These are my notes for week 3 of Harvard’s Genetics 228: Genetics in Medicine from Bench to Bedside course. I added the class late so this is just a quick review of the paper discussed in class.

X-linked adrenoleukodystrophy (ALD) [OMIM #300100] is the disease dramatized in the film Lorenzo’s Oil. It is caused by loss-of-function mutations in the ABCD1 gene. Its inheritance pattern is basically X-linked recessive, with affected boys usually presenting in childhood (mean age 7 years) with loss of myelin in the central nervous system leading to a rapid functional decline over the course of a year or two. It is not completely recessive, though - some carrier women also suffer some late-onset nerve damage leading to spastic paraparesis (stiffness and spasms of the limbs). The disease appears to result from a loss of protein function, as the causal variants include both missense [Cartier 1993] and clear loss-of-function variants such as frameshifts [Hershkovitz 2002]. The protein’s native function involves transporting very long chain fatty acids (VLCFs, ≥22 carbons) to peroxisomes for degradation.

A bizarre and unexplained aspect of ALD is the incredible phenotypic heterogeneity, even within families, where all individuals share the same causal variant. Although the typical presentation is in childhood and is rapidly progressive, some males have later onset and present with adrenal insufficiency and limited CNS involvement. There have even been fairly discordant monozygotic twins [Sobue 1994].

Lorenzo’s oil was far less effective in ALD than Hollywood would have you believe. In real life, Augusto and Michaela Odone saw their son Lorenzo’s condition stabilize, but not improve. An open-label clinical trial found that it reduced VLCF accumulation in blood, but didn’t result in any noticeable clinical improvement [Aubourg 1993].

The paper for this week covers another clinical trial by Dr. Aubourg, 16 years later [Cartier 2009]. In this trial, they sought to alleviate ALD by delivering a lentivirally vectored functional copy of the ABCD1 gene. Lots of people have tried to do gene delivery to the CNS, for instance, by direct intraparenchymal injection - for one ongoing example, see the notes on Batten disease from the ASHG2013 gene therapy session. The jury is out on whether that sort of approach will prove to yield any clinical benefits. But Cartier and Aubourg had one major advantage: they didn’t have to deliver the virus directly to the brain. During childhood, myeloid stem cells from blood will migrate to the CNS and settle there permanently, thus offering a window for peripheral intervention that doesn’t exist in many CNS diseases. Indeed, the standard of care for ALD, when a matched donor is available, is allogenic bone marrow transplantation, which gives rise to myeloid cells encoding wild-type ABCD1 which help to correct CNS deficits. Cartier and Aubourg’s trial focused on two boys for whom allogenic transplantation wasn’t an option. They isolated hematopoeitic stem cells from blood, transformed them with replication-incompetent lentivirus overexpressing ABCD1 under a foreign promoter, and transplanted them back in.

The two patients who received this therapy both had their cognitive decline largely arrested, similar to patients receiving allogenic grafts, and in sharp contrast to patients with graft rejection. So although their condition didn’t improve, the clinical evidence suggested that this therapy was better than nothing. As a result, this therapeutic approach (with some modifications to the viral vector) has now advanced to a Phase 2/3 trial sponsored by Bluebird Bio.

This paper’s impact reached far beyond ALD. Much of the text is devoted to discussing the extensive efforts they undertook to document the safety of the vector. Ever since 2000, when a clincial trial for X-linked severe combined immunodeficiency led to leukemia in a subset of patients [reviewed in Herzog 2010], the gene therapy field has been incredibly worried about the potential for retroviral integration to activate oncogenes. Cartier and Aubourg therefore went to extensive efforts to identify integration sites in the two ALD patients’ blood. No single integration site was overrepresented, meaning there was no evidence of clonal expansion due to oncogene activation. And by establishing the potential of peripheral transformed stem cell transplantation to correct CNS deficits, the trial also encouraged similar efforts in other diseases - see for instance a very similar approach being undertaken in metachromatic leukodystrophy (MLD), presented by Dr. Luigi Naldini at ASHG2013.