High methylguanine-DNA Methyltransferase Research Articles

MGMT expression predicts response to temozolomide in pancreatic neuroendocrine tumors.

Temozolomide (TEM) showed encouraging results in well-differentiated pancreatic neuroendocrine tumors (WDPNETs). Low O(6)-methylguanine-DNA methyltransferase (MGMT) expression and MGMT promoter methylation within tumors correlate with a better outcome under TEM-based chemotherapy in glioblastoma. We aimed to assess whether MGMT expression and MGMT promoter methylation could help predict the efficacy of TEM-based chemotherapy in patients with WDPNET. Consecutive patients with progressive WDPNET and/or liver involvement over 50% who received TEM between 2006 and 2012 were retrospectively studied. Tumor response was assessed according to Response Evaluation Criteria in Solid Tumors (RECIST) 1.1 guidelines. Nuclear expression of MGMT was assessed by immunochemistry (H-score, 0-300) and MGMT promoter methylation by pyrosequencing. Forty-three patients (21 men, 58years (27-84)) with grade 1 WDPNET (n=6) or 2 (n=36) were analyzed. Objective response, stable disease, and progression rates were seen in 17 patients (39.5%), 18 patients (41.9%), and 8 patients (18.6%), respectively. Low MGMT expression (≤50) was associated with radiological objective response (P=0.04) and better progression-free survival (PFS) (HR=0.35 (0.15-0.81), P=0.01). Disease control rate at 18months of treatment remained satisfying with an MGMT score up to 100 (74%) but dropped with a higher expression. High MGMT promoter methylation was associated with a low MGMT expression and longer PFS (HR=0.37 (0.29-1.08), P=0.05). Low MGMT score (≤50) appears to predict an objective tumor response, whereas an intermediate MGMT score (50-100) seems to be associated with prolonged stable disease.

MGMT in colorectal cancer: a promising component of personalized treatment.

The identification of new, effective drugs is a pressing need in colorectal cancer (CRC) rescue therapy. Data examining O (6)-methylguanine-DNA-methyl transferase (MGMT) and its predictive role in temozolomide (TMZ) treatment in CRC are scarce. In this study, the effect of MGMT status on the cytotoxic sensitivity caused by TMZ was analyzed using cytology proliferation assays in colon cancer cell lines. MGMT protein expression was assessed with immunohistochemistry in 385 patients. Concordance between primary and metastatic sites and the role of MGMT status on survival were statistically analyzed. TMZ sensitivity was significantly affected by the level of MGMT protein expression. Of 385 cases, 13 (3.4%) demonstrated loss of MGMT expression. However, low MGMT expression levels were significantly more common in signet ring cell carcinomas (p = 0.011). In 111 of 385 cases, the overall concordance of MGMT status between primary tumor and metastatic sites was 66.67% (κ = 0.271, p < 0.001). The median progression-free survival was significantly different between groups with low or high MGMT expression for the irinotecan-based regimen (p = 0.025), but MGMT protein expression was not observed to be a prognostic factor. In conclusion, MGMT was an important in vitro predictor of TMZ activity in CRC. The rate of MGMT protein loss was low in metastatic CRC patients from China, and MGMT might be more commonly lost in signet ring cell carcinoma. The MGMT status at primary and metastatic sites was consistent, but the power of concordance was poor. Further study into these topics is warranted.

O(6) -methylguanine DNA methyltransferase repairs platinum-DNA adducts following cisplatin treatment and predicts prognoses of nasopharyngeal carcinoma.

Cisplatin (CDDP) is an important anti-cancer drug commonly used in various human cancers, including nasopharyngeal carcinoma (NPC). How to overcome the drug resistance of CDDP provides opportunities to improve clinical outcomes of NPC. O(6) -methylguanine-DNA methyltransferase (MGMT) has been well-characterized to be a therapeutic determinant of O(6) -alkylguanine alkylating drugs. However, the underlying mechanism and clinical relevance between MGMT and CDDP remain poorly defined in NPC. In this study, we showed that MGMT-proficient cells were highly resistant to the cytotoxic effects of CDDP as compared to MGMT-deficient cells. Further studies showed that the platinum level of DNA after CDDP exposure was significantly lower in MGMT-proficient cells than in MGMT-deficient cells. Host cell reactivation assay revealed that MGMT protected NPC cells from CDDP-induced DNA damage by enhancing DNA repair capacity. Importantly, we demonstrated for the first time that MGMT protein directly bound to CDDP-induced DNA damages. Subsequently, CDDP-bound MGMT protein became ubiquitinated and was degraded through ubiquitin-mediated proteasome system. We further analyzed the relationship between MGMT expression and clinical survivals in a cohort of 83 NPC patients. NPC patients who received CDDP-based concurrent chemoradiotherapy (CCRT), with high MGMT expression level, exhibited shorter progression-free survival (PFS; p = 0.022) and overall survival (OS; p = 0.015), than patients with low MGMT expression level. Furthermore, high MGMT expression level remained to be an independent prognostic factor for worse PFS (p = 0.01, hazard ratio 2.23) and OS (p = 0.018, hazard ratio 2.14). Our findings suggest that MGMT protein is important to determine the efficacy of CDDP in NPC.

