

  • Dornbusch J, Zacharis A, Meinhardt M, Erdmann K, Wolff I, Froehner M, Wirth MP, Zastrow S, & Fuessel S (2013). Analyses of potential predictive markers and survival data for a response to sunitinib in patients with metastatic renal cell carcinoma. PloS one, 8 (9) PMID: 24086736
  • Garcia-Donas J, Esteban E, Leandro-García LJ, Castellano DE, del Alba AG, Climent MA, Arranz JA, Gallardo E, Puente J, Bellmunt J, Mellado B, Martínez E, Moreno F, Font A, Robledo M, & Rodríguez-Antona C (2011). Single nucleotide polymorphism associations with response and toxic effects in patients with advanced renal-cell carcinoma treated with first-line sunitinib: a multicentre, observational, prospective study. The Lancet. Oncology, 12 (12), 1143-50 PMID: 22015057
  • Moore LE, Nickerson ML, Brennan P, Toro JR, Jaeger E, Rinsky J, Han SS, Zaridze D, Matveev V, Janout V, Kollarova H, Bencko V, Navratilova M, Szeszenia-Dabrowska N, Mates D, Schmidt LS, Lenz P, Karami S, Linehan WM, Merino M, Chanock S, Boffetta P, Chow WH, Waldman FM, & Rothman N (2011). Von Hippel-Lindau (VHL) inactivation in sporadic clear cell renal cancer: associations with germline VHL polymorphisms and etiologic risk factors. PLoS genetics, 7 (10) PMID: 22022277
  • Motzer et al., PMID: 25100134
  • Muriel López C, Esteban E, Astudillo A, Pardo P, Berros JP, Izquierdo M, Crespo G, Fonseca PJ, Sanmamed M, & Martínez-Camblor P (2012). Predictive factors for response to treatment in patients with advanced renal cell carcinoma. Investigational new drugs, 30 (6), 2443-9 PMID: 22644070
  • Schneider BP, Wang M, Radovich M, Sledge GW, Badve S, Thor A, Flockhart DA, Hancock B, Davidson N, Gralow J, Dickler M, Perez EA, Cobleigh M, Shenkier T, Edgerton S, Miller KD, & ECOG 2100 (2008). Association of vascular endothelial growth factor and vascular endothelial growth factor receptor-2 genetic polymorphisms with outcome in a trial of paclitaxel compared with paclitaxel plus bevacizumab in advanced breast cancer: ECOG 2100. Journal of clinical oncology : official journal of the American Society of Clinical Oncology, 26 (28), 4672-8 PMID: 18824714

www.bhdsyndrome.org – the primary online resource for anyone interested in BHD Syndrome.

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Guidelines for HLRCC kidney cancer risk, surveillance and treatment published

Hereditary leiomyomatosis and renal cell cancer (HLRCC) is a rare kidney cancer susceptibility syndrome caused by autosomal dominant mutations in the FH gene. The three main symptoms of HLRCC are red skin papules called cutaneous piloleiomyomas; multiple early-onset uterine leiomyomas; and susceptibility to type 2 papillary renal cell carcinoma.

Clinical guidelines for HLRCC were the subject of a panel discussion at the 5th BHD and 2nd HLRCC Symposium in Paris last summer. The panel was led by Professor Fred Menko, and included Professor Eamonn Maher, Professor Stéphane Richard, Dr W. Marston Linehan, Dr Laura Schmidt and Graham Lovitt, chairman of the HLRCC Family Alliance. The results of this discussion have now been published in Familial Cancer (Menko et al., 2014).

Menko et al. suggest that patients who have histologically confirmed multiple cutaneous piloleiomyomas, or at least two of the following – symptomatic uterine leiomyomas before age 40, type 2 papillary carcinoma before age 40, or a first-degree relative who meets one of these criteria –  meet clinical diagnostic criteria and should be  referred for genetic testing where possible. Up to 24% of families with clinical features of HLRCC have been reported to not have a detectable FH mutation. In these families, immunohistochemical staining of tumours to demonstrate increased protein succination can confirm a diagnosis of HLRCC.

