Slide 1

Well, thank you very much Mr Chairman, Doctor Grinyo and Doctor Sever dear colleagues, it’s a great pleasure to have this opportunity to give an update on management of chronic allograft nephropathy.

Slide 2

I would like to make the following disposition in my talk today. A few slides on epidemiology, more slides on pathogenesis leading up to what can we possibly do to prevent or treat chronic allograft nephropathy today?

Slide 3

Now, we all know from several investigations that much of the improvement in renal transplantation results has occurred during the first 6-12 months, as depicted in this slide from Hariharan. Whereas the long-term graft survival seemed to be parallel irrespective of what time point you’re looking at.

Slide 4

It’s actually a difference in the actuarial graft survival or half life, as shown in this study by Meier Kriesche from a few years back with an improvement by only 1 or 2 years.

Slide 5

But that’s mainly dependent upon the improved results in re-transplantation,

Slide 6

whereas primary transplantation, long term results or other long term graft dysfunction are basically the same.

Slide 7

This is our own picture of factors contributing to development of chronic allograft nephropathy today. Divided into primary injury and secondary injury and also divided into alloimmune response factors, innate immune response and non-immune reactivity.

Slide 8

I will go over a few of these as a background for what my proposal will be for prevention and treatment of chronic allograft nephropathy.

Slide 9

Starting off with a few words on clinical features of CAN. Proteinuria and serum-creatinine have turned out to be two of the most important risk factors and these patients very often present with increased serum creatinine or reduced renal function and a significant degree of proteinuria.

Slide 10

In this study which is from the ALERT trial both proteinuria and serum creatinine were independent risk factors for long-term graft loss and 83% of these patients lost their grafts due to CAN.

Slide 11

The histopathology has partly been covered in a very nice way by Doctor Mihatsch.

Slide 12

But only to repeat was is significant or typical for CAN patients is the arterial intimal proliferation, as demonstrated on this slide.

Slide 13

Interstitial fibrosis, tubular atrophy and early glomerulosclerosis where glomerular basement membrane and tubular basement membrane duplication has been described by several investigators shown in this slide from an experimental model of CAN by Joosten and collaborators in Leiden in Holland.

Slide 14

Also a couple of years back in this very nice study by Nankivell, Jeremy Chapman and collaborators in Australia they could very nicely demonstrate that in their single centre study of close to one thousand biopsies or patients followed over 10 years that there was a steady increase of these changes in the grafts from days 0 until 10 years follow up.

Slide 15

And the degree of chronic BANFF graded chronic changes also increased over time, so that at 10 years in this study almost all patients had grade I chronic allograft nephropathy changes.

Slide 16

Now with regards to pathogenesis I want to divide this into primary and secondary risks starting off with primary injury, focusing on acute rejection and ischemia reperfusion injury.

Slide 17

Now, acute rejection episodes have been shown by many, many investigators to have a strong impact upon the development of CAN later on. This is just an example from one of our own studies where a previous treatment for acute rejection had a significant impact upon graft loss due to CAN.

Slide 18

But as pointed out by Doctor Opelz some 10 years back, if you are successful in treatment of your acute rejection, it seems to be irrespective of what kind of treatment you use when you treat the acute rejection and get a good renal function after treatment of acute rejection. It does not seem to play any role for the graft survival in the years to come following a treatment. They fall into the same sort of graft survival rate as patients without any previous acute rejection.

Slide 19

Several mechanisms are involved in the tissue injury by an acute rejection leading to these chronic changes. Some of these are listed on this slide where I think that the upregulation of certain cytokines and growth factors and EMT transition and deposition of extracellular matrix proteins play a significant role.

Slide 20

A few words about secondary injury focusing on some of these factors that are considered to be progression factors and chronic CNI toxicity since that is something that we are targeting to quite a large extent today in order to try to prevent or reduce CAN.

Slide 21

Now, renal transplant function has been demonstrated by several investigators to be one of the strongest risk factors for graft loss in CAN.

Slide 22

This is again from the ALERT trial showing a relative risk of 5.4 per hundred micromolar per litre of losing the graft during a 5 year follow up with the highest risk in patients with the highest creatinine level.

