Fluorescence Lifetime Measurement of Prefibrillar Sickle Hemoglobin Oligomers as a Platform for Drug Discovery in Sickle Cell Disease

Nagamani Vunnam; Scott Hansen; Maryjane Olivia Been; Chih Hung Lo; Anil K. Pandey; Carolyn N. Paulsen; John A. Rohde; David D. Thomas; Jonathan N. Sachs; David K. Wood

doi:10.1021/acs.biomac.2c00671

Fluorescence Lifetime Measurement of Prefibrillar Sickle Hemoglobin Oligomers as a Platform for Drug Discovery in Sickle Cell Disease

Nagamani Vunnam, Scott Hansen, Maryjane Olivia Been, Chih Hung Lo, Anil K. Pandey, Carolyn N. Paulsen, John A. Rohde, David D. Thomas, Jonathan N. Sachs, David K. Wood

Research output: Contribution to journal › Article › peer-review

5 Scopus citations

Abstract

The molecular origin of sickle cell disease (SCD) has been known since 1949, but treatments remain limited. We present the first high-throughput screening (HTS) platform for discovering small molecules that directly inhibit sickle hemoglobin (HbS) oligomerization and improve blood flow, potentially overcoming a long-standing bottleneck in SCD drug discovery. We show that at concentrations far below the threshold for nucleation and rapid polymerization, deoxygenated HbS forms small assemblies of multiple α₂β₂tetramers. Our HTS platform leverages high-sensitivity fluorescence lifetime measurements that monitor these temporally stable prefibrillar HbS oligomers. We show that this approach is sensitive to compounds that inhibit HbS polymerization with or without modulating hemoglobin oxygen binding affinity. We also report the results of a pilot small-molecule screen in which we discovered and validated several novel inhibitors of HbS oligomerization.

Original language	English (US)
Pages (from-to)	3822-3830
Number of pages	9
Journal	Biomacromolecules
Volume	23
Issue number	9
DOIs	https://doi.org/10.1021/acs.biomac.2c00671
State	Published - Sep 12 2022

Bibliographical note

Funding Information:
We thank the patients at Children’s Hospital and Clinics of Minnesota for their generous blood donations and the sickle care team, including Dr. Steve Nelson, Ashley Kinsella, Pauline Mitby, Ali Koste, Rachel Hinsch, and Emily Olson for blood sample collection. We acknowledge funding under Grants R01HL132906 (D.K.W.), R21HL152313 (J.N.S., D.K.W.), and R35GM131814 (J.N.S.). Portions of this work were conducted in the Minnesota Nano Center, which is supported by the National Science Foundation through the National Nanotechnology Coordinated Infrastructure (NNCI) under Award Number ECCS-2025124.

Publisher Copyright:
© 2022 American Chemical Society. All rights reserved.

Keywords

Anemia, Sickle Cell/drug therapy
Drug Discovery
Hemoglobin, Sickle/chemistry
Hemoglobins
Humans
Oxygen/metabolism

PubMed: MeSH publication types

Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, N.I.H., Extramural

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10.1021/acs.biomac.2c00671

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title = "Fluorescence Lifetime Measurement of Prefibrillar Sickle Hemoglobin Oligomers as a Platform for Drug Discovery in Sickle Cell Disease",

abstract = "The molecular origin of sickle cell disease (SCD) has been known since 1949, but treatments remain limited. We present the first high-throughput screening (HTS) platform for discovering small molecules that directly inhibit sickle hemoglobin (HbS) oligomerization and improve blood flow, potentially overcoming a long-standing bottleneck in SCD drug discovery. We show that at concentrations far below the threshold for nucleation and rapid polymerization, deoxygenated HbS forms small assemblies of multiple α2β2tetramers. Our HTS platform leverages high-sensitivity fluorescence lifetime measurements that monitor these temporally stable prefibrillar HbS oligomers. We show that this approach is sensitive to compounds that inhibit HbS polymerization with or without modulating hemoglobin oxygen binding affinity. We also report the results of a pilot small-molecule screen in which we discovered and validated several novel inhibitors of HbS oligomerization.",

keywords = "Anemia, Sickle Cell/drug therapy, Drug Discovery, Hemoglobin, Sickle/chemistry, Hemoglobins, Humans, Oxygen/metabolism",