O6-methylguanine-DNA methyltransferase (MGMT) immunohistochemistry as a predictor of resistance to temozolomide in primary CNS lymphoma

Temozolomide, an alkylating agent, has shown promise in treating primary central nervous system lymphoma (PCNSL). The enzyme O(6)-methylguanine-DNA methyltransferase (MGMT) repairs alkylating damage, such as that induced by temozolomide. We hypothesized that MGMT immunohistochemistry would predict resistance to temozolomide in PCNSL. A retrospective study of newly-diagnosed and recurrent PCNSL patients treated at our institution was conducted to study the predictive value of MGMT immunohistochemistry for response to temozolomide. 20 patients who were treated with temozolomide as a single agent were identified during the study time period. 6/20 patients demonstrated a response, corresponding to an objective response rate of 30 % (95 % CI 8-52). Five patients with low MGMT level (<30 %) showed a response to temozolomide. Only one of 10 patients (10 %) with high MGMT level (≥30 %) exhibited a response to temozolomide. Small sample numbers precluded formal statistical comparisons. Two patients with complete response remain alive without progressive disease 6.7 and 7.2 years after temozolomide initiation. Immunohistochemistry can be performed on small biopsies to selectively assess MGMT status in tumor versus surrounding inflammation. MGMT analysis by immunohistochemistry may predict response to temozolomide in PCNSL and should be prospectively investigated.

Triazene Compounds in the Treatment of Acute Myeloid Leukemia: A Short Review and a Case Report

Acute myeloid leukemia (AML) is a highly lethal disease, especially in old patients. Chemoresistance and the absence of host immune responses against autochthonous malignancy play a major role in the poor prognosis of AML. The triazene compounds Dacarbazine and Temozolomide are monofunctional alkylators that donate methyl groups to many sites in DNA, including the O(6)-position of guanine producing O(6)-methylguanine (O(6)-MeG). If not repaired, O(6)-MeG frequently mispairs with thymine during DNA duplication. O(6)-MeG:T mismatches can be recognized by the mismatch repair (MMR) system which activates a cascade of molecular events leading to cell cycle arrest and cell death. If MMR is defective, cells continue to divide and GC → AT transition mutations occur. In preclinical models, such mutations can lead to the appearance of abnormal proteins containing non-self peptides ("chemical xenogenization" CX) that can be recognized by host cell-mediated immunity. Repair of O(6)-MeG is achieved by the DNA repair protein, O(6)-methylguanine-DNA methyltransferase (MGMT), which removes the methyl adduct in an autoinactivating stoichiometric reaction. High MGMT levels attenuate the pharmacodynamic effects of triazenes. In the last few years, triazenes, alone or with MGMT inhibitors, have been tested in AML. In view of their potential activity as CX inducers, triazenes could offer the additional advantage of host anti-leukemia immune responses. The present paper describes several studies of leukemia treatment with triazenes and a case of acute refractory leukemia with massive skin infiltration by malignant cells. Treatment with Temozolomide and Lomeguatrib, a potent MGMT inhibitor, produced a huge, although transient, blastolysis and complete disappearance of all skin lesions.

Methylguanine DNA methyl transferase (MGMT) expression to predict response to temozolomide (TMZ) in patients with digestive neuroendocrine tumors (NETs).

