LRRK2 Inhibitor Hits Target, Appears Safe for Parkinson’s

Mutations in leucine-rich repeat kinase 2 increase a person's risk of developing Parkinson's disease. In the June 8 Science Translational Medicine, Danna Jennings and co-workers at Denali Therapeutics, South San Francisco, published preclinical and the first clinical trial data on DNL201, a LRRK2 inhibitor.

  • DNL201 quelled LRRK2 kinase activity in blood cells.
  • It crossed the blood-brain barrier and elicited no serious side effects.
  • A Phase 2 trial of a modified version began in April.

In cell culture, the small molecule blocked LRRK2 activity and restored lysosomal function. In macaques, DNL201 reduced LRRK2 activity in peripheral blood monocytes without apparent toxicity.

In a Phase 1 study in healthy adults and a Phase 1b in people with PD, DNL201 reduced the amount of phosphorylated LRRK2 in the blood, normalized levels of a lysosomal marker in the urine, and reached the same drug concentration in the cerebrospinal fluid as in the blood. Over the course of both 28-day trials, the drug caused mild to moderate side effects.

“[This] study represents a major step forward for LRRK2 kinase inhibition as a therapeutic strategy in humans, demonstrating target engagement and, crucially, safety and tolerability,” Patrick Lewis, University College London, wrote in an accompanying editorial.

Others were cautiously optimistic. “As with many other drugs under investigation, DNL201 has promising results in preclinical models and appears relatively safe in short-term human studies, but it still has a long way to go,” Warren Olanow, Clintrex Research Corp, Sarasota, Florida, told Alzforum. Dario Alessi, University of Dundee, Scotland, agreed. “Though this is magnificent preclinical and early clinical work, we don't yet know whether this drug will slow disease progression and if it will benefit patients without a LRRK2 mutation,” he told Alzforum.

In PD, overactive LRRK2 disrupts lysosomes, promoting the spread of misfolded α-synuclein, the principal component of Lewy bodies (Sep 2018 news). In rat models of the disease, genetically knocking down this kinase, or calming it with the inhibitor PF-06447475, reduced α-synuclein aggregation and dopaminergic neuron loss (Daher et al., 2014; Daher et al., 2015).

Now, Jennings and co-first authors Sarah Huntwork-Rodriguez and Anastasia Henry of Denali report that DNL201 tempered LRRK2 activity and normalized lysosomal autophagy in mouse and human cell cultures. Treated cortical neurons from mutant LRRK2 knock-in mice had less phosphorylated LRRK2 and less phosphorylated Rab10, an LRRK2 substrate. A decrease in both indicates a drop in the kinase’s activity. In astrocytes from the knock-ins, the inhibitor restored normal lysosomal protein degradation. In human neuroglioma cells overexpressing mutant LRRK2, DNL201 shrank bloated lysosomes back to their normal shape and size.

The scientists saw similar effects in nonhuman primates. Macaques intravenously injected with 16 mg/kg of DNL201 daily for 28 days had 80 percent less p-LRRK2 and p-Rab12, another LRRK2 substrate, in their blood cells and brain tissue. The drug reached a similar concentration in cerebrospinal fluid as in the blood, indicating that DNL201 easily entered the brain.

In a 39-week toxicity study, monkeys given as much as 32 mg/kg daily retained normal vital signs and kidney function. However, some cells in their lungs and kidneys became swollen. These cells returned to normal after the researchers stopped giving the drug to the animals. This cellular side effect has been reported in monkeys after systemic LRRK2 inhibition, and does not appear to affect breathing (Apr 2020 news). “Blocking 50-80 percent of LRRK2 activity, rather than all of it, may prevent some of the lung cell effects by allowing the kinase to maintain its housekeeping duties,” suggested Alessi.

How did DNL201 fare in people? It was tested in two trials: a Phase 1 in 122 healthy adults, ages 18-70, and a Phase 1b in 28 people who had mild to moderate PD. Of the latter, eight carried the common G2019S LRRK2 variant, which increases risk for the disease, though it is not 100 percent penetrant.

Healthy volunteers took 25, 50, 80, or 100 mg of DNL201 twice daily for 10 days. They gave samples of blood, CSF, and urine. The drug easily crossed the BBB, as indicated by a CSF-to-blood drug ratio of about one. In people who took the highest dose, plasma levels of p-LRRK2 and p-Rab10 dropped 80 percent and urinary levels of bis(monoacylglycerol) phosphate fell about 75 percent. This membrane lipid is a marker of lysosomal distress, and BMP in urine ticks up in LRRK2 mutation carriers (Alcalay et al., 2020).