Expert opinion suggests that 15% of HLRCC patients are at risk of developing kidney cancer, most commonly type 2 papillary renal cell carcinoma. The mean age of diagnosis is 41 years, with a range of 11 to 90 years of age. While the risk is low, an estimated 1-2% of patients of HLRCC patients developing kidney cancer before the age of 20, and HLRCC patients as young as 10 have presented with kidney cancer.

Given the aggressive nature of HLRCC-associated kidney cancer, and the fact it can develop at a young age, Menko et al. suggest that DNA testing should be considered from the age of 8-10, although decisions should be made on an individual basis in collaboration with the family. Tumour surveillance should be offered annually from this age to children with a confirmed mutation and to those who are at risk of inheriting HLRCC, but who have not undergone gene testing. MRI is the preferred screening method, using 1-3mm slices through the kidneys in order to find small tumours.

HLRCC renal tumours are usually unilateral and solitary. Tumours tend to be more aggressive, with an increased chance of metastasising, even when the tumour is small, meaning that the 3cm rule and nephron sparing surgery used to manage BHD and VHL tumours is not appropriate. In HLRCC, once a tumour is found, the tumour should be promptly resected with wide surgical margins, and retroperitoneal lymphadenectomy should be considered. Where there is doubt that a partial nephrectomy would be curative, radical nephrectomy should be performed. In the authors’ experience, patients had a good prognosis and showed no evidence of disease when tumours were found early and managed surgically.

Loss of FH leads to dysregulation of the TCA cycle and glycolysis and several therapies targeting these pathways have been recently developed, which may be appropriate systemic treatments for HLRCC patients with metastatic disease. However, access to these treatments is currently only available through clinical trials.

While publication of these guidelines will help clinicians diagnose HLRCC patients and to manage their renal tumours optimally, the authors discuss the need for clinicians to share data internationally in order to ensure that these guidelines meet the needs of HLRCC patients, and can be refined if necessary. In particular, more data about childhood cases of HLRCC renal cancer would help determine the best age to perform germline genetic testing and to start tumour surveillance.

 

  • Menko FH, Maher ER, Schmidt LS, Middelton LA, Aittomäki K, Tomlinson I, Richard S, & Linehan WM (2014). Hereditary leiomyomatosis and renal cell cancer (HLRCC): renal cancer risk, surveillance and treatment. Familial cancer PMID: 25012257

www.bhdsyndrome.org – the primary online resource for anyone interested in BHD Syndrome.

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Doctors should consider germline genetic testing in kidney cancer patients under 46

It is estimated that between 5 and 8% of kidney cancers are hereditary (Linehan et al., 2010). There are several clues that help clinicians diagnose these cases: patients who present with multifocal and/or bilateral tumours; who have a family history of the disease; or who are younger than the average age of onset are more likely to have inherited a genetic susceptibility to their cancer.

Correctly diagnosing hereditary cancer cases is important as it may change the chosen treatment pathway, suggest a patient’s prognosis and future risk of recurrence, it allows other family members at risk of developing the disease to be identified, and has implications for family planning.

For some cancers, specific guidelines exist to help doctors identify hereditary cancer cases. For example, in the UK, if a patient, or one of their first degree relatives, presents with breast cancer before the age of 40, the patient should be referred for genetic testing (NICE guidelines, 2013). Currently there are no similar guidelines for hereditary kidney cancers, meaning that referral for genetic testing relies on the patient having a switched on clinician.

A recent study from Dr W. Marston Linehan’s group at the National Cancer Institute (NCI), in the States suggests that germline genetic testing should be considered for all patients who present with kidney cancer before the age of 46 (Shuch et al., 2014).

The authors first analysed the mean age of initial diagnosis in over 100,000 kidney cancer cases from the Surveillance, Epidemiology and End Results (SEER) Program database held at the NCI. The mean age of first kidney cancer diagnosis was found to be 63.4 years old. Subtype analysis of gender, tumour histology, and race showed slight variances in mean age of diagnosis, but these were not significantly different from the overall mean, thus Shuch et al. did not include these factors in their subsequent analyses to define an age threshold for genetic testing.

