Special feature: The legacy of Malcolm Simons, the junk DNA genius

This feature appeared in the March/April 2012 issue of Australian Life Scientist. To subscribe to the magazine, go here.

Less than four months after the completion of the Human Genome Project (HGP), at the opening ceremony of the 23rd International Congress of Genetics (ICoG) in Melbourne in July 2003, two geneticists – one a devout Christian, the other an atheist and Humanist – took the stage to condemn patenting of human genes.

They were British Nobel laureate Sir John Sulston, who coordinated Britain’s contribution, and US geneticist Francis Collins, director of the Human Genome Research Institute. They used the “Olympics of Genetics” as a platform to warn that unrestrained commercialism could enmesh medical research and the biosciences in a sticky web of patents and exorbitant licence fees.

The next day Collins stunned his audience and the media by ending his plenary address on the HGP with a scathing attack on a small Melbourne biotechnology company, Genetic Technologies. He accused the company of trying to hijack drug research by enforcing its claim to what he called a “flimsy patent” over vast tracts of the human genome – the 98.5 per cent of the genome that does not code for proteins.

The 1993 patent was far too broad, and should never have been issued, he said. Genetic Technologies was pursuing an aggressive licensing strategy that was harming genetics research and drug development.

Collins invited questions. A slightly stooped man, bald and dressed in black, rose and said: “I happen to be the inventor of that patent, and it’s not wise of you to be taking pot-shots at Genetic Technologies, and certainly not at the inventor.”

Malcolm Simons introduced himself, and calmly thanked his discomforted critic for drawing attention to the alleged problems with his patents, but he begged to differ: the patents were well-founded and their novelty was not in question. After all, the Patent and Trademark Office in Collins’ own country had found them valid.

Several weeks before his ICoG encounter with Collins, the 63-year-old New Zealand-born immunogeneticist had received a bone-marrow transplant at Royal Melbourne Hospital. High-dose chemotherapy was the cause of his hair loss.

At ICoG the next day, ABC-TV documentary producer Sonya Pemberton previewed her new 30-minute documentary for the media, The Genius of Junk reviewing Simons’ achievements, and his controversial patents. In one scene, Simons’ oncologist displays an X-ray of his patient’s brain on a light box, revealing scores of small, dark spots scattered through healthy neural tissue. Each spot was a small tumour. Simons’ dismay was palpable.

A year earlier, in the US, he had developed severe pain in his rib cage, centred on his sternum. He suspected multiple myeloma; later medical tests in Hong Kong confirmed the diagnosis.

The cancer of plasma cells has a median survival time of five years. Simons’ mutation was towards the less aggressive end of the spectrum, and with high-dose chemotherapy and experimental drugs, he lived nine and a half years before finally succumbing on January 25 of this year.

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Aggressive strategy

The ICoG experience left Simons aggrieved that his reputation was being sullied by Genetic Technologies’ aggressive commercial strategy, and its pursuit of lucrative deals with companies and health agencies it considered were infringing its patents covering the “junk DNA” diagnostic techniques he had invented.

The patents originated from a moment of deep insight in San Francisco on December 23, 1987. Simons was walking to lunch at a Chinese restaurant with geneticist Roger Lebo, of the University of California, San Francisco , Mervyn Jacobson and David Cunningham, manager of Jacobson’s US company.

Lebo was searching for mutant alleles involved in Charcot-Marie Tooth Syndrome, a group of hereditary neuropathies affecting the brain and spinal cord. Simons was chatting with him about haplotypes: clusters of alleles inherited en bloc on the same chromosome segment, between recombination loci.

Several months earlier, Simons had painstakingly analysed dozens of individual patterns of restriction fragment length polymorphisms (RFLPs) in the Major Histocompatability Complex (MHC), a highly polymorphic region of the genome central to our immune system.