The authors believe the drop in BMP indicates improved lysosome function. Alessi agreed. “BMP may be a very useful, noninvasive marker," he said. Olanow was more guarded. “While the peripheral biomarkers show target engagement, this doesn't guarantee that you will see the same effect in the brain,” he noted.

As for side effects, volunteers seemed to tolerate the drug well, with the most common adverse events being headache, dizziness, and nausea. Five volunteers withdrew; four had taken the drug, and three of them dropped out because of side effects.

In the Phase 1b trial, 12 PD participants took 50 mg, nine took 30 mg DNL201, and seven took placebo three times daily for 28 days. Blood levels of p-LRRK2 fell 55 and 85 percent after the low or high doses, respectively, while p-Rab10 levels plummeted 80 and 87 percent (see image below). Because LRRK2 mutation carriers and people with sporadic PD have twice as much p-Rab10 in their blood cells as do controls, the authors think LRRK2 activity should be at least halved to achieve a therapeutic effect. Urinary BMP dropped about 60 percent after high-dose DNL201, but a smaller decline after the low dose was not statistically significant.

LRRK2 Dynamics. In people with PD, baseline levels (BL) of p-Rab10 fell on day 1 of dosing with 30 mg (yellow) or 50 mg (orange) DNL201, and remained low on day 28. In people on placebo (gray line), p-Rab10 levels rose over the course of the trial. [Courtesy of Jennings et al., Science Translational Medicine, 2022.]

Most adverse events were mild or moderate; as with the other trial, the most common was headache. One PD volunteer withdrew due to moderate headache and low blood pressure. Lung and kidney function remained normal. As might be expected during this short trial, neither motor symptoms nor cognition changed, as judged by the Unified Parkinson’s Disease Rating Scale (UPDRS), the Non-Motor Symptoms Scale, or the Montreal Cognitive Assessment.

Denali has since dropped DNL201 for DNL151, which the company says has better pharmacokinetics. Two Phase 1 trials, one in 186 healthy adults and another in 36 people with PD, finished in February 2021 and December 2020, respectively, though no data has yet been published.

A Phase 2 trial of DNL151 began in April. About 640 people with early stage PD will take 225 mg of the drug, or placebo, daily for 48 to 144 weeks. As a primary outcome, the trial will measure time until symptoms worsen on the UPDRS. Secondary endpoints include adverse events, change from baseline UPDRS score, and time to worsening of daily activities. The trial is slated to wrap up in 2025.

Blocking LRRK2 kinase activity may also correct lysosome dysfunction caused by mutations other than those in the kinase. Jennings and colleagues report that DNL201 partially restored lysosome protein turnover in fibroblasts from a person with Gaucher disease, a lysosomal storage disorder. Gaucher’s can be caused by variants in the lysosomal enzyme glucocerebrosidase (GBA). People with GBA mutations are also at increased risk of PD, and there is some evidence that inhibiting LRRK2 boosts glucocerebrosidase activity, though it is not clear if that’s what DNL201 did in this case (Sanyal et al., 2020; Ysselstein et al., 2019).—Chelsea Weidman Burke

Comments

    • User Profile Image Brian Fiske
      The Michael J. Fox Foundation for Parkinson's Research
    • Posted:

    The findings reported by Jennings et al. represent a comprehensive summary of data so far on DNL201, a leading treatment in development for Parkinson’s disease. Given their pioneering role in moving therapies targeting LRRK2 into clinical testing, Denali and its collaborators should be congratulated for providing this detailed look across both nonclinical and clinical experience with DNL201. Many in the field, including in the patient community, are eagerly watching the progress of these important studies.

    As the authors point out (and as does Lewis in the companion perspective), many questions still remain, and more data will be needed to further assess longer-term safety of LRRK2 inhibitors, along with future readouts on possible efficacy in slowing disease progression. That LRRK2 kinase inhibition may also influence biology linked to other forms of PD associated with GBA1 mutations, as reported by Jennings et al., is intriguing and a topic of emerging interest among the research community.

    At The Michael J. Fox Foundation for Parkinson’s Research, we have made significant investments over the years to drive increased understanding and better measurement of LRRK2 and its cellular substrates and worked closely with Denali and other industry leaders to understand the needs for enabling informative, careful, and scientifically rigorous clinical testing of LRRK2 inhibitors. I am grateful to Denali and all those in the research and drug development communities who are working hard to translate LRRK2 into potentially promising treatments for Parkinson’s disease.