Next they reviewed the average age of diagnosis of 608 patients with hereditary kidney cancers, recruited through the Clinical Manifestations and Molecular Bases of Heritable Urologic Malignant Disorders trial at the NCI. The dataset comprised 387 VHL patients, 127 BHD patients, 56 HLRCC patients, 25 HPRC patients, and 13 SDH patients, and to be included in the trial, patients needed to have renal cell carcinoma, a clinical diagnosis of one of these syndromes and/or a genetic diagnosis. Overall, the mean age of this cohort’s first kidney cancer diagnosis was 39.2 years, meaning that, on average, patients with hereditary kidney cancers get their first tumour nearly 25 years earlier than patients with sporadic cancers.

In order to define an age threshold before which genetic testing should be performed, Shuch et al. analysed what percentage of hereditary kidney cancer patients would be identified and the number of patients needed to test in order to find a single patient with hereditary kidney cancer. Testing at an earlier age means that while a larger proportion of those tested are hereditary cancer patients, any hereditary cases that develop after that age will be missed. Conversely, testing at a later age means that although more hereditary cases will have had time to manifest, the concomitant increase in sporadic cases means that more patients would have to undergo unnecessary genetic testing in order to identify the hereditary cases.

Thus, the authors found that testing all patients at the age of 46 and under struck an optimal balance between sensitivity whilst ensuring the number of patients tested unnecessarily remained low. With a cut off age of 46, 70% of hereditary cases had manifested by this age, and an estimated 7- 28 individuals are needed to test in order to find a single case of hereditary kidney cancer.

There are a number of confounding factors in this study. The hereditary cancer cohort only included five types of kidney cancer syndrome – and in particular renal cell carcinoma syndromes – meaning that this study is not directly relevant to other kidney cancer syndromes such as TSC. Additionally, it is possible that some cases in the control cohort were undiagnosed cases of familial kidney cancers, which would artificially lower the average age of cancer onset in this cohort. Furthermore, subtype analysis of the hereditary kidney cancer cohort showed that the average age of onset in BHD is later than the others at 50.3, meaning that many BHD cases might not get picked up with routine screening.

Thus, although this study gives clinicians a starting point at which to consider genetic testing in younger kidney cancer patients, a cut of age of 46 should only be considered to be a rough guide.

 

  • Linehan WM, Srinivasan R, & Schmidt LS (2010). The genetic basis of kidney cancer: a metabolic disease. Nature reviews. Urology, 7 (5), 277-85 PMID: 20448661
  • Shuch B, Vourganti S, Ricketts CJ, Middleton L, Peterson J, Merino MJ, Metwalli AR, Srinivasan R, & Linehan WM (2014). Defining early-onset kidney cancer: implications for germline and somatic mutation testing and clinical management. Journal of clinical oncology : official journal of the American Society of Clinical Oncology, 32 (5), 431-7 PMID: 24378414

www.bhdsyndrome.org – the primary online resource for anyone interested in BHD Syndrome.

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Investing in cancer research pays off

In the UK, a significant amount of medical research funding – an estimated £2.9 billion in 2012/13 – comes from the public purse through taxes and charitable donations, or from the Wellcome Trust, so it is important to see whether the resultant health benefits justify this spending. However, conclusively proving that funding medical research benefits society financially is incredibly difficult.

In 2008, the Health Economics Research Group, Office of Health Economics and RAND Europe published the results of a study investigating the financial benefits of investing in cardiovascular disease research, and in mental health research in the UK. This study found that the rate of return was 9% for cardiovascular disease and 7% for mental health, meaning that for each £1 spent on cardiovascular or mental health research, benefits to health costs – for example cheaper treatment, or more effective disease prevention so services were ultimately used less – were 9p and 7p every year in perpetuity respectively. These were both at least double the UK government’s minimum recommended threshold of 3.5%, indicating that the government should view investment in these areas positively.

Building on the methodologies developed for the 2008 study, Glover et al. (2014) investigated the economic value of investing in cancer research. This study was funded by the Wellcome Trust, Cancer Research UK, the National Institute for Health Research, and the Academy of Medical Sciences.

