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Nitrogen Mustard Sensitivity and Choline Transport in Walker 256 Carcinosarcoma Cells in Vitro

Gerald J. Goldenberg and B. K. Sinha
Gerald J. Goldenberg
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B. K. Sinha
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DOI:  Published November 1973
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Summary

Single cell suspension cultures of Walker 256 carcinosarcoma were established in vitro in Fischer's medium supplemented with 10% horse serum. The cells grew exponentially with a doubling time of 16 to 17 hours, had a cell volume (mean ± S.E.) of 2313 ± 89 cu µm, and were cloned in soft agar with a cloning efficiency of 68.7 ± 6.3%. Sensitivity to the alkylating agent nitrogen mustard (HN2) was comparable to that observed previously with L5178Y murine lymphoblasts; the D0 (the dose of drug reducing survival to 37% of the initial cell population) was 5.35 ng/ml, and the extrapolation number, n, was 1.27. An investigation of the interaction of hydrolyzed nitrogen mustard and choline transport revealed that uptake of choline-14C obeyed simple Michaelis-Menten kinetics, proceeded “uphill” against a concentration gradient of over 30-fold, and showed a decrease in distribution ratio as choline concentration increased and choline transport was competitively inhibited by hydrolyzed NH2. The Michaelis constant, Km, for choline transport was 2.90 × 10−5 m, the transport capacity, Vmax, was 4.83 × 10-17 moles/min/cell, and the inhibition constant, K1, with hydrolyzed HN2 as inhibitor, was 7.83 × 10−5 m. These findings indicate that choline transport by Walker carcinosarcoma cells is an active carrier-mediated process and that choline and hydrolyzed HN2 compete for the same transport mechanism.

Footnotes

  • ↵1 This work was supported by a grant from the National Cancer Institute of Canada.

  • Received June 5, 1973.
  • Accepted July 9, 1973.
  • ©1973 American Association for Cancer Research.
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November 1973
Volume 33, Issue 11
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Nitrogen Mustard Sensitivity and Choline Transport in Walker 256 Carcinosarcoma Cells in Vitro
Gerald J. Goldenberg and B. K. Sinha
Cancer Res November 1 1973 (33) (11) 2584-2587;

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Nitrogen Mustard Sensitivity and Choline Transport in Walker 256 Carcinosarcoma Cells in Vitro
Gerald J. Goldenberg and B. K. Sinha
Cancer Res November 1 1973 (33) (11) 2584-2587;
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Cancer Research Online ISSN: 1538-7445
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