AASK: a cautionary tale for bardoxolone?

Robert Leversee had some questions regarding my presentation on diabetic nephropathy. You can see his concerns in the comments after the post. he was specifically concerned about this slide.

Robert felt it minimized the GFR gains found with bardoxolone. What is not clear from the deck is that 56 weeks, represents the GFR one month after stopping the drug. In the lecture, I pointed out that patients that were on bardoxolone all had a higher GFR than at baseline, while patients randomized to placebo had a lower GFR.

As a reminder, the primary end-point of the study was the change in GFR at 24 weeks and that was dramatic.

The reason I included the slide showing the 56 week data was my concern that bardoxolone may be pulling a creatinine slight of hand. My personal concern is that the changes in GFR are due to simple hemodynamic changes like were seen with amlodipine in AASK.

AASK was a trial of hypertension therapy in African Americans with a renal end-point rather than a cardiovascular end-point that are more common in hypertension trials. The trial is a two by three design with two blood pressure targets (MAP 102-107 vs <92) and three blood pressure medications (amlodipine, ramipril, metoprolol).

The data is difficult to interpret because the amlodipine caused an acute hemodynamic-related bump in the GFR, but after 12 months the loss of GFR in the amlodipine group was faster than with ramipril. The study designers designated co-primary end points, a total change in GFR and a chronic change in GFR that ignored the initial 3 months.

Ramipril was superior to amlodipine in the chronic phase but not in the total change in GFR. Though this ambiguity was not represented in the conclusions of the trial:

The fact that amlodipine improved renal function for one year makes me nervous about the one year duration of the bardoxolone study. Thankfully BEACON is in full swing enrolling patients so a definitive answer is just ahead.

ASN Renal Week day 4: APOL1 the best medical science story of 2010

The Renal Fellow network may have ranked APOL1 as the fourth biggest story of 2010 but I think it is actually the best story in all of medicine, not just nephrology.

When I went to ASN Renal Week I stayed at Castle Marne, an idiosynchratic bed and breakfast about a mile and a half from the conference center. The other people staying at the Castle were a rogues gallery of interesting conference participants.

One of the breakfast crew was David J. Friedman, the second author on the Science paper blowing the lid off of APOL1.

The APOL1 story begins in 2008 with the discovery of MYH9. Scientists were doing whole genome analysis to find a genetic explanation for the excess renal risk African Americans face. This excess risk is seen in the dialysis population, where African Americans are over represented. This is particularly true in patients with ESRD due to hypertension. African Americans represent 13% of the U.S. population but represent 48% of the patients on dialysis due to hypertension (HA-ESRD).
Data from USRDS and US Census
The two other places that the increased renal risk of African Americans is seen is in FSGS and HIV associated nephropathy (HIVAN). FSGS is 5 times more likely in young African American males than in age-matched Caucasians. HIVAN is nearly unheard of among people of European ancestry. The only cause renal failure more specific to black patients than HIVAN is sickle cell nephropathy.
The genetic locus 22q13.1 was found to convey phenomenal excess risk of FSGS. The excess risk was 400-700% (OR5-8). In whole genome analysis, researchers are delighted to find odds ratios of 1.1-1.2. Finding ORs of this magnitude is nearly unheard of. Poking around the genetic neighborhood, the researchers found a likely genetic target, MYH9. 
MYH9 codes for an intra-cellular myoglobin. MYH9 was an especially appealing candidate gene because it is expressed in podocytes and mutations of the gene had previously been found to be associated with glomerular pathology. Quickly MYH9 was declared the genetic explanation for excess risk of renal disease among African Americans and the scientific nephrology community geared up to crack every mystery related to MYH9. 
It was the gene that launched a 1,000 RO1s.
The NIH and NIDDK sponsored symposia to get scientists up to speed with breakthrough discovery
Unfortunately, no one was able to find the specific genetic mutation that led to these renal complications. From the discussion of the original paper:

A limitation of our study is that we have not yet identified the causal sequence variation in MYH9 that is associated with FSGS

Then in August 2010, David Friedman and his team identified APOL1 as the gene that actually was associated with FSGS, HA-ESRD and HIVAN. The association was discovered after new genetic material was made available in the 1000 Genomes Project, a public database of genetic information from individuals around the world including a number of Africans.

APOL1 lives just to the centromere side of MYH9. Friedman et al showed a tighter association with APOL1 than MYH9 and when they controlled for APOL1, MYH9 was no longer significantly associated with renal disease.

As the scientific community began to feel the rumbles of truth emerge about MYH9 and APOL1, researchers hitching their wagon to MYH9, prayed they were funded before the NIH scorers realized that MYH9 was the wrong gene. Scientists with research proposals on MYH9 that were too late would have to rewrite the grant to focus on the new target, APOL1.

Friedman’s team didn’t just identify APOL1 they told a fascinating story involving parasitology, evolution and human migration.

In 2003 APOL1 was identified as the genetic source for an immunity factor which protected people from African sleeping sickness. 95% of African sleeping sickness (I refuse to use the 3-letter acronym) is caused by Trypanosoma brucei gambiense.

