Convergence of acquired mutations and alternative splicing of CD19 enables resistance to CART-19 immunotherapy

Elena Sotillo, David M. Barrett, Kathryn L. Black, Asen Bagashev, Derek Oldridge, Glendon Wu, Robyn Sussman, Claudia Lanauze, Marco Ruella, Matthew R. Gazzara, Nicole M. Martinez, Colleen T. Harrington, Elaine Y. L. Chung, Jessica Perazzelli, Ted J. Hofmann, Shannon L. Maude, Pichai Raman, Alejandro Barrera, Saar Gill, Simon F. Lacey & 10 others Jan J. Melenhorst, David Allman, Elad Jacoby, Terry Fry, Crystal Mackall, Yoseph Barash, Kristen W. Lynch, John M. Maris, Stephan A. Grupp, Andrei Thomas-Tikhonenko

Research output: Contribution to journalArticle

286 Citations (Scopus)

Abstract

The CD19 antigen, expressed on most B-cell acute lymphoblastic leukemias (B-ALL), can be targeted with chimeric antigen receptor–armed T cells (CART-19), but relapses with epitope loss occur in 10% to 20% of pediatric responders. We detected hemizygous deletions spanning the CD19 locus and de novo frameshift and missense mutations in exon 2 of CD19 in some relapse samples. However, we also discovered alternatively spliced CD19 mRNA species, including one lacking exon 2. Pull-down/siRNA experiments identified SRSF3 as a splicing factor involved in exon 2 retention, and its levels were lower in relapsed B-ALL. Using genome editing, we demonstrated that exon 2 skipping bypasses exon 2 mutations in B-ALL cells and allows expression of the N-terminally truncated CD19 variant, which fails to trigger killing by CART-19 but partly rescues defects associated with CD19 loss. Thus, this mechanism of resistance is based on a combination of deleterious mutations and ensuing selection for alternatively spliced RNA isoforms. Significance: CART-19 yield 70% response rates in patients with B-ALL, but also produce escape variants. We discovered that the underlying mechanism is the selection for preexisting alternatively spliced CD19 isoforms with the compromised CART-19 epitope. This mechanism suggests a possibility of targeting alternative CD19 ectodomains, which could improve survival of patients with B-cell neoplasms.

Original languageEnglish (US)
Pages (from-to)1282-1295
Number of pages14
JournalCancer Discovery
Volume5
Issue number12
DOIs
StatePublished - Dec 1 2015
Externally publishedYes

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Alternative Splicing
Immunotherapy
Exons
B-Lymphocytes
Precursor Cell Lymphoblastic Leukemia-Lymphoma
Mutation
Epitopes
CD19 Antigens
RNA Isoforms
Recurrence
Frameshift Mutation
Missense Mutation
Small Interfering RNA
Protein Isoforms
Pediatrics
T-Lymphocytes
Antigens
Messenger RNA
Survival
Neoplasms

ASJC Scopus subject areas

  • Oncology

Cite this

Sotillo, E., Barrett, D. M., Black, K. L., Bagashev, A., Oldridge, D., Wu, G., ... Thomas-Tikhonenko, A. (2015). Convergence of acquired mutations and alternative splicing of CD19 enables resistance to CART-19 immunotherapy. Cancer Discovery, 5(12), 1282-1295. https://doi.org/10.1158/2159-8290.CD-15-1020

Convergence of acquired mutations and alternative splicing of CD19 enables resistance to CART-19 immunotherapy. / Sotillo, Elena; Barrett, David M.; Black, Kathryn L.; Bagashev, Asen; Oldridge, Derek; Wu, Glendon; Sussman, Robyn; Lanauze, Claudia; Ruella, Marco; Gazzara, Matthew R.; Martinez, Nicole M.; Harrington, Colleen T.; Chung, Elaine Y. L.; Perazzelli, Jessica; Hofmann, Ted J.; Maude, Shannon L.; Raman, Pichai; Barrera, Alejandro; Gill, Saar; Lacey, Simon F.; Melenhorst, Jan J.; Allman, David; Jacoby, Elad; Fry, Terry; Mackall, Crystal; Barash, Yoseph; Lynch, Kristen W.; Maris, John M.; Grupp, Stephan A.; Thomas-Tikhonenko, Andrei.

In: Cancer Discovery, Vol. 5, No. 12, 01.12.2015, p. 1282-1295.