Using data from the National Cancer Research Institute (NCRI) Cancer Research Database, they found that 10 funders, including the Medical Research Council and the Wellcome Trust, accounted for over 95% of the research conducted at the NCRI between 2002 and 2011. Funding data covering the 30 year period between 1970 and 2009 were compiled for these 10 funders, and showed that in this time together they had spent roughly £15 billion (adjusted to 2011/2012 prices) on all cancer-related  research activity.

Next the authors determined which cancer interventions had yielded the greatest difference to health outcomes in the last 20 years. They found the seven major interventions were smoking cessation/ prevention programmes; breast, cervical and bowel cancer screening programmes; and improved breast, prostate and colorectal cancer treatments. In particular, anti-smoking schemes provided 65% of the net monetary benefit seen in this time, making it the most effective cancer intervention analysed.

In order to determine what contribution UK research had to these interventions, the authors analysed the citations used in 31 clinical guidelines commonly used in the UK, and assessed which citations resulted from UK funding. The proportion of financial health gains due to UK-funded research was found to average 17%. However, this analysis does not take into government schemes, such as awareness campaigns or anti-smoking messages on cigarette packages.

The net monetary benefit (NMB) of the seven interventions was sensitive to the quality-adjusted life year (QALY) value.  When the QALY was set at £25,000, all seven interventions were found to show a NMB. However, when the QALY were reduced to £20,000, prostate and colorectal cancer treatments, and breast cancer screening did not yield financial gains on investment. This indicates that the financial gains of these interventions are more marginal, and also illustrates the sensitivity of the analysis.

The overall rate of return for UK government and charitable investment in cancer research was estimated to be 10.1%. However, the average time elapsed between investment and patient benefit was 15 years, meaning that investment in cancer research is a long term one.

This study focusses on common cancers and lifestyle factors, such as breast cancer and smoking. However, research on rare diseases can yield benefits for both rare and common diseases. Rare diseases are often more extreme forms of common disorders, and usually have a more straightforward aetiology, making them easier to investigate than their common counterparts. Research on the rare disease hypercholesterolemia led to the development of statins, which are now commonly prescribed in the UK for high cholesterol, and provide sales revenues worth billions of dollars; and the rare disease alkaptonuria is a good model for  osteoarthritis (Taylor et al., 2011), which affects 1 in 3 people over the age of 45. Additionally, rare diseases are often chronic and disabling, meaning that medical and social care can be costly, both to the patient and to the NHS.

Investment in cardiovascular, mental health and cancer research have all proven to have a positive rate of return on investment. Additionally, increased public investment in research often yields sufficient intellectual property or innovation to attract private investment, further increasing the health benefits gained from public investment (Haskel et al., 2014, a report for the Campaign for Science and Engineering).

Thus, increased spending on rare disease research may prove to be a wise investment for the UK government and charity sector, as developing treatments for rare diseases will not only benefit rare diseases patients themselves, but also has the potential to benefit the wider population with more common forms of those diseases.

 

  • Glover M, Buxton M, Guthrie S, Hanney S, Pollitt A, & Grant J (2014). Estimating the returns to UK publicly funded cancer-related research in terms of the net value of improved health outcomes. BMC medicine, 12 PMID: 24930803
  • Taylor AM, Boyde A, Wilson PJ, Jarvis JC, Davidson JS, Hunt JA, Ranganath LR, & Gallagher JA (2011). The role of calcified cartilage and subchondral bone in the initiation and progression of ochronotic arthropathy in alkaptonuria. Arthritis and rheumatism, 63 (12), 3887-96 PMID: 22127706

www.bhdsyndrome.org – the primary online resource for anyone interested in BHD Syndrome.

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BHD causes pneumothoraces during childhood in rare cases

Birt-Hogg-Dubé Syndrome is caused by inactivating mutations in the FLCN gene, characterised by skin lesions on the face and upper body; lung cysts and predisposition to pneumothorax; and kidney cancer. Although symptoms typically appear in the third and fourth decade of life, Johannesma et al. (2014) suggest that pneumothorax in patients under the age of 18 might be underdiagnosed.

In two large cohort studies, 24% were found to have had one or more episodes of pneumothorax, with a predicted lifetime risk of29% of developing a pneumothorax (Houweling et al., 2011, Toro et al., 2007). The age range of patients’ initial pneumothorax episodes was 18 – 74 years, with a median age of 36 – 38 years, suggesting that the age of onset of this symptom can vary widely (Houweling et al., 2011, Toro et al., 2007). However, the majority of families in both of these studies were recruited via dermatology clinics, meaning that there might be some ascertainment bias in these cohorts.

