BHD Toolkit: Understanding your Genetic Test Result – Amino Acid Mutations

Our last BHD toolkit provided you with all the information needed to understand and interpret your DNA sequencing result, however it didn’t go into details on how to understand the mutation at the protein level.

On your test result this may look something like p.(Trp511*) or something a little more complicated like p.(His429Profs*27).

Please note that your result may look different depending on the test provider. If there is something on your result you don’t understand, please ask your clinician or genetic counsellor, or contact us for further help.

From DNA to Protein

DNA is a unique code that contains all the instructions required for making the proteins needed for the development, growth and function of our bodies. The building blocks of our DNA are called nucleotides. There are four of these made of the bases: adenine, cytosine, guanine and thymine. The sequence of these determines our unique genetic code and tells the machinery in our cells what proteins to make, through an intermediate molecule called RNA. RNA carries the message of the DNA and allows it to be translated into proteins. It is very similar to DNA, however a base called uracil, replaces the base thymine.  

Image shows the flow of information from DNA to RNA (transcription) and RNA to Protein (translation).

Like DNA and RNA, proteins are also made of building blocks. These are not like those in DNA and RNA and are called amino acids. Our DNA is read in triplets: 3 bases (also called a codon) code for 1 amino acid. There are 20 different amino acids that can be made from the following different combinations:

Image shows which combination of nucleotides produce each amino acid. A key explaining the abbreviations for the amino acids is on the right.

There are also 3 codons that do not code for an amino acid and instead signify for the protein to stop being made.

Interpreting Protein Mutations
Interpreting your protein mutation is very similar to interpreting your DNA mutation – you just need to know how to break it down into pieces!

Firstly the p. at the beginning tells us that this result is at the protein level.

The next three letters refer to the amino acid that is normally there in a non-mutated version of folliculin (see the above image).

The numbers that follow this refer to the position of the amino acid in the sequence.

The next bit(s) tell us what has happened to that particular amino acid at the position, for which there are a few different options

  1. The numbers may be followed by ‘del’, which doesn’t stand for an amino acid and means deletion.
    e.g. p.(Phe157del) – the amino acid phenylalanine at position 157 has been deleted.
  2. It is also possible for there to be a larger deletion.
    e.g. p.(His111_Gln116del) – the amino acids from position 111 through 116 have been deleted.
  3. There may also be another three letters referring to a different amino acid.
    e.g. p.(Glu434Lys) – a Glutamine at position 434 has been swapped to a Lysine.
  4. There also may be no letters after the numbers and there is only an *.
    e.g. p.(Trp511*) – the amino acid tryptophan at position 511 has been mutated to a stop codon and the rest of the protein will not be made.

It also possible that the result will look even more complicated as suggested above such as p.(His429Profs*27). Based on the above information it can be inferred that the amino acid histidine (His) at position 429 has been changed to a proline (Pro). However, the remaining fs*27 is yet to be explained.

The fs stands for frameshift. As our DNA is read in triplets, if a single amino acid is deleted or inserted it knocks the reading of triplets out of frame and means that the triplets are not read as they were designed. This can result in the introduction of a premature stop codon, as is the case with the p.(His429Profs*27). The fs*27 indicates that a frameshift has introduced a stop codon (*) 27 amino acids after the mutation.

FLCN mutation
DNA: c.1285dup
Protein: p.(His429Profs*27)

FLCN Genetic Code: 
An image of the genetic code of folliculin from amino acids 420 to 430, with amino acid 429 (Histidine) highlighted.
Mutated FLCN Genetic Code:
An image of the genetic code of folliculin from amino acids 420 to 430, with amino acid 429 (Proline) highlighted. 
Accompanying text: The  C at nucleotide position 1285 has been duplicated which has resulted in the 429th amino acid changing from a histidine to a proline.
The next image shows the FLCN genetic code from 445 - 455.
Accompanying text: Looking further down the sequence we can see that a stop codon (*) has been introduced at amino acid 455 - 27 amino acids down from the original histidine (inclusive of the histidine itself).

It is also possible that your sequencing result may look like p.? – in which case the change at the protein level is unknown and cannot be predicted. This occurs when the type of DNA mutation is a splice-site mutation. You can read our previous BHD Toolkit post which explains the different types of DNA mutations here.

We hope you enjoy these toolkit posts and find them useful. If there is any aspect of BHD you would like us to do a deep dive on and explain in more detail, please let us know by email.

Winter BHD Case Report Round-Up

This week’s blog post contains a round-up of the latest case reports for Birt-Hogg-Dubé Syndrome (BHD) in the literature.

Case Report 1

Firstly, a case report by Degheili et al., documents BHD in 2 individuals with kidney cysts but no kidney tumours. As opposed to tumours, which are solid masses of unusual tissue, cysts are small sacs normally filled with air or fluid. Kidney cysts do not normally impact the function of the kidney but in rare cases can grow large enough to cause pain or discomfort and can block the flow of urine through the kidneys.

The first case was a 70-year-old woman referred for investigation of cysts on her lungs. The patient also had innumerable pale skin bumps on her face and upper torso which were later confirmed to be fibrofolliculomas. A CT scan of her kidneys revealed bilateral cysts and were considered simple. The patient was followed for four years and both lung and kidney cysts remained stable, with the formation of no new cysts.

The second case was a 50-year-old woman with a personal and family history of recurrent pneumothoraces, as well as several skin lesions characteristic of BHD. Genetic testing of the folliculin gene confirmed the diagnosis of BHD. CT scans revealed cysts on both her lungs and kidneys with no kidney tumours.

