Mutant PIK3CA is a targetable driver alteration in histiocytic neoplasms

Abstract

Langerhans cell histiocytosis (LCH) is an inflammatory myeloid neoplasm characterized by the accumulation of clonal mononuclear phagocyte system cells expressing CD1a and CD207. In the past decade, molecular profiling of LCH as well as other histiocytic neoplasms demonstrated that these diseases are driven by MAPK activating alterations, with somatic BRAFV600E mutations in >50% of patients with LCH, and clinical inhibition of MAPK signaling has demonstrated remarkable clinical efficacy. At the same time, activating alterations in kinase-encoding genes, such as PIK3CA, ALK, RET, and CSF1R, which can activate mitogenic pathways independent from the MAPK pathway, have been reported in a subset of histiocytic neoplasms with anecdotal evidence of successful targeted treatment of histiocytoses harboring driver alterations in RET, ALK, and CSF1R. However, evidence supporting the biological consequences of expression of PIK3CA mutations in hematopoietic cells has been lacking, and whether targeted inhibition of PI3K is clinically efficacious in histiocytic neoplasms is unknown. Here, we provide evidence that activating mutations in PIK3CA can drive histiocytic neoplasms in vivo using a conditional knockin mouse expressing mutant PIK3CAH1047R in monocyte/dendritic cell progenitors. In parallel, we demonstrate successful treatment of PIK3CA-mutated, multisystemic LCH using alpelisib, an inhibitor of the alpha catalytic subunit of PI3K. Alpelisib demonstrated a tolerable safety profile at a dose of 750 mg per week and clinical and metabolic complete remission in a patient with PIK3CA-mutated LCH. These data demonstrate PIK3CA as a targetable noncanonical driver of LCH and underscore the importance of mutational analysis-based personalized treatment in histiocytic neoplasms.

Graphical abstract

Figure 1.

Recurrent activating PIK3CA mutations drives histiocytic neoplasms in vivo. (A) Diagram of PIK3CA alterations across cancer showing hotspots in the helical and kinase domains, the locations of activating mutations. (B) Protein diagram summarizing recurrent PIK3CA mutations across histiocytic neoplasm subtypes. (C) Kaplan-Meier curve of primary CD11c-cre Pik3ca^H1047R and littermate Pik3ca control mice; n = 15-20 mice; ∗∗∗∗P < .0001. Log-rank (Mantel-Cox) test. (D) Representative histological images of bone marrow showing trilineage hematopoiesis in control mice with no evidence of histiocytosis compared with the bone marrow of CD11c-cre Pik3ca^H1047R that shows involvement of the bone marrow by increased CD68⁺, large, foamy histiocytes, and multinucleated giant cells reminiscent of a human histiocytic neoplasm (H&E stain and murine CD68 immunohistochemistry; 600× magnification). (E) Bar plots of percentage of B- (B220⁺) and T- (CD3⁺) lymphocytes among CD45⁺ cells in blood (left), bone marrow (middle), and spleen (right) in control vs mutant mice by flow cytometry. (F) As in panel E but for myelomonocytic (CD11b⁺) and dendritic (CD11c⁺ MHCII⁺) cells in blood. (G) Box-and-whisker plots of hemoglobin in mice. Mean ± SD. n = 3 mice; ∗P<.01; 1-way ANOVA. ANOVA, analysis of variance; H&E, hematoxylin and eosin; SD, standard deviation.

Figure 2.

Pathological evaluation of a patient with PIK3CA^M1043V-mutated Langerhans cell histiocytosis. (A) Lung wedge biopsy showing infiltration of lung parenchyma by Langerhans cell histiocytosis (red oval) (H&E; 100× magnification). (B) CD1a immunohistochemistry confirming infiltration of lung parenchyma by Langerhans cell histiocytosis (red oval) (CD1a immunohistochemistry; 100× magnification). (C) Phospho-ERK1/2 (pERK1/2) immunohistochemistry demonstrating a lack of expression of pERK1/2 by the Langerhans cell histiocytosis of the patient (pERK1/2 immunohistochemistry; 100× magnification). (D) PIK3CA c.3127A>G; p.M1043V missense mutation detected by targeted next-generation sequencing of the Langerhans cell histiocytosis involving the lung biopsy of the patient that was absent in paired normal fingernail DNA.

Figure 3.

Response dynamics of pulmonary LCH involvement sites upon PI3Kα inhibition. (A) PET, CT, and fusion axial data layers at the level of the lungs were derived from PET/CT studies performed before and during systemic treatment with alpelisib. (B) Quantification of lesion SUVmax values as apparent in panel A. Abnormal FDG uptake in multiple lung nodules on the diagnostic PET image (white arrows). CT, computed tomography; FDG, fluorodeoxyglucose; PET, positron emitted tomography; SUVmax, maximum standard unit value.

Figure 4.

Response dynamics of skeletal and nodal LCH involvement sites upon PI3Kα inhibition. (A-C) panels depicting imaging data relevant to the lytic lesion in T11. (A) PET, CT, and fusion axial data layers at the level of vertebra T11 derived from PET/CT studies performed before and during systemic treatment with alpelisib. (B-C) Quantification of lesion (B) SUVmax values and (C) HU as apparent in panel A increase in 9 m; HU: P < .001. (D-G) Panels depicting imaging data relevant to the right-sided cervical lymphadenopathy. (D) PET, CT, and fusion axial data layers at the level of vertebra C1 derived from PET/CT studies performed before and during systemic treatment with alpelisib. (E-G) Quantification of lesion (E) SUVmax values, (F) HU and dimensions (G) as apparent in panel D. Abnormal FDG uptake foci in the diagnostic PET and fusion images in A and D are indicated by white arrows. CT, computed tomography; FDG, fluorodeoxyglucose; HU, Hounsfield units; PET, positron emitted tomography; SUVmax, maximum standard unit value.

Figure 5.

Expression levels of PTEN and its regulating miRNAs measured by qRT-PCR before and after treatment with alpelisib. (A) PTEN expression levels in PBMCs. PTEN expression was normalized to HPRT1 endogenous controls. (B) miR-21-5p, miR-26a-5p, and miR-181c-5p expression in PBMCs and (C) plasma samples. MiRNA expression was normalized to U6 endogenous control and spike-in control cel-miR-39, respectively. The histograms represent the relative expression from at least 3 experiments. ∗P < .05; ∗∗P < .001; ∗∗∗P < .0001. HPRT1, hypoxanthine phosphoribosyltransferase 1.