4133 Background: TMZ is an orally given drug used in patients (pts) with NETs, with encouraging results in pancreatic NETs in a recent study (Strosberg et al, 2011). Higher treatment efficacy seems to be correlated to MGMT tumor deficiency (Kulke et al, 2010). Aim-To assess MGMT expression in digestive NETs and to correlate with the efficacy of TMZ-based therapies. Methods: All pts with well differentiated, progressive, non resectable NETs treated with TMZ-based chemotherapy were included. Treatment efficacy was defined according to RECIST. Pts with progression or only stable disease were considered as “non-responders”. Nuclear expression of MGMT was assessed by immunohistochemistry on primary tumors or metastases and graded according the product of the intensity of staining (0 to 3) and the rate of positive cells (%), leading to a score comprised between 0 and 300. A score ≥ 80 was defined as “high” staining. Results: 22 pts (age 59 years (36-81)) with pancreatic (14 pts), small bowel (5 pts) or other (3 pts) NETs, grade 1 (5 pts) or 2 (17 pts) (WHO 2010 classification) were included. They received TMZ alone (19 pts) or combined with capecitabine (3 pts) as first line (3 pts) or 2+ line (19 pts).After a median of 6 cycles (3-16), objective response, stable disease and progression rates were seen in 32 % (7 pts, all with pancreatic NETs), 41% (9 pts, 5 pancreatic) and 27% (6 pts, 4 small bowel), respectively. Median (range) MGMT score was 10 (0-300). A “High” MGMT score was seen in 36% of pts (small bowel 4, pancreas 3 pts) ; it was correlated with primary tumor location (more frequent in small bowel NETs, p=0.02) and predictive of the absence of response (p=0.02). A “Low “MGMT score tended to be associated with objective response (p=0.06) whereas none of the pts with “high” score had tumor response. Response rate in pts with “low” MGMT score was 50%. Conclusions: MGMT deficiency is more frequent in pancreatic than small bowel NETs. Patients with pancreatic NET and low MGMT score are good candidates for TMZ, whereas those with high score should be treated with other drug in first intention. In patients with small bowel NET with most often high MGMT score, tumor stabilization using TMZ seems to be rare.

O6-Methylguanine-DNA Methyltransferase (MGMT) Promoter Methylation and Low MGMT-Encoded Protein Expression as Prognostic Markers in Glioblastoma Patients Treated with Biodegradable Carmustine Wafer Implants after Initial Surgery Followed by Radiotherapy with Concomitant and Adjuvant Temozolomide (P06.004)

Objective: To study the prognostic impact of O 6 -methylguanine-DNA methyltransferase (MGMT) on newly diagnosed glioblastoma patients being treated with carmustine-releasing wafers (Gliadel®) along with temozolomide (TMZ). Background MGMT promoter methylation status was proposed as a prognostic biomarker for glioblastoma patients. However, the prognostic impact of MGMT on newly diagnosed glioblastoma patients treated using surgical resection and Gliadel® implants followed by Stupp9s protocol is still unknown. Design/Methods: MGMT promoter methylation status and protein expression were analyzed in formalin-fixed, paraffin-embedded tumor specimens obtained from 111 French patients with newly diagnosed glioblastoma. Patients received Gliadel® followed by radiotherapy plus concomitant and adjuvant TMZ chemotherapy, while being enrolled in a French multicenter prospective study. Results: For the whole cohort, median overall survival (OS) and progression-free survival (PFS) were 17.5 and 10.3 months, respectively. Patients with tumors harboring MGMT methylation displayed a significantly longer OS compared to those with wild-type MGMT (21.7 vs . 15.1 months, p=0.025). Similarly, patients with low MGMT protein expression (≤15%) exhibited significantly improved OS compared to those with high MGMT expression (27.0 vs. 15.1 months, p=0.021). The extent of resection was the strongest clinical outcome predictor. In multivariate Cox models adjusted for gender, performance status, and extent of surgery, both MGMT methylation and protein expression were found to be independent prognosticators, which was internally validated using bootstrap resampling techniques. However, no significant correlation was found between protein expression and MGMT methylation status, thus suggesting that the two assays probably assess different biological features. Conclusions: MGMT promoter methylation status and low MGMT expression were both found to be positive prognosticators in newly diagnosed glioblastoma patients treated using surgical resection and Gliadel® implants followed by Stupp9s protocol. Disclosure: Dr. Guillamo has nothing to disclose. Dr. Levallet has nothing to disclose. Dr. Dugue has nothing to disclose. Dr. Vital has nothing to disclose. Dr. Diebold has nothing to disclose. Dr. Menei has nothing to disclose. Dr. Colin has nothing to disclose. Dr. Peruzzy has nothing to disclose. Dr. Emery has nothing to disclose. Dr. Bernaudin has nothing to disclose. Dr. Chapon has nothing to disclose. Dr. Lechapt-Zalcman has nothing to disclose.