Slide 23

Similar findings by other investigators like in this slide from one of the publications by Hariharan in the United States that the higher creatinine at 1 year, the higher the risk of losing the graft.

Slide 24

Proteinuria, again in this time the ALERT trial that if you had more than 400 mg/24 hours, 30% risk of losing the graft.

Slide 25

Taking together proteinuria and a high creatinine level led to 60% risk of losing the graft within 4 years, if you had more than 400 mg of albumin and a creatinine above 170.

Slide 26

Nephrotoxicity due to immunosuppressive agents involves vasoconstriction, relative ischemia, EMT transition, profibrotic signalling and contributes to the development of CAN.

Slide 27

Complicated slide stating some of the mechanisms by which calcineurin inhibitors contribute to the development of interstitial fibrosis and the arteriolar hyalinosis that Doctor Mihatsch also mentioned in his previous talk.

Slide 28

Again back to the study by Nankivell and Jeremy Chapman and collaborators, they could demonstrate in their cohort of renal transplanted patients that at 10 years there were calcineurin nephrotoxicity signs in almost all their patients. So this is an important target we believe today for prevention of CAN.

Slide 29

Hypertension, several different mechanisms by which hypertension occurs in renal transplanted patients. Both have to do with renal allograft disease but also with the immunosuppressive therapy that we are using where corticosteroids and calcineurin inhibitors are the main contributors to the development of post transplant hypertension.

Slide 30

Classical study by Opelz, Ritz and collaborators regarding hypertension that the higher degree of hypertension, the greater the risk to lose their graft irrespective of what time point is considered that they compare with. This is apparently independent of renal function.

Slide 31

We did the same finding that hypertension was a clear risk factor to lose the graft during the follow up in the ALERT trial.

Slide 32

Ischemia reperfusion injury is a problem that contributes to some extent to the development of CAN by induction of inflammatory mediators, upregulation of cytokines and ILs, oxidative stress, development of apoptosis and possibly induction of senescence.

Slide 33

Delayed graft function has an impact on graft survival.

Slide 34

Figures from the UNOS showing that with DGF 33% 10 year survival, without DGF 53% graft survival and cold ischemia time and ischemic injury influences the relative risk of delayed graft function very clearly.

Slide 35

Senescence has been put forward by several investigators to be an important phenomenon, epiphenomenon or not, I don’t know that for sure. Phil Hariharan and others have investigated that clearly. It is related to oxidative stress and ischemia reperfusion injury, leads to a telomere shortening in experimental renal transplants and CAN has been stated to be a formal accelerated type of ageing.

Slide 36

A few slides from experimental studies showing that with time telomere length decreases in experimental renal transplants.

Slide 37

This is also related to the degree of ischemia reperfusion injury.

Slide 38

In our own hands in leukocytes from dialysis patients we could also show that the degree of systemic inflammation is related to telomere length.

Slide 39

The more inflammation, shorter telomere lengths and that translates into reduced patient survival.

Slide 40

A paper will be presented tomorrow by Carrero.

Slide 41

Finally, with regards to pathogenesis what is the influence of BKV and CMV?

Slide 42

Recently we have looked with a very new sensitive technique into the presence of the CMV antigens in cases with CAN and have been able to show that in the majority of these lost grafts there was positive reactivity for CMV early after transplantation and also late. We could also with insight to methodology show that CMV was present in transplants that were lost due to CAN.

Slide 43

Over to prevention and treatment starting off with prevention of primary injury, acute rejection.

Slide 44

Well as mentioned earlier, with new combination of immunosuppressive regimens and the use of induction therapy the rate of acute rejection has improved a lot so that now we’re down to almost around 15% with some of the protocols that are being used, 15% of acute rejections in renal transplantation.

Slide 45

Different combinations have also been related to the risk of later graft loss in this study by Cherikh the lowest rate in the combination of tacrolimus and MMF regimen.

Slide 46

IR, or how to prevent ischemia reperfusion injury?

Slide 47

A few novel approaches have been tried mainly in experimental models such as the use of costimulatory blockade, the use of P-selectin, IGF-1, induction of HO-1 which is a protective gene and also the use of Bcl2 another protective gene which leads to reduced apoptosis. Some of them could be demonstrated in experimental transplantation, again to reduce the risk of chronic allograft nephropathy.