It is questioned whether the presence of kidney cysts is an incidental finding, or a true feature of BHD. Renal cysts are found in as many as 4 in 10 people in the general population. However, these tend to be isolated, unilateral and simple, compared with the multiple bilateral cysts found in these patients, and in the reported literature on kidney cysts in BHD. However, without further documentation of kidney cysts on a larger scale, an association cannot yet be made.

Case Report 2

In this case report by Lakhani et al., BHD was diagnosed through incidental findings on a lung CT scan. A 59-year-old male presented to the hospital following a CT scan of the lung, abdomen and kidney as part of a trauma workup which revealed the presence of lung cysts. A second CT scan of the chest showed multiple, thin-walled, cysts with a basal distribution (i.e. cysts were located at the bottom of the lungs). The patient had no personal or family history of spontaneous pneumothoraces or kidney cancer. Despite this, and due to the characteristics of the cysts in the lung, BHD was suspected and was confirmed by genetic testing.

BHD is thought to remain underdiagnosed, partly due to variation in clinical presentation as well as lack of awareness. This case report highlights the importance of spreading awareness of BHD within the medical community, as lung cysts found incidentally on imaging studies may be the first sign of BHD in many cases. Early diagnosis of BHD can be critical in minimising the risk of kidney cancer. The BHD Foundation is creating a BHD awareness leaflet that will be circulated to the doctors who are likely to come across the first signs of BHD including dermatologists (skin doctors), radiologists (doctors who review images) and lung doctors,

Case Report 3

The final case report documents a 37-year-old male who was admitted to hospital with lower back pain and hematuresis (blood in the urine). The patient had a personal and family history of spontaneous pneumothorax. A CT scan of the kidney showed a mass on the right kidney and robotic-assisted surgery was performed to remove the tumour. It was noted that the tumour had an unusual shape, and upon further investigation of a sample of the tumour tissue, the tumour was considered as an unclassified type of renal cell carcinoma (RCC). This is unusual for individuals with BHD who normally present with chromophobe, oncocytoma or a hybrid chromophobe/oncocytoma type of RCC.

Genetic testing was performed and a mutation in the folliculin gene was found. In BHD, mutation of a single copy of folliculin is sufficient to cause the skin and lung manifestations, but it is thought that a mutation in the second copy is required for the development of kidney cancer. In this report, it appears that they did not observe a mutation in the second copy of folliculin in the kidney tumour tissue, however it is unclear whether any other genes known to be drivers of cancer development were analysed for mutations. If there were no other mutations and only a single copy of folliculin was mutated this report would represent a rare finding of kidney cancer in and individual with BHD.

Case reports such as these are invaluable additions to the BHD scientific literature that can raise questions about uncommon manifestations or highlight different routes to diagnosis that can increase our knowledge and understanding of the condition. As well as this, they can also help to raise awareness of BHD among clinicians which is an important aspect of the work of the BHD Foundation.

BHD Foundation 2021 Highlights

For our first blog post of 2022 we thought we would share some of our personal highlights from last year and how we plan to continue to support the BHD community throughout the next year.

January – March

After a quiet period for the Myrovlytis Trust and BHD Foundation, 2021 marked a new beginning as Anna Webb was appointed as Charity Director in January. Charity Officer Jazzmin Huber joined in February and soon after weekly blog posts and monthly newsletters were re-established.

We also started getting to know some of the BHD community through written and video interviews.

April – June

2 new members of staff joined in this period, Charity Officer Katie Nightingale joined in May, and our Office Manager, Katie Honeywood, completed the team in June.

April marked the first Meet the Expert event with genetic counsellor Lindsay Middleton. This was followed by a second Meet the Expert event in June with Professor Stefan Marciniak who answered all your pneumothorax-related questions.

June also marked the launch of our BHD explainer video (watch it here) and the First World Pneumothorax Day.

July – September

Over the summer months the team were finishing updates to the BHD Foundation website including new printable information leaflets, a BHD medical card and a customisable letter to help you tell family members about BHD.

We also held another Meet the Expert event with Professor Gennady Bratslavsky who shared his clinical and research perspectives on BHD and the kidneys.

October – December

October was a particularly exciting month as we held our first virtual BHD Symposium. Held across 2 days we held both research- and patient-focused sessions and were delighted to have over 240 people register across all sessions. You can watch recordings of our patient sessions here.

Additionally, through the Myrovlytis Trust, researchers and clinicians were able to apply for funding to carry out new research about BHD. Awards were made in November and we will be blogging about our grant awardees very soon!

Personal Highlights

Anna Webb

It has been a great honour to lead the Myrovlytis Trust and BHD Foundation since 2021, reinvigorating our offering and embarking on several new and exciting projects. The BHD community has and always will be at the heart of everything we do, and I am really looking forward to working with clinicians, researchers and families to move the field towards new therapies and eventually a cure. We have big plans for 2022 and are excited to let you know all about them very soon!

Jazzmin Huber

I was delighted to join the Myrovlytis Trust Team last February and the year has flown by. Some of my favourite moments included our first Meet the Expert event. It was so lovely to see the BHD community coming together and to hear about the positive impact Lindsay Middleton had on so many lives. Also, despite never leaving my desk I feel I have travelled all over the world. It has been an absolute pleasure talking with researchers, clinicians and families across the globe who all share a common goal of raising awareness of BHD.