Johannesma et al. report the cases of two unrelated Dutch BHD patients who suffered their first pneumothorax at the age of 14. Both patients were given chest CT scans, as they suffered recurrent episodes as teenagers, and revealed the presence of lung cysts in the lower lobes of the lungs. Genetic testing confirmed each had a pathogenic FLCN mutation, confirming a diagnosis of BHD.

This report follows three other reports of BHD patients suffering pneumothoraces as teenagers: a 7 year old French boy (Bessis et al., 2006); a 16 year old Japanese girl (Gunji et al., 2007); and an 18 year old Dutch girl (Houweling et al., 2011). Thus, although pneumothorax during childhood is rare in BHD patients, both clinicians and BHD patients with children should be aware that it is a possibility.

It is possible that BHD is underdiagnosed in cases of paediatric spontaneous pneumothorax as BHD is normally considered to be an adult-onset disease. Additionally, CT scans – which would help diagnose BHD if lung cysts were found – are not normally administered to paediatric patients, unless deemed to be absolutely necessary, due to the low dose of radiation patients are exposed to during the scan (Balfour-Lynn et al., 2005).

Given that both patients reported by Johannesma et al. were found to have lung cysts as teenagers but neither had yet developed skin or kidney symptoms, it is possible that lung cysts are the first symptom of BHD to develop, but are not normally found unless the patient receives a chest CT for some other reason. Indeed, pneumothorax risk is strongly correlated with increased number and size of lung cysts (Toro et al., 2007), and it has subsequently been hypothesised that lung collapses are caused by air-filled cysts rupturing and releasing air into the chest cavity (Furuya and Nakatani, 2013). Thus it is likely that all the reported paediatric patients also developed lung cysts in childhood.

Four of the five paediatric BHD patients did have recurrent episodes, with one patient having four recurrent lung collapses before the age of 18 (Johannesma et al. 2014), and another having six episodes between the ages of 16 and 38 (Gunji et al., 2007). Given that the average number of pneumothoraces per patient was found to be two in the cohort studied by Toro et al. this does suggest that BHD patients who have their first pneumothorax in childhood might be at higher risk of recurrent episodes than other BHD patients.

Thus, although BHD patients suffer pneumothoraces during childhood only in rare cases, as childhood pneumothorax is itself rare, only affecting 4 in 100,000 boys and 1 in 100,000 girls, Johannesma et al. recommend that a thorough family history, a low dose CT scan, and genetic testing should be considered for paediatric patients who experience repeated episodes of spontaneous pneumothorax.

 

  • Balfour-Lynn IM, Abrahamson E, Cohen G, Hartley J, King S, Parikh D, Spencer D, Thomson AH, Urquhart D, & Paediatric Pleural Diseases Subcommittee of the BTS Standards of Care Committee (2005). BTS guidelines for the management of pleural infection in children. Thorax, 60, i1-21 PMID: 15681514
  • Bessis D, Giraud S, & Richard S (2006). A novel familial germline mutation in the initiator codon of the BHD gene in a patient with Birt-Hogg-Dubé syndrome. The British journal of dermatology, 155 (5), 1067-9 PMID: 17034545
  • Furuya M, & Nakatani Y (2013). Birt-Hogg-Dube syndrome: clinicopathological features of the lung. Journal of Clinical Pathology, 66 (3), 178-86 PMID: 23223565
  • Gunji Y, Akiyoshi T, Sato T, Kurihara M, Tominaga S, Takahashi K, Seyama K. (2007) Mutations of the Birt Hogg Dube gene in patients with multiple lung cysts and recurrent pneumothorax. Journal of Medical Genetics, 44 (9), 588-93 PMID: 17496196
  • Houweling AC, Gijezen LM, Jonker MA, van Doorn MB, Oldenburg RA, van Spaendonck-Zwarts KY, Leter EM, van Os TA, van Grieken NC, Jaspars EH, de Jong MM, Bongers EM, Johannesma PC, Postmus PE, van Moorselaar RJ, van Waesberghe JH, Starink TM, van Steensel MA, Gille JJ, & Menko FH (2011). Renal cancer and pneumothorax risk in Birt-Hogg-Dubé syndrome; an analysis of 115 FLCN mutation carriers from 35 BHD families. British journal of cancer, 105 (12), 1912-9 PMID: 22146830
  • Johannesma PC, van den Borne BE, Gille JJ, Nagelkerke AF, van Waesberghe JT, Paul MA, van Moorselaar RJ, Menko FH, & Postmus PE (2014). Spontaneous pneumothorax as indicator for Birt-Hogg-Dubé syndrome in paediatric patients. BMC pediatrics, 14 PMID: 24994497
  • Toro JR, Pautler SE, Stewart L, Glenn GM, Weinreich M, Toure O, Wei MH, Schmidt LS, Davis L, Zbar B, Choyke P, Steinberg SM, Nguyen DM, & Linehan WM (2007). Lung cysts, spontaneous pneumothorax, and genetic associations in 89 families with Birt-Hogg-Dubé syndrome. American journal of respiratory and critical care medicine, 175 (10), 1044-53 PMID: 17322109

