# Presentation: 498615

## Furan-Induced Cytotoxicity, Cell Proliferation, and Tumorgenicity in Mouse Liver

- Dr. Glenda Moser

## 2 Year NTP Bioassay

- Non-neoplastic
- Rats
- Kidney
- Biliary Tract
- Liver
- Mice
- Biliary Tract
- Liver
- Neoplasms
- Rats
- Cholangiosarcoma – 2, 4, 8 mg/kg
- Mononuclear cell leukemia
- Mice
- Benign pheochromocytoma
- Hepatocellular – 8 mg/kg

## Slide 3

## Cancer Multistage

**Genotoxic**
- Direct DNA damage
- Single exposure
- Metabolism
**Mechanisms of Genotoxicity**
- Mutations
- Chromosomal translocations or loss
- Gene amplification; fusion of viral promoters
- DNA adducts
- Promoter insertion

## Assays for Genotoxicity after Furan Exposure

## Cancer Multistage

**Non-Genotoxic or Epigenetic**
- Clonal expansion
- Sustained exposure, high doses
- Reversible
- Specificity
- B6C3F1 mouse
- Liver

## Cancer Multistage

**Non-genotoxic Mechanisms**
- Activated oncogenes, loss of suppressor genes-ras
- Methylations
- Immunosuppression
- Alteration of signal transduction and gene expression
- Growth factors
- Receptors
- Cell-cell communication
- Receptor mediated-receptor
- Oxidative stress
- Altered DNA repair
- Differentiation
- Apoptosis
- Cytotoxicity/Cell proliferation
- Secondary mutations

## Liver Cytotoxicity 

**Clinical Chemistry**
- ALT
**Histopathologically**
- Pynotic nuclei/ Condensed chromatin
- Inflammatory response- left and caudate lobes
- Degenerated hepatocytes/cytoplasmic vacuolation

## Approach

- Male B6C3F1
- 10/Group
- 13 Weeks
- Gavage
- 5X/wk
- Dose levels 0, 0.5, 2, 4, 8, and 15 mg/kg
- BrdU
- Osmotic pump
- 7-day

## Slide 10

## Slide 11

## Cell proliferation 

**LI**
**Methodologies**
- BrdU thymidine analog *in vivo*
- IHC for BrdU
- Routes of administration
- Pulse
- Cumulative
**Quantifying**

## Slide 13

## Slide 14

## Slide 15

## Slide 16

## Carcinogenic Study

- Female B6C3F1
- Dose levels of 0.0, 0.5, 1.0, 2.0, 4.0, 	and 8.0 mg/kg body weight
- 50- 100 per Group
- 2 years
- Gavage
- 5X/wk

## Slide 18

## Slide 19

## Incidence of Cytotoxicity in Cancer Study

## Slide 21

## Slide 22

## Liver Findings in Male and Female Mice Exposed to to Furan 

| mg/kg | 8.0  | 4.0  | 2.0  | 1.0 | 0.5  |
| --- | --- | --- | --- | --- | --- |
| Cytotoxicity |     | +/-   - | -      - | -      - | -     - |
| ALT/SDH (IU) |       | -    +/-
 | -      - | -      - | -      - |
| LI (%) |     | -      - | -      - | -      - | -      - |
| Incidence of  Liver lesions |    
 |  +/- |     - |  - | - |

## Conclusion

- The study demonstrated a dose-dependent increase in furan-induced liver tumors in female B6C3F1 mice and a relationship between dose, cytotoxicity, and tumor induction for a model cytotoxic hepatocarcinogen.

## Overview of Furan

- Reproduced results of NTP study
- Threshold for liver tumors
- At 13 weeks liver cytotoxicity and cell proliferation reversible
- Other chemicals with same proposed mechanisms
- Mutations and other events may be secondary to hepatocyte cytolethality or increased cell proliferation
- Biologically plausible model
- Furan-induced effects after short term exposure are inhibited by p450 Inhibitors
- Similar pharmokinetics in mouse and human in vitro; rat metabolizes furan slower

## Areas of Interest

- Do p450 inhibitors decrease liver tumorigenesis in mouse liver?
- Are liver tumors due to bolus dose?
- Are the positive genotoxic results due to direct damage to DNA, high doses tested and/or secondary to cell proliferation?
- What are the molecular or gene expression changes in liver tumors? Do they occur in cholangiosarcomas?
- Is there a threshold for cholangiosarcomas or mononuclear leukemia?  Is the mode of action similar to that of liver tumors?
- Do biliary tract epithelial cells have pharmokinetic parameters similar to that of hepatocytes?
- What is the relevance of mouse liver findings to cholangiosarcomas and leukemia or humans? PK
- Are there populations of humans that are susceptible to furan-induced effects?  Is age a factor?

## Acknowledgement

- Special thanks to the
- ILS toxicology techs animal care, laboratory assistants,
- ILS histology department,
- Dr. Robert Maronpot, NIEHS
- Julie Foley, NIEHS
- Dr. Tom Goldsworthy.

## Questions?