Cancer Investigation, 10(4), 323-326 (1992)

ED IT0 R I AL

Back to the Future: New Theories on 5-Fluorouracil/Interferon Interactions

Scott Wadler, M.D.

Department of Oncology Monteflore Medical Center Bronx, New York 10467 Albert Einstein Cancer Center Bronx, New York 10461

Biological agents, including the interferons, the inter- leukins, colony-stimulating factors, tumor necrosis fac- tor, and monoclonal antibodies, have become an increas- ingly important focus of both preclinical and clinical in- vestigations. Interest in these agents derives from their higher degree of specificity as compared with chemo- therapy or radiation therapy, with relative sparing of nor- mal host tissues and diminution of myelosuppression, mucosal damage, and renal insufficiency. While anti- cancer effects of these agents have been modest, their higher degree of selectivity has led to novel applications. For example, colony-stimulating factors stimulate bone marrow progenitor cells ameliorating the myelosup- pressive effects of chemotherapy. Interleukin-2, employed ex vivo with peripheral blood mononuclear cells, expands the lymphokine-activated killer cell compartment, which can then be reinfused. Monoclonal antibodies offer a multitude of novel opportunities for anticancer effects by producing highly specific spatial apposition of cancer cells with toxins, lethal radioisotopes, or killer T cells.

Despite these novel applications, incorporation of biologic agents into the cancer armamentarium remains a unique challenge for the clinician. The assumptions for their use differ from those for cytotoxic drugs because of their unique properties including a spectrum of

pleiotropic effects not always appreciated prior to their initial clinical testing, the wide range of biologically ef- fective doses, and their poorly understood mechanism of action. For the majority of biologic agents, the optimal conditions for their administration have not been fully defined. As clinical researchers have gained greater in- sight into their biologic properties, new clinical indica- tions have evolved.

The interferons ( IFNs) are a good example of this. First identified in 1957 by Isaacs and Lindenmann because of their antiviral properties (l), it soon was discovered that IFNs also possessed important antiproliferative effects against tumor cells, and in addition, were capable of augmenting the activity of natural killer cells and other components of the immune system (2-4). Based on their immunostimulatory properties, IFNs were introduced into clinical trials as single agents against such “immunologi- cally sensitive” tumors as renal cell carcinoma and melanoma (S,6), and also against AIDS-related Kaposi’s sarcoma (7) where it was hypothesized that IFNs offered the opportunity not only for direct antiproliferative effects against the tumor target, but also immunopotentiating ef- fects in compromised patients and antiviral effects against the human immunodeficiency virus. Modest, but reproducible activity was demonstrated against these

323

Can

cer

Inve

st D

ownl

oade

d fr

om in

form

ahea

lthca

re.c

om b

y U

nive

rsity

of

Nor

th C

arol

ina

on 1

1/06

/14

For

pers

onal

use

onl

y.

324 Wader

tumors; nevertheless, this antitumor activity has never been conclusively correlated with augmentation of any im- mune function.

Incorporation of IFNs into chemotherapeutic regimens against refractory solid tumors was based in part on the hypothesis that cytotoxic agents were effective for “chemically debulking” large solid tumors, but not for eradicating micmscopic residual disease, and furthermore that IFNs with their higher degree of immune specifici- ty, would effectively eradicate microscopic residual disease, but would likely be ineffective against bulky tumors with large antigexdantibody ratios (8). It was soon realized, however, that simultaneous administration of IFNs with chemotherapeutic drugs was as effective as se- quential administration. In an elegant series of experiments against human breast and lung tumors explanted to an im- munodeprived mouse model, IFNs markedly augmented the antitumor effects of cyclophosphamide, doxorubicin, and cisplatin, casting further doubt on the hypothesis that IFN was acting via an imtnune effector cell and suggesting that IFNs were acting by modulating the actions of the cytotoxic drug albeit by an undefined mechanism (9,lO).