Trypanosomes cause the mortal disease African Sleeping Sickness

Trypanosome lytic factor (TLF) protected humans from sleeping sickness until Trypanosoma brucei rhodesiense and gambiense evolved a protein, Serum Resistance Associate Protein, that deactivated TLF. This adaoptive response by the trypanosome made humans susceptible to infection. This was the state until about 10,000 years ago when variants of APOL1 appeared and restored the protective action of TLF and made the carrier of even a single copy immune to African sleeping sickness.

These genetic variants of APOL1 appeared in Africa 10,000 years ago, but much of the human race had already left Africa to spread across six continents. Additionally, regions that did not have the Tze Tze fly didn’t have trypanosomes and hence didn’t have selective pressure for the APOL1 variants. In regions endemic to Tze Tze fly, the selective pressure for these mutations was immense. In the U.S. 30% of African Americans carry APOL1. Heterozygotes are immune to trypanosomes and may have a modest increase in the risk of HA-ESRD (OR 1.26, no risk for FSGS) . Homozygotes for APOL1 are equally immune to trypanosomes but unluckily have a sky high risk of renal disease.

So APOL1 behaves like sickle cell anemia and malaria. Heterozygotes are immune but homozygotes suffer from  devastating disease. Balanced polymorphism.

The last twist is the mystery of HIVAN in Africa. HIVAN is found in western, Sub-Saharan Africa. Eastern Africa has a lower rate of HIVAN than would be expected. This data comes from cohort studies done in Kenya and Ethiopia. The risk of HIVAN is associated with APOL1. The Tze Tze fly is not endemic to Eastern Africa, hence no trypanosomes, so no selective pressure for APOL1, so few people are homozygotes for the variant of APOL1 that predisposes to HIVAN.

This story was one of many that were batted around the breakfast table at Castle Marne and served to show that I found the perfect place to stay during ASN Renal Week.

HeLa, Salk and the Tuskegee Institute

The Immortal Life of Henrietta Lacks is a multi-headed beast. The story is structured around three discernable plot lines. The first, is the history of the Lacks Family from slavery war up through present day. Another, is the author’s story of  how she met the family and uncovered the history. And the last leg is the history of science and how it relates to human cell culture and the HeLa cells.

This last story line is amazing, but I’m a little cooler to the other two. So far I’d say the third line is strong enough to justify reading the whole book but this is no The Checklist Manifesto.

One of the most interesting stories is regarding the first scientific win for the brand new science of human cell culture. HeLa cells were instrumental in the widespread testing of the Salk Polio vaccine. (All of the following excerpts from the book come from here)

“..in April 1952, [George] Gey and one of his colleagues from the NFIP* advisory committee –William Scherer, a young postdoctoral fellow from the University of Minnesota– tried infecting Henrietta’s cells with poliovirus. Within days they found that HeLa was, in fact, more susceptible to the virus than any cultured cells had ever been… they knew they’d found exactly what the NFIP was looking for”… “On Memorial Day 1952, Gey…sent Mary to the post office…When the package arrived in Minneapolis about four days later, Scherer put the cells in an incubator and they began to grow. It was the first time live cells had ever been successfully shipped in the mail.” …”When the NFIP heard the news that HeLa was susceptible to poliovirus and could grow in large quantities for little money, it immediately contracted Scherer to oversee development of a HeLa Distribution Center at the Tuskegee Institute… [p95] …it was the first-ever cell-production factory and it started with a single vial of HeLa that Gey had sent Scherer in their first shipping experiment, not long after Henrietta’s death. [p96]

George Gey was the original scientist who created the immortal HeLa cell line.
*NFIP was the  National Foundation for Infantile Paralysis, the organization now known as the March of Dimes, created by FDR to fight polio.

I had never known that the Tuskegee Institue had a role in the war on Polio and development of the Salk Vaccine. The HeLa cells were used in wide spread testing of the vaccine to make sure it was immunogenic. Since HeLa were able to be infected and killed by the Polio virus, they became a convenient means of testing the vaccine. The vaccine was administered to volunteers and six weeks later if that patient’s serum protected HeLa cells from Polio infection that alloquot of vaccine and its administration technique was immunogenic.

The only thing I knew of the Tuskegee Institute was its role in medicine’s most horrific racial crime, the studying of 400 African American men with syphillis without telling them they were infected or offerring treatment. This deception lasted for forty years. From Wikipedia:

By 1947 penicillin had become the standard treatment for syphilis. Choices might have included treating all syphilitic subjects and closing the study, or splitting off a control group for testing with penicillin. Instead, the Tuskegee scientists continued the study, withholding penicillin and information about it from the patients. In addition, scientists prevented participants from accessing syphilis treatment programs available to others in the area. The study continued, under numerous supervisors, until 1972, when a leak to the press resulted in its termination. Victims included numerous men who died of syphilis, wives who contracted the disease, and children born with congenital syphilis.[4]

So finding out that Tuskegee had a role in Polio was interesting, but discovering that the technicians and scientists in Tuskegee were all African American and that the Tuskegee institue had won the contract to produce the cultures in a form of proto-afirmative action blew my mind. An afirmative action program was happening at the same place, and at the same time, as one of the darkest moments in the mistreatment of African Americans.

…”Black scientists and technicians, many of them women, used cells from a black woman to help save the lives of millions of Americans, most of them white. And they did so on the same campus –and at the very same time– that state officials were conducting the infamous Tuskegee syphilis studies.