 www.bhdsyndrome.org – the primary online resource for anyone interested in BHD Syndrome.

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Intratumoral peripheral small papillary tufts might be a diagnostic clue of BHD

Contrary to other hereditary kidney cancer conditions, BHD-associated renal tumours are known to be histologically diverse and discordant within families, meaning that members of the same family do not necessarily get the same type of kidney tumour (Pavlovich et al., 2005). Kuroda et al. (2014) recently reported on the histology of tumours from six BHD patients, and found that, despite being histologically diverse, all had intratumoral peripheral small papillary tufts – clusters of papillary cells – suggesting that these microscopic lesions might be a hallmark of BHD.

The authors selected BHD kidney tumours from six patients who were clinically diagnosed with BHD at Kochi Red Cross Hospital in Japan, between January 2010 and October 2013. These six cases correspond to five families and three of these cases have been previously reported (Furuya et al., 2012, Nagashima et al., 2012). Germline DNA samples were available for genetic analysis for five cases, and a pathogenic  FLCN mutation was found in all five. No sample was available for the sixth patient, so a genetic diagnosis could not be made for this patient, although she has been clinically diagnosed with BHD.

The series consisted of three men and three women, and the average age at diagnosis was 60. One patient had a history of spontaneous pneumothorax, three patients had skin lesions, and all six had lung cysts. All patients are alive without disease following treatment, and show no signs of recurrence or disease in a follow up period ranging between 10 and 46 months.

Five patients had multi-focal kidney tumours, and two had bilateral kidney tumours. Only one patient had a solitary tumour, which may be because she was the youngest patient in this cohort at 46 years old.  The tumours analysed in this study consisted of three hybrid oncocytic/chromophobe tumours; a tumour of unclassified histology, but with features resembling hybrid chromophobe/ clear cell histology; a collision tumour consisting of chromophobe, clear cell and papillary primary tumours; a chromophobe tumour; and a clear cell tumour. This wide range of tumour histologies in such a small sample series supports previous observations that BHD renal tumours are highly variable.

However, small papillary tufts were found in all samples. These papillary tufts were present mainly at the interface between the tumour and normal kidney tissue, or in the case of the collision tumour, at the interface between the chromophobe and clear cell tumours. Given their histology and their presence at the periphery of tumours, the authors termed these microscopic lesions intratumoral peripheral small papillary tufts (ITPSPTs). The authors hypothesise that these microscopic lesions could be precursor lesions that subsequently develop into tumours.

Although ITPSPTs have not been previously reported, similar lesions were been reported in patient with acquired renal cystic disease (Cheuk et al., 2002). Although cytogenetic analysis was performed on these papillary tufts, germline or tumour DNA sequencing for FLCN mutations was not performed. Thus it is possible this patient was an undiagnosed BHD patient, as they had multiple kidney cysts and a tumour with both clear cell and papillary histology.