Slide 48

With regards to senescence there is no definite way to deal with that but it could be assumed that the reduction of stressors, a removal of nephrotoxic agents and possible use of antioxidative measures may be a way of reducing the degree of senescence and improving the premature ageing of the kidney but very little has been demonstrated in actual transplantation so far.

Slide 49

With regards to improvement of renal transplant function by reduction of CNI nephrotoxicity, 3 different approaches are currently being used, selection of CNI there are small differences between the different types of calcineurin inhibitors, reduction or withdrawal of CNI and introduction of mTOR inhibitors or MMF.

Slide 50

This slide from the --- study illustrates very clearly I think how randomisation at 3 months to continued use of calcineurin inhibitor or the use of sirolimus leads to different glomerular filtration rates at 12 months.

Slide 51

Several switch studies are ongoing presently. CONVERT trial, we’ve heard a few presentations on the CONVERT trial but not the final publication. ASCERTAIN is a switch study where CNI is reduced or withdrawn and certican is introduced. TRANCEPT is a retrospective study with CNI reduced or withdrawn and introduction of MMF. NOCTET is a similar study in heart and lung transplanted patients.

Slide 52

I think for the sake of time an intermediate presentation was done on the TRANCEPT study showing that the retrospective reduction of the glomerular filtration rate by 10 ml/minute until the time of switch whereby this reduction of GFR seemed to have levelled off after CNI was withdrawn or reduced and MMF was introduced.

Slide 53

But we have another couple of years to go until the TranCept will give its final report.

Slide 54

What about antiproliferative approaches?

Slide 55

Well, what we have access to is sirolimus and everolimus where there is quite a lot of quite interesting data on reduction of intimal hyperplasia in grafts.

Slide 56

This is from one of the older studies by Randal Morris

Slide 57

showing that the use of rapamune in heart transplants in monkeys led to a strong reduction of intimal hyperplasia compared to the control individuals.

Slide 58

The same has been shown in other models,

Slide 59

Use of everolimus in this case much less intimal hyperplasia and similar in clinical heart transplantation using IVUS instrument that the use of everolimus much less intimal hyperplasia compared with a control arm.

Slide 60

So this seems quite promising with regards to what’s happening in a vascular wall in these transplants.

Slide 61

Another very intriguing approach which has been tried only in experimental transplantation is the use of a PDGF signalling inhibitor, Imatinib. Acute rejection increases PDGF induction during development of CAN.

Slide 62

And this has been tried in experimental transplantation by Saveko in Finland showing that 90 days after experimental kidney transplantation there was a high expression both of inflammation, fibrosis, intimal proliferation and glomerular matrix increase. Whereas in cases that were treated also with this PDGF blocker these changes were heavily reduced. We have tried to get hold of this compound for clinical use but not yet been all that successful.

Slide 63

Finally, what about ACE inhibitors and angiotensin receptor blockers?

Slide 64

There is intriguing experimental work on the use of enalapril in experimental kidney transplantations showing a reduction of glomerulosclerosis, lymphocyte infiltration, macrophage infiltration and expression of adhesion molecules.

Slide 65

Similarly we did with an ARB in an aortic transplant model showing a reduction of intimal hyperplasia in the candersatan both in syngeneic and allogeneic transplants.

Slide 66

From the group in Austria with Doctor Oberbauer they presented a couple of years ago that the use of ACE and ARB led to improved graft survival compared to cases without such treatment.

Slide 67

In this study there was also an improved patient survival.

Slide 68

Whereas other studies like the one by Doctor Opelz, which is a register study then, he could not demonstrate any difference at all with regards to graft survival or patient survival in patients that were treated with ACE inhibitors or ARB.

Slide 69

Neither could we see any difference in the 520 patients on such treatment compared with those without ACE inhibitors with regards to graft loss, just the same frequency of graft losses in that study.

Slide 70

Finally, what about statin treatment?

Slide 71

We know from cardiac transplant studies by Kobashigawa that the use of pravastatin leads to a reduction in intimal hyperplasia in coronary arteries compared to control cases.

Slide 72

In heart transplantation this leads or readily translates into patient survival being better in pravastatin treated cases compared to control cases.

