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Administrative data

additional toxicological information
Type of information:
other: Pathology working group review of histological changes in kidneys of male rats from 2-year inhalation carcinogenicity study
Adequacy of study:
supporting study
Study period:
3 March 2009 to 25 March 2009
Rationale for reliability incl. deficiencies:
other: The study represents an extensive re-analysis of kidney pathology observed in male rats assigned to a 2-year carcinogenesis bioassay (NTP Study No. 05181-03).

Data source

Reference Type:
other: Pathology review of selected changes in the kidneys of male rats from a 2-year carcinogenicity study.
Report Date:

Materials and methods

Type of study / information:
This report represents an in-depth re-analysis of the histopathological changes noted in the kidneys of male rats from a 2-year inhalation carcinogenicity study conducted by the US NTP (Study No. 05181-03) conducted by a panel of 5 senior pathologists (Pathology Working Group) with extensive experience in the interpretation of chemically-induced nephrotoxicity and neoplasia in laboratory animals. The objectives of the PWG were:1) Establish the most appropriate diagnoses for all proliferative changes observed in the kidneys of male rats assigned to the 2-year carcinogenesis bioassay (NTP Study No. 05181-03).2) Provide comment relative to potential pathogenic mechanisms that likely resulted in male rat kidney tumors.3) Provide perspective concerning the risks associated with potential human exposures to tetrahydrofuran based upon findings in laboratory rodents.
Principles of method if other than guideline:
There is no applicable guideline for this study.
GLP compliance:

Test material


Results and discussion

Any other information on results incl. tables

Table 1: Comparison of Renal Proliferative Changes Observed by NTP and PWG Pathologists in a 2 -Year Inhalation Study with Tetrahydrofuran in F344 Rats (NTP Study No. 05181 -03)

 Review Pathologists     NTP     PWG
             Group 1 (N = 50) 0 ppm
 Animal No.  Neoplasm  Hyperplasia*  Neoplasm  ATH**
 001 - 4 - Present
 008 -  4  -  -
 009  -  -  Present
 025  -  -  Present
 029  -  4  -  -
 038  Adenoma  Adenoma
 049  -  3  Adenoma  Present
             Group 2 (N = 50) 200 ppm
 206  3  -
 215  -  4  -  Present
 236  Present
 237  -
 240  4  -
             Group 3 (N = 50) 600 ppm
 401  -  2  Present
 409  Adenoma  -  Present
 410  -  1  -  -
 414  Adenoma  -  Adenoma  -
 423 -  1  -  Present
 431  4  -  Present
 432 Adenoma  4  -
 444  Adenoma  -  Adenoma  -
 450  -  3  -
             Group 4 (N = 50) 1800 ppm
 611  3 Present 
 616  4 Adenoma 
 620  Carcinoma  -  Adenoma  -
 624  Adenoma  4  Adenoma  -
 631  -  3  -
 633  -  4  -  -
 636  3  Adenoma  -
 643  Adenoma  2 Adenoma 
 647 Adenoma   - Adenoma   -
 649 Carcinoma   - Adenoma   -

* Includes both "simple" and "atypical" tubular hyperplasia.

** Atypical Tubular Hyperplasia (regarded by the PWG as pre-neoplastic).

The PWG results indicated that there was little difference in the incidence of proliferative changes (neoplastic and pre-neoplastic combined) between the high concentration group (1800 ppm) and the control group. Although 7 benign adenomas were observed in the high concentration males as compared with 2 in the control group, 4 control group males displayed foci of atypical tubular hyperplasia as compared with 1 male in the high concentration group. These atypical hyperplastic foci were considered to be pre-neoplastic alterations on a biologic continuum with adenomas. It was important to recognize that proliferative changes in most animals were represented by small, focal, hypercellular (intratubular) clusters where the distinction between hyperplasia and neoplasia was often difficult. The overall incidences of proliferative changes in the control group compared with the 1800 ppm group (6 versus 8) did not suggest a test article effect. Noteworthy was the unequivocal PWG consensus that renal cell carcinomas observed by NTP pathologists in two 1800 ppm males (nos. 620 and 649) were, in fact, benign adenomas rather than malignant tumors.

Mean incidences and severity grades for CPN were similar when the high concentration males were compared to the control group, indicating that an exposure-related exacerbation of CPN was not present in the 2 -year bioassay. Mean severity grades for CPN, as determined by both NTP and PWG pathologists, have been summarized in the following table. It should be noted that the PWG examined only kidneys that displayed proliferative lesions from male rats assigned to Groups 2 and 3. Therefore, mean severity grades for CPN were not determined for these 2 groups.

       Comparison of Mean Severity Grades for Chronic Progressive Nephropathy (CPN) as Observed by NTP and PWG Pathologists
Review Pathologists   NTP  PWG
Group 1 (N = 50) 0 ppm  2.88  2.84 
Group 2 (N = 50) 200 ppm  2.86 Not determined 
Group 3 (N = 50) 600 ppm  3.12  Not determined 
Group 4 (N = 50) 1800 ppm   3.0 2.88 

In addition to morphological alterations relating to tubular cell proliferation and CPN, the PWG recognized that histologic changes associated with alpha-2U-globulin nephropathy (e.g., increased hyaline droplets, casts of exfoliated tubular cells and linear patterns of papillary mineralization) were not present in the sections examined. This "non-finding" was in agreement with the NTP interpretations.

14 -Week Inhalation Study (NTP Study No. 05181 -01):

The PWG confirmed that hyalin droplets (consistent with alpha-2U-globulin) were present in the proximal tubular epithelium of males assigned to the high dose concentration in the 14 -week sub-chronic study; however, similar deposits were present in control males, and it was not possible to distinguish differences between the two groups by routine light microscopic examinations. Although hyaline droplets may have been slightly increased in the high concentration males, histologic evidence of alpha-2U-globulin-related cytotoxicity in the form of casts of exfoliated tubular cells at the junction of inner and outer stripe of the outer medulla could not be confirmed.