Katie Nightingale

I trained as a scientist and worked in a lab doing research before starting at the Myrovlytis Trust as a Charity Officer in May. I was thrilled to be able to use my scientific training to directly impact individuals affected by BHD. I have really enjoyed leading the development of the BHD patient registry and uniting clinicians, researchers and individuals living with BHD to work collaboratively on this project with the Trust. It is difficult to choose just one highlight from 2021 but I think the BHD Symposium, and particularly the patient session focusing on BHD research was my favourite moment!

Katie Honeywood

I was so pleased to be appointed as Office Manager for the Trust in June 2021. I have really enjoyed getting involved behind the scenes to ensure robust processes are in place to help both staff and Trustees get on with their day-to-day roles. One of the highlights of working for the Trust is patient advocacy which can include anything from helping someone find a specialist to creating the monthly newsletter or even assisting with running the BHD Symposium. There is nothing quite like knowing your work has helped someone to improve and manage their own health conditions.

2022 Plans

This coming year we will continue to support the BHD community as we have done in 2021 through blog posts, patient interviews and Meet the Expert events and build upon our platform with new patient-focused events and resources.

Work started on the development of a BHD patient registry last year, and we will be launching the platform in the coming months. This is a very exciting project for us, and we hope that the data collected through the registry will enable us to help researchers and clinicians answer some of the unknown questions about BHD including determining the prevalence of BHD and the creation of standardised diagnostic and management guidelines.

To find out about upcoming events, receive updates on projects such as the BHD patient registry and discover how you can get involved with our work, sign up to our monthly newsletter.

Our first event of 2022 is a Meet the Expert event with us! On the 15th of January 2022, we will be sharing more of our 2021 achievements as well as exciting projects we have planned throughout 2022 and beyond. Importantly, this is also your chance to share your thoughts on what you would like to see more of in the future to best suit your needs as an individual with BHD. Tickets are limited, so please secure your place by registering for the event now.

The BHD Foundation

BHD Toolkit: Understanding Your Genetic Test Result

This guide will help you understand your genetic test result and how to interpret the “code” you are given when you get your DNA sequencing results back. Please note that your result may look different depending on the test provider. If there is something on your result you don’t understand, please ask your clinician or genetic counsellor, or contact us for further help.

What do the numbers mean?

  • Your DNA sequencing result may look something like c.1285del, c.1429C>T, c.927_954dup, c.610_611delinsTA, or perhaps even something like c.872-?_1062+?del.
  • This “code” relates to the particular mutation in the folliculin (FLCN) gene and enables the classification of the variant as e.g. pathogenic or a variant of unknown significance (VUS).
  • The numbers in the sequencing result relate to how far along in the FLCN gene the mutation occurs e.g. c.1285del indicates the mutation is at nucleotide 1285.
  • Nucleotides are the building blocks of our DNA, and are represented by the letters ‘A’ (adenine) ‘C’ (cytosine) ‘G’ (guanine) and ‘T’ (thymine).

What do the letters mean?

  • There are a few different types of letter(s) found in the sequencing result, which have different meanings.
  • Firstly, the small ‘c.’ at the beginning of the result means a coding DNA reference sequence was used. Not all the nucleotides in a gene are used to make (code) the protein. Some bits, such as introns, are cut out. A coding DNA reference therefore refers to only the nucleotides used to make the protein.
  • If there is an ‘A’, ‘C’, ‘G’ and/or ‘T’ in your result this refers to which nucleotide(s) are altered (e.g. deleted or inserted).
  • Finally, if there is a short word such as ‘del’ or ‘dup’ then this indicates what has happened to cause the mutation. These are explained in the section below.

What do the words mean?

  • del: one or more letters of the DNA code are missing (deleted).
  • dup: one or more letters of the DNA code are present twice (duplicated).
  • ins: one or more letters of the DNA code have been added (inserted).
  • delins: one or more letters of the DNA code are missing (deleted) and have been replaced by one or more new letters (inserted).

What do the symbols mean?

  • There are several different symbols you might see in your sequencing result and have various meanings. The most common ones are as follows:
  • _ (underscore) indicates a range.
  • > (greater than) indicates a substitution from one nucleotide to another one.
  • + (plus) or (minus) indicate the mutation occurs within an intron.
  • ? (question mark) indicates unknown positions.

How do I put it all together?

  • Now you have all the pieces of the puzzle to understand your sequencing result!
  • Here are some examples of FLCN variants found in the LOVD database:
  • c.1285del: The nucleotide at position 1285 has been deleted.
  • c.1429C>T: The nucleotide at position 1429 has been swapped from a C to a T.
  • c.927_954dup: The nucleotides between position 927 and 954 (inclusive) have been duplicated.
  • c.610_611delinsTA: The nucleotides at position 610 and 611 have been deleted and a T and an A have been inserted.
  • c.872-?_1062+?del: There has been a deletion of several nucleotides, although the exact location has not been identified. The start point is located in the intron following the nucleotide at position 872, and the end point is in the intron before nucleotide at position 1062.

What does this all mean?

  • Now you can understand your sequencing result, what does it actually mean?
  • You may see the one of the following terms on your test result.
  • Pathogenic: There is sufficient evidence that this variant is capable of causing the manifestations seen in BHD.
  • Likely Pathogenic: There is strong evidence that this variant is capable of causing manifestations seen in BHD.
  • Variant of Unknown Significance: There is currently not sufficient evidence to suggest this variant is capable of causing the manifestations seen in BHD.
  • These terms do not determine whether or not you will develop any of the manifestations associated with BHD and reflect the degree of evidence for the development of BHD symptoms in individuals with specific variants.
  • It is therefore recommended to inform your family members that you have BHD and undergo regular monitoring of your kidneys.
  • The BHD Foundation website has a letter you can use to inform your family members, and can answer any questions you have surrounding BHD and testing.