An individual’s DNA, when cleaved by bacterial restriction enzymes, forms distinctive patterns of bands on an electrophoresis gel. While individual RFLP patterns are unique, even unrelated individuals share certain fragments; they might be the same length, carry the same charge or they migrate to the same position on a gel. Family members share a higher proportion of RFLPs, according to their degree of relatedness.

RFLP technology was developed by UK geneticist Alec Jeffrey in 1985 and allowed geneticists to compare DNA profiles of individuals, with the technique rapidly becoming a standard tool for genetic research. Simons established Australia’s first paternity-testing laboratory in Melbourne in 1988 using Jeffreys’ technique.

Simons “junk DNA” patents emerged from his realisation that, if protein-coding DNA sequences – i.e. genes – accounted for less than five per cent of the human genome, most of the cleavage sites targeted by restriction enzymes had to lie within the non-coding 95 per cent: the so-called “junk” DNA within and between genes.

If fragments shared by close relatives were being cleaved at the same sites, and most cleavage sites lay in “junk DNA”, it implied they must be highly conserved – even more so than protein-coding sequences. That implied the “junk” served some important function, even if he had no idea what that function might be.

Simons knew that particular combinations of alleles in the MHC on chromosome 6 are inherited en bloc, as haplotypes. In fact, haplotype matching underpins the matching of donors with recipients of organ transplants. But he knew that unrelated individuals often share RFLP fragments, even across different racial and ethnic groups.

At the 10th International HLA and Immunogenetics Workshop in Salt Lake City in 1988, Simons reported on how he’d manually analysed a large RFLP data set from cell lines contributed by laboratories around the world, from patients with autoimmune disorders including ankylosing spondylitis and rheumatoid arthritis.

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Shared haplotypes

Simons was intrigued to find that patients from ethnic groups that had diverged around 60,000 years ago – Caucasians, Japanese, Chinese, African, Amerindian and Caucasians – shared certain haplotypes that were essentially identical to those previously identified from studies of large, multi-generational families with the same disorders.

The standard approach to tracking down disease-causing mutant genes was to compare RFLP patterns in large, multi-generation family pedigrees, and identify shared fragments using genetic markers. The approach was impracticable for rare genetic disorders, where there are too few affected individuals in affected families to establish a statistically significant association between recurring genetic markers and a potential disease locus, indicating they are co-inherited at high frequency because of their proximity on the same chromosome.

Then came Simons’ brainwave. He asked Lebo: why do we limit ourselves to studying inherited disorders in large, multi-generation families? Why not compare RFLP patterns in large numbers of unrelated individuals with the same genetic disorder, and use the polymerase chain reaction to identify shared “junk DNA” markers linked to candidate disease haplotypes?

After the ICoG episode in 2003, where Collins and Sulston claimed that Simons had patented “prior art” because the RFLP-based technique was already widely used in the late 1980s, I interviewed eminent British geneticist and cystic fibrosis expert Professor Bob Williamson. I asked him whether, at the time, he had thought of using unrelated individuals to locate disease-causing alleles within shared, highly conserved haplotypes. Williamson said the idea had not occurred to him.

Given his prominence in the field and familiarity with the state-of-the-art science, his response suggests the view of critics like Collins and Sulston that Simons had opportunistically patented a technique already in general use was unsubstantiated.

Simons lodged his first patent application for his “junk DNA” technique in August 1989. It related only to the Major Histocompatibility Complex; he later generalised the principle of using non-coding DNA to identify disease alleles to the entire genome, and lodged further patent applications covering all human, animal and plant genomes in 1990.

He scoured the literature for prior art. The first research paper to “reinvent” his approach was not published until 1993. Its authors were clearly unaware of Simons’ patent. Others were published in 1996 and 1997, nearly a decade after his original filing.

Interviewed in September 2011, Simons said he believed his second, genome-wide, patent, filed in June 1990, was his most important contribution to science. “It described how you can use this non-randomness and the haplotypes that you define [with DNA markers] to map disease related genes, without using family pedigrees. My claim is that nobody else thought of it before me.