4 -Week Inhalation Study (BASF Project 9910151/99007):

In contrast to the 14 -week study conducted by the NTP, the PWG recognized that hyaline droplets were distinctly increased in numbers and extent in the proximal tubular cells of male rats exposed to the high concentration (1800 ppm) of tetrahydrofuran for 4 weeks; immunohistochemistry confirmed that the hyalin droplets were alpha-2U-globulin. The PWG also recognized that Bromo-deoxyuridine labeling conducted by BASF for this study identified that "hot-spots" of accelerated cell proliferation were present in the cortex of rats exposed to 1800 ppm for 20 days. Although the PWG confirmed that tetrahydrofuran exposures were associated with increased deposits of alpha-2U-globulin and acknowledged that "hot spots" of accelerated tubular cell proliferation had been associated with this finding, it could not confirm the presence of distinct cytotoxicity by routine light microscopic examinations.


Diagnostic Criteria: Although the differences were not statistically significant, histopathologic assessments by both the NTP and PWG confirmed that renal cell adenomas were slightly more prevalent in high exposure male kidneys (7 tumors) when compared with the control group (2 tumors). When all proliferative lesions (preneoplastic and neoplastic) were considered, however, incidence values were very similar between the 2 groups. This determination was fascilated by the PWG's application of refined diagnostic criteria that distinguished between "reactive" tubular hyperplasia (associated with CPN) and atypical tubular hyperplasia that was considered to represent a likely preneoplastic change. Original NTP data did not make a distinction between reactive and atypical hyperplasia. Thus, summation of overall proliferative changes that were likely to progress to neoplasia was not included in the initial NTP pathology report.

Potential Pathogenic Mechanisms: The routine, NTP-sponsored mutagenicity tests for tetrahydrofuran were negative. Furthermore, histologic features such as metaplasia, anaplasia, dysplasia and increased hyperplasia that are sometimes indicative of exposure to genotoxic chemicals were not observed. Accordingly, the PWG discounted tetrahydrofuran as a likely genotoxic chemical, and focused upon non-genotoxic mechanisms that may have contributed to the slightly higher numbers of tubular adenomas in the 1800 ppm exposure group. In this regard, essentially all adenomas and atypical tubular hyperplasias were present in kidneys that displayed advanced (severe end stage) CPN, and the PWG considered that accelerated tubular cell degeneration and regeneration associated with CPN was likely responsible for the development of most proliferative lesions. In this connection, however, a tetrahydrofuran-induced exacerbation of CPN was not considered to be contributory because mean severity grades for CPN were similar between the control and the high concentration groups. With routine light microscopic procedures, the PWG could not confirm the deposits of alpha-2U-globulin were directly associated with tubular cell cytotoxicity. Exfoliation of proximal tubular cells with the formation of intratubular casts at the junction of the inner and outer stripe of the outer medulla was not observed; and linear mineral deposits were not present in medullary tubules following prolonged exposures. The PWG confirmed, however, that deposits of alpha-2U-globulin were mildly increased in the proximal tubular cells of rats assigned to the 4 -week BASF investigations. Furthermore, the PWG accepted the BASF report that detailed accelerated tubular proliferation for male rats assigned to the high concentration group. Taken together, these observations indicated that inhalation of tetrahydrofuran, at a concentration level of 1800 ppm, did induce tubular cell proliferation via the alpha-2U-globulin mechanism; however, overt (necrotizing) tubular cell damage could not be detected by routine microscopic observations. In turn, slight increases in tubular cell turnover associated with alpha-2U-globulin may have contributed to the development of some of the adenomas in males assigned to the 1800 ppm group of the 2 -year bioassay.

Risk for Humans: Important findings were the lack of evidence of early occurrence of tumors or preneoplastic lesions in the test article-exposed groups, and no evidence of tumor progression to carcinoma, both of which would be expected with compound -induced kidney tumors. Although slightly more adenomas were observed inthe 1800 ppm males when compared with the control group, the difference was not statistically significant; and when all proliferative changes were considered, incidence values were similar between groups. Perhaps of primary importance was the recognition that both mechanisms that likely resulted in renal proliferative changes (advanced CPN and alpha-2U-globulin nephropathy) have no counterpart in humans. These observations, along with an absence of renal tumors in female rats, indicate that the male rat tumors have no relevance for humans, and that it would be inappropriate to extrapolate these findings to humans.

Applicant's summary and conclusion

The consensus diagnoses of the PWG confirmed that there was no difference in the incidence of renal tubular proliferative changes (neoplastic and pre-neoplastic combined) when control and tetrahydrofuran-exposed groups were compared for the 2-year bioassay. Although the incidence of tubular adenomas was slightly higher in rats exposed to the 1800 ppm tetrahydrofuran, the difference was not statistically significant when compared with the control group, and there was no evidence of early tumor occurrence or of tumor progression to carcinoma. Furthermore, PWG members agreed that, based upon the absence of experimental results that would implicate genotoxic mechanisms for the observed proliferative changes, it was likely that atypical hyperplasias and adenomas in the control group resulted from regenerative processes associated with advanced CPN and/or low-grade alpha-2U-globulin nephropathy. In this connection, it was important to recognize that CPN in aging rats and alpha-2U-globulin nephropathy have no known human counterparts. Therefore, it was the consensus of the PWG that mechanisms that likely contributed to the formation of renal tubular adenomas in male rats assigned to the 2-year carcinogenesis bioassay with tetrahydrofuran (NTP Study No. 05181-03) pose no risk for humans.