O6‐methylguanine‐DNA methyltransferase (MGMT) promoter methylation and low MGMT‐encoded protein expression as prognostic markers in glioblastoma patients treated with biodegradable carmustine wafer implants after initial surgery followed by radiotherapy with concomitant and adjuvant temozolomide

O(6) -methylguanine-DNA methyltransferase (MGMT) promoter methylation status was proposed as a prognostic biomarker for patients with glioblastoma. However, the prognostic impact of MGMT in patients with newly diagnosed glioblastoma who receive carmustine-releasing wafers (Gliadel) along with temozolomide (TMZ) is still unknown. MGMT promoter methylation status and protein expression were analyzed in formalin-fixed, paraffin-embedded tumor specimens obtained from 111 French patients with newly diagnosed glioblastoma. Patients received the Gliadel wafers followed by radiotherapy plus concomitant and adjuvant TMZ chemotherapy while they were enrolled in a French multicenter prospective study. For the whole cohort, the median overall survival (OS) was 17.5 months, and the progression-free survival was 10.3 months. Patients with tumors that harbored MGMT methylation had a significantly longer OS compared with patients who had wild-type MGMT (21.7 months vs 15.1 months; P = .025). Similarly, patients who had low MGMT protein expression (≤15%) had a significantly improved OS compared with patients who had high MGMT expression (27.0 months vs 15.1 months; P = .021). The extent of resection was the strongest clinical predictor of outcome. In multivariate Cox models that were adjusted for sex, performance status, and extent of surgery, both MGMT methylation and protein expression were identified as independent prognosticators, and the finding was validated internally using a bootstrap resampling technique. Discrepancies were identified between protein expression and MGMT methylation status, thus suggesting that the 2 assays probably assess different biologic features. MGMT promoter methylation status and low MGMT expression both were identified as positive prognosticators in patients with newly diagnosed glioblastoma who underwent surgical resection and received Gliadel wafer implants followed by adjuvant radiotherapy and concomitant oral TMZ chemotherapy (the Stupp protocol).

Low O6‐methylguanine‐DNA methyltransferase (MGMT) expression and response to temozolomide in aggressive pituitary tumours

Recent case reports detail the successful use of temozolomide in the management of aggressive pituitary tumours. O(6)-methylguanine-DNA methyltransferase (MGMT) is a DNA repair protein that counteracts the effect of temozolomide. To study MGMT expression in pituitary tumours and consider whether MGMT expression is associated with response to temozolomide therapy in aggressive pituitary tumours. We report two patients with aggressive pituitary tumours treated with temozolomide, one who responded to temozolomide and the other who did not. MGMT expression was assessed in a further 88 archived pituitary tumour samples. MGMT expression was assessed by immunohistochemistry. MGMT promoter methylation was studied by methylation-specific polymerase chain reaction (MSP), sequencing of MGMT was performed and loss of heterozygosity (LOH) analysis undertaken. Low MGMT expression and MGMT promoter methylation were found in the pituitary tumour of the patient who responded to temozolomide. Conversely, high MGMT expression was seen in the patient demonstrating a poor response to temozolomide. Eleven out of 88 archived tumour samples (13%) had low MGMT expression. Prolactinomas were more likely to have low MGMT expression compared with other pituitary tumour subtypes (P < 0.001). There was no significant difference in MGMT expression between invasive and noninvasive tumours, or between recurrent and nonrecurrent tumours. A significant inverse correlation was found between MGMT expression and promoter methylation (P = 0.012). MGMT expression as assessed by immunohistochemistry may predict response to temozolomide therapy in patients with aggressive pituitary tumours. MGMT promoter methylation is likely to explain low MGMT expression in some, but not all, pituitary tumours.