    View all comments by Brian Fiske

  2. At present, there are no available therapies that can slow the progression of Parkinson’s disease. Pathogenic mutations in the LRRK2 gene increase protein kinase activity and represent the most common genetic risk factor for Parkinson’s disease. Importantly, a large body of work has demonstrated that pharmacological inhibition of LRRK2 can limit the detrimental consequences of mutations across a variety of experimental models. In this study, Jennings et al. share their findings from the first-in-human clinical trials of the LRRK2 kinase inhibitor, DNL201.

    Their preclinical data builds upon previous work linking LRRK2 mutations with lysosomal dysfunction (Eguchi et al., 2018). The authors demonstrate that DNL201 is capable of reversing stress-associated lysosomal enlargements in a cell line expressing the LRRK2 G2019S mutation. Interestingly, the authors expand the scope of their study to investigate fibroblasts derived from Gaucher’s patients carrying GBA1 mutations. Treatment with DNL201 appeared to partially correct dysfunctional lysosomal protein turnover. These findings are consistent with recent data suggesting inhibition of LRRK2 kinase activity can treat lysosomal dysfunction that is not caused by LRRK2-mutations (Sanyal et al., 2020). 

    Their study also examined PBMCs derived from G2019S mutation carriers as well as patients with sporadic PD and healthy controls. Jennings et al. report a roughly twofold increase in pT73 Rab10 in patients with sporadic PD and G2019S mutation carriers alike. It is noteworthy, however, that the primary data, by the way of blots, is absent from this manuscript. Findings surrounding blood-based analysis of Phospho-Rab10 from LRRK2-mutation carriers are mixed. Fan et al. (2021) recently reported elevated Phospho-Rab10 are observed in carriers of the R1441C mutation but not G2019S.

    A promising finding from this study is that DNL201 is well-tolerated in both PD patients and healthy controls. Furthermore, Jennings et al. demonstrate a dose-dependent inhibition of LRRK2 kinase activity (in whole blood). Their data represents an important milestone in the development of the first disease-modifying therapeutic for Parkinson’s disease. A recurring concern from studies of previous LRRK2 inhibitors is the presence of lung pathology in rodents and nonhuman primates (Baptista et al., 2020). Encouragingly, Jennings et al. demonstrate that DNL201 did not appear to have any impact on lung function across all doses investigated during the trial period. As the authors note, longer-term monitoring will be necessary to understand the impact of chronic dosing in follow-up studies. Additional studies are also needed to address target engagement of DNL201 in the human brain and the consequences of LRRK2 kinase inhibition and reversal of disrupted lysosomal functions.

    References:

    Eguchi T, Kuwahara T, Sakurai M, Komori T, Fujimoto T, Ito G, Yoshimura SI, Harada A, Fukuda M, Koike M, Iwatsubo T. LRRK2 and its substrate Rab GTPases are sequentially targeted onto stressed lysosomes and maintain their homeostasis. Proc Natl Acad Sci U S A. 2018 Sep 12; PubMed.

    Sanyal A, Novis HS, Gasser E, Lin S, LaVoie MJ. LRRK2 Kinase Inhibition Rescues Deficits in Lysosome Function Due to Heterozygous GBA1 Expression in Human iPSC-Derived Neurons. Front Neurosci. 2020;14:442. Epub 2020 May 15 PubMed.

    Fan Y, Nirujogi RS, Garrido A, Ruiz-Martínez J, Bergareche-Yarza A, Mondragón-Rezola E, Vinagre-Aragón A, Croitoru I, Gorostidi Pagola A, Paternain Markinez L, Alcalay R, Hickman RA, Düring J, Gomes S, Pratuseviciute N, Padmanabhan S, Valldeoriola F, Pérez Sisqués L, Malagelada C, Ximelis T, Molina Porcel L, Martí MJ, Tolosa E, Alessi DR, Sammler EM. R1441G but not G2019S mutation enhances LRRK2 mediated Rab10 phosphorylation in human peripheral blood neutrophils. Acta Neuropathol. 2021 Sep;142(3):475-494. Epub 2021 Jun 14 PubMed.

