# BIOCHEMISTRY AND PATHOLOGY OF 5-FU/FOLATE THERAPY

> **NIH NIH R01** · UNIVERSITY OF SOUTHERN CALIFORNIA · 1992 · $190,515

## Abstract

This Proposal will study the hypothesis that pharmacodynamic studies of
whole extracts of tumor specimens of patients treated with 5-FU plus pure
(/)-leucovorin will be improved by our newly-developed methods for
quantitative histopathology.  Our biochemical data suggest that the goals
of 5-FU/leucovorin therapy are decreases in intratumoral thymidylate
synthase (TS) and expansion of H4PteGlu pools.  Tumors of patients with
breast and gastrointestinal cancer will be biopsied before and after 5-
FU(/)-leucovorin therapy and at tumor progression.  Equal hemi-cylinder
tumor biopsies will be taken for surgical pathology and for tissue
homogenization assays of TS, tetrahydrofolate, 5,10-
methylenetetrahydrofolate, dUMP, and FdUMP, by established [3H]FdUMP
ligand-binding assays, and for TS mRNA by the Danenberg assay.  TS protein
will also be quantitated by ELISA assay using a rabbit polyclonal anti-
human TS antibody preparation.  Pathology studies of 4-micron sections will
include immunoperoxidase antibody staining for TS, nuclear staining for
cell counts, common-leukocyte-antigen staining for white cells, and Ki 67
staining for S-phase and growth fraction determination.  Initial studies
will include comparison of fresh frozen vs. paraffin-embedded tissues.  The
antibodies used for TS immunostaining may also enable visualization of
H4PteGlun and 5,10-CH2-H4PteGlun pools, by forming TS-FdUMP-folate ternary
complexes in situ, using exogenous TS addition.  Quantitation of the
numbers, intensities, and areas of immunostaining will be done by digital
image analysis using video microscopy and JAVA software; a novel method for
validation of this approach will sue common-embedding of reference cells in
the specimen blocks for same-frame calibration of parameters.  Double-
labeling, such as immunofluorescence combined with immunoperoxidase
staining for TS will permit quantitation of TS antigen levels by cell type.
In addition, the image analysis system should be useful for quantitation of
cells visualized by in situ mRNA hybridization of transcripts of interest.

## Key facts

- **NIH application ID:** 3178852
- **Project number:** 2R01CA039629-07A2
- **Recipient organization:** UNIVERSITY OF SOUTHERN CALIFORNIA
- **SAM.gov UEI:** G88KLJR3KYT5
- **Principal Investigator:** COLIN PAUL SPEARS
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 1992
- **Award amount:** $190,515
- **Award type:** 2
- **Project period:** 1985-05-01 → 1995-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/3178852

## Citation

> US National Institutes of Health, RePORTER application 3178852, BIOCHEMISTRY AND PATHOLOGY OF 5-FU/FOLATE THERAPY (2R01CA039629-07A2). Retrieved via MORVS 2026-10-25 from https://api.morvs.ai/grant/nih/3178852. Licensed CC0.

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