# Elucidating Mechanisms of Therapy-Resistance to Interferon-alfa in Myeloproliferative Neoplasm Stem Cells

> **NIH NIH R01** · WEILL MEDICAL COLL OF CORNELL UNIV · 2024 · $686,979

## Abstract

PROJECT SUMMARY
Recombinant interferon-alpha (IFN) remains a highly effective therapy for patients with myeloproliferative
neoplasms (MPN). We recently identified that patients with CALR-mutated MPN frequently exhibit normalization
of blood counts (i.e. clinical response), but often do not exhibit a decrease in tumor burden (i.e. molecular
response), providing an informative model to decipher the mechanisms of therapy-resistance to IFN.
Interrogating the molecular impact of IFN on human MPN stem cells may reveal critical insights into mechanisms
of therapy-resistance. Thus, we applied our innovative Genotyping of Transcriptomes (GoT) platform – that
captures the mutation status and single-cell whole transcriptomes (scRNA-seq) within the same cells – CD34+
cells from serial bone marrow (BM) aspirates from patients with CALR-mutated MPN treated with IFN. Strikingly,
we observed that IFN caused major shifts in the differentiation landscapes, distinctly in the mutated and wildtype
progenitors: IFN exposure on wildtype cells resulted in a large expansion of lymphoid progenitors, while the
mutated cells, in contrast, displayed an expansion of the granulo-monocytic (GM) progenitors (with a less striking
expansion of the lymphoid compartment). Our preliminary data indicate that (1) the GM differentiation bias of
CALR-mutated stem cells may underlie therapy-resistance, and that (2) the CALR-mutation induced UPR may
prime the mutated stem cells toward the GM lineage and play a role in therapy-resistance. To interrogate these
hypotheses, we will determine the transcription factor (TF) networks that govern the IFN-induced differentiation
shifts by applying a novel single-cell multi-omics platform that captures RNA-seq, chromatin accessibility and
somatic genotyping within the same thousands of single cells (GoT-ATAC) to the same IFN-treated cohort (Aim
1a), and by targeting these TF networks in mouse models (Aim 1b). We will define the role of UPR in therapy-
resistance in treated CALR-mutated cells through GoT-ATAC and chromatin binding assays (Aim 2a) and by
assessing perturbations to the UPR pathways in mouse models (Aim 2b). Finally, we will determine the impact
of co-mutations in DNMT3A or ASXL1 in therapy-resistance to IFN in CALR-mutated MPN via application of
single-cell multi-omics platforms to clinical samples (Aim 3a) and interrogation of IFN effects on novel mouse
models with double mutations (Aim 3b). The project is centered on a conceptually innovative framework in which
we superimpose neoplastic and normal hematopoietic development within the same individuals to define how
therapy reshapes differentiation topographies, as a function of mutation status and cell identity. This conceptual
innovation is enabled by technical innovations in single-cell multi-omics platforms applied to compelling clinical
cohorts, coupled with functional assessments in novel mouse models. These studies have the potential to
uncover new insights into the mechanisms of...

## Key facts

- **NIH application ID:** 10901939
- **Project number:** 5R01HL167139-02
- **Recipient organization:** WEILL MEDICAL COLL OF CORNELL UNIV
- **Principal Investigator:** Ann Mullally
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $686,979
- **Award type:** 5
- **Project period:** 2023-08-15 → 2027-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10901939

## Citation

> US National Institutes of Health, RePORTER application 10901939, Elucidating Mechanisms of Therapy-Resistance to Interferon-alfa in Myeloproliferative Neoplasm Stem Cells (5R01HL167139-02). Retrieved via AI Analytics 2026-08-11 from https://api.ai-analytics.org/grant/nih/10901939. Licensed CC0.

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