    Baptista MA, Merchant K, Barrett T, Bhargava S, Bryce DK, Ellis JM, Estrada AA, Fell MJ, Fiske BK, Fuji RN, Galatsis P, Henry AG, Hill S, Hirst W, Houle C, Kennedy ME, Liu X, Maddess ML, Markgraf C, Mei H, Meier WA, Needle E, Ploch S, Royer C, Rudolph K, Sharma AK, Stepan A, Steyn S, Trost C, Yin Z, Yu H, Wang X, Sherer TB. LRRK2 inhibitors induce reversible changes in nonhuman primate lungs without measurable pulmonary deficits. Sci Transl Med. 2020 Apr 22;12(540) PubMed.

    View all comments by George Heaton View all comments by Zhenyu Yue

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References

Therapeutics Citations

  1. DNL201
  2. DNL151

News Citations

  1. Does LRRK2 Sweep α-Synuclein from the Cell?
  2. Sigh of Relief? Lung Effects of LRRK2 Inhibitors are Mild.

Research Models Citations

  1. LRRK2 G2019S KI Mouse

Paper Citations

  1. Daher JP, Volpicelli-Daley LA, Blackburn JP, Moehle MS, West AB. Abrogation of α-synuclein-mediated dopaminergic neurodegeneration in LRRK2-deficient rats. Proc Natl Acad Sci U S A. 2014 Jun 24;111(25):9289-94. Epub 2014 Jun 9 PubMed.
  2. Daher JP, Abdelmotilib HA, Hu X, Volpicelli-Daley LA, Moehle MS, Fraser KB, Needle E, Chen Y, Steyn SJ, Galatsis P, Hirst WD, West AB. Leucine-rich Repeat Kinase 2 (LRRK2) Pharmacological Inhibition Abates α-Synuclein Gene-induced Neurodegeneration. J Biol Chem. 2015 Aug 7;290(32):19433-44. Epub 2015 Jun 15 PubMed.
  3. Alcalay RN, Hsieh F, Tengstrand E, Padmanabhan S, Baptista M, Kehoe C, Narayan S, Boehme AK, Merchant K. Higher Urine bis(Monoacylglycerol)Phosphate Levels in LRRK2 G2019S Mutation Carriers: Implications for Therapeutic Development. Mov Disord. 2020 Jan;35(1):134-141. Epub 2019 Sep 10 PubMed.
  4. Sanyal A, DeAndrade MP, Novis HS, Lin S, Chang J, Lengacher N, Tomlinson JJ, Tansey MG, LaVoie MJ. Lysosome and Inflammatory Defects in GBA1-Mutant Astrocytes Are Normalized by LRRK2 Inhibition. Mov Disord. 2020 May;35(5):760-773. Epub 2020 Feb 8 PubMed.
  5. Ysselstein D, Nguyen M, Young TJ, Severino A, Schwake M, Merchant K, Krainc D. LRRK2 kinase activity regulates lysosomal glucocerebrosidase in neurons derived from Parkinson's disease patients. Nat Commun. 2019 Dec 5;10(1):5570. PubMed.

External Citations

  1. Phase 1
  2. Phase 1b
  3. one
  4. another
  5. Phase 2

Further Reading

Papers

  1. Wojewska DN, Kortholt A. LRRK2 Targeting Strategies as Potential Treatment of Parkinson's Disease. Biomolecules. 2021 Jul 26;11(8) PubMed.
  2. Tolosa E, Vila M, Klein C, Rascol O. LRRK2 in Parkinson disease: challenges of clinical trials. Nat Rev Neurol. 2020 Feb;16(2):97-107. Epub 2020 Jan 24 PubMed.

Primary Papers

  1. Jennings D, Huntwork-Rodriguez S, Henry AG, Sasaki JC, Meisner R, Diaz D, Solanoy H, Wang X, Negrou E, Bondar VV, Ghosh R, Maloney MT, Propson NE, Zhu Y, Maciuca RD, Harris L, Kay A, LeWitt P, King TA, Kern D, Ellenbogen A, Goodman I, Siderowf A, Aldred J, Omidvar O, Masoud ST, Davis SS, Arguello A, Estrada AA, de Vicente J, Sweeney ZK, Astarita G, Borin MT, Wong BK, Wong H, Nguyen H, Scearce-Levie K, Ho C, Troyer MD. Preclinical and clinical evaluation of the LRRK2 inhibitor DNL201 for Parkinson's disease. Sci Transl Med. 2022 Jun 8;14(648):eabj2658. PubMed.
  2. Lewis PA. A step forward for LRRK2 inhibitors in Parkinson's disease. Sci Transl Med. 2022 Jun 8;14(648):eabq7374. PubMed.

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