Slide 73

But when we did a similar study in the ALERT trial, there was absolutely no difference with regards to graft loss, doubling of serum creatinine in cases treated with fluvastatin compared to placebo.

Slide 74

So our impression is that there isn’t really any benefit with statin treatment in renal transplantation when it comes to renal development of CAN and renal graft loss.

Slide 75

Finally, only speculative again, whether BKV or CMV turns out to be important in the long run may be antiviral treatment may also have a role in the prevention of CAN.

Slide 76

Several other novel strategies could also be anticipated and has partly been shown already, tolerance induction, several different models where the investigators have been able to demonstrate that it leads to reduced chronic scarring of the transplant, for example. Induction of protective genes have also been in experimental transplantation again, demonstrated to reduce the degree of CAN changes.

Slide 77

So, in summary and conclusion long term graft survival remains basically unchanged over time in the first graft of renal transplants and late graft losses are largely due to CAN. Complex pathogenesis of CAN with both initial injury and progression factors that play a role, donor antigen dependent and independent factors such as acute rejection, renal dysfunction, hypertension, proteinuria and the choice of an immunosuppressive regimens play an important role.

Slide 78

Prevention of primary injury and attenuation of progression factors are targets for management of CAN in renal transplantation but there is absolutely no silver bullet available but what we can do is to better prevent from acute rejections eliminate HLA and non-HLA antibodies, reduce ischemia reperfusion injury, oxidative stress and possibly senescence in the graft, use of immunosuppressive regimens that are less nephrotoxic, improvement of renal transplant function but whatever means that is apart form reduction of nephrotoxic agents, treatment of hypertension and proteinuria and possibly again, prevention and treatment of BKV and CMV may also turn out to be future options. Thank you very much for your attention.

Slide 79

Chairman: Thank you Bengt. We have time for two questions.

Question: So, I’m Doctor – from Helsinki. I would ask you about the mTOR inhibitors. What would be the best time to introduce them if you wish to reduce CNI inhibitors? And regarding that when it’s over proteinuria they are contra indicated and one of the hallmarks of CAN is the development of proteinuria. So when would be the right time to introduce them and do you see a role of protocol biopsies to diagnose and to introduce them in the right time?

Dr. Fellstrom Well, with regards to your last question, most definitely I believe as Doctor Mihatsch also said that protocol biopsies will give, will, are of great importance to identify these changes at another stage. We have been doing that for a long time. Protocol biopsies at 6 months. Believe me we get very valuable information form that. Right now this is not ongoing, it was part of a project when we did that but we will most likely go back to doing that. With regards to reduction or withdrawal of CNI and the ideal or optimal time point, I don’t think anybody has an answer to that but there are a number of studies, a few studies ongoing which are addressing that question and one is the ASCERTAIN that will be finished by September of next year, where we will possibly be able to find out if there is an ideal point for this switch. There’s also another study SEMPER study going on where a switch is done according to the protocol between 2-3 months following transplantation in a randomised fashion they switch from continued CNI over to everolimus. So that study may also be able to answer that question.

Question: Thank you.

Chairman: Doctor Ponticelli.

Question: Ponticelli Milan, congratulations Bengt for your presentation. However, the data you presented in particular the data of Nankivell and co-workers refer to old times when cyclosporine and tacrolimus were given at very high doses. Today we have learnt to give low doses of CNI and I’m not convinced that CNI toxicity represents one of the most important triggers for so-called CAN. I feel and this is my question that probably the quality of the kidney is probably more important because today at least in Italy, we have a lot of older cadaveric donors and most of these kidneys have a low GFR and already have some kidney lesions and there is evidence for example, that in the long-term kidneys from unrelated living donors have a better outcome than kidneys from cadaveric donors. So what is your impression about the quality of kidneys as one of the most important triggers of CAN?

Dr. Fellstron I agree with you entirely that the quality of the kidney plays a significant role and it’s one of the jokers, one of the question marks you have to take into account if you make protocol biopsies at 6 months, how much of these changes came through the kidney from the very beginning and how much is something that has developed since transplantation but I mean our own data also points in the same direction as so many other investigators that the quality of the kidney at the time of transplantation has a very important impact for sure.

Chairman: Thank you Bengt these last questions and comments are related to the next presentation.