Quantitation of T-cell receptor frequencies by competitive polymerase chain reaction: Dynamics of T-cell clonotype frequencies in an expanding tumor-infiltrating lymphocyte culture

Mark D. McKee, Timothy M. Clay, Rochelle A. Diamond, Steven A. Rosenberg, Michael I. Nishimura

Research output: Contribution to journalArticle

Abstract

The use of T-cell receptor (TCR) genes as markers for antigen-reactive T cells is dependent on the ability of the TCR genes to rapidly identify antigen-reactive T-cell clonotypes in patient samples. We recently reported a competitive reverse-transcriptase polymerase chain reaction (cRT-PCR) method that can measure the frequency of individual TCRBV subfamilies and clonotypes in mixed lymphocyte populations more accurately than other semiquantitative PCR assays. However, it is impractical to measure changes in the absolute frequency of each TCRBV subfamily to identify those T cells with increasing frequency after antigen stimulation in vivo or in vitro. Therefore, we have modified our cRT-PCR method to more rapidly identify expanding T-cell populations by combining all of the TCRBV subfamily-specific competitors into a single sample to determine the relative abundance of each TCRBV subfamily. Using an expanding TIL 620 culture, we identified four TCRBV (BV2, BV12, BV17, and BV23) subfamilies that expanded over a 23-day period. These subfamilies accounted for 23% of the T cells in the day 35 culture and increased to 57%, 92%, and 80% of the days 44, 51, and 58 cultures respectively. Analysis of DNA sequences demonstrated that the observed expansion was caused primarily by a single clonotype within each subfamily. T cells expressing BV17 and BV23 recognized gp100 and MART-1 respectively. Therefore. this cRT-PCR method can detect expanding T-cell populations based solely on their TCRBV subfamily expression. Furthermore, T-cell expansion in a mixed TIL population was a good predictor of antigen reactivity.

Original languageEnglish (US)
Pages (from-to)419-429
Number of pages11
JournalJournal of Immunotherapy
Volume23
Issue number4
DOIs
StatePublished - Jul 2000
Externally publishedYes

Fingerprint

Tumor-Infiltrating Lymphocytes
T-Cell Antigen Receptor
T-Lymphocytes
Polymerase Chain Reaction
Reverse Transcriptase Polymerase Chain Reaction
T-Cell Receptor Genes
Antigens
Population
Differentiation Antigens
DNA Sequence Analysis
Lymphocytes

Keywords

  • Competitive PCR
  • Melanoma
  • T-cell receptor
  • TIL

ASJC Scopus subject areas

  • Cancer Research
  • Pharmacology
  • Immunology

Cite this

Quantitation of T-cell receptor frequencies by competitive polymerase chain reaction : Dynamics of T-cell clonotype frequencies in an expanding tumor-infiltrating lymphocyte culture. / McKee, Mark D.; Clay, Timothy M.; Diamond, Rochelle A.; Rosenberg, Steven A.; Nishimura, Michael I.

In: Journal of Immunotherapy, Vol. 23, No. 4, 07.2000, p. 419-429.

Research output: Contribution to journalArticle

McKee, Mark D. ; Clay, Timothy M. ; Diamond, Rochelle A. ; Rosenberg, Steven A. ; Nishimura, Michael I. / Quantitation of T-cell receptor frequencies by competitive polymerase chain reaction : Dynamics of T-cell clonotype frequencies in an expanding tumor-infiltrating lymphocyte culture. In: Journal of Immunotherapy. 2000 ; Vol. 23, No. 4. pp. 419-429.
@article{424d5d56261341c99964e7f9976e7a29,
title = "Quantitation of T-cell receptor frequencies by competitive polymerase chain reaction: Dynamics of T-cell clonotype frequencies in an expanding tumor-infiltrating lymphocyte culture",
abstract = "The use of T-cell receptor (TCR) genes as markers for antigen-reactive T cells is dependent on the ability of the TCR genes to rapidly identify antigen-reactive T-cell clonotypes in patient samples. We recently reported a competitive reverse-transcriptase polymerase chain reaction (cRT-PCR) method that can measure the frequency of individual TCRBV subfamilies and clonotypes in mixed lymphocyte populations more accurately than other semiquantitative PCR assays. However, it is impractical to measure changes in the absolute frequency of each TCRBV subfamily to identify those T cells with increasing frequency after antigen stimulation in vivo or in vitro. Therefore, we have modified our cRT-PCR method to more rapidly identify expanding T-cell populations by combining all of the TCRBV subfamily-specific competitors into a single sample to determine the relative abundance of each TCRBV subfamily. Using an expanding TIL 620 culture, we identified four TCRBV (BV2, BV12, BV17, and BV23) subfamilies that expanded over a 23-day period. These subfamilies accounted for 23{\%} of the T cells in the day 35 culture and increased to 57{\%}, 92{\%}, and 80{\%} of the days 44, 51, and 58 cultures respectively. Analysis of DNA sequences demonstrated that the observed expansion was caused primarily by a single clonotype within each subfamily. T cells expressing BV17 and BV23 recognized gp100 and MART-1 respectively. Therefore. this cRT-PCR method can detect expanding T-cell populations based solely on their TCRBV subfamily expression. Furthermore, T-cell expansion in a mixed TIL population was a good predictor of antigen reactivity.",
keywords = "Competitive PCR, Melanoma, T-cell receptor, TIL",
author = "McKee, {Mark D.} and Clay, {Timothy M.} and Diamond, {Rochelle A.} and Rosenberg, {Steven A.} and Nishimura, {Michael I.}",
year = "2000",
month = "7",
doi = "10.1097/00002371-200007000-00005",
language = "English (US)",
volume = "23",
pages = "419--429",
journal = "Journal of Immunotherapy",
issn = "1524-9557",
publisher = "Lippincott Williams and Wilkins",
number = "4",