You can also download this information as an infographic.

Dieke’s Story: Diagnosis

Dieke was recently diagnosed with BHD, after initially being diagnosed with LAM. In this interview she shares her diagnosis journey, how she manages her condition and what advice she would give to others in similar situations.

You can read this interview in English below, or download a copy in Dutch (Dieke’s first language) here.

When were you first diagnosed with BHD? 

I was diagnosed March 2021. 

How did you get diagnosed and what was the process/how long did it take? 

In 2018 I had stabbing pains in my lower abdomen. Those are still unexplained, but on a CT scan, lung cysts were found by coincidence. The first diagnosis was probable LAM, but I read in my medical report that they could not rule out BHD. I asked my pulmonologist about that, and he agreed to do a genetic test which turned out to be positive for BHD. 

What impact has the diagnosis had on you and your family members?

For me it was a relief to have a definite diagnosis. I always doubted LAM. 

How did you tell your family members about BHD and what was their response? 

I told my family I was getting tested, and that BHD is hereditary. Because my maternal grandfather died of bilateral kidney cancer, I supposed it came from my mother’s side. I did ask my father’s family, but there was nobody who had issues related to BHD. I think my mother only realised it was true when she heard she had indeed BHD herself. My sister tested negative, and my brother is still considering getting tested. My children decided to postpone testing for a while.

What were your first symptoms?  

My first symptoms were the lung cysts.

What other symptoms have you had?  

I do have some fibrofolliculomas on my face, but I just noticed them with my glasses on after diagnosis.

What has been the most challenging aspect of being recently diagnosed BHD? 

It makes a bit of a hypochondriac and insecure about my health sometimes.

How do you manage your symptoms of BHD?  

I do cardiotraining 2-3 times a week and eat healthy, to keep me as fit as possible.

What treatments have you had/are having? 

I was having treatment for LAM until recently, which meant regular blood and pulmonary tests. After my BHD diagnosis, I had a CT scan of my kidneys. Because that showed no issues it was planned that I would have yearly ultrasounds. However, I heard professor Bratslavsky explain at the “Meet the Expert” event, that people with BHD should have MRIs instead. I discussed that with my urologist, and he agreed to that. I will still be having pulmonary tests also. The fibrofolliculomas are almost invisible and do not need treatment at the moment.

What advice would you give to people who have also been recently diagnosed or are in the process of being diagnosed?  

I think it is important to be on top on your own medical situation. I my case, asking about a loose comment about BHD in my medical file led to my diagnosis. Especially with BHD being such a rare disease it is important to inform yourself, because chances are that your doctors have never heard about BHD. The BHD Foundation is a very good source!

What would you like to see for the future of BHD to increase patient quality of life? 

Early diagnosis is key with kidney cancer, so awareness among clinicians is very important. For people with severe issues with fibrofolliculomas it would increase quality of life to have a method to erase or at least make them less visible.

What can the BHD Foundation do to better support the BHD patient community? 

I think the BHD Foundation is doing a great job!

How can we raise awareness of BHD among clinicians? 

We as BHD patients can do our (small) part by pointing clinicians to the BHD Foundation. The BHD Foundation could probably send information about BHD to associations of doctors like genetic counsellors, GP’s, dermatologists etc. They should be able to spread the information.

Is there a Beneficial Role for Loss of FLCN in the Liver?

Birt-Hogg-Dubé Syndrome (BHD) is caused by mutations in the gene folliculin (FLCN). We have previously covered some of the roles FLCN plays within our cells that contributes to the manifestations seen in BHD (link to blogs). Most of the research done on FLCN has been focused on the role of FLCN in the kidneys or lungs as these are the tissues most relevant to BHD. However, FLCN is expressed in the majority of our tissues and so it may be important to understand the role FLCN plays in other parts of the body. A recent study by Paquette et al., has uncovered a surprising role for FLCN in the liver in the context of fatty liver disease progression.

Non-alcoholic fatty liver disease (NAFLD) is the most frequent liver disease worldwide and is associated with obesity, type 2 diabetes and dyslipidemia (unhealthy levels of fat in the blood e.g. high cholesterol). Around one third of patients will progress to non-alcoholic steatohepatitis (NASH), which is characterised by liver damage, persistent inflammation fibrosis (scarring) due to accumulation of lipids (fats) in the liver. Around one third of NASH cases will lead to liver cirrhosis and liver failure. There are no approved drugs to treat NASH or to slow its progression.

How does FLCN play a role in fatty liver disease?

As it is accumulation of lipids in the liver that drives damage, regulation of lipid levels is crucial to maintain a healthy liver. FLCN is a known regulator of a protein called AMPK (AMP-activated protein kinase), a master regulator of cellular energy metabolism. Importantly, AMPK controls the switch from the generation of fatty acids to their breakdown. Loss of FLCN leads to a chronic activation of AMPK, which could be beneficial in the context of fatty liver disease. To study the role of FLCN in the role of liver fibrosis progression, the authors of the study generated a mouse model that lacked FLCN specifically in the liver and examined fatty acid accumulation and liver damage.

Firstly, they fed a high-fat diet to the mice lacking FLCN in the liver, or control mice and found that the control mice gained significantly more weight. When they looked at the livers of these mice, they could also see a reduction in fat droplets in the mice lacking liver FLCN compared to control. 