In the early 198Os, following reports of in vitro synergy between partially purified IFNs and other cytotoxic agents, studies fiom Japan demonstrated synergy for the combination of 5-FU and IFN against tumor cell lines derived from refractory gastrointestinal malignancies (1 1,12), In vitro studies from Albert Einstein confirmed the synergistic interaction between 5-FU and recombinant IFNs-a and -& demonstrating a 10- to 100-fold increase in maximal cytotoxic e f k t at concentrations of IFN that were themselves nontoxic, and demonstrating that this did not result from an antipmliferative or cytokinetic effect of the combination (13). Based on these studies a Phase 11 trial was initiated which demonstrated an objective response rate of 63 % among previously untreated patients (14). When this clinical trial was duplicated in the multi- institutional setting by the Eastern Cooperative Group, the objective response rate was 42% with projected me- dian survival by Kaplan-Meier analysis of over 18 months (15). Thus, these trials supported the in vitro data sug- gesting a positive interaction between 5-FU and IFN. To confirm that the combination of 5-FU and rIFNa is truly more efficacious than 5-FW alone, a randomized trial will be required; several are currently ongoing, including a five-arm trial of 5-FU/IFN versus high-dose 5-FU, 5-FU/leucovorin, and 5-FU/PALA being conducted by the Eastern Cooperative Oncology Group,

Unlike interactions between 5-FU and other biochemical modulating agents such as leucovorin or methotrexate, for which well-described in vitro interactions were exploited

in the design of clinical trials, the mechanism of interac- tion between 5-FW and IFN is not understood. This results in part from a spectrum of pleiotropic actions on both malignant and normal tissues, including host effects, exhibited by IFN.

Early clues to the mechanism of interaction came from Elias and Crissman, who demonstrated reversal of the cytotoxic effects in the presence of dThd, suggesting that the cytotoxicity of the combination resulted from inhibi- tion of thymidylate synthase (TS) (16). This observation was confirmed by observations that, while levels of TS mRNA were not affected by incubation with IFN, levels of TS protein decreased relative to cells treated with 5-FU alone (17). Chu et al. have demonstrated in human colon carcinoma H630 cells that the effects of IFN on TS oc- cur at the translational level (18). Thus, the presence of IFN prevents tumor cells from circumventing the blockade at the level of TS by synthesizing additional TS.

Whether these effects are sufficient to account for a 10- to 100-fold increase in the cytotoxicity of 5-FU as ob- served in vitro is unclear. Furthermore, other biochemi- cal effects of IFN have been identified, including enhance- ment of the anabolism of 5-FU to its active metabolite, FdUMP, and inhibition of thymidine salvage pathways (19). The relative importance of each these remains unknown. Undoubtedly, other biochemical effects of the combination will be discovered.

A very important clue to the interaction of 5-FU and IFN has come fmm a series of Phase I/pharmacokinetic studies which demonstrate that IFN decreased the clearance of 5-FU raising the area under the curve of pa- tients receiving the combination (20-23). This pharmaco- kinetic advantage for the combination may account for the augmentation in 5-FU activity in the presence of IFN.

In this issue of Cancer Znvestigution Dim Rubio and colleagues make another interesting observation concern- ing the mechanism of interaction of the combination of 5-FU/IFN (24). They treated 33 evaluable patients with a regimen similar to the Albert Einstein regimen and observed a statistically significant increase in responses among patients with liver metastases as opposed to those with metastases in other sites. This included 4 complete and 3 partial responses in 23 evaluable liver metastases as opposed to only 1 complete response among 33 non- hepatic sites. These investigators conclude that modula- tion takes place in an “oriented organ” fashion, and sug- gest that activation of Kupffer cells in the liver may be a mechanism by which the combination acts.

Ample in vitro evidence suggests that Kupffer ells may play an active antitumor role against metastatic adeno- carcinoma (25). Kupffer cells elaborate a variety of

Can

cer

Inve

st D

ownl

oade

d fr

om in

form

ahea

lthca

re.c

om b

y U

nive

rsity

of

Nor

th C

arol

ina

on 1

1/06

/14

For

pers

onal

use

onl

y.