}

TY - JOUR

T1 - Quantitation of T-cell receptor frequencies by competitive polymerase chain reaction

T2 - Dynamics of T-cell clonotype frequencies in an expanding tumor-infiltrating lymphocyte culture

AU - McKee, Mark D.

AU - Clay, Timothy M.

AU - Diamond, Rochelle A.

AU - Rosenberg, Steven A.

AU - Nishimura, Michael I.

PY - 2000/7

Y1 - 2000/7

N2 - The use of T-cell receptor (TCR) genes as markers for antigen-reactive T cells is dependent on the ability of the TCR genes to rapidly identify antigen-reactive T-cell clonotypes in patient samples. We recently reported a competitive reverse-transcriptase polymerase chain reaction (cRT-PCR) method that can measure the frequency of individual TCRBV subfamilies and clonotypes in mixed lymphocyte populations more accurately than other semiquantitative PCR assays. However, it is impractical to measure changes in the absolute frequency of each TCRBV subfamily to identify those T cells with increasing frequency after antigen stimulation in vivo or in vitro. Therefore, we have modified our cRT-PCR method to more rapidly identify expanding T-cell populations by combining all of the TCRBV subfamily-specific competitors into a single sample to determine the relative abundance of each TCRBV subfamily. Using an expanding TIL 620 culture, we identified four TCRBV (BV2, BV12, BV17, and BV23) subfamilies that expanded over a 23-day period. These subfamilies accounted for 23% of the T cells in the day 35 culture and increased to 57%, 92%, and 80% of the days 44, 51, and 58 cultures respectively. Analysis of DNA sequences demonstrated that the observed expansion was caused primarily by a single clonotype within each subfamily. T cells expressing BV17 and BV23 recognized gp100 and MART-1 respectively. Therefore. this cRT-PCR method can detect expanding T-cell populations based solely on their TCRBV subfamily expression. Furthermore, T-cell expansion in a mixed TIL population was a good predictor of antigen reactivity.

AB - The use of T-cell receptor (TCR) genes as markers for antigen-reactive T cells is dependent on the ability of the TCR genes to rapidly identify antigen-reactive T-cell clonotypes in patient samples. We recently reported a competitive reverse-transcriptase polymerase chain reaction (cRT-PCR) method that can measure the frequency of individual TCRBV subfamilies and clonotypes in mixed lymphocyte populations more accurately than other semiquantitative PCR assays. However, it is impractical to measure changes in the absolute frequency of each TCRBV subfamily to identify those T cells with increasing frequency after antigen stimulation in vivo or in vitro. Therefore, we have modified our cRT-PCR method to more rapidly identify expanding T-cell populations by combining all of the TCRBV subfamily-specific competitors into a single sample to determine the relative abundance of each TCRBV subfamily. Using an expanding TIL 620 culture, we identified four TCRBV (BV2, BV12, BV17, and BV23) subfamilies that expanded over a 23-day period. These subfamilies accounted for 23% of the T cells in the day 35 culture and increased to 57%, 92%, and 80% of the days 44, 51, and 58 cultures respectively. Analysis of DNA sequences demonstrated that the observed expansion was caused primarily by a single clonotype within each subfamily. T cells expressing BV17 and BV23 recognized gp100 and MART-1 respectively. Therefore. this cRT-PCR method can detect expanding T-cell populations based solely on their TCRBV subfamily expression. Furthermore, T-cell expansion in a mixed TIL population was a good predictor of antigen reactivity.

KW - Competitive PCR

KW - Melanoma

KW - T-cell receptor

KW - TIL

UR - http://www.scopus.com/inward/record.url?scp=0033936778&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0033936778&partnerID=8YFLogxK

U2 - 10.1097/00002371-200007000-00005

DO - 10.1097/00002371-200007000-00005

M3 - Article

C2 - 10916751

AN - SCOPUS:0033936778

VL - 23

SP - 419

EP - 429

JO - Journal of Immunotherapy

JF - Journal of Immunotherapy

SN - 1524-9557

IS - 4

ER -