“Bob Williamson was a world expert in linkage disequilibrium, and so was Roger Lebo, and they both used family pedigrees to discover disease alleles. I told Lebo we could use combinations of markers to define particular haplotypes, but we could only identify haplotypes by using highly conserved markers in non-coding regions.

“When you’re dealing with multiple gene variants, you must determine the specific variation in the protein-coding exons that associates with a particular set of markers.”

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Novelty issue

However, Sulston and Collins created some uncertainty in Simons’ mind as to whether his broader junk DNA patent, lodged in the US in 1990 and issued in 1998, met the criteria of being novel, non-obvious and free of prior art. He felt Genetic Technologies was exploiting the patent in ways he had not intended, and his own reputation and credibility were suffering because of his former association with the company.

Jacobson’s acquisition of Australasian rights to use Salt Lake City-based Myriad Technologies’ tests for mutations in the BRCA1 and BRCA2 breast-cancer tumour suppressor genes earned particularly strong criticism medical researchers, clinicians, and the media.

Simons was convinced that Genetic Technologies was taking an unconscionably broad view of the scope of his patents, despite its success in defending them in the US courts. He even acted as an adviser to big US gene-testing company Applera, in its unsuccessful 2003 court challenge to the validity of Genetic Technologies patents.

Simons’ technique involved probing a junk DNA site within a haplotype with a set of PCR primers and checking the intervening DNA sequence for single nucleotide polymorphisms (SNPs) that uniquely “marked” that haplotype, and no other.

“In other words, the unique SNP allows the amplified sequence to be used as a surrogate marker for all the alleles in that haplotypes,” he said. “Genetic Technologies’ method is to place primers in the non-coding DNA, without necessarily covering a SNP site, and use them to amplify every allele at that locus.

“Not only is that not what I invented, Professor Henry Ehrlich was using a 330-nucleotide amplicon at the HLA-DP beta locus to identify every allele at that locus before I came up with my concept – and that certainly constitutes prior art.”

“If you use a primer covering a non-coding variant lacking a unique SNP, you’re not getting one unique allele to serve as proxy for every unique allele within that particular haplotype. Potentially, you get the entire ‘family tree’ of haplotypes, from the ancestral haplotypes down to every variant that has evolved since through mutation.

“The fact is that almost all these alleles have close relationships with others, and they fall into distinct ancestral groups. Group 2 might include alleles 15 and 16, Group 5 might contain alleles 11 and 12, and so on,” said Simons.

“The ancestral group gets you into the ballpark, and as you progressively resolve the alleles, you eventually get down to the point of doing DNA sequencing, which gives you the unique allele within a particular lineage.

“We now know of more than a thousand HLA-DP beta alleles, that can be assigned to about 20 ancestral alleles. In each group, subsequent mutations have produced more and more discrete, distinguishable alleles. They all have ancestral SNPs in common, but we can actually track the evolution of an allele lineage back through time. That’s what I invented,” he said.

“My invention, and the way Genetic Technologies is applying my patent, are starkly and easily distinguishable. I believe – and some of my peers in the HLA community believe – that Genetic Technologies is overreaching the scope of my invention, and charging licensing fees on techniques that I knew at the time were prior art.

“I formally assigned my ‘junk DNA’ patents to the company on October 30, 1999, and don’t want to be seen to be obtaining any benefit from the licensing fees they’re charging. Because of their actions, I’ve been cast as a pariah for the past decade, and that has prevented my contributions to haploid genomics being recognised at the highest level.”

Malcolm Simons, DSc, (1939-2012) was born in New Zealand, trained as an immunologist. He was a member of both the New Zealand Davis Cup squad, and the New Zealand national squash team in the 1970s before emigrating to Australia. He founded Australia’s first DNA paternity-testing laboratory in Melbourne, and was the inventor of patents that employ highly conserved sequences in non-coding DNA as proxy markers for haplotypes associated with inherited genetic disorders.