New Theories on 5-PU-IFN Interactions 325

autocrine and paracrine factors that influence the growth of adjacent hepatocytes, lipocytes, and potentially cancer cells (26,27). Furthermore, various immune mediators, including interferon-gamma, interleukin-2, lipopolysac- charide, and muramyl dipptide, augment Kupffer cell activation (28-31). Finally, one intriguing study in a nude mouse model demonstrated that addition of human IFN- y to 5-FU did not enhance the cytotoxic effects of 5-FU against an explanted human colon cancer metastatic to the liver, but that muse IFN-y did enhance the effects of 5-FU, suggesting that synergy was mediated via a mouse immune effector cell (32).

The observations by Diaz Rubio and colleagues bring us full circle back to original theories of combined immune/cytotoxic action against tumor cells. These in- teresting observations warrant a return to animal models to determine whether activation of Kupffer cells plays an important role in augmentation of 5-FU activity in the liver.

12.

13.

14.

15.

16.

17.

18.

REFERENCE3

1. Isaacs H, Lindaunann J: Virus interference: 1. The interferon. Proc R Soc London (Biol), 147:257-262, 1957.

2. Pestka S, Langer JA, Zoon KC, Samwl CE: Interferons and their actions. Annu Rev Biochem 56:727-777, 1987.

3. Pfeffer LM, Murphy JS, Tamm I: Intaferon effects on the growth and division ofhuman fibroblasts. Exp Cell Res 121:lll-115, 1979.

4. Grossberg S: Interferons: an overview of their biological and biochemical properties. In MecMsms of hterfcron Actions. Vol I. Edited by LM Pfeffer. CRC Press, Boca Raton, FL, 1987, pp

5. Kirkwood JM, Emtoff MS, Davis CA et al: Comparison of intra- muscular and intravenous recombinant alpha-2 interferon in melanoma and other cancers. AM Int Med 103:32-36, 1985.

6. Quesada JR, Rios A, Swanson D et al: hitumor activity of recombinant-dcrived interferon alpha in metastatic renal cell carcinoma. J Clin Oncol 3:1522-1528, 1985.

7. 0I.oOpman JE, aoaleib MS, Goodman J et al: Recombinant alpha4 interferon therapy for Kaposi’s sarcma associated with the ac- quired immunodeficiency syndrome. AM Int Med 100:671-676, 1984.

8. Gutterman JU, Hersh EM: Immunotherapy. In Cancer Medicine. Wted by JF Holland, E Frei. Lea and Febiger, Philadelphia, 1982,

9. Balkwill FR. Moodie EM: Positive interactions between human interferon and cyclophosphamidc or Adriamycin in a human tumor model system. Cancer Res 44:5Q4-908, 1984.

10. Carmichael J, Fergusaon RJ, Wolf CR et al: Augmentation of cytotoxicity of chemokrapy by human a-interferons in human non-small cell lung canax xenograb. Canca Res 46:4916-4920, 1986.

11. Kimoto Y: Antitumor effects of interferons with chemotherapeutic agents. Jpn J Cancer fiemother 13293-301, 1986.

1-32.

p~ 1100-1132.

19.

20.

21.

22.

23.

24.

25.

26.

27.

28.

Namba M, Yarrmmoto S, Tanaka H et al: In vitro and in vivo studies on potentiation of cytotoxic effects of anticancer drugs or cobalt 60 gamma-ray by interferon on human neoplastic cells. Cancer

