Botensilimab/Balstilimab Elicits Major Pathologic Responses in Colon Cancer

Findings from the phase 2 NEST trial (NCT05571293), published in Clinical Cancer Research, showed that neoadjuvant botensilimab plus balstilimab yielded encouraging pathologic responses and safety in patients with non-metastatic mismatch repair–proficient (pMMR) and mismatch repair–deficient (dMMR) colorectal cancer.¹ The dual checkpoint inhibitor combination did not delay planned surgery in any of the 24 enrolled patients, meeting the trial’s primary feasibility objective.

What were the pathologic response and survival findings?

Among 22 evaluable pMMR tumors pooled across cohorts A and B, the pathologic response rate, defined as achieving at least 50% tumor regression, was 59% (n = 13/22 patients; 95% CI, 36%-79%). The major pathologic response (MPR) rate was 41% (n = 9/22), and the pathologic complete response (pCR) rate was 32% (n = 7/22; 95% CI, 14%-55%).

All 4 dMMR tumors achieved MPR (MPR rate, 100%; 95% CI, 40%-100%), including 2 pCRs, with the remaining 2 tumors each showing 98% and 99% treatment effect.

At a median follow-up of 32.2 months for cohort A and 23.5 months for cohorts B and C, no patients had disease recurrence, translating to a disease-free survival rate of 100% at data cutoff. Seven of 8 evaluable patients (88%) cleared circulating tumor DNA (ctDNA) before surgery, and ctDNA remained undetectable in all patients following resection. Tumors with KRAS G12D or an activating BRAF mutation showed no pathologic response, while only 3 of 19 tumors (14%) with other KRAS/BRAF alterations or wild-type status had no response (P = .048). According to investigators, this association requires validation in larger cohorts.

Previous NEST data were presented at the 2025 ASCO Gastrointestinal Cancers Symposium, where the complete MPR rate improved with extended time to surgery, from 14% with 2 balstilimab doses to 40% with 4 doses.² The mature results confirm and extend that trend with translational data.

How did treatment remodel the tumor immune microenvironment?

Multiplex immunofluorescence of matched baseline and resected tumors, performed using the Lunaphore COMET platform across more than 286,000 identified immune cells, showed a 13-fold increase in CD8-positive T-cell density from baseline to resection. A numerical decrease in FOXP3-positive regulatory T cells (Tregs) was also observed. Immune cell density in resection specimens was significantly higher in responding vs nonresponding tumors (3.1 cells/µm² vs 0.52 cells/µm²; P = .0048). This was driven largely by CD8-positive T cells and B cells, which together comprised 34% of the immune infiltrate in responders compared with 15% and 6%, respectively, in nonresponders.

Responding tumors showed organized, colocalized immune infiltrates, including B cell– and CD8-positive T-cell–enriched cellular neighborhoods, while nonresponding tumors retained spatially isolated infiltrates dominated by epithelial, stromal, and macrophage-rich niches. These compositional and spatial patterns were broadly similar between pMMR and dMMR responders, suggesting a shared mechanism of immune activation regardless of mismatch repair status.

What did histologic review of resected tumors show?

Post-treatment tumor beds in patients who achieved MPR were characterized by dense mixed inflammatory infiltrates of lymphocytes, plasma cells, and histiocytes, along with mucosal granulation tissue, acellular mucin pools, and submucosal fibrosis. A prominent Crohn’s-like lymphoid reaction, indicative of a robust antitumor immune response, was present in 73% of pMMR tumors with a pathologic response greater than 50% compared with 13% of nonresponding tumors (P = .028). In tumors with a partial response, residual viable glands frequently showed disrupted crypts and luminal microabscesses consistent with ongoing immune-mediated destruction.

What is the design of the NEST trial?

NEST was a single-center, open-label, non-randomized phase 2 study conducted at Weill Cornell Medicine/New York-Presbyterian Hospital in patients 18 years and older with resectable, non-metastatic colorectal cancer. Those in cohort A (NEST-1; n = 10, enrolled March to September 2023) received botensilimab at 75 mg on day 1 plus balstilimab at 240 mg on days 1 and 15, followed by surgery. Patients in cohorts B and C, the amended NEST-2 portion that opened in February 2024, received the same botensilimab dose plus 4 balstilimab doses on days 1, 15, 29, and 43.

The median time to surgery was 29 days (range, 21-37) for cohort A and 55 days (range, 21-104) for cohorts B and C. No grade 4 toxicities occurred. In cohort A, 1 patient had grade 2 diarrhea/colitis that resolved with infliximab (Remicade), and 5 patients experienced a common symptom pattern of fever, chills, and fatigue.

In the NEST-2 cohorts, 5 patients developed immune-mediated colitis or myositis requiring high-dose steroids, and 1 required infliximab, with all events resolving without delaying surgery.

References

1. Shah MA, Hissong E, Jafari MD, et al. Neoadjuvant botensilimab/balstilimab for localized mismatch repair proficient and deficient colon cancer: results of the NEST phase 2 clinical trial. Clin Cancer Res. Published online 2026. doi:10.1158/1078-0432.CCR-26-0998
2. Hissong E, Jafari D, Khan S, et al. Neoadjuvant botensilimab (BOT) plus balstilimab (BAL) in resectable mismatch repair proficient (pMMR) and deficient (dMMR) colorectal cancer (CRC). J Clin Oncol. 2025;43(suppl 4):207. doi:10.1200/JCO.2025.43.4_suppl.207