MPN Driver Mutations Can Be Acquired as Early as in Utero, Study Shows

A study presented during the 2020 ASH Annual Meeting has suggested that certain driver mutations for myeloproliferative neoplasms (MPNs) can be traced back to when they were acquired as early as in utero.

“If you can not only detect clones early but then calculate their rates of growth with a repeat sample you can then plot the growth trajectory of these clones and estimate the latency to a potential clinical manifestation, thus offering opportunities for early preventive strategies,” said Jyoti Nangalia, MBBChir, senior study author, and cancer research UK clinician scientist at Sanger Institute, in a virtual presentation of the data.

The study comprised a cohort of 10 patients with essential thrombocythemia (ET), polycythemia vera (PV), and myelofibrosis (MF), with a median age of 48.5 (range, 20-76).

Each patients’ peripheral blood and bone marrow samples were grown into single cell–derived hematopoietic colonies. Each colony underwent whole-genome sequencing. A total of 952 whole-genome sequences were produced, each reflecting that of the single cell from which the colony was derived.

“Right from the start of life, as our cells are dividing, mutations are being acquired, and they’re being passed down from generation to generation such that at any one time, the mutations within individual cells represent natural bar codes that can be used to trace back the ancestry of those cells right to the start of life, and so we did this in MPNs,” said Nangalia.

Phylogenetic trees were drawn based on the somatic mutations that had been identified. Driver mutations were subsequently assigned to the tree and evaluated for appearance patterns across each colony, reflecting the relative development of the driver mutations in each patient.

Because the total number of somatic mutations in an individual colony was shown to correlate with age, investigators converted the relative development of mutations to absolute development to more accurately understand clonal evolution.

“Our blood stem cells require mutations throughout life, roughly 18 mutations across the genome per year. Therefore, by applying patient-specific mutation rates and clone-specific mutation rates, we were able to put a start time and an end time to each individual branch across the phylogenetic trees in our cohort,” said Nangalia.

In the first patient who had been diagnosed with ET at the age of 21, the JAK2 mutation was acquired early, timing between 6.2 weeks post-conception and 1.3 years of age. In the phylogenetic tree, the branching downstream of JAK2 demonstrated how the single stem cell that acquired JAK2 expanded into a clone of stem cells in rapid succession.

Similarly, in the second patient diagnosed with PV at the age of 31, the JAK2 mutation was acquired early, timing between 4.2 weeks post-conception and 8.6 years of age. Clonal expansion also demonstrated a DNMT3A mutation, which is the most common mutation in age-related clonal hematopoiesis, said Nangalia. However, in this patient, the mutation was also acquired early, timing between 4.6 weeks post-conception and 7.8 years of age, growing slowly before reaching a detectable clonal fraction.

In some patients, the JAK2 and DNMT3A mutations were acquired very early, including in utero. In one patient diagnosed with PV at age 33, the JAK2 mutation was acquired between 9.1 weeks post-conception and 4.1 months after birth, and the DNMT3A mutation was acquired between 19.4 weeks and 22.2 weeks post-conception. Another patient with PV showed that the DNMT3A mutation was acquired as early as between 1.2 weeks and 7.9 weeks post-conception.

In the patient with PV diagnosed at age 33, evidence of clonal evolution in the MPN clone was shown over 3 decades, cascading from a JAK2 mutation 4.1 months post-birth to homozygous JAK2 at 17.8 years to 1q amplification at 33 years.

In other patients, JAK2 was acquired as the second driver mutation in an already expanded DNMT3A-mutated clone.

Notably, a patient diagnosed with ET at age 54 had acquired DNMT3A R882H by 2 years of age, which is one of the most common mutations found in acute myeloid leukemia, said Nangalia. However, in another patient who had been diagnosed with ET at 76 years of age, there was a driverless clonal expansion.

“We know that clonal hematopoiesis can often lack driver mutations and is thought to be the case in up to 50% of patients with clonal hematopoiesis. Here, we showed that that also has a single-cell origin. What is driving clonal expansion in this individual in that particular clone, we do not know,” said Nangalia.

Across the cohort, recurrent or similar genetic aberrations were found in individual patients. A patient diagnosed with PV at age 53, who acquired a JAK2 exon 12 mutation in childhood, developed 3 independent DNMT3A mutations and 4 independent homozygous acquisitions stemming from their JAK2 exon 12 mutation.

Other patients demonstrated evidence of independent acquisitions of 1q amplifications, leading to myelofibrotic transformation. Another patient demonstrated evidence of multiple independent acquisitions of JAK2 V617F, suggesting that factors other than driver mutations, such as the patient’s germline or microenvironment within the bone marrow, also influence clonal evolution.

In the second portion of the study, investigators revisited the patients for whom phylogenetic trees had been drawn and re-sequenced the mutations that had been identified in the phylogenetic trees in the whole blood.

Combining the mutant clonal fractions in blood with the pattern of branching in the trees, investigators calculated the rate at which the clones had been growing over each patient’s lifespan.

Clone rates varied significantly. For example, in a patient with 3 mutant clones that had been acquired in utero––DNMT3A, JAK2, and JAK2/TET2––the rates of annual growth were 9% (95% CI, 5%-25%), 67% (95% CI, 6%-246%), and 233% (95% CI, 143%-360%), respectively, the latter of which translates to a doubling-in-size time of every 7 months.

Regarding clones that had the same genetic makeup, such as clones consisting solely of the JAK2 V617F driver mutation, the annual growth rate varied among individual patients, ranging from 18% (95% CI, 13%-23%) to 68% (95% CI, 41%-95%).

“This again suggests that there are factors other than JAK2 that determine the consequences of acquiring it in individual patients,” said Nangalia.

Additional results revealed that the rate of growth was associated with the time of diagnosis. In patients with slow growth rates of less than 50%, over 50 years had gone by prior to diagnosis, whereas in patients with growth rates over 100%, it took less than 10 years before a diagnosis was made.

In retroactively calculating what the clonal fractions would have been leading up to diagnosis, the slow growing JAK2 clones could have been detected with sensitive assays 40 years prior to diagnosis and up to 10 years before diagnosis for faster growing clones.

Providing additional perspective during a press briefing, Robert Brodsky, MD, moderator, and director of the Division of Hematology at Johns Hopkin Medicine stated, “These results suggest that there may be untapped opportunities to detect these conditions much earlier and potentially intervene and prevent disease development.”

_Reference

_{Williams N, Lee J, Moore L, et al. Driver mutation acquisition in utero and childhood followed by lifelong clonal evolution underlie myeloproliferative neoplasms. Presented at: 2020 ASH Annual Meeting & Exposition; December 5-8, 2020; virtual. Abstract LBA-1.}