Secondly, they fed the same mice a specific diet that results in the onset of NASH-like symptoms such as liver fibrosis. Unlike the high fat diet in the first experiment, this diet-induced model of fibrosis does not lead to weight gain. Again, they saw significant reduction in fat accumulation in the livers lacking FLCN compared to control. They also saw reduced markers of liver fibrosis and inflammation in the absence of FLCN in the liver, demonstrating that a loss of FLCN protects against liver damage in this context. 


This is the first time a protective role for FLCN has been demonstrated in the liver. The authors of the study also saw no harmful effect of FLCN deletion in the liver which means there is potential for FLCN to be targeted therapeutically for fatty liver disease. It may be possible to develop a drug that is able to inhibit FLCN in the liver and could represent a new treatment route for NASH and even obesity. However, extreme care would have to be taken so that the loss of FLCN is specific to the liver, and would not be able to result in any of the manifestations seen in BHD.

What does this mean for individuals with BHD?
This was not addressed in the current study and so it is currently unclear if there are any consequences for individuals with BHD. It would be extremely interesting to look at samples of the liver in BHD patients or to examine the prevalence of fatty liver diseases within the BHD community. However, it is important to remember that these studies were conducted in mice lacking FLCN in only the liver, and were fed particular diets to induce a model of liver fibrosis and disease and as such may not be reflective of what happens in an individual with BHD.

BHD Toolkit: Getting your Diagnosis

We know that receiving a diagnosis for a rare condition can sometimes be a confusing and overwhelming process. Following on from our first BHD Toolkit, we spoke to Dr Derek Lim, Consultant Clinical Geneticist at Birmingham Women’s and Children’s Hospital, UK to help understand what happens at an appointment with a clinical geneticist. Although this pathway is specific to the UK, we hope this provides a general idea of what to expect when you see a clinical geneticist.

How do you get an appointment with a clinical geneticist?

Dr Lim described the three main routes by which individuals are referred to his monthly BHD genetics clinic.

  1. Individuals who have already had a positive diagnostic genetic test for BHD and are referred to discuss the diagnosis, management, surveillance and genetic counselling.
  2. Individuals who are referred to him for a possible diagnosis of BHD (i.e. they have not yet had a genetic test).
  3. Individuals who have a family member who has been diagnosed with BHD.

What happens during your appointment?

What happens during the appointment will depend on whether an individual has already had a genetic test for BHD.

If this is the case (i.e. route number 1), then the genetic test result is explained to them along with an explanation of BHD including the different manifestations and recommended management and surveillance strategies. Dr Lim explained he would also take a thorough family history, examine individuals for skin bumps and refer them for a CT scan of their lungs if they have not already had one and arrange a scan of the kidneys.

If an individual has not had a genetic test for folliculin (FLCN) and are being referred for a possible BHD diagnosis (i.e. route number 2), then Dr Lim provides an explanation of BHD and offers a skin examination to check for skin bumps. If there is sufficient evidence to suspect BHD, a genetic test to look for FLCN variants will be offered. In this instance, the results are often relayed to the individual in a letter as BHD and the management of the condition have already been explained. There is, however, less opportunity to offer further explanation of the sequencing result itself but a follow-up consultation can be arranged if required.

If an individual has been referred to the clinic because a family member has been diagnosed with BHD (i.e. route number 3), a genetic test for the specific variant identified in the family is offered in the following cases: If there are clinical signs of BHD (e.g. skin bumps, or history of pneumothorax/lung cysts or kidney tumours) – this is called a “diagnostic test” or, if there are no clinical signs  a “pre-symptomatic test” is offered. Sometimes it may be appropriate to test another family member first (e.g. a parent of the individual being seen in clinic).

Why is it important to know the specific FLCN variants?

There are several reasons why knowing the specific variant an affected individual has is important. Firstly, from a clinical perspective, the test required for other family members is simpler and cheaper than having to test for variants in the entire FLCN gene, or potentially panel of genes. Dr Lim likened this to finding a spelling mistake in a book. It is a much easier task to find the spelling mistake once you have been able to narrow down the page, paragraph and line the spelling mistake is in. Secondly, knowing the specific variant is also useful to predict the effect of the mutation on the function of the FLCN protein. In turn, this allows the prediction of whether the FLCN variant will be pathogenic, in other words, how likely it is to develop any symptoms associated with BHD. Finally, it is useful for scientists to know what different variants exist so that research can be done to identify if there are any correlations between genotype (the genetic code) and phenotype (the symptoms of BHD), e.g. whether having a particular FLCN variant increases the risk of developing kidney cancer.

What are some of the common questions asked during an appointment?

During appointments where an individual is discussing their diagnosis, a lot of information can be given in a short space of time, and it may be difficult to think of questions that might be relevant.  We asked Dr Lim the most common questions he is asked in his clinic and here are his top 3:

  1. When should my children be tested?
    In the UK, monitoring of the kidneys (preferably a MRI scan) begins at 18 years of age and so appointments to discuss genetic testing are usually made between the ages of 16 and 18.
  2. How often do I need lung scans?
    In the UK, regular monitoring of the lungs isn’t recommended due to repeated radiation exposure through CT imaging. A CT scan is recommended at diagnosis if the individual has not already had one. Importantly, individuals are made aware of the symptoms of a pneumothorax and what to do if an individual experiences one.
  3. Will I get skin bumps and what are the available treatments?
    Although skin bumps are the most common manifestation of BHD, it is not guaranteed an individual will develop them Some individuals may only have a few whereas others may have hundreds. There are treatments available, however they do not prevent the formation of new skin bumps. You can find a list of possible treatments here.