Targeting Methylguanine-DNA Methyltransferase in the Treatment of Neuroblastoma

The combination of temozolomide and irinotecan has preclinical schedule-dependent synergy against neuroblastoma but is not curative for relapsed high-risk patients. We hypothesized that the DNA repair protein methylguanine-DNA methyltransferase (MGMT) is an important resistance factor, and that inactivation of MGMT would sensitize neuroblastoma cells to these agents. MGMT protein expression was assessed in 74 primary neuroblastoma tumors. Growth inhibition assays were done to determine the IC(50) and the extent of synergy observed with various concentrations of temozolomide, irinotecan, and the MGMT-inactivating agent O(6)-benzylguanine, using cultured syngeneic neuroblastoma cells with either low or high levels of MGMT expression. We then assessed efficacy in a mouse xenograft model of metastatic neuroblastoma. MGMT was expressed by all 74 tumors evaluated. Pretreatment of neuroblastoma cells with O(6)-benzylguanine reduced the IC(50) of temozolomide by 10-fold regardless of level of MGMT expression, and pretreatment with BG followed by temozolomide + irinotecan further reduced the IC(50) in cells with high MGMT expression another 10-fold, to well below clinically achievable concentrations. The combination index was 0.27 to 0.30 for all three drugs in both cell lines, indicating strong synergy. Survival at 100 days for mice with metastatic neuroblastoma was 56% with three-drug treatment, compared with untreated controls (0%, P < 0.001) or temozolomide + irinotecan (10%, P = 0.081). MGMT is widely expressed in primary neuroblastoma tumors, and is a relevant therapeutic target. Both in vitro and in vivo studies suggest inactivation of MGMT with O(6)-benzylguanine may increase the activity of temozolomide and irinotecan against neuroblastoma.

O6-Methylguanine-DNA methyl transferase gene expression and prognosis in breast carcinoma

O(6)-methylguanine-DNA methyl transferase (MGMT) in human carcinomas has been associated with tumor resistance to alkylating agents. The aims of this study were: i) to correlate tumor MGMT expression and patient and tumor characteristics in malignant breast carcinomas treated with induction chemotherapy including cyclophosphamide (CPM) and ii) to study the predictive and prognostic values of tumor MGMT gene expression. We used RT-PCR to measure the levels of tumor MGMT expression in 107 patients with breast carcinomas prior to neoadjuvant chemotherapy. Sixty patients (56%) received anthracyclines and CPM and 47 (44%) received only anthracyclines. Low levels of MGMT expression correlated with Scarff-Bloom-Richardson grade III (p<0.005), elevated S-phase (p<0.05), negative estrogen receptors (p<0.05), metastatic status (p<0.05) and occurrence of death (p=0.01). MGMT expression was not predictive of treatment response. Unexpectedly, survival was longer when tumor MGMT expression was high (p<0.005). The 4-year survival rate was 76% for high level MGMT patients and only 55% for others. This difference is also significant using the COX model (p<0.05). In breast cancer, tumor MGMT expression was not predictive of response to CPM. A low MGMT expression was significantly related to poor survival.

Acquired resistance of melanoma cells to the antineoplastic agent fotemustine is caused by reactivation of the DNA repair gene mgmt

Acquired resistance to antineoplastic agents is a frequent obstacle in tumor therapy. Malignant melanoma cells are particularly well known for their unresponsiveness to chemotherapy; only about 30% of tumors exhibit a transient clinical response to treatment. In our study, we investigated the molecular mechanism of acquired resistance of melanoma cells (MeWo) to the chloroethylating drug fotemustine. Determination of O(6)-methylguanine-DNA methyltransferase (MGMT) activity showed that MeWo cells that acquired resistance to fotemustine upon repeated treatment with the drug display high MGMT activity, whereas the parental cell line had no detectable MGMT. The resistant cell lines exhibit cross-resistance to other O(6)-alkylating agents, such as N-methyl-N'-nitro-N-nitrosoguanidine. Acquired resistance to fotemustine was alleviated by treatment with the MGMT inhibitor O(6)-benzylguanine demonstrating that reactivation of MGMT is the main underlying cause of acquired alkylating drug resistance. As compared with control cells, both MGMT mRNA and MGMT protein were expressed at a high level in fotemustine resistant cells. Southern blot analysis proved that the MGMT gene was not amplified. There was also only an insignificant difference in the CpG methylation pattern of the MGMT promoter whereas a clear hypermethylation in the body of the gene was observed in fotemustine resistant cells. The conclusion that hypermethylation is responsible for reactivation of the MGMT gene gained support by the finding that MGMT activity significantly declined and cells reverted (partially) to the parental sensitive phenotype upon treatment with 5-azacytidine. This is the first report of acquired resistance to a chloroethylating antineoplastic drug of melanoma cells due to gene hypermethylation.

Correlation of tumor O6 methylguanine-DNA methyltransferase levels with survival of malignant astrocytoma patients treated with bis-chloroethylnitrosourea: a Southwest Oncology Group study.