As ITPSPTs were found in all tumours in their cohort, Kuroda et al. suggest that the presence of ITPSPTs might be a hallmark of BHD tumours, and therefore a diagnostic clue. Analysing ITPSPTs in a larger series of BHD renal tumours and control non-BHD kidney tumours, is required to confirm that ITPSPTs are specific to BHD tumours. They also suggest that patients who present with collision tumours or multiple tumours of different histologies may have BHD, and that these patients should be referred for genetic testing.

 

  • Cheuk W, Lo ES, Chan AK, & Chan JK (2002). Atypical epithelial proliferations in acquired renal cystic disease harbor cytogenetic aberrations. Human pathology, 33 (7), 761-5 PMID: 12196929
  • Furuya M, Tanaka R, Koga S, Yatabe Y, Gotoda H, Takagi S, Hsu YH, Fujii T, Okada A, Kuroda N, Moritani S, Mizuno H, Nagashima Y, Nagahama K, Hiroshima K, Yoshino I, Nomura F, Aoki I, & Nakatani Y (2012). Pulmonary cysts of Birt-Hogg-Dubé syndrome: a clinicopathologic and immunohistochemical study of 9 families. The American journal of surgical pathology, 36 (4), 589-600 PMID: 22441547
  • Kuroda N, Furuya M, Nagashima Y, Gotohda H, Moritani S, Kawakami F, Imamura Y, Bando Y, Takahashi M, Kanayama HO, Ota S, Michal M, Hes O, & Nakatani Y (2014). Intratumoral peripheral small papillary tufts: a diagnostic clue of renal tumors associated with Birt-Hogg-Dubé syndrome. Annals of diagnostic pathology, 18 (3), 171-6 PMID: 24767893
  • Nagashima Y, Furuya M, Gotohda H, Takagi S, Hes O, Michal M, Grossmann P, Tanaka R, Nakatani Y, & Kuroda N (2012). FLCN gene-mutated renal cell neoplasms: mother and daughter cases with a novel germline mutation. International journal of urology : official journal of the Japanese Urological Association, 19 (5), 468-70 PMID: 22211584
  • Pavlovich CP, Grubb RL 3rd, Hurley K, Glenn GM, Toro J, Schmidt LS, Torres-Cabala C, Merino MJ, Zbar B, Choyke P, Walther MM, & Linehan WM (2005). Evaluation and management of renal tumors in the Birt-Hogg-Dubé syndrome. The Journal of urology, 173 (5), 1482-6 PMID: 15821464

www.bhdsyndrome.org – the primary online resource for anyone interested in BHD Syndrome.

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FLCN is important for cardiomyocyte development

Cardiac hypertrophy is an adaptive response that occurs following increased stress on the heart wall, and can be caused by strenuous exercise, hypertension, heart attack or heart valve disease. In some cases, this can lead to heart failure.

Although the biological mechanism underlying cardiac hypertrophy is not fully elucidated, dysregulation of the AMPK-mTOR signalling pathway is known to play a role (Maillet et al., 2013). Hasumi et al. (2014) decided to investigate the role of FLCN in cardiac hypertrophy, given FLCN’s role in AMPK and mTOR signalling, by generating mice with both copies of FLCN specifically deleted in heart and skeletal muscle using the CKM-Cre driver mouse strain.

Mice lacking FLCN in heart and skeletal muscle had cardiac hypertrophy as shown by enlarged hearts, severe cardiac dysfunction and reduced lifespan. In vitro data from mouse embryonic fibroblasts (MEFs), showed that loss of FLCN led to increased mTOR signalling, increased protein synthesis and decreased autophagy, suggesting that dysregulated mTOR signalling may be responsible for the heart pathology in the FLCN knockout mice. In support of this hypothesis, Rapamycin treatment significantly reduced heart size and improved cardiac function.

However, mice lacking both the FLCN and the PCG1a gene showed no cardiac hypertrophy, indicating that the phenotype was mediated entirely through PCG1a. Increased PGC1a expression lead to increased mitochondrial respiration, with increased the amount of intracellular ATP, which  inhibited AMPK activity, and ultimately led to mTOR dysregulation.