author = "Nagamani Vunnam and Scott Hansen and Been, {Maryjane Olivia} and Lo, {Chih Hung} and Pandey, {Anil K.} and Paulsen, {Carolyn N.} and Rohde, {John A.} and Thomas, {David D.} and Sachs, {Jonathan N.} and Wood, {David K.}",

note = "Funding Information: We thank the patients at Children{\textquoteright}s Hospital and Clinics of Minnesota for their generous blood donations and the sickle care team, including Dr. Steve Nelson, Ashley Kinsella, Pauline Mitby, Ali Koste, Rachel Hinsch, and Emily Olson for blood sample collection. We acknowledge funding under Grants R01HL132906 (D.K.W.), R21HL152313 (J.N.S., D.K.W.), and R35GM131814 (J.N.S.). Portions of this work were conducted in the Minnesota Nano Center, which is supported by the National Science Foundation through the National Nanotechnology Coordinated Infrastructure (NNCI) under Award Number ECCS-2025124. Publisher Copyright: {\textcopyright} 2022 American Chemical Society. All rights reserved.",

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doi = "10.1021/acs.biomac.2c00671",

language = "English (US)",

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AU - Vunnam, Nagamani

AU - Hansen, Scott

AU - Been, Maryjane Olivia

AU - Lo, Chih Hung

AU - Pandey, Anil K.

AU - Paulsen, Carolyn N.

AU - Rohde, John A.

AU - Thomas, David D.

AU - Sachs, Jonathan N.

AU - Wood, David K.

N1 - Funding Information: We thank the patients at Children’s Hospital and Clinics of Minnesota for their generous blood donations and the sickle care team, including Dr. Steve Nelson, Ashley Kinsella, Pauline Mitby, Ali Koste, Rachel Hinsch, and Emily Olson for blood sample collection. We acknowledge funding under Grants R01HL132906 (D.K.W.), R21HL152313 (J.N.S., D.K.W.), and R35GM131814 (J.N.S.). Portions of this work were conducted in the Minnesota Nano Center, which is supported by the National Science Foundation through the National Nanotechnology Coordinated Infrastructure (NNCI) under Award Number ECCS-2025124. Publisher Copyright: © 2022 American Chemical Society. All rights reserved.

PY - 2022/9/12

Y1 - 2022/9/12

N2 - The molecular origin of sickle cell disease (SCD) has been known since 1949, but treatments remain limited. We present the first high-throughput screening (HTS) platform for discovering small molecules that directly inhibit sickle hemoglobin (HbS) oligomerization and improve blood flow, potentially overcoming a long-standing bottleneck in SCD drug discovery. We show that at concentrations far below the threshold for nucleation and rapid polymerization, deoxygenated HbS forms small assemblies of multiple α2β2tetramers. Our HTS platform leverages high-sensitivity fluorescence lifetime measurements that monitor these temporally stable prefibrillar HbS oligomers. We show that this approach is sensitive to compounds that inhibit HbS polymerization with or without modulating hemoglobin oxygen binding affinity. We also report the results of a pilot small-molecule screen in which we discovered and validated several novel inhibitors of HbS oligomerization.

AB - The molecular origin of sickle cell disease (SCD) has been known since 1949, but treatments remain limited. We present the first high-throughput screening (HTS) platform for discovering small molecules that directly inhibit sickle hemoglobin (HbS) oligomerization and improve blood flow, potentially overcoming a long-standing bottleneck in SCD drug discovery. We show that at concentrations far below the threshold for nucleation and rapid polymerization, deoxygenated HbS forms small assemblies of multiple α2β2tetramers. Our HTS platform leverages high-sensitivity fluorescence lifetime measurements that monitor these temporally stable prefibrillar HbS oligomers. We show that this approach is sensitive to compounds that inhibit HbS polymerization with or without modulating hemoglobin oxygen binding affinity. We also report the results of a pilot small-molecule screen in which we discovered and validated several novel inhibitors of HbS oligomerization.

KW - Anemia, Sickle Cell/drug therapy

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KW - Hemoglobin, Sickle/chemistry

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KW - Oxygen/metabolism

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ER -

Fluorescence Lifetime Measurement of Prefibrillar Sickle Hemoglobin Oligomers as a Platform for Drug Discovery in Sickle Cell Disease

Abstract

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Keywords

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