Prior studies show that increased levels of the DNA repair protein O6 methylguanine-DNA methyltransferase (MGMT), also referred to as O6-alkylguanine-DNA alkyltransferase (AGT) correlate with the resistance of glioma cell lines to nitrosoureas. The observed nitrosourea sensitivity of MGMT-deficient lines (methyl excision repair negative [MER-]) and those repair-proficient lines pretreated with MGMT-specific inhibitors (eg, O6 benzylguanine) has raised the possibility that tumor MGMT levels may be an important predictor of survival in patients with gliomas. We correlated the MGMT level in malignant astrocytoma tissues, obtained from patients treated with radiotherapy and bis-chloroethylnitrosourea (BCNU) on a prior prospective trial (Southwest Oncology Group [SWOG] 8737), with overall and failure-free survival. Of 64 assessable patients with malignant astrocytoma (63% glioblastoma, 37% anaplastic astrocytoma), 64% had high (> 60,000 molecules/nucleus) MGMT levels. The overall median survival for patients with high versus low MGMT levels was 8 and 29 months, respectively (P=.0002), and median failure-free survival 3 and 6 months, respectively (P=.008). Subset analysis by histology (high v low MGMT levels) for anaplastic astrocytoma was 14 versus 62 months (n=24) and for glioblastoma was 7 versus 12 months (n=40). The overall hazards ratio (risk for death) for high versus low MGMT levels was 3.41; in young patients, the hazards ratio was higher (age 18 to 40 years, 4.19; age 41 to 60 years, 3.08) but became equal by MGMT level at age older than 60 years (1.11). Multivariate analysis showed that MGMT was independent of other known prognostic factors (age, performance status, histology). The MGMT level in tumor tissue specimens may be a predictive marker of survival in patients with malignant astrocytoma that is independent of other previously described prognostic variables.

Transfection and expression of human O6-methylguanine-DNA methyltransferase (MGMT) cDNA in Chinese hamster cells: the role of MGMT in protection against the genotoxic effects of alkylating agents

O6-Methylguanine-DNA methyltransferase (MGMT) is responsible for removal of O6-alkylguanine from DNA induced by alkylating mutagens/carcinogens. To analyze the involvement of O6-alkylguanine in the generation and MGMT in avoidance of various genotoxic effects of alkylating agents, we transfected Chinese hamster ovary (CHO) cells that lack MGMT activity with human MGMT cDNA cloned into a mammalian expression vector (pSV2MGMT). A high proportion (60-80%) of transfectants selected for a cotransfected neo gene survived treatment with high doses of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and N-hydroxyethyl-N-chloroethylnitrosourea (HeCNU). Parallel transfections with an expression vector containing the bacterial ada gene (pSV2ada) showed the human MGMT to be more effective than the ada expression vector in mediating alkylation resistance. Various clonal CHO cell lines have been established stably transfected with the human MGMT cDNA. The transfectants expressed human MGMT at levels ranging from 8600 to 210,000 molecules per cell. The high MGMT expressors became strongly resistant to the killing effects of MNNG, HeCNU, N-methyl-N-nitrosourea (MNU) and, to a significant lesser degree, methyl methanesulfonate (MMS) and ethyl methanesulfonate (EMS). No killing resistance was observed to N-ethyl-N-nitrosourea (ENU), though the MGMT and ada transfectants showed reduction in mutation frequency induced by this agent. Protection from mutation induction by MGMT (and ada) expression was also demonstrated for MNNG. The transfectants were also protected from the sister chromatid exchange (SCE) inducing and, to a lesser degree, clastogenic effect of MNNG and MNU, and slightly to EMS and MMS. Again no protection was observed towards ENU. Correlations between MGMT activity and resistance to a given end point suggest that, for MNNG, O6-methylguanine is the preponderant toxic, mutagenic and SCE inducing lesion. About 90% of MNNG (and MNU) induced SCEs and nearly all of the MNNG-induced gene mutations seem to be due to this adduct. For alkylation-induced chromosomal aberrations, however, and for cell killing and SCEs induced by MMS, EMS and ENU, other lesions than O6-alkylguanine appear to be of major importance. The data strongly support the view that O6-methylguanine is a genotoxic lesion and MGMT a function decisively involved in avoidance of genotoxic effects in cells exposed to MNNG and related compounds. They indicate also that it is important to take into account the property and mode of action of any given alkylating agent in assessing the protective role of MGMT against alkylation-induced genotoxicity.