We are extremely thankful to Dr Lim for providing us with his insight into what happens during a clinical genetics appointment. If you have any further questions about genetic testing for BHD please get in touch with us by email and we will be happy to help. Our next toolkit will delve deeper into the actual sequencing results to help you understand what they mean!

RAREsummit21 Conference Report

The Myrovlytis Trust and BHD Foundation attended RAREsummit21 on 7th October 2021. Organised by Cambridge Rare Disease Network, this one-day event brought together patients, advocates and experts to discuss the challenges facing the rare disease community.

The first session focused on equality, diversity and inclusion (EDI) in rare diseases and featured a panel discussion in which individuals shared their personal experiences of EDI barriers, particularly on the journey to diagnosis. Your ethnic background, gender and where you live can all be a barrier to diagnosis and an individual affected by a rare disease often has to persist to be listened to compared to those without such barriers. The lack of diversity in clinical trials was also spoken about, and the need to identify ways to increase trial participation among ethnic minorities. There was also a discussion on health literacy, and the need for patient advocacy groups to create resources that are suitable for the target audience.

The discussion of health literacy led nicely into the second session entitled “Rare Disease Health Information Delivered Well”, which underlined the importance of providing clear reliable knowledge to enable informed decisions. Getting a rare disease diagnosis can be an extremely overwhelming situation. Individuals or their carers are often given a lot of information which is not always related in the most accessible format, for example the use of very technical or medical language. It has been shown that 40-80% of information discussed in a medical consultation is immediately forgotten and therefore a great deal of thought needs to go into providing this information in the most sensitive and accessible formats. There is also the additional complication of accessing high quality, reliable information due to increasing amounts of information available on the internet, some of which will be outdated or incorrect.

Following on from the discussion on delivering health information, was a session on amplifying the patient voice through patient voice publications. It was agreed that this is extremely important to better understand the disease and burden on the community and that it is the patient voice that can drive engagement with other stakeholders e.g. industry. It was noted that needs outside of healthcare are often unheard and unmet, and a patient voice publication can help address this issue. It is not just other stakeholders that can benefit from a patient voice publication, they can be useful for other individuals affected by rare diseases and provide a reassuring voice that you are not alone in your rare disease journey. However, there needs to be a considerate strategy for how and where to publish a patient voice publication. For example, publication in a medical or scientific journal is probably not the best approach as some of them are not accessible to the public, and even if they are, there are still barriers to finding them in the first place. It is also not sufficient to just create a public voice publication without an appropriate strategy of how to effectively use it and engage with other patients, industry and healthcare professionals.

Engaging with industry can be a challenging task for a small patient advocacy group. The afternoon session entitled “Patient Group and Industry Partnering: What are the right ingredients to help catalyse new treatments?” gave an excellent overview of what a successful industry partnership looks like. The discussion focused on being ‘industry ready’ and that partnering with industry is a long-term goal. The first steps towards this include raising an awareness and having an in-depth understanding of the condition. A clear goal and outcome for patients is required and industry need to understand the immediate benefits to patients. Partnering with industry can be a long road, but persistence from the patient group is key, and making those connections is important. Most conversations can be beneficial, even if it is not appropriate to form a partnership, patient groups are raising awareness and representing their community through these conversations. Additionally, it may open doors for other opportunities with different companies, and an opportunity to have another discussion in the future.

The day ended by featuring a successful partnership with the rare disease patient organisation Ring20 which provides support for an ultra-rare form of epilepsy, and their partnering with the company Illumina. This was a really uplifting way to close RAREsummit, and demonstrated that hard work and persistence, along with an engaged patient community and voice can make a real difference to the lives of individuals affected by Ring Chromosome 20 Syndrome.

Take home messages

The underlying theme of the day was that patients are central to, and critical for, the success of a patient advocacy group. The mission of the Myrovlytis Trust and BHD Foundation is to improve the quality of life for individuals with rare conditions. To enable us to do this, we need to raise awareness of BHD, drive research and engage with industry, all of which require a strong patient voice. Going forwards we would like to further engage with the BHD community and create more patient-centred resources to raise awareness among clinicians. We would also like to take steps towards partnering with industry such as creating a patient registry to increase our understanding of the full range of symptoms associated with BHD.

Who should be referred for genetic testing for inherited kidney cancer syndromes?

It is well known that mutations in certain genes can contribute to the development of kidney cancer. The Cancer Genome Atlas and studies investigating germline mutations in adult cancers identified mutations in 6 – 16% (i.e. 6 – 16  out of 100 people) of kidney cancer cases and there are now 15 genes associated with hereditary kidney cancer syndromes (including folliculin, the gene mutated in Birt-Hogg-Dubé Syndrome, BHD). Therefore, identification of individuals who require genetic risk assessment is important to ensure they receive the most appropriate care. However, clinical guidelines on genetic risk assessment are lacking. Clear guidelines are required to aid clinicians to decide who requires further evaluation, how genetic counselling and testing should be performed, and which genes should be considered. This has particular relevance in the US where differing insurance policies on coverage for genetic testing can result in barriers in to access care. To address this a panel of experts, including Professor Gennady Bratslavsky (the invited speaker at our most recent Meet the Expert event), discussed several questions regarding genetic risk assessment for hereditary kidney cancer to develop consensus clinical guidelines (Please note this paper isn’t open access).

A series of questions were curated by a steering committee and grouped into 5 different categories:

  1. Who should undergo genetic risk assessment?
  2. When should genetic risk assessment be performed?
  3. What testing should be performed?
  4. How should germline risk assessment be conducted?
  5. Testing in cases of isolated extrarenal lesions associated with known syndromes.

