Clinical Utilization Pattern of Liquid Biopsies (LB) to Detect Actionable Driver Mutations, Guide Treatment Decisions and Monitor Disease Burden During Treatment of 33 Metastatic Colorectal Cancer (mCRC) Patients (pts) at a Fox Chase Cancer Center GI Oncology Subspecialty Clinic

Background: Liquid biopsy (LB) captures dynamic genomic alterations (alts) across metastatic colorectal cancer (mCRC) therapy and may complement tissue biopsy (TB). We sought to describe the utility of LB and better understand mCRC biology during therapy.Methods: Thirty-three patients (pts) with mCRC underwent LB. We used permutation-based t-tests to assess associations between alts, and clinical variables and used Kendall's tau to measure correlations.Results: Of 33 pts, 15 were women; 22 had colon, and the rest rectal cancer. Pts received a median of two lines of therapy before LB. Nineteen pts had limited testing on TB (RAS/RAF/TP53/APC), 11 extended NGS, and 3 no TB. Maxpct and alts correlated with CEA (p < 0.001, respectively). In 3/5 pts with serial LB, CEA correlated with maxpct trend, and CT tumor burden. In 6 pts, mutant RAS was seen in LB and not TB; 5/6 had received anti-EGFR therapy prior to LB, suggesting RAS alts developed post-therapy. In two pts RAS-mutated by TB, no RAS alts were detected on LB; these pts had low disease burden on CT at time of LB that also did not reveal APC or TP53 alts. In six patients who were KRAS wt based on TB, post anti-EGFR LB revealed subclonal KRAS mutations, likely a treatment effect. The median number of alts was higher post anti-EGFR LB (n = 12) vs. anti-EGFR naïve LB (n = 22) (9.5 vs. 5.5, p = 0.059) but not statistically significant. More alts were also noted in post anti-EGFR therapy LB vs. KRAS wt anti-EGFR-naïve LB (n = 6) (9.5 vs. 5) among patients with KRAS wild-type tumors, although the difference was not significant (p = 0.182).Conclusions: LB across mCRC therapy detects driver mutations, monitors disease burden, and identifies sub-clonal alts that reflect drug resistance, tumor evolution, and heterogeneity. Interpretation of LB results is impacted by clinical context

Successful Inhibition of Tumor Development by Specific Class-3 Semaphorins Is Associated with Expression of Appropriate Semaphorin Receptors by Tumor Cells

The class-3 semaphorins (sema3s) include seven family members. Six of them bind to neuropilin-1 (np1) or neuropilin-2 (np2) receptors or to both, while the seventh, sema3E, binds to the plexin-D1 receptor. Sema3B and sema3F were previously characterized as tumor suppressors and as inhibitors of tumor angiogenesis. To determine if additional class-3 semaphorins such as sema3A, sema3D, sema3E and sema3G possess anti-angiogenic and anti-tumorigenic properties, we expressed the recombinant full length semaphorins in four different tumorigenic cell lines expressing different combinations of class-3 semaphorin receptors. We show for the first time that sema3A, sema3D, sema3E and sema3G can function as potent anti-tumorigenic agents. All the semaphorins we examined were also able to reduce the concentration of tumor associated blood vessels although the potencies of the anti-angiogenic effects varied depending on the tumor cell type. Surprisingly, there was little correlation between the ability to inhibit tumor angiogenesis and their anti-tumorigenic activity. None of the semaphorins inhibited the adhesion of the tumor cells to plastic or fibronectin nor did they modulate the proliferation of tumor cells cultured in cell culture dishes. However, various semaphorins were able to inhibit the formation of soft agar colonies from tumor cells expressing appropriate semaphorin receptors, although in this case too the inhibitory effect was not always correlated with the anti-tumorigenic effect. In contrast, the anti-tumorigenic effect of each of the semaphorins correlated very well with tumor cell expression of specific signal transducing receptors for particular semaphorins. This correlation was not broken even in cases in which the tumor cells expressed significant concentrations of endogenous semaphorins. Our results suggest that combinations of different class-3 semaphorins may be more effective than single semaphorins in cases in which tumor cells express more than one type of semaphorin receptors

Lymphocyte Exhaustion in AML Patients and Impacts of HMA/Venetoclax or Intensive Chemotherapy on Their Biology

Acute myeloid leukemia (AML) is an aggressive malignancy that requires rapid treatment with chemotherapies to reduce tumor burden. However, these chemotherapies can compromise lymphocyte function, thereby hindering normal anti-tumor immune responses and likely limiting the efficacy of subsequent immunotherapy. To better understand these negative impacts, we assessed the immunological effects of standard-of-care AML therapies on lymphocyte phenotype and function over time. When compared to healthy donors, untreated AML patients showed evidence of lymphocyte activation and exhaustion and had more prevalent CD57+NKG2C+ adaptive NK cells, which was independent of human cytomegalovirus (HCMV) status. HMA/venetoclax treatment resulted in a greater fraction of T cells with effector memory phenotype, inhibited IFN-γ secretion by CD8+ T cells, upregulated perforin expression in NK cells, downregulated PD-1 and 2B4 expression on CD4+ T cells, and stimulated Treg proliferation and CTLA-4 expression. Additionally, we showed increased expression of perforin and CD39 and enhanced IFN-γ production by T cells from pre-treatment blood samples of venetoclax-resistant AML patients. Our results provide insight into the lymphocyte status in previously untreated AML patients and the effects of standard-of-care treatments on their biology and functions. We also found novel pre-treatment characteristics of T cells that could potentially predict venetoclax resistance