Acquired Resistance to KRAS (G12C) Inhibition in Cancer

BACKGROUND: Clinical trials of the KRAS inhibitors adagrasib and sotorasib have shown promising activity in cancers harboring KRAS glycine-to-cysteine amino acid substitutions at codon 12 (KRAS(G12C)). The mechanisms of acquired resistance to these therapies are currently unknown. METHODS: Among patients with KRAS(G12C) -mutant cancers treated with adagrasib monotherapy, we performed genomic and histologic analyses that compared pretreatment samples with those obtained after the development of resistance. Cell-based experiments were conducted to study mutations that confer resistance to KRAS(G12C) inhibitors. RESULTS: A total of 38 patients were included in this study: 27 with non-small-cell lung cancer, 10 with colorectal cancer, and 1 with appendiceal cancer. Putative mechanisms of resistance to adagrasib were detected in 17 patients (45% of the cohort), of whom 7 (18% of the cohort) had multiple coincident mechanisms. Acquired KRAS alterations included G12D/R/V/W, G13D, Q61H, R68S, H95D/Q/R, Y96C, and high-level amplification of the KRAS(G12C) allele. Acquired bypass mechanisms of resistance included MET amplification; activating mutations in NRAS, BRAF, MAP2K1, and RET; oncogenic fusions involving ALK, RET, BRAF, RAF1, and FGFR3; and loss-of-function mutations in NF1 and PTEN. In two of nine patients with lung adenocarcinoma for whom paired tissue-biopsy samples were available, histologic transformation to squamous-cell carcinoma was observed without identification of any other resistance mechanisms. Using an in vitro deep mutational scanning screen, we systematically defined the landscape of KRAS mutations that confer resistance to KRAS(G12C) inhibitors. CONCLUSIONS: Diverse genomic and histologic mechanisms impart resistance to covalent KRAS(G12C) inhibitors, and new therapeutic strategies are required to delay and overcome this drug resistance in patients with cancer. (Funded by Mirati Therapeutics and others; ClinicalTrials.gov number, NCT03785249.)

DW-MRI as a Biomarker to Compare Therapeutic Outcomes in Radiotherapy Regimens Incorporating Temozolomide or Gemcitabine in Glioblastoma

The effectiveness of the radiosensitizer gemcitabine (GEM) was evaluated in a mouse glioma along with the imaging biomarker diffusion-weighted magnetic resonance imaging (DW-MRI) for early detection of treatment effects. A genetically engineered murine GBM model [Ink4a-Arf−/− PtenloxP/loxP/Ntv-a RCAS/PDGF(+)/Cre(+)] was treated with gemcitabine (GEM), temozolomide (TMZ) +/− ionizing radiation (IR). Therapeutic efficacy was quantified by contrast-enhanced MRI and DW-MRI for growth rate and tumor cellularity, respectively. Mice treated with GEM, TMZ and radiation showed a significant reduction in growth rates as early as three days post-treatment initiation. Both combination treatments (GEM/IR and TMZ/IR) resulted in improved survival over single therapies. Tumor diffusion values increased prior to detectable changes in tumor volume growth rates following administration of therapies. Concomitant GEM/IR and TMZ/IR was active and well tolerated in this GBM model and similarly prolonged median survival of tumor bearing mice. DW-MRI provided early changes to radiosensitization treatment warranting evaluation of this imaging biomarker in clinical trials

Extracellular matrix protection factor 1 (ECPF-1): A novel osteoarthritis therapeutic demonstrates chondroprotective properties in a rat model of osteoarthritis; a quantitative micro computed tomography study of the tibia and femur

Introduction: Osteoarthritis (OA) is one of the most prevalent joint diseases, affecting millions of people and yet there is currently no cure. Finding a therapeutic that can cure OA would be beneficial for millions of people and those prone to a future degenerative disease. Objective: Current therapeutics for OA are focused on relieving symptoms for late stages of the disease. Extracellular Matrix Protection Factor-1 (ECPF-1), is a novel, highly specific Matrix metalloprotease-13 (MMP-13) inhibitor that blocks extracellular matrix degradation. Methods: A chemically-induced rat model of knee OA was used to study the effects of ECPF-1 in early stage OA progression. Micro computed tomography (µCT) images of the rat knee joints were quantified by measuring bone volume, spacing and total joint volume and trabecular spacing, thickness and number. Results: Data collected focuses on the treatment effects of ECPF-1 in the acute stage of OA after a loading phase (4 weekly injections of ECPF-1) and an 8-week protection-extension phase. For the femur, all ECPF-1 treated µCT measurements trended toward the values in the normal age-matched rat at both 4 and 8 weeks. For the tibia, all ECPF-1 treated µCT measurements trended toward the values in the normal age-matched rat at 8 weeks. The trabecular number values in both the femur and the tibia were most prominent in the 8-week samples of animals treated with ECPF-1 and exhibited the most progress toward normal, age-matched rat readings. Conclusions: This model showed that using an MMP inhibitor such as ECPF-1 could help in treating acute, post-traumatic OA. This is a potential new treatment for OA that indicates the ability to slow the disease progression