Wadler S, Wersto R, Weinberg V et al: Interaction of fluorouracil and interferon in human colon CBllCer cell lines: cytotoxic and cytokinetic effects. Cancer Res 505735-5739, 1990. Wadler S, Wiemik PH: CliNcal update on the role of fluorouracil and recombinant interferon alfa-la m the treatment of colorectal carcinoma. Semin Oncol 17:16-21, 1990. Wadler S, Lembersky B, Kirkwood JM et al: Phase II trial of fluorouracil (5FU) and recombinant alfa-2 interferon (PN) in pa- tients @@) with advanced colorectal cancer: An Eastern Cooperative Oncology Group (ECOG) Study. Roc Am Soc Clin Oncol, 1991. Elias L, Crissman HA: Jnterfmn effects u p the adenocarciaoma 38 and HL60 cell lines: antiproliferative responses and synergistic interactions with halogenated pyrimidine antimetabolites. Cancer Res 48:4868-4873, 1988. Wadler S, Schwartz EL, Thompson D, Wiernik PH: Effects of recombinant interfemn on thymidylate synthase expression and ter- nary complex famation in human tumr cell lints. Roc Am Assoc Cancer Res 31:402, 1990. Chu E, Zinu S, Auegra C: Translatid regulation of thymidylate synthase in response to 5-fluorouracil and gamma-interferon in a human colon cancer cell line (H630). Proc Am Assoc Cancer Res 32:416, 1991. Hoffman M, Wadler S, Schwartz EL, W i d PH: Effects of recombinant interferon-alpha on anabolism of (6-3H) 5-fluorouracil in human colon cancer cell lines. Proc Am Assoc Cancer Res 31:408, 1990. Danhauser L, (filchrist T, Friemann J et al: Effects of recombi- nant interferon-a 2b on the plasma phamacokinetics of fluomuracil in patients with advanced cancer. Proc Am Assoc Cancer Res 32:176, 1991. Grem JL, McAtee N, Murphy RF et al: A pilot study of interferon alfa-2a in combination with 5-fluorouracil plus highdose leucovorin in metastatic gastrointestinal carcinoma. Proc Am Soc Clin Oncol 9:70, 1990. Lindley C, Bcmard S, Gavigan M et al: Interferon-alpha increases 5-fluorouracil(5FU) plasma levels &fold within one hour: results of a phase I study. J Interferon Res 10:S132, 1990. Schuller J, Czejka M, Miksche M et al: Mluence of interferon a 2b (IFN) f leucovorin (LV) on pharnmcokinetics (PK) of 5-flUOrOUraCil. Roc Am Soc Clin Oncol 10:98, 1991. Diaz Rubio E, Jimeno J, Camps C et al: Treatment of advanced colorectal cancer with recombinant interferon alpha and fluo-il: activity in liver metastases. Canc~r Invest 10:257-262, 1992. Roh MS, Wang L, Oyedeji C et al: Human Kupffer cells are cytotoxic against human colon adenocarcinoma. Surgery

Kuiper J, Kamp JAAM, Van Berkd TJC: Induction of odthine decarboxylase in rat liver by phorbol ester is mediated by pros- tanoids. J Biol Chem 264:6874-6878, 1989. Friedman S, Arthur MJP Activation of cultured rat hepatic lipocytcs by Kupffer cell conditioned medium. J Clin Invest 84:1780-1785, 1989. Curran RD, Billiar TR, West MA et al: Effect of interleukin 2 on Kupffer cell activation. Arch Surg 123:1373-1378, 1988.

54~2262-2267, 1984.

108:400-405, 1990.

Can

cer

Inve

st D

ownl

oade

d fr

om in

form

ahea

lthca

re.c

om b

y U

nive

rsity

of

Nor

th C

arol

ina

on 1

1/06

/14

For

pers

onal

use

onl

y.

326 Wadler

29. Phillips NG, Tsao MS: Inhibition of expaimental liver tumor p w t h in mice. by liposanes containing a lipoFhilic muramyl dipep-

30. Stukart MJ, Rijnsent A, Roos E: Induction oftumoricidal activity in rat liver macrophages by liposomes containing recombinant rat gamma-interferon SuFplemented with lipopolysaccharide or muramyl dipeptide. Cancer Res 47:388&3885, 1987.

31. Curran RD, Billiar TR, Kispert PH et al: Hepatocytes enhance Kupffer cell-mediated tumor cell cytostasis in vim. Surgery

32. Morikawa K, Fan D, Denkin8 YM U al: Mechanisms of combined effects of gamma-interferon and 5-fluorouracil on human colon cancers implanted into nude mice. Cancer Res 49:799-805,1989.

ti& derivative. Cancer Res 49:936-939, 1989. 106:126-132, 1990.

Can

cer

Inve

st D

ownl

oade

d fr

om in

form

ahea

lthca

re.c

om b

y U

nive

rsity

of

Nor

th C

arol

ina

on 1

1/06

/14

For

pers

onal

use

onl

y.