Molecular Oncology Programme

Experimental Oncology Group

Group Leader:  Mariano Barbacid
Research highlights
Identification of novel therapeutic targets for the treatment of K-Ras driven lung adenocarcinoma

The recent discovery that lung tumours display significant levels of clonal heterogeneity (Govindan, Science, 2014) implies that effective therapies must target early oncogenic events/alterations present in all tumour cells and not only in clonal variants that appear during tumour development. To provide potential solutions to this key issue we decided to search for novel therapeutic targets present in the earliest stages of lung tumour development, expecting that such targets will be present in the entire tumour population including the putative cancer initiating/stem cells. Among the most highly expressed druggable genes we identified Ddr1, a locus that encodes a tyrosine protein kinase receptor. As reported early this year (Ambrogio et al., Nat Med, 2016), genetic and pharmacological inhibition of Ddr1 prevented progression of K-Ras driven p53 wild type, but not p53 mutant tumours. Yet concomitant inhibition of Ddr1 and Notch, a downstream mediator of Ddr1 activity, led to a significant anti-tumour effect even in aggressive K-RasG12V; p53 mutant adenocarcinomas. More importantly, this treatment induced regression of K-RAS; p53 mutant patient-derived lung orthoxenografts (PDX) with a therapeutic efficacy superior to standard chemotherapy. Identification of additional targets present in these early K-Ras mutant driven lung cells should expand the therapeutic opportunities to treat K-RAS mutant tumours in the clinic, thus by-passing the challenges derived from the development of intra-tumour heterogeneity.

Lack of selective advantage for lung cells expressing K-RAS and EGFRAC oncogenes

Activating mutations in KRAS and EGFR, the 2 most frequent oncogenic drivers in human lung adenocarcinoma, occur in a mutually exclusive manner suggesting functional redundancy and implying lack of positive selection. By means of a mouse model engineered to induce expression of mutant EGFRL858R in advanced tumours driven by a resident KrasG12V oncogene, we show that, instead, their co-expression is detrimental for the progression of lung denocarcinoma. In vivo expression of EGFRL858R in KrasG12V-driven tumours triggers an immediate response with hallmarks of replicative stress resulting in apoptosis. Yet, a fraction of tumour cells survive, but enter a transient cytostatic state incompatible with tumour development that is fully reversible upon discontinuation of EGFRL858R expression. Ultimately, continuous co-expression of both mutants results in the attenuation of the overall oncogenic signalling to levels compatible with cell proliferation and tumour growth. In sum, our results indicate that the mutual exclusivity of KRAS and EGFR activating mutations occurs as a combination of cellular toxicity and signal adjustment that results in the lack of selective advantage for those cells expressing both oncogenes.

Whereas the wild type H-Ras and K-Ras proteins are bioequivalent, their oncogenic isoforms H-RasG12V and K-RasG12V induce different tumour spectra

We have provided genetic evidence demonstrating that the H-Ras and K-Ras proteins are fully bioequivalent in mice. Previous studies have shown that replacement of the K-Ras alleles by H-Ras coding sequences resulted in viable mice (Potenza et al., EMBO Rep, 2005). Yet, these mice displayed cardiovascular defects. Now, we have shown that these defects were due to the presence of the 4 H-Ras expressing alleles in these mice. Ablation of the 2 endogenous H-Ras alleles, hence generating mice that only express the H-Ras protein from the 2 targeted K-Ras alleles, is absolutely normal.

These results appear to be at variance with the well-established observation that H-Ras and K-Ras oncogenes are involved in different human tumour types. To determine whether the oncogenic versions of the H-Ras and K-Ras proteins are also bioequivalent, we knocked-in H-RasG12V oncogene sequences into the K-Ras locus. Germline expression of H-RasG12V or K-RasG12V from the K-Ras locus resulted in equal embryonic lethality. However, their expression in adult mice led to different tumour phenotypes. Whereas H-RasG12V elicited papillomas and haematopoietic tumours, K-RasG12V induced lung tumours and gastric lesions. The reason why H-RasG12V expression failed to cause lung tumours is due to the induction of a senescence-like state due to excessive MAP kinase signalling. Likewise, H-RasG12V but not K-RasG12V induced oncogene-induced senescence in mouse embryonic fibroblasts (MEFs). Label-free quantitative analysis revealed that minor differences in H-RasG12V expression levels led to drastically different biological outputs, suggesting that subtle differences in MAP kinase signalling influence the differential tumour spectra induced by RAS oncoproteins.