The questions were reviewed by the expert panel which included urologists, medical oncologists, genetic counsellors, clinical geneticists, and patient advocates. Uniform consensus was defined as ≥85% agreement.

1. Who should undergo genetic risk assessment?

Uniform consensus on who should undergo genetic risk assessment was reached for the following conditions:

  • Individuals with or without kidney tumours with a personal or family history of classic manifestations associated with hereditary kidney cancer (e.g. spontaneous pneumothorax).
  • Individuals with a 1st degree relative with a documented germline mutation (or 2nd degree relative if 1st degree is not available – see family tree diagram below ).
  • Individuals with kidney cancer who also have a 1st or 2nd degree relative with kidney cancer.
  • Individuals with kidney cancer that shows a specific histology (such as hybrid oncocytoma tumours – common in BHD).
  • Individuals with bilateral or multifocal kidney tumours (also common in BHD).

A consensus could not be agreed regarding a cut-off age for genetic risk assessment (i.e. whether age alone was a sufficient criterion to recommend genetic risk assessment in an individual with a kidney tumour).

2. When should genetic risk assessment be performed?

Uniform consensus on when genetic risk assessment should be performed was reached for the following condition:

  • Individuals with a renal lesion less than 3 cm and a strong suspicion for a hereditary cancer syndrome.

It was also agreed that a skin biopsy is not necessary to guide genetic risk assessment in individuals with a kidney tumour and skin lesions resembling those associated with a hereditary cancer syndrome. Other questions, including whether histologic diagnosis (i.e. confirming the type of kidney cancer) is required before genetic risk assessment was more contentious and the panel did not reach a consensus.

3. What testing should be performed?

A uniform consensus was agreed that multi-gene testing should be considered for individuals with more than 1 risk factor for a hereditary kidney cancer syndrome. It was generally agreed (although a uniform consensus was not reached) that individuals with a suspicion for a particular syndrome with a defined gene should be considered for single gene testing.

4. How should germline risk assessment be conducted?

Uniform consensus on how genetic risk assessment should be conducted was reached for the following conditions:

  • Genetic testing should not be performed without prior genetic counselling.
  • Genetic counselling may be offered by a clinician with expertise in hereditary kidney cancer syndromes.
  • A telehealth consultation with a licensed counsellor is also sufficient before genetic testing.

Some of the panel agreed that a standardised video covering the essentials of pre-test genetic counselling was sufficient, however many were concerned about the lack of opportunity for discussion with a qualified provider.

5. Testing in cases of isolated extrarenal lesions associated with known syndromes

This category covered whether genetic risk assessment should be considered for individuals with no kidney lesions or a family history of kidney cancer but have other manifestations outside of the kidney associated with known hereditary kidney cancer syndromes. It was agreed that genetic testing should be offered in the following cases:

  • A single pheochromocytoma or paraganglioma (adrenal gland tumours).
  • A single endolymphatic sac tumour (a tumour in the ear strongly associated with Von Hippel-Lindau syndrome).
  • Uveal melanoma (cancer of the eye).
  • A single fumarate hydratase-deficient uterine fibroid (associated with hereditary leiomyomatosis and renal cell carcinoma (HLRCC)).

Regarding BHD, a consensus was not reached on individuals with a history of spontaneous pneumothorax or skin fibrofolliculomas.

Summary

Genetic testing is an important factor in assuring an individual receives appropriate care and management. For example, in BHD individuals should monitor their kidneys regularly and if the need for surgery arises, preserving as much of the kidney as possible is essential.  It was noted that there are several barriers to initiating genetic testing including a lack of confidence of clinicians to discuss the risks and benefits and interpretation and explanation of genetic testing results. Consensus guidelines on genetic risk assessment may aid clinicians to overcome the barriers of initiating genetic testing. As such, the findings here represent the first consensus guidelines for genetic risk assessment in hereditary kidney cancer. However, it should be noted that all panel members were from North America and therefore these findings may not necessarily be applicable worldwide. Additionally, the statements here are relatively broad given there are no previous consensus statements that can be further refined. Therefore, it is vital that follow up meetings are held to further refine and update these guidelines, particularly where there was a lack of consensus.

At the BHD Foundation, we think this is a really important step towards implementing standardised guidelines for the diagnosis and management of BHD. We are happy to further advise on genetic counselling and testing and to help locate specialists in your area. Please do email us with all your BHD related queries.

What are the Different Types of Kidney Cancer?

A recent study estimated that there were 431,000 new cases of renal cell carcinoma (RCC – kidney cancer) and an estimated 179,000 deaths in 2020 worldwide. In non-metastatic settings (i.e., where the cancer is localised in the kidney and has not spread beyond) more than 90% of patients survive at least 5 years. However, in patients with metastatic RCC (where the cancer has spread to other parts of the body) the outcome is poorer.

RCC can be categorised into 2 main groups – clear cell RCC, which accounts for 75-80% of all cases, and non-clear cell RCC (nccRCC), which can be further subdivided into smaller groups based on the differences in histology, for example cell shape and cellular alterations. Around 5% of nccRCC cases have a genetic disposition and, like in Birt-Hogg-Dubé syndrome (BHD), these tumours are commonly multiple and bilateral and have an earlier age of onset compared to nccRCC not associated with a genetic condition. In general, kidney cancer associated with BHD is normally slow growing and rarely metastasises. This blog post will focus on the features and treatment of nccRCC. However, it does not discuss oncocytoma or hybrid RCC, 2 of the most common types of BHD-associated kidney cancer.