Research output: Contribution to journalArticle

Sotillo, E, Barrett, DM, Black, KL, Bagashev, A, Oldridge, D, Wu, G, Sussman, R, Lanauze, C, Ruella, M, Gazzara, MR, Martinez, NM, Harrington, CT, Chung, EYL, Perazzelli, J, Hofmann, TJ, Maude, SL, Raman, P, Barrera, A, Gill, S, Lacey, SF, Melenhorst, JJ, Allman, D, Jacoby, E, Fry, T, Mackall, C, Barash, Y, Lynch, KW, Maris, JM, Grupp, SA & Thomas-Tikhonenko, A 2015, 'Convergence of acquired mutations and alternative splicing of CD19 enables resistance to CART-19 immunotherapy', Cancer Discovery, vol. 5, no. 12, pp. 1282-1295. https://doi.org/10.1158/2159-8290.CD-15-1020
Sotillo, Elena ; Barrett, David M. ; Black, Kathryn L. ; Bagashev, Asen ; Oldridge, Derek ; Wu, Glendon ; Sussman, Robyn ; Lanauze, Claudia ; Ruella, Marco ; Gazzara, Matthew R. ; Martinez, Nicole M. ; Harrington, Colleen T. ; Chung, Elaine Y. L. ; Perazzelli, Jessica ; Hofmann, Ted J. ; Maude, Shannon L. ; Raman, Pichai ; Barrera, Alejandro ; Gill, Saar ; Lacey, Simon F. ; Melenhorst, Jan J. ; Allman, David ; Jacoby, Elad ; Fry, Terry ; Mackall, Crystal ; Barash, Yoseph ; Lynch, Kristen W. ; Maris, John M. ; Grupp, Stephan A. ; Thomas-Tikhonenko, Andrei. / Convergence of acquired mutations and alternative splicing of CD19 enables resistance to CART-19 immunotherapy. In: Cancer Discovery. 2015 ; Vol. 5, No. 12. pp. 1282-1295.
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abstract = "The CD19 antigen, expressed on most B-cell acute lymphoblastic leukemias (B-ALL), can be targeted with chimeric antigen receptor–armed T cells (CART-19), but relapses with epitope loss occur in 10{\%} to 20{\%} of pediatric responders. We detected hemizygous deletions spanning the CD19 locus and de novo frameshift and missense mutations in exon 2 of CD19 in some relapse samples. However, we also discovered alternatively spliced CD19 mRNA species, including one lacking exon 2. Pull-down/siRNA experiments identified SRSF3 as a splicing factor involved in exon 2 retention, and its levels were lower in relapsed B-ALL. Using genome editing, we demonstrated that exon 2 skipping bypasses exon 2 mutations in B-ALL cells and allows expression of the N-terminally truncated CD19 variant, which fails to trigger killing by CART-19 but partly rescues defects associated with CD19 loss. Thus, this mechanism of resistance is based on a combination of deleterious mutations and ensuing selection for alternatively spliced RNA isoforms. Significance: CART-19 yield 70{\%} response rates in patients with B-ALL, but also produce escape variants. We discovered that the underlying mechanism is the selection for preexisting alternatively spliced CD19 isoforms with the compromised CART-19 epitope. This mechanism suggests a possibility of targeting alternative CD19 ectodomains, which could improve survival of patients with B-cell neoplasms.",
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AU - Sotillo, Elena

AU - Barrett, David M.

AU - Black, Kathryn L.

AU - Bagashev, Asen

AU - Oldridge, Derek

AU - Wu, Glendon

AU - Sussman, Robyn

AU - Lanauze, Claudia

AU - Ruella, Marco

AU - Gazzara, Matthew R.

AU - Martinez, Nicole M.

AU - Harrington, Colleen T.

AU - Chung, Elaine Y. L.

AU - Perazzelli, Jessica

AU - Hofmann, Ted J.

AU - Maude, Shannon L.

AU - Raman, Pichai

AU - Barrera, Alejandro

AU - Gill, Saar

AU - Lacey, Simon F.

AU - Melenhorst, Jan J.

AU - Allman, David

AU - Jacoby, Elad

AU - Fry, Terry

AU - Mackall, Crystal

AU - Barash, Yoseph

AU - Lynch, Kristen W.

AU - Maris, John M.

AU - Grupp, Stephan A.

AU - Thomas-Tikhonenko, Andrei

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