The results of this study correspond with those of several others. Goncharova et al. found that loss of FLCN in alveolar type II cells lead to a decrease in E-cadherin expression, reduced LKB1 signalling, and a consequent reduction in AMPK activity. Interestingly, mice lacking functional LKB1 also develop cardiac hypertrophy (Ikeda et al., 2009). Furthermore, Kumasaka et al. recently suggested that heterozygous loss of FLCN in the lung makes alveoli weaker and susceptible to mechanical stress during breathing, which may cause cysts to develop. Taken together, is it possible that reduced E‑cadherin expression and subsequent reduced AMPK activity via reduced LKB1 activity caused by FLCN depletion make cardiac and lung tissues vulnerable to mechanical stress, thus causing cardiac hypertrophy and lung cysts to develop.

Recently Yan et al. also noted increased PCG1a expression, increased mitochondrial mass, and increased ATP production in FLCN-null cells. Yan et al., however, found constitutive AMPK activity to be upstream of PCG1a hyperactivity, rather than downstream. Furthermore, conversely to the data reported here by Hasumi et al., which suggest that FLCN inhibits mTOR signalling, last year two studies reported that FLCN activates mTOR signalling at the lysosome following amino acid restimulation.

The role of FLCN in AMPK-mTOR signalling has been difficult to define, with a number of contradicting reports. While some of these differences are likely to be due to cell specific effects, due to the role of AMPK, mTOR and FLCN in energy sensing, a concerted effort to investigate different culture conditions on FLCN-null cells may also shed light on this issue. Furthermore, systematic analysis of sub-cellular FLCN localisation and which proteins FLCN interacts with in different cell types, in different sub-cellular locations and under different culture conditions will also be valuable to delineate this pathway.

Cardiac hypertrophy is not a symptom of BHD, and the mice studied by Hasumi et al. had both copies of FLCN deleted, whereas BHD patients are heterozygous for FLCN inactivation. Thus BHD patients should not be concerned about developing cardiac hypertrophy. However, it would be interesting to determine the proportion of cardiac hypertrophy patients with sporadic FLCN inactivation to assess the clinical significance of this finding.

 

  • Goncharova EA, Goncharov DA, James ML, Atochina-Vasserman EN, Stepanova V, Hong SB, Li H, Gonzales L, Baba M, Linehan WM, Gow AJ, Margulies S, Guttentag S, Schmidt LS, & Krymskaya VP (2014). Folliculin controls lung alveolar enlargement and epithelial cell survival through E-cadherin, LKB1, and AMPK. Cell reports, 7 (2), 412-23 PMID: 24726356
  • Hasumi Y, Baba M, Hasumi H, Huang Y, Lang M, Reindorf R, Oh HB, Sciarretta S, Nagashima K, Haines DC, Schneider MD, Adelstein RS, Schmidt LS, Sadoshima J, & Marston Linehan W (2014). Folliculin (Flcn) inactivation leads to murine cardiac hypertrophy through mTORC1 deregulation. Human molecular genetics PMID: 24908670
  • Ikeda Y, Sato K, Pimentel DR, Sam F, Shaw RJ, Dyck JR, & Walsh K (2009). Cardiac-specific deletion of LKB1 leads to hypertrophy and dysfunction. The Journal of biological chemistry, 284 (51), 35839-49 PMID: 19828446
  • Kumasaka T, Hayashi T, Mitani K, Kataoka H, Kikkawa M, Tobino K, Kobayashi E, Gunji Y, Kunogi M, Kurihara M, & Seyama K (2014). Characterization of pulmonary cysts in Birt-Hogg-Dubé syndrome: histopathological and morphometric analysis of 229 pulmonary cysts from 50 unrelated patients. Histopathology, 65 (1), 100-10 PMID: 24393238
  • Maillet M, van Berlo JH, & Molkentin JD (2013). Molecular basis of physiological heart growth: fundamental concepts and new players. Nature reviews. Molecular cell biology, 14 (1), 38-48 PMID: 23258295
  • 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 (2014). The tumor suppressor folliculin regulates AMPK-dependent metabolic transformation. The Journal of clinical investigation, 124 (6), 2640-50 PMID: 24762438

www.bhdsyndrome.org – the primary online resource for anyone interested in BHD Syndrome.

pf button FLCN is important for cardiomyocyte development