Types of nccRCC

  • Papillary RCC – the most frequent type of nccRCC and 2nd most common type of kidney cancer. Several genetic mutations are associated with papillary nccRCC, but this type is not normally associated with BHD.
  • Chromophobe RCC – the next most frequent type of nccRCC and one of the most common types associated with BHD. Chromophobe RCC is also associated with a number of other rare genetic syndromes including tuberous sclerosis and Cowden syndrome.
  • Other types of nccRCC include collecting duct RCC, MiT family translocation RCC and renal medullary carcinoma. None of these have been previously associated with BHD and are all very rare, aggressive forms with poor prognoses.

Although the treatments for advanced clear cell RCC have significantly evolved, little progress has been made in metastatic nccRCC due to the low incidence rates and the clinical and molecular diversity of this cancer, resulting in limited numbers of therapies available. Over the past decade, researchers have produced the cancer genome atlas, an in-depth molecular understanding of key cancer-causing alterations in different cancers. Importantly, this includes some types of RCC including clear cell, chromophobe and papillary and has revealed specific spectrums of molecular features for each type to improve the therapeutic management.

A recent paper by Marchetti et al., describes the most important signalling pathways involved in different nccRCC subtypes and includes an overview of ongoing/recently published clinical trials involving nccRCC patients (1).

nccRCC signalling pathways

There are 3 main pathways that are important in driving tumourigenesis (cancer formation) in nccRCC.

1. VEGF axis pathway

This pathway is involved in angiogenesis (the formation of new blood vessels). This is an important pathway in driving the growth of tumours which need a good network of blood vessels to provide oxygen and essential nutrients. There are several existing anticancer drugs that target this pathway to slow the growth of tumours.

2. Mesenchymal-Epithelial Transition (MET) pathway

This pathway is associated with tumour growth, metastasis and malignant cell infiltration (spread of the cancer). This pathway is particularly important in papillary RCC in which the MET gene is often mutated. A number of MET-directed inhibitory cancer drugs have been developed and some, such as cabozantinib target both the MET and VEGF pathways.

3. mTOR pathway

We have recently discussed the mTOR pathway and its role in BHD-related kidney cancer. Although there are several mTOR inhibitors available which are used in other clinical settings, they have not shown to be effective in the setting of BHD.

Immunotherapies

Generally, RCCs are characterised by a dysfunctional immune cell infiltrate leading to immune suppression in the tumour microenvironment allowing growth of the tumour. This makes it targetable by immunotherapy drugs that work against the major proteins that mediate the immune suppressive environment in the tumour microenvironment. These major proteins are called PD-1, PD-L1 and CTLA-4.

Although the use of immunotherapies in clear cell RCC was assessed in 2015, it has only recently been studied in the context of nccRCC and there are now several recently published and ongoing clinical trials (see our clinical trials page for more information on how clinical trials work).

  • The recent KEYNOTE-427 phase II clinical trial looked at the efficacy and safety of the immunotherapy pembrolizumab (a PD-1 inhibitor) as a monotherapy in advanced nccRCC (types papillary, chromophobe and unclassified). The study showed promising clinical activity however the response rates were higher for papillary and unclassified RCC than chromophobe.
  • Another phase II clinical trial tested nivolumab (another PD-1 inhibitor) in combination with cabozantinib in 2 cohorts of patients. The first had papillary, MiT family translocation or unclassified RCC and the second had chromophobe RCC. Like the KEYNOTE-427 trial, this showed promising clinical efficacy in the first cohort but there was no response in the second cohort of chromophobe RCC patients.
  • Several other trials assessing immunotherapies or combination immunotherapies are underway and are showing promising preliminary results.

The International Kidney Cancer Coalition have a very user-friendly clinical trials search tool that gives up-to-date information on current clinical trials.

Various novel strategies for the treatment of nccRCC are also emerging and are under investigation (2). These include combination therapy of the PD-1 inhibitor camrelizumab and cytokine-induced killer cells. These are immune cells that can recognise and specifically kill cancer cells. The field of individualised DNA plasmid cancer vaccines is also progressing and are used to induce an anti-tumour immune response.

Summary

nccRCC is very diverse and complex but we are constantly improving our knowledge and understanding of these cancers and therefore increasing the number of therapeutic options. However, there is still lots of research to be done including the identification of biomarkers for the prediction of progression and response to treatment for nccRCC. Recruitment of sufficient numbers of patients to clinical trials for nccRCC is still an issue due to the rarity and diversity of this group of cancers. Innovation in clinical trial design will be key to the success of future clinical trials in this context.

One of the most striking observations from this review was the lack of response to immunotherapies in chromophobe RCC. As this is one of the most common types of kidney cancer in BHD patients, it is particularly important to us as a charity that further work can be performed to understand why this is the case. This knowledge is greatly needed to open up the therapeutic potential of these immunotherapy drugs which made huge advances in the treatment of other cancers.

References

1.        Marchetti A, Rosellini M, Mollica V, Rizzo A, Tassinari E, Nuvola G, et al. Molecular Sciences The Molecular Characteristics of Non-Clear Cell Renal Cell Carcinoma: What’s the Story Morning Glory? 2021 [cited 2021 Sep 10]; Available from: https://doi.org/10.3390/ijms22126237

2.        M S, F M, G A, V M, A C, A L-B, et al. Designing novel immunocombinations in metastatic renal cell carcinoma. Immunotherapy [Internet]. 2020 Dec 1 [cited 2021 Sep 10];12(17):1257–68. Available from: https://pubmed.ncbi.nlm.nih.gov/32998603/