MR, histological images and western blots are presented from representative animals in <i>Study 1</i> treatment groups.

<p>(A) MRI data consists of anatomical contrast-enhancing T1-weighted images and ADC maps. Histological stains provide information on tumor cellularity (H&E) and apoptosis (cleaved Caspase-3). All data were acquired at day 7 post-treatment initiation. (B) Representative western blot for the detection of cleaved Caspase 3 in tumor tissue from all treatment groups. B-Actin was used as a loading control to ensure proper loading of the protein samples. The tumor tissue from all groups was acquired at day 2 post-treatment initiation.</p

MR and histological images and western blots are presented from representative animals in <i>Study 2</i> treatment groups.

<p>(A) MRI data consists of anatomical contrast-enhancing T1-weighted images and ADC maps. Histological stains provide information on tumor cellularity (H&E) and apoptosis (caspase-3). All data were acquired at day 7 post-treatment initiation. (B) Tumor tissue from animals left untreated or treated with GEM, IR and GEM+IR at day two post-treatment initiation was assessed for cleaved Caspase 3. Western blot of representative animal tissue is shown and proper loading of protein samples was ensured by probing for Gapdh.</p

Plots of the percent change in tumor volume and normalized ADC (ADC) for each of the treatment groups in <i>Study 2</i>.

<p>Presented are treatment groups (A) Control, (B) irradiation (IR), (C) gemcitabine (GEM) and (D) combination gemcitabine and irradiation (GEM+IR). Data is presented over the first week of the study as the mean ± SEM.</p

Treatment schedule and Kaplan-Meier survival plots are presented for each therapy in <i>Study 2</i>.

<p>(A) Treatment schedule schematic for <i>Study 2</i>. Animals were randomized into four groups: control, IR, GEM and GEM+IR. Animals of the control group received vehicle 2 days a week for 2 weeks. Animals in the IR group received 1 Gy for 5 days as week with a two day break between treatment blocks for 2 weeks. The GEM group received 10 mg/kg GEM in saline i.p., and GEM+IR received GEM i.p. followed by 1 Gy with a 3 hour lag time between treatments. Control vehicle and GEM administration occurred every third day for a total of four doses. Arrows indicate the day of treatment. (B) Treatment groups are Controls, irradiation (IR), gemcitabine (GEM) and combination gemcitabine and irradiation (GEM+IR).</p

Adenosine 2A receptor blockade as an immunotherapy for treatment-refractory renal cell cancer

Adenosine mediates immunosuppression within the tumor microenvironment through triggering adenosine 2A receptors (A2AR) on immune cells. To determine whether this pathway could be targeted as an immunotherapy, we performed a phase I clinical trial with a small-molecule A2AR antagonist. We find that this molecule can safely block adenosine signaling in vivo. In a cohort of 68 patients with renal cell cancer (RCC), we also observe clinical responses alone and in combination with an anti-PD-L1 antibody, including subjects who had progressed on PD-1/PD-L1 inhibitors. Durable clinical benefit is associated with increased recruitment of CD8+ T cells into the tumor. Treatment can also broaden the circulating T-cell repertoire. Clinical responses are associated with an adenosine-regulated gene-expression signature in pretreatment tumor biopsies. A2AR signaling, therefore, represents a targetable immune checkpoint distinct from PD-1/PD-L1 that restricts antitumor immunity. SIGNIFICANCE: This first-in-human study of an A2AR antagonist for cancer treatment establishes the safety and feasibility of targeting this pathway by demonstrating antitumor activity with single-agent and anti-PD-L1 combination therapy in patients with refractory RCC. Responding patients possess an adenosine-regulated gene-expression signature in pretreatment tumor biopsies.See related commentary by Sitkovsky, p. 16.This article is highlighted in the In This Issue feature, p. 1